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| ARCHIVE | Friday, August 30th, 2024 - Disease Models |
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What's hot today
September 2024 July 2024 June 2024 May 2024 April 2024 March 2024 February 2024 January 2024 December 2023 November 2023 October 2023 September 2023 August 2023 July 2023 June 2023 May 2023 April 2023 March 2023 February 2023 January 2023 December 2022 December 2021 December 2020 December 2019 December 2018 | Deshpande, P., Chen, C. Y., Chimata, A. V., Li, J. C., Sarkar, A., Yeates, C., Chen, C. H., Kango-Singh, M., Singh, A. (2024). miR-277 targets the proapoptotic gene-hid to ameliorate Abeta42-mediated neurodegeneration in Alzheimer's model. Cell Death Dis, 15(1):71 PubMed ID: 38238337
Summary: Alzheimer's disease (AD), an age-related progressive neurodegenerative disorder, exhibits reduced cognitive function with no cure to date. One of the reasons for AD is the accumulation of Amyloid-beta 42 (Aβ42) plaque(s) that trigger aberrant gene expression and signaling, which results in neuronal cell death by an unknown mechanism(s). Misexpression of human Aβ42 in the developing retina of Drosophila exhibits AD-like neuropathology. Small non-coding RNAs, microRNAs (miRNAs), post-transcriptionally regulate the expression of their target genes and thereby regulate different signaling pathways. In a forward genetic screen, miR-277 (human ortholog is hsa-miR-3660) was identified as a genetic modifier of Aβ42-mediated neurodegeneration. Loss-of-function of miR-277 enhances the Aβ42-mediated neurodegeneration. Whereas gain-of-function of miR-277 in the GMR > Aβ42 background downregulates cell death to maintain the number of neurons and thereby restores the retinal axonal targeting defects indicating the functional rescue. In addition, gain-of-function of miR-277 rescues the eclosion- and climbing assays defects observed in GMR > Aβ42 background. Thus, gain-of-function of miR-277 rescues both structurally as well as functionally the Aβ42-mediated neurodegeneration. Furthermore, head involution defective (hid), an evolutionarily conserved proapoptotic gene, was detected as one of the targets of miR-277 and validated these results using luciferase- and qPCR -assays. In the GMR > Aβ42 background, the gain-of-function of miR-277 results in the reduction of hid transcript levels to one-third of its levels as compared to GMR > Aβ42 background alone. This study provides a novel molecular mechanism where miR-277 targets and downregulates proapoptotic gene, hid, transcript levels to rescue Aβ42-mediated neurodegeneration by blocking cell death. These studies shed light on molecular mechanism(s) that mediate cell death response following Aβ42 accumulation seen in neurodegenerative disorders in humans and provide new therapeutic targets for neurodegeneration. | Le, M. U. T., Park, J. H., Son, J. G., Shon, H. K., Joh, S., Chung, C. G., Cho, J. H., Pirkl, A., Lee, S. B., Lee, T. G. (2024). Monitoring lipid alterations in Drosophila heads in an amyotrophic lateral sclerosis model with time-of-flight secondary ion mass spectrometry. The Analyst, PubMed ID: 38167886
Summary: Lipid alterations in the brain are well-documented in disease and aging, but understanding of their pathogenic implications remains incomplete. Recent technological advances in assessing lipid profiles have enabled intricate examination of the spatiotemporal variations in lipid compositions within the complex brain characterized by diverse cell types and intricate neural networks. This study coupled time-of-flight secondary ion mass spectrometry (ToF-SIMS) to an amyotrophic lateral sclerosis (ALS) Drosophila model to elucidate changes in the lipid landscape and investigate their potential role in the disease process, serving as a methodological and analytical complement to a prior approach that utilized matrix-assisted laser desorption/ionization mass spectrometry. The expansion of G(4)C(2) repeats in the C9orf72 gene is the most prevalent genetic factor in ALS. The findings indicate that expressing these repeats in fly brains elevates the levels of fatty acids, diacylglycerols, and ceramides during the early stages (day 5) of disease progression, preceding motor dysfunction. Using RNAi-based genetic screening targeting lipid regulators, it was found that reducing fatty acid transport protein 1 (FATP1) and Acyl-CoA-binding protein (ACBP) alleviates the retinal degeneration caused by G(4)C(2) repeat expression and also markedly restores the G(4)C(2)-dependent alterations in lipid profiles. Significantly, the expression of FATP1 and ACBP is upregulated in G(4)C(2)-expressing flies, suggesting their contribution to lipid dysregulation. Collectively, this novel use of ToF-SIMS with the ALS Drosophila model, alongside methodological and analytical improvements, successfully identifies crucial lipids and related genetic factors in ALS pathogenesis. |
| Hunt, L. C., Nyamkondiwa, K., Stephan, A., Jiao, J., Kavdia, K., Pagala, V., Peng, J., Demontis, F. (2023). The ubiquitin-conjugating enzyme UBE2D/eff maintains a youthful proteome and ensures protein quality control during aging. bioRxiv, PubMed ID: 38168249
Summary: Ubiquitin-conjugating enzymes (E2s) are key for regulating protein function and turnover via ubiquitination but it remains undetermined which E2s maintain proteostasis during aging. This study found that E2s have diverse roles in handling a model aggregation-prone protein (huntingtin-polyQ) in the Drosophila retina: while some E2s mediate aggregate assembly, UBE2D/effete (eff) and other E2s are required for huntingtin-polyQ degradation. UBE2D/eff is key for proteostasis also in skeletal muscle: Eff protein levels decline with aging, and muscle-specific eff knockdown causes an accelerated buildup in insoluble poly-ubiquitinated proteins (which progressively accumulate with aging) and shortens lifespan. Transgenic expression of human UBE2D2, homologous to eff, partially rescues the lifespan and proteostasis deficits caused by muscle-specific eff(RNAi) by re-establishing the physiological levels of eff(RNAi)-regulated proteins. Interestingly, UBE2D/eff knockdown in young age reproduces many of the proteomic changes that normally occur in old muscles, suggesting that the decrease in UBE2D/Eff protein levels that occurs with aging contributes to reshaping the composition of the muscle proteome. Altogether, these findings indicate that UBE2D/Eff is a key E2 ubiquitin-conjugating enzyme for maintaining a youthful proteome and for ensuring protein quality control during aging. | Brown, J. C., McMichael, B. D., Vandadi, V., Mukherjee, A., Salzler, H. R., Matera, A. G. (2023). Lysine-36 of Drosophila histone H3.3 supports adult longevity. bioRxiv, PubMed ID: 38196611
Summary: Aging is a multifactorial process that disturbs homeostasis, increases disease susceptibility, and ultimately results in death. Although the definitive set of molecular mechanisms responsible for aging remain to be discovered, epigenetic change over time is proving to be a promising piece of the puzzle. Several posttranslational histone modifications (PTMs) have been linked to the maintenance of longevity. This study focused on lysine-36 of the replication-independent histone protein, H3.3 (H3.3K36). To interrogate the role of this residue in Drosophila developmental gene regulation, a lysine to arginine mutant was generated that blocks the activity of its cognate modifying enzymes. An H3.3B(K36R) mutation was found to cause a significant reduction in adult lifespan, accompanied by dysregulation of the genomic and transcriptomic architecture. Transgenic co-expression of wild-type H3.3B completely rescues the longevity defect. Because H3.3 is known to accumulate in non-dividing tissues, transcriptome profiling of young vs aged adult fly heads was carried out. The data show that loss of H3.3K36 results in age-dependent misexpression of NF-κB and other innate immune target genes, as well as defects in silencing of heterochromatin. It is proposed H3.3K36 maintains the postmitotic epigenomic landscape, supporting longevity by regulating both pericentric and telomeric retrotransposons and by suppressing aberrant immune signaling. |
| Narwal, S., Singh, A., Tare, M. (2023). Analysis of alpha-syn and parkin interaction in mediating neuronal death in Drosophila model of Parkinson's disease. Frontiers in cellular neuroscience, 17:1295805 PubMed ID: 38239290
Summary: One of the hallmarks of Parkinson's Disease (PD) is aggregation of incorrectly folded α-synuclein (SNCA) protein resulting in selective death of dopaminergic neurons. Another form of PD is characterized by the loss-of-function of an E3-ubiquitin ligase, parkin. Mutations in SNCA and parkin result in impaired mitochondrial morphology, causing loss of dopaminergic neurons. Despite extensive research on the individual effects of SNCA and parkin, their interactions in dopaminergic neurons remain understudied. This study employed a Drosophila model to study the effect of collective overexpression of SNCA along with the downregulation of parkin in the dopaminergic neurons of the posterior brain. Overexpression of SNCA along with downregulation of parkin causes a reduction in the number of dopaminergic neuronal clusters in the posterior region of the adult brain, which is manifested as progressive locomotor dysfunction. Overexpression of SNCA and downregulation of parkin collectively results in altered mitochondrial morphology in a cluster-specific manner, only in a subset of dopaminergic neurons of the brain. Further, it was found that SNCA overexpression causes transcriptional downregulation of parkin. However, this downregulation is not further enhanced upon collective SNCA overexpression and parkin downregulation. This suggests that the interactions of SNCA and parkin may not be additive. This study thus provides insights into a potential link between α-synuclein and parkin interactions. These interactions result in altered mitochondrial morphology in a cluster-specific manner for dopaminergic neurons over a time, thus unraveling the molecular interactions involved in the etiology of Parkinson's Disease. | Tener, S. J., Lin, Z., Park, S. J., Oraedu, K., Ulgherait, M., Van Beek, E., Martínez-Muñiz, A., Pantalia, M., Gatto, J. A., Volpi, J., Stavropoulos, N., Ja, W. W., Canman, J. C., Shirasu-Hiza, M. (2024). Neuronal knockdown of Cullin3 as a Drosophila model of autism spectrum disorder. Sci Rep, 14(1):1541 PubMed ID: 38233464
Summary: Mutations in Cullin-3 (Cul3), a conserved gene encoding a ubiquitin ligase, are strongly associated with autism spectrum disorder (ASD). This study characterize ASD-related pathologies caused by neuron-specific Cul3 knockdown in Drosophila. Neuronal Cul3 knockdown causes short sleep, paralleling sleep disturbances in ASD. Because sleep defects and ASD are linked to metabolic dysregulation, the starvation response of neuronal Cul3 knockdown flies was tested; they starved faster and had lower triacylglyceride levels than controls, suggesting defects in metabolic homeostasis. ASD is also characterized by increased biomarkers of oxidative stress; neuronal Cul3 knockdown increased sensitivity to hyperoxia, an exogenous oxidative stress. Additional hallmarks of ASD are deficits in social interactions and learning. Using a courtship suppression assay that measures social interactions and memory of prior courtship, it was found that neuronal Cul3 knockdown reduced courtship and learning compared to controls. Finally, it was found that neuronal Cul3 depletion alters the anatomy of the mushroom body, a brain region required for memory and sleep. Taken together, the ASD-related phenotypes of neuronal Cul3 knockdown flies establish these flies as a genetic model to study molecular and cellular mechanisms underlying ASD pathology, including metabolic and oxidative stress dysregulation and neurodevelopment. |
Thursday, August 29th - Larval and Adult Neural Structure, Development and Function |
| Xu, C., Ramos, T. B., Marshall, O. J., Doe, C. Q. (2024). Notch signaling and Bsh homeodomain activity are integrated to diversify Drosophila lamina neuron types. Elife, 12 PubMed ID: 38193901
Summary: Notch signaling is an evolutionarily conserved pathway for specifying binary neuronal fates, yet how it specifies different fates in different contexts remains elusive. In an accompanying paper, using the Drosophila lamina neuron types (L1-L5) as a model, it was shown that the primary homeodomain transcription factor (HDTF) Bsh activates secondary HDTFs Ap (L4) and Pdm3 (L5) and specifies L4/L5 neuronal fates. This study tested the hypothesis that Notch signaling enables Bsh to differentially specify L4 and L5 fates. Asymmetric Notch signaling is shown between newborn L4 and L5 neurons, but they are not siblings; rather, Notch signaling in L4 is due to Delta expression in adjacent L1 neurons. While Notch signaling and Bsh expression are mutually independent, Notch is necessary and sufficient for Bsh to specify L4 fate over L5. The Notch(ON) L4, compared to Notch(OFF) L5, has a distinct open chromatin landscape which allows Bsh to bind distinct genomic loci, leading to L4-specific identity gene transcription. A novel model is proposed in which Notch signaling is integrated with the primary HDTF activity to diversify neuron types by directly or indirectly generating a distinct open chromatin landscape that constrains the pool of genes that a primary HDTF can activate. | Perlegos, A. E., Durkin, J., Belfer, S. J., Rodriguez, A., Shcherbakova, O., Park, K., Luong, J., Bonini, N. M., Kayser, M. S. (2024). TDP-43 impairs sleep in Drosophila through Ataxin-2-dependent metabolic disturbance. Sci Adv, 10(2):eadj4457 PubMed ID: 38198547
Summary: Neurodegenerative diseases such as amyotrophic lateral sclerosis and frontotemporal dementia are associated with substantial sleep disruption, which may accelerate cognitive decline and brain degeneration. This study defines a role for trans-activation response element (TAR) DNA binding protein 43 (TDP-43), a protein associated with human neurodegenerative disease, in regulating sleep using Drosophila. Expression of TDP-43 severely disrupts sleep, and the sleep deficit is rescued by Atx2 knockdown. Brain RNA sequencing revealed that Atx2 RNA interference regulates transcripts enriched for small-molecule metabolic signaling in TDP-43 brains. Focusing on these Atx2-regulated genes, suppressors of the TDP-43 sleep phenotype were identified enriched for metabolism pathways. Knockdown of Atx2 or treatment with rapamycin attenuated the sleep phenotype and mitigated the disruption of small-molecule glycogen metabolism caused by TDP-43. Fhese findings provide a connection between toxicity of TDP-43 and sleep disturbances and highlight key aspects of metabolism that interplay with TDP-43 toxicity upon Atx2 rescue. |
| Iyer, A. R., Scholz-Carlson, E., Bell, E., Biondi, G., Richhariya, S., Fernandez, M. P. (2024). The Circadian Neuropeptide PDF has Sexually Dimorphic Effects on Activity Rhythms. bioRxiv, PubMed ID: 38352594
Summary: The circadian system regulates the timing of multiple molecular, physiological, metabolic, and behavioral phenomena. In Drosophila as in other species, most of the research on how the timekeeping system in the brain controls timing of behavioral outputs has been conducted in males, or sex was not included as a biological variable. The main circadian pacemaker neurons in Drosophila release the neuropeptide Pigment Dispersing Factor (PDF), which functions as a key synchronizing factor in the network with complex effects on other clock neurons. Lack of Pdf or its receptor, PdfR, results in most flies displaying arrhythmicity in activity-rest cycles under constant conditions. However, results of this study show that female circadian rhythms are less affected by mutations in both Pdf and PdfR. Crispr-Cas9 mutagenesis of Pdf specifically in the ventral lateral neurons (LN (v) s) also has a greater effect on male rhythms. The influence of the M-cells over the circadian network was tested; speeding up the molecular clock specifically in the M-cells leads to sexually dimorphic phenotypes, with a more pronounced effect on male rhythmic behavior. These results suggest that the female circadian system is more resilient to manipulations of the PDF pathway and that circadian timekeeping is more distributed across the clock neuron network in females. | Brezovec, B. E., Berger, A. B., Hao, Y. A., Chen, F., Druckmann, S., Clandinin, T. R. (2024). Mapping the neural dynamics of locomotion across the Drosophila brain. Curr Biol, PubMed ID: 38242122
Summary: Locomotion engages widely distributed networks of neurons. However, understanding of the spatial architecture and temporal dynamics of the networks that underpin walking remains incomplete. This study used volumetric two-photon imaging to map neural activity associated with walking across the entire brain of Drosophila. Spatially clustered neural signals selectively associated with changes in either forward or angular velocity are defined, demonstrating that neurons with similar behavioral selectivity are clustered. These signals reveal distinct topographic maps in diverse brain regions involved in navigation, memory, sensory processing, and motor control, as well as regions not previously linked to locomotion. Temporal trajectories of neural activity were identified that sweep across these maps, including signals that anticipate future movement, representing the sequential engagement of clusters with different behavioral specificities. Finally, these maps were registered to a connectome and neural networks were identified that are proposed to underlie the observed signals, setting a foundation for subsequent circuit dissection. Overall, this work suggests a spatiotemporal framework for the emergence and execution of complex walking maneuvers and links this brain-wide neural activity to single neurons and local circuits. |
| Dopp, J., Ortega, A., Davie, K., Poovathingal, S., Baz, E. S., Liu, S. (2024). Single-cell transcriptomics reveals that glial cells integrate homeostatic and circadian processes to drive sleep-wake cycles. Nat Neurosci, 27(2):359-372 PubMed ID: 38263460
Summary: The sleep-wake cycle is determined by circadian and sleep homeostatic processes. However, the molecular impact of these processes and their interaction in different brain cell populations are unknown. To fill this gap, this study profiled the single-cell transcriptome of adult Drosophila brains across the sleep-wake cycle and four circadian times. Cell type-specific transcriptomic changes were detected, with glia displaying the largest variation. Glia are also among the few cell types whose gene expression correlates with both sleep homeostat and circadian clock. The sleep-wake cycle and sleep drive level affect the expression of clock gene regulators in glia, and disrupting clock genes specifically in glia impairs homeostatic sleep rebound after sleep deprivation. These findings provide a comprehensive view of the effects of sleep homeostatic and circadian processes on distinct cell types in an entire animal brain and reveal glia as an interaction site of these two processes to determine sleep-wake dynamics. | Lim-Kian-Siang, G., Izawa-Ishiguro, A. R., Rao, Y. (2024). Neurexin-1-dependent circuit activity is required for the maintenance of photoreceptor subtype identity in Drosophila. Mol Brain, 17(1):2 PubMed ID: 38167109
Summary: In the human and Drosophila color vision system, each photoreceptor neuron (cone cell in humans and R7/R8 photoreceptor cell in Drosophila) makes a stochastic decision to express a single photopigment of the same family with the exclusion of the others. While recent studies have begun to reveal the mechanisms that specify the generation of cone subtypes during development in mammals, nothing is known about how the mosaic of mutually exclusive cone subtypes is maintained in the mammalian retina. In Drosophila, recent work has led to the identification of several intrinsic factors that maintain the identity of R8 photoreceptor subtypes in adults. Whether and how extrinsic mechanisms are involved, however, remain unknown. This study presents evidence that supports that the Drosophila transsynaptic adhesion molecule Neurexin 1 (Dnrx-1) is required non-cell autonomously in R8p subtypes for the maintenance of R8y subtype identity. Silencing the activity of R8p subtypes caused a phenotype identical to that in dnrx-1 mutants. These results support a novel role for Nrx-1-dependent circuit activity in mediating the communication between R8 photoreceptor subtypes for maintaining the subtype identity in the retina. |
Wednesday, August 28th - RNA and RNAi |
| Jang, W., Kim, C. (2023). A Pumilio Activity Sensor Reveals Bag-of-Marbles Inhibition of Pum Activity in the Drosophila Ovary. Development & reproduction, 27(1):39-46 PubMed ID: 38075440
Summary: Pumilio (Pum) is an RNA-binding protein and translational repressor important to diverse biological processes. In the Drosophila ovary, Pum is expressed in female germline stem cells (GSCs), wherein it acts as an intrinsic stem cell maintenance factor via repressing target mRNAs that are as yet mostly unknown. Pum recognizes the Pum binding sequence (PBS) in the mRNA 3'UTR through its C-terminus Puf domain. Translational repression is mediated through its N-terminal domain, but the molecular mechanism remains largely unknown. Previous work showed that Bag-of-marbles, a critical differentiation-promoting factor of female GSCs, physically interacts with the N-terminus of Pum. It was further shown that this interaction is critical to Bam inhibition of Pum repressive action in cultured cells, but the physiological relevance was not addressed. This study designed an in vivo GFP translational reporter bearing the PBS in its 3'UTR and showed that GFP expression is reduced in cells wherein Pum is known to be active. Furthermore, it was demonstrated in pum mutant ovary that this GFP repression requires Pum, and also that the sensor faithfully monitors Pum activity. Finally, it was shown that forced expression of Bam inhibits Pum-mediated repression, validating that Bam inhibits Pum activity in vivo. | Peng, Q., Wang, Y., Xiao, Y., Chang, H., Luo, S., Wang, D., Rong, Y. S. (2023). Drosophila Amus and Bin3 methylases functionally replace mammalian MePCE for capping and the stabilization of U6 and 7SK snRNAs. Sci Adv, 9(50):eadj9359 PubMed ID: 38100593
Summary: U6 and 7SK snRNAs have a 5' cap, believed to be essential for their stability and maintained by mammalian MePCE or Drosophila Bin3 enzymes. Although both proteins are required for 7SK stability, loss of neither destabilizes U6, casting doubts on the function of capping U6. This study shows that the Drosophila Amus protein, homologous to both proteins, is essential for U6 but not 7SK stability. The loss of U6 is rescued by the expression of an Amus-MePCE hybrid protein harboring the methyltransferase domain from MePCE, highlighting the conserved function of the two proteins as the U6 capping enzyme. These investigations in human cells establish a dependence of both U6 and 7SK stability on MePCE, resolving a long-standing uncertainty. While uncovering a division of labor of Bin3/MePCE/Amus proteins, a "Bin3-Box" domain was found to be present only in enzymes associated with 7SK regulation. Targeted mutagenesis confirms its importance for Bin3 function, revealing a possible conserved element in 7SK but not U6 biology. |
| Barrington, C. L., Galindo, G., Koch, A. L., Horton, E. R., Morrison, E. J., Tisa, S., Stasevich, T. J., Rissland, O. S. (2023). Synonymous codon usage regulates translation initiation. JCell Rep, 42(12):113413 PubMed ID: 38096059
Summary: Nonoptimal synonymous codons repress gene expression, but the underlying mechanisms are poorly understood. Previous work has shown that nonoptimal codons slow translation elongation speeds and thereby trigger messenger RNA (mRNA) degradation. Nevertheless, transcript levels are often insufficient to explain protein levels, suggesting additional mechanisms by which codon usage regulates gene expression. Using reporters in human and Drosophila cells, this study fond that transcript levels account for less than half of the variation in protein abundance due to codon usage. This discrepancy is explained by translational differences whereby nonoptimal codons repress translation initiation. Nonoptimal transcripts are also less bound by the translation initiation factors eIF4E and eIF4G1, providing a mechanistic explanation for their reduced initiation rates. Importantly, translational repression can occur without mRNA decay and deadenylation, and it does not depend on the known nonoptimality sensor, CNOT3. These results reveal a potent mechanism of regulation by codon usage where nonoptimal codons repress further rounds of translation. | Ge, S., Wang, X., Wang, Y., Dong, M., Li, D., Niu, K., Wang, T., Liu, R., Zhao, C., Liu, N., Zhong, M. (2024). Hidden features of NAD-RNA epitranscriptome in Drosophila life cycle. iScience, 27(1):108618 PubMed ID: 38197055
Summary: Nicotinamide adenine dinucleotide (NAD), a nucleotide-containing metabolite, can be incorporated into the RNA 5'-terminus to result in NAD-capped RNA (NAD-RNA). Since NAD has been heightened as one of the most essential metabolites in cells, its linkage to RNA represents a critical but poorly studied modification at the epitranscriptomic level. A highly sensitive method, DO-seq, was designed to capture NAD-RNAs. Using Drosophila, thousands of previously unexplored NAD-RNAs were discoved, along with their dynamics in the fly life cycle, from embryo to adult. Evidence is shown that chromosomal clustering might be the structural basis by which co-expression can couple with NAD capping on physically and functionally linked genes. Furthermore, it is noted that NAD capping of cuticle genes inversely correlates with their gene expression. Combined, it is proposed NAD-RNA epitranscriptome as a hidden layer of regulation that underlies biological processes. DO-seq empowers the identification of NAD-capped RNAs, facilitating functional investigation into this modification. |
| Zhang, B., Duan, H., Kavaler, J., Wei, L., Eberl, D. F., Lai, E. C. (2023). A nonneural miRNA cluster mediates hearing via repression of two neural targets. Genes Dev, 37(21-24):1041-1051 PubMed ID: 38110249
Summary: We show here that mir-279/996 are absolutely essential for development and function of Johnston's organ (JO), the primary proprioceptive and auditory organ in Drosophila. Their deletion results in highly aberrant cell fate determination, including loss of scolopale cells and ectopic neurons, and mutants are electrophysiologically deaf. In vivo activity sensors and mosaic analyses indicate that these seed-related miRNAs function autonomously to suppress neural fate in nonneuronal cells. Finally, genetic interactions pinpoint two neural targets (elav and insensible) that underlie miRNA mutant JO phenotypes. This work uncovers how critical post-transcriptional regulation of specific miRNA targets governs cell specification and function of the auditory system. | Babosha, V. A., Georgiev, P. G., Maksimenko, O. G. (2024). Study of the Role of Long Noncoding roX RNA in Maintaining of the Dosage Compensation Complex in Drosophila melanogaster. Doklady Biochemistry and biophysics, PubMed ID: 38189885
Summary: The proteins MSL1, MSL2, MSL3, MLE, and MOF and noncoding RNAs roX1 and roX2 form the Drosophila dosage compensation complex (DCC), which specifically binds to the X chromosome of males. It is known that noncoding RNA roX are primary component of the DCC in the process of assembly and spreading of the complex among the X chromosome of males. However, the role of this RNA in maintaining the structure of the already assembled complex remains unclear. This work has shown that the full-assembled dosage compensation complex dissociates rather weakly when treated with RNases: the MLE helicase is effectively released from the complex, and the remaining protein components (MSL1, MSL2, and MSL3) undergo partial disassembly and continue to be part of subcomplexes. The results confirm the importance of the noncoding roX2 RNA not only in the processes of initiation of DCC assembly but also at the stage of maintaining the structure of the already assembled complex. |
Tuesday, August 27th - Embryonic Development |
| Baumgartner, S. (2024). Revisiting bicoid function: complete inactivation reveals an additional fundamental role in Drosophila egg geometry specification. Hereditas, 161(1):1 PubMed ID: 38167241
Summary: The bicoid (bcd) gene in Drosophila has served as a paradigm for a morphogen in textbooks for decades. Discovered in 1986 as a mutation affecting anterior development in the embryo, its expression pattern as a protein gradient later confirmed the prediction from transplantation experiments. These experiments suggested that the protein fulfills the criteria of a true morphogen, with the existence of a homeodomain crucial for activation of genes along the anterior-posterior axis, based on the concentration of the morphogen. The bcd gene undergoes alternative splicing, resulting in, among other isoforms, a small and often neglected isoform with low abundance, which lacks the homeodomain, termed small bicoid (smbcd). Most importantly, all known classical strong bcd alleles used in the past to determine bcd function apparently do not affect the function of this isoform. To overcome the uncertainty regarding which isoform regulates what, The bcd locus entirely using CRISPR technology. bcd(CRISPR) eggs exhibited a short and round appearance. The phenotype could be ascribed to smbcd because all bcd alleles affecting the function of the major transcript, termed large bicoid (lgbcd) showed normally sized eggs. Several patterning genes for the embryo showed expression in the oocyte, and their expression patterns were altered in bcd(CRISPR) oocytes. In bcd(CRISPR) embryos, all downstream segmentation genes showed altered expression patterns, consistent with the expression patterns in "classical" alleles; however, due to the altered egg geometry resulting in fewer blastoderm nuclei, additional constraints came into play, further affecting their expression patterns. This study unveils a novel and fundamental role of bcd in shaping the egg's geometry. This discovery demands a comprehensive revision of understanding of this important patterning gene and prompts a reevaluation of past experiments conducted under the assumption that bcd mutants were bcd(null)-mutants. | Dermady, A., DeFazio, D., Hensley, E., Ruiz, D., Chavez, A., Iannone, S., Dermady, N., Grandel, L., Hill, A. S. (2024). Neuronal excitability modulates developmental time of Drosophila melanogaster. Dev Biol, PubMed ID: 38224932
Summary: Developmental time is a fundamental life history trait that affects the reproductive success of animals. Developmental time is known to be regulated by many genes and environmental conditions, yet mechanistic understandings of how various cellular processes influence the developmental timing of an organism are lacking. The nervous system is known to control key processes that affect developmental time, including the release of hormones that signal transitions between developmental stages. This study shows that the excitability of neurons plays a crucial role in modulating developmental time. Genetic manipulation of neuronal excitability in Drosophila melanogaster alters developmental time, which is faster in animals with increased neuronal excitability. Selectively modulating the excitability of peptidergic neurons is sufficient to alter developmental time, suggesting the intriguing hypothesis that the impact of neuronal excitability on DT may be at least partially mediated by peptidergic regulation of hormone release. This effect of neuronal excitability on developmental time is seen during embryogenesis and later developmental stages. Observed phenotypic plasticity in the effect of genetically increasing neuronal excitability at different temperatures, a condition also known to modulate excitability, suggests there is an optimal level of neuronal excitability, in terms of shortening DT. Together, these data highlight a novel connection between neuronal excitability and developmental time, with broad implications related to organismal physiology and evolution. |
| Andreas, E., Cummins, B., Gedeon, T. (2024). Quantifying robustness of the gap gene network. Journal of theoretical biology:111720 PubMed ID: 38211890
Summary: Early development of Drosophila melanogaster (fruit fly) facilitated by the gap gene network has been shown to be incredibly robust, and the same patterns emerge even when the process is seriously disrupted. This study investigate this robustness using a previously developed computational framework called DSGRN (Dynamic Signatures Generated by Regulatory Networks). Mathematical innovations include the conceptual extension of this established modeling technique to enable modeling of spatially monotone environmental effects, as well as the development of a collection of graph theoretic robustness scores for network models. This allows rank ordering of the robustness of network models of cellular systems where each cell contains the same genetic network topology but is demonstrated to operate under a parameter regime that changes continuously from cell to cell. The power of this method by comparing the robustness of two previously introduced network models of gap gene expression along the anterior-posterior axis of the fruit fly embryo, both to each other and to a random sample of networks with same number of nodes and edges. It was observed that there is a substantial difference in robustness scores between the two models. The biological insight is that random network topologies are in general capable of reproducing complex patterns of expression, but that using measures of robustness to rank order networks permits a large reduction in hypothesis space for highly conserved systems such as developmental networks. | Farkas, D., Szikora, S., Jijumon, A. S., Polgar, T. F., Patai, R., Toth, M., Bugyi, B., Gajdos, T., Bíro, P., Novak, T., Erdelyi, M., Mihaly, J. (2024). Peripheral thickening of the sarcomeres and pointed end elongation of the thin filaments are both promoted by SALS and its formin interaction partners. PLoS Genet, 20(1):e1011117 PubMed ID: 38198522
Summary: During striated muscle development the first periodically repeated units appear in the premyofibrils, consisting of immature sarcomeres that must undergo a substantial growth both in length and width, to reach their final size. Beyond its well established role in sarcomere elongation, the Sarcomere length short (SALS) protein is involved in Z-disc formation and peripheral growth of the sarcomeres. Protein localization data and loss-of-function studies in the Drosophila indirect flight muscle strongly suggest that radial growth of the sarcomeres is initiated at the Z-disc. As to thin filament elongation, a powerful nanoscopy approach was used to reveal that SALS is subject to a major conformational change during sarcomere development, which might be critical to stop pointed end elongation in the adult muscles. In addition, it was demonstrated that the roles of SALS in sarcomere elongation and radial growth are both dependent on formin type of actin assembly factors. Unexpectedly, when SALS is present in excess amounts, it promotes the formation of actin aggregates highly resembling the ones described in nemaline myopathy patients. Collectively, these findings helped to shed light on the complex mechanisms of SALS during the coordinated elongation and thickening of the sarcomeres, and resulted in the discovery of a potential nemaline myopathy model, suitable for the identification of genetic and small molecule inhibitors. |
| Tah, I., Haertter, D., Crawford, J. M., Kiehart, D. P., Schmidt, C. F., Liu, A. J. (2023). Minimal vertex model explains how the amnioserosa avoids fluidization during Drosophila dorsal closure. bioRxiv, PubMed ID: 38187730
Summary: Dorsal closure is a process that occurs during embryogenesis of Drosophila melanogaster. During dorsal closure, the amnioserosa (AS), a one-cell thick epithelial tissue that fills the dorsal opening, shrinks as the lateral epidermis sheets converge and eventually merge. During this process, the aspect ratio of amnioserosa cells increases markedly. The standard 2-dimensional vertex model, which successfully describes tissue sheet mechanics in multiple contexts, would in this case predict that the tissue should fluidize via cell neighbor changes. Surprisingly, however, the amnioserosa remains an elastic solid with no such events. This study presents a minimal extension to the vertex model that explains how the amnioserosa can achieve this unexpected behavior. Continuous shrinkage of the preferred cell perimeter and cell perimeter polydispersity lead to the retention of the solid state of the amnioserosa. This model accurately captures measured cell shape and orientation changes and predicts non-monotonic junction tension that was confirmed with laser ablation experiments. | Peng, D., Jackson, D., Palicha, B., Kernfeld, E., Laughner, N., Shoemaker, A., Celniker, S. E., Loganathan, R., Cahan, P., Andrew, D. J. (2024). Organogenetic transcriptomes of the Drosophila embryo at single cell resolution. Development, 151(2) PubMed ID: 38174902
Summary: To gain insight into the transcription programs activated during the formation of Drosophila larval structures, ingle cell RNA sequencing was carried out during two periods of Drosophila embryogenesis: stages 10-12, when most organs are first specified and initiate morphological and physiological specialization; and stages 13-16, when organs achieve their final mature architectures and begin to function. The data confirm previous findings with regards to functional specialization of some organs - the salivary gland and trachea - and clarify the embryonic functions of another - the plasmatocytes. In addition two early developmental trajectories in germ cells were identified, and a potential role was uncovered for proteolysis during germline stem cell specialization. The likely cell type of origin for key components of the Drosophila matrisome (proteins that contribute to the extracellular matrix) and several commonly used Drosophila embryonic cell culture lines was identified. Finally, these findings were compared with other recent related studies and with other modalities for identifying tissue-specific gene expression patterns. These data provide a useful community resource for identifying many new players in tissue-specific morphogenesis and functional specialization of developing organs. |
Monday, August 26th - Drosophila as a Model For Human Diseases |
| Zanon, A., Guida, M., Lavdas, A. A., Corti, C., Castelo Rueda, M. P., Negro, A., Pramstaller, P. P., Domingues, F. S., Hicks, A. A., Pichler, I. (2024). Intracellular delivery of Parkin-RING0-based fragments corrects Parkin-induced mitochondrial dysfunction through interaction with SLP-2. Journal of translational medicine, 22(1):59 PubMed ID: 38229174
Summary: Loss-of-function mutations in the PRKN gene, encoding Parkin, are the most common cause of autosomal recessive Parkinson's disease (PD). Mitochondrial Stomatin-like protein 2 (SLP-2), which functions in the assembly of respiratory chain proteins, has been identified as a Parkin-binding protein. Selective knockdown of either Parkin or SLP-2 led to reduced mitochondrial and neuronal function in neuronal cells and Drosophila, where a double knockdown led to a further worsening of Parkin-deficiency phenotypes. This study investigated the minimal Parkin region involved in the Parkin-SLP-2 interaction and explored the ability of Parkin-fragments and peptides from this minimal region to restore mitochondrial function. In fibroblasts, human induced pluripotent stem cell (hiPSC)-derived neurons, and neuroblastoma cells the interaction between Parkin and SLP-2 was investigated, and the Parkin domain responsible for the binding to SLP-2 was mapped. High resolution respirometry, immunofluorescence analysis and live imaging were used to analyze mitochondrial function. Using a proximity ligation assay, the Parkin-SLP-2 interaction was quantitatively assessed in skin fibroblasts and hiPSC-derived neurons. When PD-associated PRKN mutations were present,a significantly reduced interaction between the two proteins was detected. A preferential binding was detected of SLP-2 to the N-terminal part of Parkin, with a highest affinity for the RING0 domain. Computational modeling based on the crystal structure of Parkin protein predicted several potential binding sites for SLP-2 within the Parkin RING0 domain. Amongst these, three binding sites were observed to overlap with natural PD-causing missense mutations, which were demonstrated to interfere substantially with the binding of Parkin to SLP-2. Finally, delivery of the isolated Parkin RING0 domain and a Parkin mini-peptide, conjugated to cell-permeant and mitochondrial transporters, rescued compromised mitochondrial function in Parkin-deficient neuroblastoma cells and hiPSC-derived neurons with endogenous, disease causing PRKN mutations. These findings place further emphasis on the importance of the protein-protein interaction between Parkin and SLP-2 for the maintenance of optimal mitochondrial function. The possibility of restoring an abolished binding to SLP-2 by delivering the Parkin RING0 domain or the Parkin mini-peptide involved in this specific protein-protein interaction into cells might represent a novel organelle-specific therapeutic approach for correcting mitochondrial dysfunction in Parkin-linked PD. | Zhao, B., Cowan, C. M., Coutts, J. A., Christy, D. D., Saraph, A., Hsueh, S. C. C., Plotkin, S. S., Mackenzie, I. R., Kaplan, J. M., Cashman, N. R. (2023). Targeting RACK1 to alleviate TDP-43 and FUS proteinopathy-mediated suppression of protein translation and neurodegeneration. Acta neuropathologica communications, 11(1):200 PubMed ID: 38111057
Summary: TAR DNA-binding protein 43 (TDP-43) and Fused in Sarcoma/Translocated in Sarcoma (FUS) are ribonucleoproteins associated with pathogenesis of amyotrophic lateral sclerosis (ALS) and frontotemporal lobar degeneration (FTLD). Under physiological conditions, TDP-43 and FUS are predominantly localized in the nucleus, where they participate in transcriptional regulation, RNA splicing and metabolism. In disease, however, they are typically mislocalized to the cytoplasm where they form aggregated inclusions. A number of shared cellular pathways have been identified that contribute to TDP-43 and FUS toxicity in neurodegeneration. This study reports a novel pathogenic mechanism shared by these two proteins. Pathological FUS was found to co-aggregate with a ribosomal protein, the Receptor for Activated C-Kinase 1 (RACK1), in the cytoplasm of spinal cord motor neurons of ALS, as previously reported for pathological TDP-43. In HEK293T cells transiently transfected with TDP-43 or FUS mutant lacking a functional nuclear localization signal (NLS; TDP-43(ΔNLS) and FUS(ΔNLS)), cytoplasmic TDP-43 and FUS induced co-aggregation with endogenous RACK1. These co-aggregates sequestered the translational machinery through interaction with the polyribosome, accompanied by a significant reduction of global protein translation. RACK1 knockdown decreased cytoplasmic aggregation of TDP-43(ΔNLS) or FUS(ΔNLS) and alleviated associated global translational suppression. Surprisingly, RACK1 knockdown also led to partial nuclear localization of TDP-43(ΔNLS) and FUS(ΔNLS) in some transfected cells, despite the absence of NLS. In vivo, RACK1 knockdown alleviated retinal neuronal degeneration in transgenic Drosophila melanogaster expressing hTDP-43(WT) or hTDP-43(Q331K) and improved motor function of hTDP-43(WT) flies, with no observed adverse effects on neuronal health in control knockdown flies. In conclusion, these results revealed a novel shared mechanism of pathogenesis for misfolded aggregates of TDP-43 and FUS mediated by interference with protein translation in a RACK1-dependent manner. This study provides proof-of-concept evidence for targeting RACK1 as a potential therapeutic approach for TDP-43 or FUS proteinopathy associated with ALS and FTLD. |
| Fedina, T. Y., Cummins, E. T., Promislow, D. E. L., Pletcher, S. D. (2023). The neuropeptide drosulfakinin enhances choosiness and protects males from the aging effects of social perception. Proc Natl Acad Sci U S A, 120(51):e2308305120 PubMed ID: 38079545
Summary: The motivation to reproduce is a potent natural drive, and the social behaviors that induce it can severely impact animal health and lifespan. Indeed, in Drosophila males, accelerated aging associated with reproduction arises not from the physical act of courtship or copulation but instead from the motivational drive to court and mate. To better understand the mechanisms underlying social effects on aging, male choosiness for mates was studied. Increased activity of insulin-producing cells (IPCs) of the fly brain was found to potentiated choosiness without consistently affecting courtship activity. Surprisingly, this effect was not caused by insulins themselves, but instead by drosulfakinin (DSK), another neuropeptide produced in a subset of the IPCs, acting through one of the two DSK receptors, CCKLR-17D1. Activation of Dsk(+) IPC neurons also decreased food consumption, while activation of Dsk(+) neurons outside of IPCs affected neither choosiness nor feeding, suggesting an overlap between Dsk(+)neurons modulating choosiness and those influencing satiety. Broader activation of Dsk(+) neurons (both within and outside of the IPCs) was required to rescue the detrimental effect of female pheromone exposure on male lifespan, as was the function of both DSK receptors. The same broad set of Dsk(+) neurons was found to reinforce normally aversive feeding interactions, but only after exposure to female pheromones, suggesting that perception of the opposite sex gates rewarding properties of these neurons. It is speculated that broad Dsk(+) neuron activation is associated with states of satiety and social experience, which under stressful conditions is rewarding and beneficial for lifespan. | Farkas, A., Zsindely, N., Nagy, G., KovAcs, L., Deák, P., Bodai, L. (2023). The ubiquitin thioesterase YOD1 ameliorates mutant Huntingtin induced pathology in Drosophila. Sci Rep, 13(1):21951 PubMed ID: 38081944
Summary: Huntington's disease (HD) is a neurodegenerative disorder caused by a dominant gain-of-function mutation in the huntingtin gene, resulting in an elongated polyglutamine repeat in the mutant Huntingtin (mHtt) that mediates aberrant protein interactions. Previous studies implicated the ubiquitin-proteasome system in HD, suggesting that restoring cellular proteostasis might be a key element in suppressing pathology. This study applied genetic interaction tests in a Drosophila model to ask whether modulating the levels of deubiquitinase enzymes affect HD pathology. By testing 32 deubiquitinase genes, it was found that overexpression of Yod1 ameliorated all analyzed phenotypes, including neurodegeneration, motor activity, viability, and longevity. Yod1 did not have a similar effect in amyloid beta overexpressing flies, suggesting that the observed effects might be specific to mHtt. Yod1 overexpression did not alter the number of mHtt aggregates but moderately increased the ratio of larger aggregates. Transcriptome analysis showed that Yod1 suppressed the transcriptional effects of mHtt and restored the expression of genes involved in neuronal plasticity, vesicular transport, antimicrobial defense, and protein synthesis, modifications, and clearance. Furthermore, Yod1 overexpression in HD flies leads to the upregulation of genes involved in transcriptional regulation and synaptic transmission, which might be part of a response mechanism to mHtt-induced stress. |
| Lo Piccolo, L., Yeewa, R., Pohsa, S., Yamsri, T., Calovi, D., Phetcharaburanin, J., Suksawat, M., Kulthawatsiri, T., Shotelersuk, V., Jantrapirom, S. (2023). FAME4-associating YEATS2 knockdown impairs dopaminergic synaptic integrity and leads to seizure-like behaviours in Drosophila melanogaster. Progress in neurobiology, 233:102558 PubMed ID: 38128822
Summary: Familial adult myoclonus epilepsy (FAME) is a neurological disorder caused by a TTTTA/TTTCA intronic repeat expansion. FAME4 is one of the six types of FAME that results from the repeat expansion in the first intron of the gene YEATS2. Although the RNA toxicity is believed to be the primary mechanism underlying FAME, the role of genes where repeat expansions reside is still unclear, particularly in the case of YEATS2 in neurons. This study used Drosophila to explore the effects of reducing YEATS2 expression. Two pan-neuronally driven dsDNA were used for knockdown of Drosophila YEATS2 (dYEATS2), and the resulting molecular and behavioural outcomes were evaluated. Drosophila with reduced dYEATS2 expression exhibited decreased tolerance to acute stress, disturbed locomotion, abnormal social behaviour, and decreased motivated activity. Additionally, reducing dYEATS2 expression negatively affected tyrosine hydroxylase (TH) gene expression, resulting in decreased dopamine biosynthesis. Remarkably, seizure-like behaviours induced by knocking down dYEATS2 were rescued by the administration of L-DOPA. This study reveals a novel role of YEATS2 in neurons in regulating acute stress responses, locomotion, and complex behaviours, and suggests that haploinsufficiency of YEATS2 may play a role in FAME4. | Lee, D., Yoon, E., Ham, S. J., Lee, K., Jang, H., Woo, D., Lee, D. H., Kim, S., Choi, S., Chung, J. (2024). Diabetic sensory neuropathy and insulin resistance are induced by loss of UCHL1 in Drosophila. Nat Commun, 15(1):468 PubMed ID: 38212312
Summary: Diabetic sensory neuropathy (DSN) is one of the most common complications of type 2 diabetes (T2D), however the molecular mechanistic association between T2D and DSN remains elusive. This study identified ubiquitin C-terminal hydrolase L1 (UCHL1), a deubiquitinase highly expressed in neurons, as a key molecule underlying T2D and DSN. Genetic ablation of UCHL1 leads to neuronal insulin resistance and T2D-related symptoms in Drosophila. Furthermore, loss of UCHL1 induces DSN-like phenotypes, including numbness to external noxious stimuli and axonal degeneration of sensory neurons in flies' legs. Conversely, UCHL1 overexpression improves DSN-like defects of T2D model flies. UCHL1 governs insulin signaling by deubiquitinating insulin receptor substrate 1 (IRS1) and antagonizes an E3 ligase of IRS1, Cullin 1 (CUL1). Consistent with these results, genetic and pharmacological suppression of CUL1 activity rescues T2D- and DSN-associated phenotypes. Therefore, rhwaw findings suggest a complete set of genetic factors explaining T2D and DSN, together with potential remedies for the diseases. |
Friday, August 23rd - Structure, Development, and Function of Larval and Adult Nervous System |
| Hamid, A., Gattuso, H., Caglar, A. N., Pillai, M., Steele, T., Gonzalez, A., Nagel, K., Syed, M. H. (2023). The conserved RNA-binding protein Imp is required for the specification and function of olfactory navigation circuitry in Drosophila. Curr Biol, PubMed ID: 38181792
Summary: Complex behaviors depend on the precise developmental specification of neuronal circuits, but the relationship between genetic programs for neural development, circuit structure, and behavioral output is often unclear. The central complex (CX) is a conserved sensory-motor integration center in insects, which governs many higher-order behaviors and largely derives from a small number of type II neural stem cells (NSCs). This study shows that Imp, a conserved IGF-II mRNA-binding protein expressed in type II NSCs, plays a role in specifying essential components of CX olfactory navigation circuitry. This study shows the following: (1) that multiple components of olfactory navigation circuitry arise from type II NSCs. (2) Manipulating Imp expression in type II NSCs alters the number and morphology of many of these circuit elements, with the most potent effects on neurons targeting the ventral layers of the fan-shaped body (FB). (3) Imp regulates the specification of Tachykinin-expressing ventral FB input neurons. (4) Imp is required in type II NSCs for establishing proper morphology of the CX neuropil structures. (5) Loss of Imp in type II NSCs abolishes upwind orientation to attractive odor while leaving locomotion and odor-evoked regulation of movement intact. Taken together, these findings establish that a temporally expressed gene can regulate the expression of a complex behavior by developmentally regulating the specification of multiple circuit components and provides a first step toward a developmental dissection of the CX and its roles in behavior. | Ros, I. G., Omoto, J. J., Dickinson, M. H. (2024). Descending control and regulation of spontaneous flight turns in Drosophila. Curr Biol, PubMed ID: 38228148
Summary: The clumped distribution of resources in the world has influenced the pattern of foraging behavior since the origins of locomotion, selecting for a common search motif in which straight movements through resource-poor regions alternate with zig-zag exploration in resource-rich domains. For example, during local search, flying flies spontaneously execute rapid flight turns, called body saccades, but suppress these maneuvers during long-distance dispersal or when surging upstream toward an attractive odor. This study describes the key cellular components of a neural network in flies that generate spontaneous turns as well as a specialized pair of neurons that inhibits the network and suppresses turning. Using 2-photon imaging, optogenetic activation, and genetic ablation, it was shown that only four descending neurons appear sufficient to generate the descending commands to execute flight saccades. The network is organized into two functional units-one for right turns and one for left-with each unit consisting of an excitatory (DNae014) and an inhibitory (DNb01) neuron that project to the flight motor neuropil within the ventral nerve cord. Using resources from recently published connectomes of the fly, a pair of large, distinct interneurons (VES041) was identified that form inhibitory connections to all four saccade command neurons and created specific genetic driver lines for this cell. As predicted by its connectivity, activation of VES041 strongly suppresses saccades, suggesting that it promotes straight flight to regulate the transition between local search and long-distance dispersal. These results thus identify the key elements of a network that may play a crucial role in foraging ecology. |
| Carrier, Y., Rio, L. Q., Formicola, N., de Sousa-Xavier, V., Tabet, M., Chen, Y. D., Wislez, M., Orts, L., Pinto-Teixeira, F. (2023). Biased cell adhesion organizes a circuit for visual motion integration. bioRxiv, PubMed ID: 38168373
Summary: Layer specific computations in the brain rely on neuronal processes establishing synaptic connections with specific partners in distinct laminae. In the Drosophila lobula plate neuropile, the axons of the four subtypes of T4 and T5 visual motion direction-selective neurons segregate into four layers, based on their directional preference, and form synapses with distinct subsets of postsynaptic neurons. Four bi-stratified inhibitory lobula plate intrinsic cells exhibit a consistent synaptic pattern, receiving excitatory T4/T5 inputs in one layer, and conveying inhibitory signals to an adjacent layer. This layered arrangement establishes motion opponency. This study identified layer-specific expression of different receptor-ligand pairs belonging to the Beat and Side families of Cell Adhesion Molecules (CAMs) between T4/T5 neurons and their postsynaptic partners. Genetic analysis reveals that Beat/Side mediated interactions are required to restrict T4/T5 axonal innervation to a single layer. It is proposed that Beat/Side contribute to synaptic specificity by biasing adhesion between synaptic partners before synaptogenesis. | Zhang, H., Rui, M., Ma, Z., Gong, S., Zhang, S., Zhou, Q., Gan, C., Gong, W., Wang, S. (2024). Golgi-to-ER retrograde transport prevents premature differentiation of Drosophila type II neuroblasts via Notch-signal-sending daughter cells. iScience, 27(1):108545 PubMed ID: 38213621
Summary: Stem cells are heterogeneous to generate diverse differentiated cell types required for organogenesis; however, the underlying mechanisms that differently maintain these heterogeneous stem cells are not well understood. This study identified that Golgi-to-endoplasmic reticulum (ER) retrograde transport specifically maintains type II neuroblasts (NBs) through the Notch signaling. Intermediate neural progenitors (INPs), immediate daughter cells of type II NBs, provide Delta and function as the NB niche. The Delta used by INPs is mainly produced by NBs and asymmetrically distributed to INPs. Blocking retrograde transport leads to a decrease in INP number, which reduces Notch activity and results in the premature differentiation of type II NBs. Furthermore, the reduction of Delta could suppress tumor formation caused by type II NBs. These results highlight the crosstalk between Golgi-to-ER retrograde transport, Notch signaling, stem cell niche, and fusion as an essential step in maintaining the self-renewal of type II NB lineage. |
| Xu, C., Ramos, T. B., Rogers, E. M., Reiser, M. B., Doe, C. Q. (2024). Homeodomain proteins hierarchically specify neuronal diversity and synaptic connectivity. Elife, 12 PubMed ID: 38180023
Summary: How the brain generates diverse neuron types that assemble into precise neural circuits remains unclear. Using Drosophila lamina neuron types (L1-L5), this study showed that the primary homeodomain transcription factor (HDTF) brain-specific homeobox (Bsh) is initiated in progenitors and maintained in L4/L5 neurons to adulthood. Bsh activates secondary HDTFs Ap (L4) and Pdm3 (L5) and specifies L4/L5 neuronal fates while repressing the HDTF Zfh1 to prevent ectopic L1/L3 fates (control: L1-L5; Bsh-knockdown: L1-L3), thereby generating lamina neuronal diversity for normal visual sensitivity. Subsequently, in L4 neurons, Bsh and Ap function in a feed-forward loop to activate the synapse recognition molecule DIP-β, thereby bridging neuronal fate decision to synaptic connectivity. Expression of a Bsh:Dam, specifically in L4, reveals Bsh binding to the DIP-β locus and additional candidate L4 functional identity genes. It is proposed that HDTFs function hierarchically to coordinate neuronal molecular identity, circuit formation, and function. Hierarchical HDTFs may represent a conserved mechanism for linking neuronal diversity to circuit assembly and function. | Li, S. S., Li, A. Q., Liu, Z. Y., Zhao, X. Y., Wang, G. R., Deng, Y., Wang, Q. P. (2024). Glutamine enhances sucrose taste through a gut microbiota-gut-brain axis in Drosophila. Life sciences, 339:122415 PubMed ID: 38218533
Summary: Amino acids (AAs) are known to play important roles in various physiological functions. However, their effect on sweet taste perception remains largely unknown. Drosophila was used to evaluate the effect of each AA on sucrose taste perception. Individual AA was supplemented into diets and male flies were fed on these diets for 6 days. The proboscis extension response (PER) assay was applied to assess the sucrose taste sensitivity of treated flies. The RNA-seq and germ-free (GF) flies were further used to reveal the underlying mechanisms of sucrose taste sensitization induced by glutamine (Gln). Supplementation of Gln into diets significantly enhanced sucrose taste sensitivity. This sucrose taste sensitization is dependent on gut microbiota and requires a specific gut bacterium Acetobacter tropicalis (A. tropicalis). It was further found that CNMamide (CNMa) in the gut and CNMa receptor (CNMaR) in dopaminergic neurons are required for increased sucrose taste sensitivity by Gln diet. Finally, it was demonstrated that a gut microbiota-gut-brain axis is required for Gln-induced sucrose taste sensitization. These findings can advance understanding of the complex interplay between host physiology, dietary factors, and gut microbiota. |
Thursday, August 22nd - Adult Physiology and Metabolism |
| ----- | Li, M., Macro, J., Meadows, K., Mishra, D., Martin, D., Olson, S., Huggins, B. J., Graveley, B. R., Li, J. Y. H., Rogina, B. (2023). Late-life shift in caloric intake affects fly metabolism and longevity. Proc Natl Acad Sci U S A, 120(50):e2311019120 PubMed ID: 38064506
Summary: The prevalence of obesity is increasing in older adults and contributes to age-related decline. Caloric restriction (CR) alleviates obesity phenotypes and delays the onset of age-related changes. However, how late in life organisms benefit from switching from a high-(H) to a low-calorie (L) diet is unclear. Male flies were transferred from a H to a L (HL) diet or vice versa (LH) at different times during life. Both shifts immediately change fly rate of aging even when applied late in life. HL shift rapidly reduces fly mortality rate to briefly lower rate than in flies on a constant L diet, and extends lifespan. Transcriptomic analysis uncovers that flies aged on H diet have acquired increased stress response, which may have temporal advantage over flies aged on L diet and leads to rapid decrease in mortality rate after HL switch. Conversely, a LH shift increases mortality rate, which is temporarily higher than in flies aged on a H diet, and shortens lifespan. Unexpectedly, more abundant transcriptomic changes accompanied LH shift, including increase in ribosome biogenesis, stress response and growth. These changes reflect protection from sudden release of ROS, energy storage, and use of energy to growth, which all likely contribute to higher mortality rate. As the beneficial effects of CR on physiology and lifespan are conserved across many organisms, this study provides framework to study underlying mechanisms of CR interventions that counteract the detrimental effects of H diets and reduce rate of aging even when initiated later in life. | Katheder, N. S., Browder, K. C., Chang, D., De Maziere, A., Kujala, P., van Dijk, S., Klumperman, J., Lu, T. C., Li, H., Lai, Z., Sangaraju, D., Jasper, H. (2023). Nicotinic acetylcholine receptor signaling maintains epithelial barrier integrity. Elife, 12 PubMed ID: 38063293
Summary: Disruption of epithelial barriers is a common disease manifestation in chronic degenerative diseases of the airways, lung, and intestine. Extensive human genetic studies have identified risk loci in such diseases, including in chronic obstructive pulmonary disease (COPD) and inflammatory bowel diseases. The genes associated with these loci have not fully been determined, and functional characterization of such genes requires extensive studies in model organisms. This study reports the results of a screen in Drosophila melanogaster that allowed for rapid identification, validation, and prioritization of COPD risk genes that were selected based on risk loci identified in human genome-wide association studies (GWAS). Using intestinal barrier dysfunction in flies as a readout, the results validate the impact of candidate gene perturbations on epithelial barrier function in 56% of the cases, resulting in a prioritized target gene list. The functional characterization in flies of one family of these genes, encoding for nicotinic acetylcholine receptor (nAchR) subunits is further reported. nAchR signaling in enterocytes of the fly gut promotes epithelial barrier function and epithelial homeostasis by regulating the production of the peritrophic matrix. These findings identify COPD-associated genes critical for epithelial barrier maintenance, and provide insight into the role of epithelial nAchR signaling for homeostasis. |
| Holcombe, J., Weavers, H. (2023). Functional-metabolic coupling in distinct renal cell types coordinates organ-wide physiology and delays premature ageing. Nat Commun, 14(1):8405 PubMed ID: 38110414
Summary: Precise coupling between cellular physiology and metabolism is emerging as a vital relationship underpinning tissue health and longevity. Nevertheless, functional-metabolic coupling within heterogenous microenvironments in vivo remains poorly understood due to tissue complexity and metabolic plasticity. This study established the Drosophila renal system as a paradigm for linking mechanistic analysis of metabolism, at single-cell resolution, to organ-wide physiology. Kidneys are amongst the most energetically-demanding organs, yet exactly how individual cell types fine-tune metabolism to meet their diverse, unique physiologies over the life-course remains unclear. Integrating live-imaging of metabolite and organelle dynamics with spatio-temporal genetic perturbation within intact functional tissue, this study uncovered distinct cellular metabolic signatures essential to support renal physiology and healthy ageing. Cell type-specific programming of glucose handling, PPP-mediated glutathione regeneration and FA β-oxidation via dynamic lipid-peroxisomal networks, downstream of differential ERR receptor activity, precisely match cellular energetic demands whilst limiting damage and premature senescence; however, their dramatic dysregulation may underlie age-related renal dysfunction | Camus, M. F., Inwongwan, S. (2023). Mitonuclear interactions modulate nutritional preference. Biology letters, 19(12):20230375 PubMed ID: 38053364
Summary: In nature, organisms are faced with constant nutritional options which fuel key life-history traits. Studies have shown that species can actively make nutritional decisions based on internal and external cues. Metabolism itself is underpinned by complex genomic interactions involving components from both nuclear and mitochondrial genomes. Products from these two genomes must coordinate how nutrients are extracted, used and recycled. Given the complicated nature of metabolism, it is not well understood how nutritional choices are affected by mitonuclear interactions. This is under the rationale that changes in genomic interactions will affect metabolic flux and change physiological requirements. To this end this study used a large Drosophila mitonuclear genetic panel, comprising nine isogenic nuclear genomes coupled to nine mitochondrial haplotypes, giving a total of 81 different mitonuclear genotypes. A capillary-based feeding assay was used to screen this panel for dietary preference between carbohydrate and protein. Significant mitonuclear interactions were found modulating nutritional choices, with these epistatic interactions also being dependent on sex. These findings support the notion that complex genomic interactions can place a constraint on metabolic flux. This work gives deeper insights into how key metabolic interactions can have broad implications on behaviour. |
| Matsuka, M., Otsune, S., Sugimori, S., Tsugita, Y., Ueda, H., Nakagoshi, H. (2024). Fecundity is optimized by levels of nutrient signal-dependent expression of Dve and EcR in Drosophila male accessory gland. Dev Biol, 508:8-23 PubMed ID: 38199580
Summary: Steroid hormones play various physiological roles including metabolism and reproduction. Steroid hormones in insects are ecdysteroids, and the major form in Drosophila melanogaster is ecdysone. In Drosophila males, the accessory gland is responsive to nutrient-dependent regulation of fertility/fecundity. The accessory gland is composed of two types of binucleated epithelial cells: a main cell and a secondary cell (SC). The transcription factors Defective proventriculus (Dve), Abdominal-B, and Ecdysone receptors (EcRs) are strongly expressed in adult SCs. This EcR expression is regulated by parallel pathways of nutrient signaling and the Dve activity. Induction of Dve expression is also dependent on nutrient signaling, and it becomes nutrient signal-independent during a restricted period of development. Forced dve expression during the restricted period significantly increased the number of SCs. This study provides evidence that the level of nutrient signal-dependent Dve expression during the restricted period determines the number of SCs, and that ecdysone signaling is also crucial to optimize male fecundity through nutrient signal-dependent survival and maturation of SCs. | Huang, J., Zhou, F., Zhou, H., Zheng, X., Huo, Z., Yang, M., Xu, Z., Liu, R., Wang, L., Wang, X. (2023). Systematic assessment of transcriptomic and metabolic reprogramming by blue light exposure coupled with aging. PNAS nexus, 2(12):pgad390 PubMed ID: 38059264
Summary: The prevalent use of light-emitting diodes (LEDs) has caused revolutionary changes in modern life, but the potential hazards to health of blue light are poorly understood. N(6)-methyladenosine (m(6)A) is the most prevalent posttranscriptional modification in eukaryotes and can modulate diverse physiological processes by regulating mRNA fate. To understand the effects and molecular mechanisms of daily low-intensity blue light exposure (BLE) and ascertain whether m(6)A methylation plays a role in BLE-induced phenotypes, this study constructed a series of Drosophila models under different durations of daily low-intensity BLE and multiomics profiles were obtained. These results revealed that BLE could induce transcriptomic, m(6)A epitranscriptomic, and metabolomic reprogramming in Drosophila along with aging process. Importantly, the m(6)A methylation sites enriched in the 5' untranslated regions (UTRs) of Drosophila transcripts showed strong age specificity and could be altered by BLE. It was experimentally validated that aging-related gene Tor and circadian rhythm-related gene per were regulated by 5' UTR-enriched m(6)A methylation. Overall, this study provides a systematic assessment of m(6)A RNA methylome reprogramming by BLE and aging in Drosophila model. |
Wednesday, August 21st - Gonads |
| Brown, N. C., Gordon, B., McDonough-Goldstein, C. E., Misra, S., Findlay, G. D., Clark, A. G., Wolfner, M. F. (2023). The seminal odorant binding protein Obp56g is required for mating plug formation and male fertility in Drosophila melanogaster. Elife, 12 PubMed ID: 38126735
Summary: In Drosophila melanogaster and other insects, the seminal fluid proteins (SFPs) and male sex pheromones that enter the female with sperm during mating are essential for fertility and induce profound post-mating effects on female physiology and behavior. The SFPs in D. melanogaster and other taxa include several members of the large gene family known as odorant binding proteins (Obps). Previous work in Drosophila has shown that some Obp genes are highly expressed in the antennae and can mediate behavioral responses to odorants, potentially by binding and carrying these molecules to odorant receptors. These observations have led to the hypothesis that the seminal Obps might act as molecular carriers for pheromones or other compounds important for male fertility in the ejaculate, though functional evidence in any species is lacking. This study used RNAi and CRISPR/Cas9 generated mutants to test the role of the seven seminal Obps in D. melanogaster fertility and the post-mating response (PMR). Obp56g was shown to be required for male fertility and the induction of the PMR, whereas the other six genes had no effect on fertility when mutated individually. Obp56g is expressed in the male's ejaculatory bulb, an important tissue in the reproductive tract that synthesizes components of the mating plug. Males lacking Obp56g fail to form a mating plug in the mated female's reproductive tract, leading to ejaculate loss and reduced sperm storage. The evolutionary history was examined of these seminal Obp genes, as several studies have documented rapid evolution and turnover of SFP genes across taxa. Extensive lability was found in gene copy number and evidence of positive selection acting on two genes, Obp22a and Obp51a. Comparative RNAseq data from the male reproductive tract of multiple Drosophila species revealed that Obp56g shows high male reproductive tract expression only in species of the melanogaster and obscura groups, though conserved head expression in all species tested. Together, these functional and expression data suggest that Obp56g may have been co-opted for a reproductive function over evolutionary time. | Sheahan, T. D., Grewal, A., Korthauer, L. E., Blumenthal, E. M. (2023). The Drosophila drop-dead gene is required for eggshell integrity. PLoS One, 18(12):e0295412 PubMed ID: 38051756
Summary: The eggshell of the fruit fly Drosophila melanogaster is a useful model for understanding the synthesis of a complex extracellular matrix. The eggshell is synthesized during mid-to-late oogenesis by the somatic follicle cells that surround the developing oocyte. Previously studies have shown that female flies mutant for the gene drop-dead (drd) are sterile, but the underlying cause of the sterility remained unknown. This study examined the role of drd in eggshell synthesis. Eggs laid by drd mutant females are fertilized but arrest early in embryogenesis, and the innermost layer of the eggshell, the vitelline membrane, is abnormally permeable to dye in these eggs. In addition, the major vitelline membrane proteins fail to become crosslinked by nonreducible bonds, a process that normally occurs during egg activation following ovulation, as evidenced by their solubility and detection by Western blot in laid eggs. In contrast, the Cp36 protein, which is found in the outer chorion layers of the eggshell, becomes crosslinked normally. To link the drd expression pattern with these phenotypes, drd was shown to be expressed in the ovarian follicle cells beginning in mid-oogenesis, and, importantly, that all drd mutant eggshell phenotypes could be recapitulated by selective knockdown of drd expression in the follicle cells. To determine whether drd expression was required for the crosslinking itself, in vitro activation and crosslinking experiments were. The vitelline membranes of control egg chambers could become crosslinked either by incubation in hyperosmotic medium, which activates the egg chambers, or by exogenous peroxidase and hydrogen peroxide. In contrast, neither treatment resulted in the crosslinking of the vitelline membrane in drd mutant egg chambers. These results indicate that drd expression in the follicle cells is necessary for vitelline membrane proteins to serve as substrates for peroxidase-mediated cross-linking at the end of oogenesis. |
| Rohrbach, E. W., Knapp, E. M., Deshpande, S. A., Krantz, D. E. (2024). Expression and potential regulatory functions of Drosophila octopamine receptors in the female reproductive tract. G3 (Bethesda), PubMed ID: 38244217
Summary: Aminergic signaling is known to play a critical role in regulating female reproductive processes in both mammals and insects. In Drosophila, the ortholog of noradrenaline, octopamine (OA), is required for ovulation as well as several other female reproductive processes. Two OA receptors have already been shown to be expressed in the Drosophila reproductive tract and to be required for egg-laying: OAMB and Octβ2R. The Drosophila genome contains four additional OA receptors- Octα2R, Octβ1R, Octβ3R, and Oct/TyrR-but their cellular patterns of expression in the reproductive tract and potential contribution(s) to egg-laying are not known. In addition, the mechanisms by which OAMB and Octβ2R regulate reproduction are incompletely understood. Using a panel of MiMIC Gal4 lines, this study showed that Octα2R, Octβ1R, Octβ3R, and Oct/TyrR receptors are not detectable in either epithelium or muscle but are clearly expressed in neurons within the female fly reproductive tract. Optogenetic activation of neurons that express at least three types of OA receptors stimulate contractions in the lateral oviduct. Octopamine was found to stimulate calcium transients in the sperm storage organs and that its effects in spermathecal, secretory cells can be blocked by knock-down of OAMB. These data extend understanding of the pathways by which octopamine regulates egg-laying in Drosophila and raise the possibility that multiple OA receptor subtypes could play a role in this process. | Benner, L., Muron, S., Gomez, J. G., Oliver, B. (2023). OVO Positively Regulates Essential Maternal Pathways by Binding Near the Transcriptional Start Sites in the Drosophila Female Germline. bioRxiv, PubMed ID: 38076814
Summary: Differentiation of female germline stem cells into a mature oocyte includes the expression of a number of mRNAs and proteins that drive early embryonic development in Drosophila. There has been little insight into what activates the expression of these maternal factors. One candidate is the zinc-finger protein OVO. OVO is required for female germline viability, and has been shown to positively regulate its own expression, as well as a downstream target, ovarian tumor (otu), by binding to the transcriptional start site (TSS). To find additional OVO targets in the female germline and further elucidate OVO's role in oocyte development, ChIP-seq was performed to determine genome-wide OVO occupancy, as well as RNA-seq to where OVO is required. OVO preferentially binds in close proximity to target TSSs genome-wide, is associated with open chromatin, transcriptionally active histone marks, and OVO-dependent expression. Motif enrichment analysis on OVO ChIP peaks identified a 5'-TAACNGT-3' OVO DNA binding motif near TSS, but without the precise motif spacing relative to TSS characteristic of RNA Polymerase II complex binding core promoter elements. Integrated genomics analysis showed that 525 genes that are bound and increase in expression downstream of OVO are known to be maternally loaded into eggs and early embryos. These include genes involved in anterior/posterior/germ plasm specification (bcd, exu, swa, osk, nos, pgc, gcl), egg activation (png, plu, gnu, wisp, C(3)g, mtrm), translational regulation (cup, orb, bru1, me31B), and vitelline membrane formation (fs(1)N, fs(1)M3, clos). This suggests that OVO is a master transcriptional regulator of oocyte development and is responsible for the expression of structural components of the egg as well as maternally provided RNAs that are required for early embryonic pattern formation. |
| Suyama, R., Cetraro, N., Yew, J. Y., Kai, T. (2023). Microbes control Drosophila germline stem cell increase and egg maturation through hormonal pathways. Communications biology, 6(1):1287 PubMed ID: 38123715
Summary: Reproduction is highly dependent on environmental and physiological factors including nutrition, mating stimuli and microbes. Among these factors, microbes facilitate vital functions for host animals such as nutritional intake, metabolic regulation, and enhancing fertility under poor nutrition conditions. However, detailed molecular mechanisms by which microbes control germline maturation, leading to reproduction, remain largely unknown. This study shows that environmental microbes exert a beneficial effect on Drosophila oogenesis by promoting germline stem cell (GSC) proliferation and subsequent egg maturation via acceleration of ovarian cell division and suppression of apoptosis. Moreover, insulin-related signaling is not required; rather, the ecdysone pathway is necessary for microbe-induced increase of GSCs and promotion of egg maturation, while juvenile hormone contributes only to increasing GSC numbers, suggesting that hormonal pathways are activated at different stages of oogenesis. These findings reveal that environmental microbes can enhance host reproductivity by modulating host hormone release and promoting oogenesis. | Hafezi, Y., Omurzakov, A., Carlisle, J. A., Caldas, I. V., Wolfner, M. F., Clark, A. G. (2024). The Drosophila melanogaster Y-linked gene, WDY, is required for sperm to swim in the female reproductive tract. Communications biology, 7(1):90 PubMed ID: 38216628
Summary: Unique patterns of inheritance and selection on Y chromosomes have led to the evolution of specialized gene functions. This study reports CRISPR mutants in Drosophila of the Y-linked gene, WDY, which is required for male fertility. The sperm tails of WDY mutants beat approximately half as fast as those of wild-type and that mutant sperm do not propel themselves within the male ejaculatory duct or female reproductive tract. Therefore, although mature sperm are produced by WDY mutant males, and are transferred to females, those sperm fail to enter the female sperm storage organs. This study reports genotype-dependent and regional differences in sperm motility that appear to break the correlation between sperm tail beating and propulsion. Furthermore, a significant change was identified in hydrophobicity at a residue at a putative calcium-binding site in WDY orthologs at the split between the melanogaster and obscura species groups, when WDY first became Y-linked. This suggests that a major functional change in WDY coincided with its appearance on the Y chromosome. Finally, it was shown that mutants for another Y-linked gene, PRY, also show a sperm storage defect that may explain their subfertility. Overall, this study provides direct evidence for the long-held presumption that protein-coding genes on the Drosophila Y regulate sperm motility. |
Tuesday, August 20th - Signaling |
| Masuda, W., Yamakawa, T., Ajima, R., Miyake, K., Umemiya, T., Azuma, K., Tamaru, J. I., Kiso, M., Das, P., Saga, Y., Matsuno, K., Kitagawa, M. (2023). TM2D3, a mammalian homologue of Drosophila neurogenic gene product Almondex, regulates surface presentation of Notch receptors. Sci Rep, 13(1):20913 PubMed ID: 38016980
Summary: Notch signaling is an evolutionarily conserved mechanism required for numerous types of cell fate decisions in metazoans. It mediates short-range communication between cells with receptors and ligands, both of which are expressed on the cell surfaces. In response to the ligand-receptor interaction, the ligand and the extracellular domain of the Notch receptor (NECD) in the complex are internalized into ligand-expressing cells by endocytosis, a prerequisite process for the conformational change of the membrane proximal region of Notch to induce critical proteolytic cleavages for its activation. This study reports that overexpression of transmembrane 2 (TM2) domain containing 3 (TM2D3), a mammalian homologue of Drosophila melanogaster Almondex (Amx), activates Notch1. This activation requires the ligand-binding domain in Notch1 and the C-terminal region containing TM2 domain in TM2D3. TM2D3 physically associates with Notch1 at the region distinct from the ligand-binding domain and enhances expression of Notch1 on the cell surface. Furthermore, cell surface expression of Notch1 and Notch2 is reduced in Tm2d3-deficient cells. Finally, amx-deficient Drosophila early embryos exhibit impaired endocytosis of NECD and Delta ligand, for which surface presentation of Notch is required. These results indicate that TM2D3 is an element involved in Notch signaling through the surface presentation. | Gumeni, S., Lamprou, M., Evangelakou, Z., Manola, M. S., Trougakos, I. P. (2023). Sustained Nrf2 Overexpression-Induced Metabolic Deregulation Can Be Attenuated by Modulating Insulin/Insulin-like Growth Factor. Signaling. Cells, 12(22) PubMed ID: 37998385
Summary: The modulation of insulin/insulin-like growth factor signaling (IIS) is associated with altered nutritional and metabolic states. The Drosophila genome encodes eight insulin-like peptides, whose activity is regulated by a group of secreted factors, including Ecdysone-inducible gene L2 (ImpL2), which acts as a potent IIS inhibitor. cncC (cncC/Nrf2), the fly ortholog of Nrf2, is a positive transcriptional regulator of ImpL2, as part of a negative feedback loop aiming to suppress cncC/Nrf2 activity. This finding correlated with the observation that sustained cncC/Nrf2 overexpression/activation (cncC(OE); a condition that signals organismal stress) deregulates IIS, causing hyperglycemia, the exhaustion of energy stores in flies' tissues, and accelerated aging. This study extends these studies in Drosophila by assaying the functional implication of ImpL2 in cncC(OE)-mediated metabolic deregulation. ImpL2 knockdown (KD) in cncC(OE) flies was found to partially reactivated IIS, attenuate hyperglycemia and restore tissue energetics. Moreover, ImpL2 KD largely suppressed cncC(OE)-mediated premature aging. In support, pharmacological treatment of cncC(OE) flies with Metformin, a first-line medication for type 2 diabetes, restored (dose-dependently) IIS functionality and extended cncC(OE) flies' longevity. These findings exemplify the effect of chronic stress in predisposition to diabetic phenotypes, indicating the potential prophylactic role of maintaining normal IIS functionality. |
| Zhang, S., Wu, S., Yao, R., Wei, X., Ohlstein, B., Guo, Z. (2024). Eclosion muscles secrete ecdysteroids to initiate asymmetric intestinal stem cell division in Drosophila. Dev Cell, 59(1):125-140.e112 PubMed ID: 38096823
Summary: During organ development, tissue stem cells first expand via symmetric divisions and then switch to asymmetric divisions to minimize the time to obtain a mature tissue. In the Drosophila midgut, intestinal stem cells switch their divisions from symmetric to asymmetric at midpupal development to produce enteroendocrine cells. However, the signals that initiate this switch are unknown. This study identified the signal as ecdysteroids. In the presence of ecdysone, EcR and Usp promote the expression of E93 to suppress Br expression, resulting in asymmetric divisions. Surprisingly, the primary source of pupal ecdysone is not from the prothoracic gland but from dorsal internal oblique muscles (DIOMs), a group of transient skeletal muscles that are required for eclosion. Genetic analysis shows that DIOMs secrete ecdysteroids during mTOR-mediated muscle remodeling. These findings identify sequential endocrine and mechanical roles for skeletal muscle, which ensure the timely asymmetric divisions of intestinal stem cells. | Kalodimou, K., Stapountzi, M., Vullings, N., Seib, E., Klein, T., Delidakis, C. (2023). Separable Roles for Neur and Ubiquitin in Delta Signalling in the Drosophila CNS Lineages. Cells, 12(24) PubMed ID: 38132160
Summary: The execution of a Notch signal at the plasma membrane relies on the mechanical force exerted onto Notch by its ligand. It has been appreciated that the DSL ligands need to collaborate with a ubiquitin (Ub) ligase, either Neuralized or Mindbomb1, in order to exert this pulling force, but the role of ubiquitylation per se is uncertain. Regarding the Delta-Neur pair, it is documented that neither the Neur catalytic domain nor the Delta intracellular lysines (putative Ub acceptors) are needed for activity. This study presents a dissection of the Delta activity using the Delta-Notch-dependent expression of Hey in newborn Drosophila neurons as a sensitive in vivo assay. The Delta-Neur interaction per se, rather than ubiquitylation, is needed for activity, pointing to the existence of a Delta-Neur signaling complex. The Neur catalytic domain, although not strictly needed, greatly improves Delta-Neur complex functionality when the Delta lysines are mutated, suggesting that the ubiquitylation of some component of the complex, other than Delta, can enhance signaling. Since Hey expression is sensitive to the perturbation of endocytosis, it is proposed that the Delta-Neur complex triggers a force-generating endocytosis event that activates Notch in the adjacent cell. |
| Messina, O., Raynal, F., Gurgo, J., Fiche, J. B., Pancaldi, V., Nollmann, M. (2023). 3D chromatin interactions involving Drosophila insulators are infrequent but preferential and arise before TADs and transcription. Nat Commun, 14(1):6678 PubMed ID: 37865700
Summary: In mammals, insulators contribute to the regulation of loop extrusion to organize chromatin into topologically associating domains. In Drosophila the role of insulators in 3D genome organization is, however, under current debate. This study addressed this question by combining bioinformatics analysis and multiplexed chromatin imaging. A class of Drosophila insulators enriched at regions forming preferential chromatin interactions genome-wide. Notably, most of these 3D interactions do not involve TAD borders. Multiplexed imaging shows that these interactions occur infrequently, and only rarely involve multiple genomic regions coalescing together in space in single cells. Finally, it was shown that non-border preferential 3D interactions enriched in this class of insulators are present before TADs and transcription during Drosophila development. These results are inconsistent with insulators forming stable hubs in single cells, and instead suggest that they fine-tune existing 3D chromatin interactions, providing an additional regulatory layer for transcriptional regulation (Messina, 2023). | Christensen, C. F., Laurichesse, Q., Loudhaief, R., Colombani, J., Andersen, D. S. (2024). Drosophila activins adapt gut size to food intake and promote regenerative growth. Nat Commun, 15(1):273 PubMed ID: 38177201
Summary: Rapidly renewable tissues adapt different strategies to cope with environmental insults. While tissue repair is associated with increased intestinal stem cell (ISC) proliferation and accelerated tissue turnover rates, reduced calorie intake triggers a homeostasis-breaking process causing adaptive resizing of the gut. This study shows that activins are key drivers of both adaptive and regenerative growth. Activin-β (Actβ) is produced by stem and progenitor cells in response to intestinal infections and stimulates ISC proliferation and turnover rates to promote tissue repair. Dawdle (Daw), a divergent Drosophila activin, signals through its receptor, Baboon, in progenitor cells to promote their maturation into enterocytes (ECs). Daw is dynamically regulated during starvation-refeeding cycles, where it couples nutrient intake with progenitor maturation and adaptive resizing of the gut. These results highlight an activin-dependent mechanism coupling nutrient intake with progenitor-to-EC maturation to promote adaptive resizing of the gut and further establish activins as key regulators of adult tissue plasticity. |
Monday, August 19th - Stress |
| Ok, S., Park, J. E., Byun, S., Kang, K., Son, J., Kang, M. J. (2023). ATF4 Responds to Metabolic Stress in Drosophila. . Frontiers in bioscience (Landmark edition), 28(12):344 PubMed ID: 38179767
Summary: Activating transcription factor 4 (ATF4) is a fundamental basic-leucine zipper transcription factor that plays a pivotal role in numerous stress responses, including endoplasmic reticulum (ER) stress and the integrated stress response. ATF4 regulates adaptive gene expression, thereby triggering stress resistance in cells. To characterize the metabolic status of atf4-/- Drosophila larvae, both metabolomic and microarray analyses were conducted. Metabolomic analysis demonstrated an increase in lactate levels in atf4-/- mutants when compared to wild-type flies. However, there was a significant reduction in adenosine triphosphate (ATP) synthesis in the atf4-/- flies, suggesting an abnormal energy metabolism in the mutant larvae. Microarray analysis unveiled that Drosophila ATF4 controls gene expression related to diverse biological processes, including lipase activity, oxidoreductase activity, acyltransferase, immune response, cell death, and transcription factor, particularly under nutrient-restricted conditions. In situ hybridization analysis further demonstrated specific augmentation of CG6283, classified as a gastric lipase, within the gastric caeca of nutrient-restricted flies. Moreover, overexpression of lipases, CG6283 and CG6295, made the flies resistant to starvation. These findings underscore the role of Drosophila ATF4 in responding to metabolic fluctuations and modulating gene expression associated with metabolism and stress adaptation. Dysregulation of ATF4 may detrimentally impact the development and physiology of Drosophila. | Chen, T., Zhang, M., Ding, Z., Hu, J., Yang, J., He, L., Jia, J., Yang, J., Yang, J., Song, X., Chen, P., Zhai, Z., Huang, J., Wang, Y., Qin, H. (2023). The Drosophila NPY-like system protects against chronic stress-induced learning deficit by preventing the disruption of autophagic flux. Proc Natl Acad Sci U S A, 120(51):e2307632120 PubMed ID: 38079543
Summary: Chronic stress may induce learning and memory deficits that are associated with a depression-like state in Drosophila melanogaster. The molecular and neural mechanisms underlying the etiology of chronic stress-induced learning deficit (CSLD) remain elusive. This study shows that the autophagy-lysosomal pathway, a conserved cellular signaling mechanism, is associated with chronic stress in Drosophila, as indicated by time-series transcriptome profiling. The findings demonstrate that chronic stress induces the disruption of autophagic flux, and chronic disruption of autophagic flux could lead to a learning deficit. Remarkably, preventing the disruption of autophagic flux by up-regulating the basal autophagy level is sufficient to protect against CSLD. Consistent with the essential role of the dopaminergic system in modulating susceptibility to CSLD, dopamine neuronal activity is also indispensable for chronic stress to induce the disruption of autophagic flux. By screening knockout mutants, it was found that neuropeptide F, the Drosophila homolog of neuropeptide Y, is necessary for normal autophagic flux and promotes resilience to CSLD. Moreover, neuropeptide F signaling during chronic stress treatment promotes resilience to CSLD by preventing the disruption of autophagic flux. Importantly, neuropeptide F receptor activity in dopamine neurons also promotes resilience to CSLD. Together, these data elucidate a mechanism by which stress-induced excessive dopaminergic activity precipitates the disruption of autophagic flux, and chronic disruption of autophagic flux leads to CSLD, while inhibitory neuropeptide F signaling to dopamine neurons promotes resilience to CSLD by preventing the disruption of autophagic flux. |
| Wiil, J., Sørensen, J. G., Colinet, H. (2023). Exploring cross-protective effects between cold and immune stress in Drosophila melanogaster. Parasite (Paris, France), 30:54 PubMed ID: 38084935
Summary: It is well established that environmental and biotic stressors like temperature and pathogens/parasites are essential for the life of small ectotherms. There are complex interactions between cold stress and pathogen infection in insects. Possible cross-protective mechanisms occur between both stressors, suggesting broad connectivity in insect stress responses. In this study, the functional significance of these interactions was tested, as well as the potential role of newly uncovered candidate genes, turandot. This was done using an array of factorial experiments exposing Drosophila melanogaster flies to a combination of different cold stress regimes (acute or chronic) and infections with the parasitic fungus Beauveria bassiana. Following these crossed treatments, phenotypic and molecular responses were assessed by measuring 1) induced cold tolerance, 2) immune resistance to parasitic fungus, and 3) activation of turandot genes. Various responses were found in the phenotypic outcomes according to the various treatment combinations with higher susceptibility to infection following cold stress, but also significantly higher acute cold survival in flies that were infected. Regarding molecular responses, overexpression of turandot genes was found in response to most treatments, suggesting reactivity to both cold and infection. Moreover, maximum peak expressions were distinctly observed in the combined treatments (infection plus cold), indicating a marked synergistic effect of the stressors on turandot gene expression patterns. These results reflect the great complexity of cross-tolerance reactions between infection and abiotic stress, but could also shed light on the mechanisms underlying the activation of these responses. | Fanara, J. J., Sassi, P. L., Goenaga, J., Hasson, E. (2023). Genetic basis and repeatability for desiccation resistance in Drosophila melanogaster (Diptera: Drosophilidae). Genetica, PubMed ID: 38102503
Summary: Dehydration is a stress factor for organisms inhabiting natural habitats where water is scarce. Thus, it may be expected that species facing arid environments will develop mechanisms that maximize resistance to desiccation. Insects are excellent models for studying the effects of dehydration as well as the mechanisms and processes that prevent water loss since the effect of desiccation is greater due to the higher area/volume ratio than larger animals. Even though physiological and behavioral mechanisms to cope with desiccation are being understood, the genetic basis underlying the mechanisms related to variation in desiccation resistance and the context-dependent effect remain unsolved. This study analyzed the genetic bases of desiccation resistance in Drosophila melanogaster and identified candidate genes that underlie trait variation. Quantitative genetic analysis of desiccation resistance revealed sexual dimorphism and extensive genetic variation. The phenotype-genotype association analyses (GWAS) identified 71 candidate genes responsible for total phenotypic variation in desiccation resistance. Half of these candidate genes were sex-specific suggesting that the genetic architecture underlying this adaptive trait differs between males and females. Moreover, the public availability of desiccation data analyzed on the same lines but in a different lab allows investigation of the reliability and repeatability of results obtained in independent screens. This survey indicates a pervasive micro-environment lab-dependent effect since no overlap was detected in the sets of genes affecting desiccation resistance identified between labs. |
| Owings, K. G., Chow, C. Y. (2023). A Drosophila screen identifies a role for histone methylation in ER stress preconditioning. G3 (Bethesda), PubMed ID: 38098286
Summary: Stress preconditioning occurs when transient, sublethal stress events impact an organism's ability to counter future stresses. Although preconditioning effects are often noted in the literature, very little is known about the underlying mechanisms. To model preconditioning, a panel of genetically diverse Drosophila melanogaster were exposed to a sublethal heat shock, and how well the flies survived subsequent exposure to endoplasmic reticulum (ER) stress was assessed. The impact of preconditioning varied with genetic background, ranging from dying half as fast to 4 and a half times faster with preconditioning compared to no preconditioning. Subsequent association and transcriptional analyses revealed that histone methylation, and transcriptional regulation are both candidate preconditioning modifier pathways. Strikingly, almost all subunits (7/8) in the Set1/COMPASS complex were identified as candidate modifiers of preconditioning. Functional analysis of Set1 knockdown flies demonstrated that loss of Set1 led to the transcriptional dysregulation of canonical ER stress genes during preconditioning. Based on these analyses, a preconditioning model is proposed in which Set1 helps to establish an interim transcriptional "memory" of previous stress events, resulting in a preconditioned response to subsequent stress. | Shekhar, S., Wert, K. J., Kramer, H. (2023). Visual impairment cell non-autonomously dysregulates brain-wide proteostasis. bioRxiv, PubMed ID: 37961457
Summary: Loss of hearing or vision has been identified as a significant risk factor for dementia but underlying molecular mechanisms are unknown. In different Drosophila models of blindness, Non-autonomous induction of stress granules was found in the brain and their reversal occurred upon restoration of vision. Stress granules include cytosolic condensates of Refractory to sigma P (Ref(2)p or p62), ATF4 and XRP1. This cytosolic restraint of the ATF4 and XRP1 transcription factors dampens expression of their downstream targets during cellular stress. Cytosolic condensates of p62 and ATF4 were also evident in the thalamus and hippocampus of mouse models of congenital or degenerative blindness. These data indicate conservation of the link between loss of sensory input and dysregulation of stress responses critical for protein quality control in the brain. |
Friday, August 16th - Behavior |
| Cooney, P. C., Huang, Y., Li, W., Perera, D. M., Hormigo, R., Tabachnik, T., Godage, I. S., Hillman, E. M. C., Grueber, W. B., Zarin, A. A. (2023). Neuromuscular basis of Drosophila larval rolling escape behavior. Proc Natl Acad Sci U S A, 120(51):e2303641120 PubMed ID: 38096410
Summary: When threatened by dangerous or harmful stimuli, animals engage in diverse forms of rapid escape behaviors. In Drosophila larvae, one type of escape response involves C-shaped bending and lateral rolling followed by rapid forward crawling. The sensory circuitry that promotes larval escape has been extensively characterized; however, the motor programs underlying rolling are unknown. This study characterized the neuromuscular basis of rolling escape behavior. High-speed, volumetric, Swept Confocally Aligned Planar Excitation (SCAPE) microscopy was used to image muscle activity during larval rolling. Unlike sequential peristaltic muscle contractions that progress from segment to segment during forward and backward crawling, muscle activity progresses circumferentially during bending and rolling escape behavior. It is proposed that progression of muscular contraction around the larva's circumference results in a transient misalignment between weight and the ground support forces, which generates a torque that induces stabilizing body rotation. Therefore, successive cycles of slight misalignment followed by reactive aligning rotation lead to continuous rolling motion. Supporting the biomechanical model, it was found that disrupting the activity of muscle groups undergoing circumferential contraction progression leads to rolling defects. EM connectome data was used to identify premotor to motor connectivity patterns that could drive rolling behavior, and neural silencing approaches were performed to demonstrate the crucial role of a group of glutamatergic premotor neurons in rolling. These data reveal body-wide muscle activity patterns and putative premotor circuitry organization for execution of the rolling escape response. | Huang, Y., Zhang, J., You, D., Chen, S., Lin, Z., Li, B., Ling, M., Tong, H., Li, F. (2024). Mechanisms underlying palmitic acid-induced disruption of locomotor activity and sleep behavior in Drosophila. Comparative biochemistry and physiology Toxicology & pharmacology : CBP, 276:109813 PubMed ID: 38070757
Summary: The globally prevalent of sleep disorders is partly attributed to unhealthy dietary habits. This study investigated the underlying mechanisms of elevated palmitic acid (PA) intake on locomotor activity and sleep behavior in Drosophila. The results indicate that exposure to PA significantly elevated Drosophila's daytime and nighttime locomotor activity while concurrently reducing overall sleep duration. Utilizing 16S rRNA sequencing, substantial alterations were observed in the composition of the gut microbiota induced by PA, notably, characterized by a significant reduction in Lactobacillus plantarum. Furthermore, PA significantly increased the levels of inflammatory factors Upd3 and Eiger in Drosophila intestines, and downregulated the expression of Gad and Tph, as well as 5-HT1A. Conversely, Gdh and Hdc were significantly upregulated in the PA group. Supplementation with L. plantarum or lactic acid significantly ameliorated PA-induced disruptions in both locomotor activity and sleep behavior. This supplementation also suppressed the expression of intestinal inflammatory factors, thus restoring impaired neurotransmitter-mediated sleep-wake regulation. Moreover, specific knockdown of intestinal epithelial Upd3 or Eiger similarly restored disrupted neurotransmitter expression, ultimately improving PA-induced disturbances in Drosophila locomotor activity and sleep behavior. These findings provide important insights into the intricate interplay between dietary components and essential behaviors, highlighting potential avenues for addressing health challenges associated with modern dietary habits. |
| Ahn, J. E., Amrein, H. (2023). Opposing chemosensory functions of closely related gustatory receptors. Elife, 12 PubMed ID: 38060294
Summary: In the fruit fly Drosophila melanogaster, gustatory sensory neurons express taste receptors that are tuned to distinct groups of chemicals, thereby activating neural ensembles that elicit either feeding or avoidance behavior. Members of a family of ligand -gated receptor channels, the Gustatory receptors (Grs), play a central role in these behaviors. In general, closely related, evolutionarily conserved Gr proteins are co-expressed in the same type of taste neurons, tuned to chemically related compounds, and therefore triggering the same behavioral response. This study reports that members of the Gr28 subfamily are expressed in largely non-overlapping sets of taste neurons in Drosophila larvae, detect chemicals of different valence, and trigger opposing feeding behaviors. The intrinsic properties of Gr28 neurons were determined by expressing the mammalian Vanilloid Receptor 1 (VR1), which is activated by capsaicin, a chemical to which wild-type Drosophila larvae do not respond. When VR1 is expressed in Gr28a neurons, larvae become attracted to capsaicin, consistent with reports showing that Gr28a itself encodes a receptor for nutritious RNA. In contrast, expression of VR1 in two pairs of Gr28b.c neurons triggers avoidance to capsaicin. Moreover, neuronal inactivation experiments show that the Gr28b.c neurons are necessary for avoidance of several bitter compounds. Lastly, behavioral experiments of Gr28 deficient larvae and live Ca(2+) imaging studies of Gr28b.c neurons revealed that denatonium benzoate, a synthetic bitter compound that shares structural similarities with natural bitter chemicals, is a ligand for a receptor complex containing a Gr28b.c or Gr28b.a subunit. Thus, the Gr28 proteins, which have been evolutionarily conserved over 260 million years in insects, represent the first taste receptor subfamily in which specific members mediate behavior with opposite valence | Stupski, S. D., van Breugel, F. (2023). Wind Gates Search States in Free Flight.. bioRxiv, PubMed ID: 38076971
Summary: For any organism tracking a chemical cue to its source, the motion of its surrounding fluid provides crucial information for success. For both swimming and flying animals engaged in olfactory search, turning into the direction of oncoming wind or water current is often a critical first step. However, in nature, fluid flow may be turbulent or very still, and it is unclear how organisms adjust their search strategies accordingly due to the challenges of separately controlling flow and chemical encounters. This study used the genetic toolkit of Drosophila melanogaster, a model organism for olfaction, to develop an optogenetic paradigm to deliver temporally precise virtual olfactory experiences in free-flying animals while independently manipulating the wind conditions. It was shown that in free flight, Drosophila melanogaster adopt distinct search routines that are gated by whether they are flying in laminar wind or in still air. It was first confirmed that in laminar wind flies turn upwind, and further, it was shown that they achieve this using a rapid ballistic turn. In still air, flies adopt remarkably stereotyped "sink and circle" search state characterized by 60° turns at 3-4 Hz, biased in a consistent direction. In both laminar wind and still air, immediately after odor onset, flies decelerate and often perform a rapid turn. Both maneuvers are consistent with predictions from recent control theoretic analyses for how insects may estimate properties of wind while in flight. It is suggested that flies may use their deceleration and "anemometric" turn as active sensing maneuvers to rapidly gauge properties of their wind environment before initiating a proximal or upwind search routine. |
| Davis, B. A., Mongeau, J. M. (2023). The influence of saccades on yaw gaze stabilization in fly flight. PLoS Comput Biol, 19(12):e1011746 PubMed ID: 38127819
Summary: In a way analogous to human vision, the fruit fly D. melanogaster and many other flying insects generate smooth and saccadic movements to stabilize and shift their gaze in flight, respectively. It has been hypothesized that this combination of continuous and discrete movements benefits both flight stability and performance, particularly at high frequencies or speeds. This study developed a hybrid control system model to explore the effects of saccades on the yaw stabilization reflex of D. melanogaster. Inspired from experimental data, the model includes a first order plant, a Proportional-Integral (PI) continuous controller, and a saccadic reset system that fires based on the integrated error of the continuous controller. The gain, delay and switching threshold parameter space were explored to quantify the optimum regions for yaw stability and performance. The addition of saccades to a continuous controller provides benefits to both stability and performance across a range of frequencies. This model suggests that Drosophila operates near its optimal switching threshold for its experimental gain set. Based on experimental data, D. melanogaster was shown to operate in a region that trades off performance and stability. This trade-off increases flight robustness to compensate for internal perturbations such as wing damage. | Anna, G., John, M., Kannan, N. N. (2023). miR-277 regulates the phase of circadian activity-rest rhythm in Drosophila melanogaster. Frontiers in physiology 14:1082866 PubMed ID: 38089472
Summary: Circadian clocks temporally organize behaviour and physiology of organisms with a rhythmicity of about 24 h. In Drosophila, the circadian clock is composed of mainly four clock genes: period (per), timeless (tim), Clock (Clk) and cycle (cyc) which constitutes the transcription-translation feedback loop. The circadian clock is further regulated via post-transcriptional and post-translational mechanisms among which microRNAs (miRNAs) are well known post-transcriptional regulatory molecules. This study identified and characterized the role of miRNA-277 (miR-277) expressed in the clock neurons in regulating the circadian rhythm. Downregulation of miR-277 in the pacemaker neurons expressing circadian neuropeptide, pigment dispersing factor (PDF) advanced the phase of the morning activity peak under 12 h light: 12 h dark cycles (LD) at lower light intensities and these flies exhibited less robust rhythms compared to the controls under constant darkness. In addition, downregulation of miR-277 in the PDF expressing neurons abolished the Clk gene transcript oscillation under LD. This study points to the potential role of miR-277 in fine tuning the Clk expression and in maintaining the phase of the circadian rhythm in Drosophila. |
Thursday, August 15th - Immune Response |
| Drees, L., Schneider, S., Riedel, D., Schuh, R., Behr, M. (2023). The proteolysis of ZP proteins is essential to control cell membrane structure and integrity of developing tracheal tubes in Drosophila. Elife, 12 PubMed ID: 37872795
Summary: Membrane expansion integrates multiple forces to mediate precise tube growth and network formation. Defects lead to deformations, as found in diseases such as polycystic kidney diseases, aortic aneurysms, stenosis, and tortuosity. This study identified a mechanism of sensing and responding to the membrane-driven expansion of tracheal tubes. The apical membrane is anchored to the apical extracellular matrix (aECM) and causes expansion forces that elongate the tracheal tubes. The aECM provides a mechanical tension that balances the resulting expansion forces, with Dumpy being an elastic molecule that modulates the mechanical stress on the matrix during tracheal tube expansion. The zona pellucida (ZP) domain protein Piopio was shown to interact and cooperates with the ZP protein Dumpy at tracheal cells. To resist shear stresses which arise during tube expansion, Piopio undergoes ectodomain shedding by the Matriptase homolog Notopleural, which releases Piopio-Dumpy-mediated linkages between membranes and extracellular matrix. Failure of this process leads to deformations of the apical membrane, tears the apical matrix, and impairs tubular network function. Conserved ectodomain shedding was shown of the human TGFβ type III receptor by Notopleural and the human Matriptase, providing novel findings for in-depth analysis of diseases caused by cell and tube shape changes. | Hernandez-Lopez, C., Puliafito, A., Xu, Y., Lu, Z., Di Talia, S., Vergassola, M. (2023). Two-fluid dynamics and micron-thin boundary layers shape cytoplasmic flows in early Drosophila embryos. Proc Natl Acad Sci U S A, 120(44):e2302879120 PubMed ID: 37878715
Summary: Cytoplasmic flows are widely emerging as key functional players in development. In early Drosophila embryos, flows drive the spreading of nuclei across the embryo. This study combined hydrodynamic modeling with quantitative imaging to develop a two-fluid model that features an active actomyosin gel and a passive viscous cytosol. Gel contractility is controlled by the cell cycle oscillator, the two fluids being coupled by friction. In addition to recapitulating experimental flow patterns, this model explains observations that remained elusive and makes a series of predictions. First, the model captures the vorticity of cytosolic flows, which highlights deviations from Stokes' flow that were observed experimentally but remained unexplained. Second, the model reveals strong differences in the gel and cytosol motion. In particular, a micron-sized boundary layer is predicted close to the cortex, where the gel slides tangentially while the cytosolic flow cannot slip. Third, the model unveils a mechanism that stabilizes the spreading of nuclei with respect to perturbations of their initial positions. This self-correcting mechanism is argued to be functionally important for proper nuclear spreading. Fourth, this model was used to analyze the effects of flows on the transport of the morphogen Bicoid and the establishment of its gradients. Finally, the model predicts that the flow strength should be reduced if the shape of the domain is more round, which is experimentally confirmed in Drosophila mutants. Thus, this two-fluid model explains flows and nuclear positioning in early Drosophila, while making predictions that suggest novel future experiments. |
| Sun, J., Zhang, C., Gao, F., Stathopoulos, A. (2023). Single-cell transcriptomics illuminates regulatory steps driving anterior-posterior patterning of Drosophila embryonic mesoderm. Cell Rep, 42(10):113289 PubMed ID: 37858470
Summary: Single-cell technologies promise to uncover how transcriptional programs orchestrate complex processes during embryogenesis. This study applied a combination of single-cell technology and genetic analysis to investigate the dynamic transcriptional changes associated with Drosophila embryo morphogenesis at gastrulation. The dataset encompassing the blastoderm-to-gastrula transition provides a comprehensive single-cell map of gene expression across cell lineages validated by genetic analysis. Subclustering and trajectory analyses revealed a surprising stepwise progression in patterning to transition zygotic gene expression and specify germ layers as well as uncovered an early role for ecdysone signaling in epithelial-to-mesenchymal transition in the mesoderm. Multipotent progenitors were shown to arise prior to gastrulation by analyzing the transcription trajectory of caudal mesoderm cells, including a derivative that ultimately incorporates into visceral muscles of the midgut and hindgut. This study provides a rich resource of gastrulation and elucidates spatially regulated temporal transitions of transcription states during the process. | Htet, P. H., Lauga, E. (2023). Cortex-driven cytoplasmic flows in elongated cells: fluid mechanics and application to nuclear transport in Drosophila embryos. Journal of the Royal Society, Interface, 20(208):20230428 PubMed ID: 37963561
Summary: The Drosophila melanogaster embryo, an elongated multi-nucleated cell, is a classical model system for eukaryotic development and morphogenesis. Recent work has shown that bulk cytoplasmic flows, driven by cortical contractions along the walls of the embryo, enable the uniform spreading of nuclei along the anterior-posterior axis necessary for proper embryonic development. This study proposed two mathematical models to characterize cytoplasmic flows driven by tangential cortical contractions in elongated cells. Assuming Newtonian fluid flow at low Reynolds number in a spheroidal cell, the flow field was first computed exactly, thereby bypassing the need for numerical computations. These results were then applied to recent experiments on nuclear transport in cell cycles 4-6 of Drosophila embryo development. By fitting the cortical contractions in this model to measurements, it was revealed that experimental cortical flows enable near-optimal axial spreading of nuclei. A second mathematical approach, applicable to general elongated cell geometries, exploits a long-wavelength approximation to produce an even simpler solution, with errors below compared with the full model. An application of this long-wavelength result to transport leads to fully analytical solutions for the nuclear concentration that capture the essential physics of the system, including optimal axial spreading of nuclei. |
| Popkova, A., Andrensek, U., Pagnotta, S., Ziherl, P., Krajnc, M., Rauzi, M. (2024). A mechanical wave travels along a genetic guide to drive the formation of an epithelial furrow during Drosophila gastrulation. Dev Cell, PubMed ID: 38228140
Summary: Epithelial furrowing is a fundamental morphogenetic process during gastrulation, neurulation, and body shaping. A furrow often results from a fold that propagates along a line. How fold formation and propagation are controlled and driven is poorly understood. To shed light on this, the formation was studied of the cephalic furrow, a fold that runs along the embryo dorsal-ventral axis during Drosophila gastrulation and the developmental role of which is still unknown. Evidence is provided of its function, and this study shows that epithelial furrowing is initiated by a group of cells. This cellular cluster works as a pacemaker, triggering a bidirectional morphogenetic wave powered by actomyosin contractions and sustained by de novo medial apex-to-apex cell adhesion. The pacemaker's Cartesian position is under the crossed control of the anterior-posterior and dorsal-ventral gene patterning systems. Thus, furrow formation is driven by a mechanical trigger wave that travels under the control of a multidimensional genetic guide. | Clarke, D. N., Martin, A. C. (2023). EGFR-dependent actomyosin patterning coordinates morphogenetic movements between tissues. bioRxiv, PubMed ID: 38187543
Summary: The movements that give rise to the body's structure are powered by cell shape changes and rearrangements that are coordinated at supracellular scales. How such cellular coordination arises and integrates different morphogenetic programs is unclear. Using quantitative imaging, =a complex pattern of adherens junction (AJ) levels were found in the ectoderm prior to gastrulation onset in Drosophila. AJ intensity exhibited a double-sided gradient, with peaks at the dorsal midline and ventral neuroectoderm. This dorsal-ventral AJ pattern is regulated by epidermal growth factor (EGF) signaling and that this signal is required for ectoderm cell movement during mesoderm invagination and axis extension. AJ levels and junctional actomyosin were identified as downstream effectors of EGFR signaling. Overall, this study demonstrates a mechanism of coordination between tissue folding and convergent extension that facilitates embryo-wide gastrulation movements. |
Tuesday, August 14th - Evolution |
| Nunez, J. C. B., Lenhart, B. A., Bangerter, A., Murray, C. S., Mazzeo, G. R., Yu, Y., Nystrom, T. L., Tern, C., Erickson, P. A., Bergland, A. O. (2023). A cosmopolitan inversion facilitates seasonal adaptation in overwintering Drosophila. Genetics, PubMed ID: 38051996
Summary: Fluctuations in the strength and direction of natural selection through time are a ubiquitous feature of life on Earth. One evolutionary outcome of such fluctuations is adaptive tracking, wherein populations rapidly adapt from standing genetic variation. In certain circumstances, adaptive tracking can lead to the long-term maintenance of functional polymorphism despite allele frequency change due to selection. Although adaptive tracking is likely a common process, there is still a limited understanding of aspects of its genetic architecture and its strength relative to other evolutionary forces such as drift. Drosophila melanogaster living in temperate regions evolve to track seasonal fluctuations and are an excellent system to tackle these gaps in knowledge. By sequencing orchard populations collected across multiple years, the genomic signal was characterized of seasonal demography, and it was identified that the cosmopolitan inversion In(2L)t facilitates seasonal adaptive tracking and shows molecular footprints of selection. A meta-analysis of phenotypic studies shows that seasonal loci within In(2L)t are associated with behavior, life history, physiology, and morphological traits. Candidate loci were identified and experimentally linked to phenotype. This work contributes to general understanding of fluctuating selection and highlights the evolutionary outcome and dynamics of contemporary selection on inversions. | Ayala, N. M., Genetti, M., Corbett-Detig, R. (2023). Inferring multi-locus selection in admixed populations. PLoS Genet, 19(11):e1011062 PubMed ID: 38015992
Summary: Admixture, the exchange of genetic information between distinct source populations, is thought to be a major source of adaptive genetic variation. Unlike mutation events, which periodically generate single alleles, admixture can introduce many selected alleles simultaneously. As such, the effects of linkage between selected alleles may be especially pronounced in admixed populations. However, existing tools for identifying selected mutations within admixed populations only account for selection at a single site, overlooking phenomena such as linkage among proximal selected alleles. This study developed and extensively validated a method for identifying and quantifying the individual effects of multiple linked selected sites on a chromosome in admixed populations. This approach numerically calculates the expected local ancestry landscape in an admixed population for a given multi-locus selection model, and then maximizes the likelihood of the model. After applying this method to admixed populations of Drosophila melanogaster and Passer italiae, it was found that the impacts between linked sites may be an important contributor to natural selection in admixed populations. Furthermore, for the situations that are considered, the selection coefficients and number of selected sites are overestimated in analyses that do not consider the effects of linkage among selected sites. These results imply that linkage among selected sites may be an important evolutionary force in admixed populations. This tool provides a powerful generalized method to investigate these crucial phenomena in diverse populations. |
| Harris, M., Kim, B., Garud, N. (2023). Enrichment of hard sweeps on the X chromosome compared to autosomes in six Drosophila species. bioRxiv, PubMed ID: 38106201
Summary: The X chromosome, being hemizygous in males, is exposed one third of the time increasing the visibility of new mutations to natural selection, potentially leading to different evolutionary dynamics than autosomes. Recently, an enrichment of hard selective sweeps over soft selective sweeps was found on the X chromosome relative to the autosomes in a North American population of Drosophila melanogaster. To understand whether this enrichment is a universal feature of evolution on the X chromosome, diversity patterns were analyzed across six commonly studied Drosophila species. An increased proportion of regions with steep reductions were found in diversity and elevated homozygosity on the X chromosome compared to autosomes. To assess if these signatures are consistent with positive selection, a wide variety of evolutionary scenarios were simulared spanning variations in demography, mutation rate, recombination rate, background selection, hard sweeps, and soft sweeps, and it was found that the diversity patterns observed on the X are most consistent with hard sweeps. These findings highlight the importance of sex chromosomes in driving evolutionary processes and suggest that hard sweeps have played a significant role in shaping diversity patterns on the X chromosome across multiple Drosophila species. | Ueno, T., Takenoshita, A., Hamamichi, K., Sato, M. P., Takahashi, Y. (2023). Rapid seasonal changes in phenotypes in a wild Drosophila population. Sci Rep, 13(1):21940 PubMed ID: 38114661
Summary: Seasonal environmental change is one of the most rapid and striking environmental variables. Although relatively rapid adaptation to environmental changes over several years or several decades has been described in many taxa, rapid responses to seasonal environments are delicate, and therefore, the detection of the evolutionary responses requires sensitive methods. This study examined seasonal changes in phenotypes related to thermal tolerance and morphological traits of Drosophila lutescens collected at the spring and autumn periods from a single location. It was first demonstrated that flies in the two seasonal periods were almost genetically identical using double-digest restriction site-associated DNA sequencing and analysis. Using an experimental design to eliminate the effect of possible confounding factors that influence phenotypes (i.e., maternal effects and the environmental conditions in which each phenotype was analyzed), it was shown that the heat tolerance of D. lutescens was significantly higher in the autumn population than in the spring population. Furthermore, cold tolerance was slightly higher in the spring population than in the autumn one. Although wing length and thorax length did not change significantly between seasons, the ratio of wing length to thorax length changed significantly between them. These results suggest that seasonal environmental heterogeneity induces rapid phenotypic changes within a year. Finally, the possibility of rapid evolutionary responses to seasonal changes is discusssed. |
| Xiao, C., Duarri-Redondo, S., Thorholludottir, D. A. V., Chen, Y., Schlotterer, C. (2023). Non-additive effects between genotypes: Implications for competitive fitness assays. Ecology and evolution, 13(11):e10713. PubMed ID: 37941737
Summary: Competitive fitness assays are widely used in evolutionary biology and typically rely on a reference strain to compare different focal genotypes. This approach implicitly relies on the absence of interaction between the competing genotypes. In other words, the performance of the reference strain must not depend on the competitor. This report scrutinized this assumption by competing diverged Drosophila simulans populations against a common reference strain. Strong evidence was detected for interaction between the competing genotypes: (1) Frequency-dependent selection was common with opposite effects in genetically diverged populations. (2) Temporal heterogeneity of fitness estimates, which can be partially attributed to a competitor-specific delay in the eclosion of the reference strain. It is proposed that this inconsistent behavior of the reference strain can be considered a specific case of a genotype x environment interaction. Focal populations could modify the environment of the reference strain, either indirectly by altering the microbiome composition and food availability or directly by genotype-specific cannibalism. These results provide new insights into the interaction of diverged genotypes and have important implications for the interpretation of competitive fitness assays. | Grandchamp, A., Czuppon, P., Bornberg-Bauer, E. (2024). Quantification and modeling of turnover dynamics of de novo transcripts in Drosophila melanogaster. Nucleic Acids Res, 52(1):274-287 PubMed ID: 38000384
Summary: Most of the transcribed eukaryotic genomes are composed of non-coding transcripts. Among these transcripts, some are newly transcribed when compared to outgroups and are referred to as de novo transcripts. De novo transcripts have been shown to play a major role in genomic innovations. However, little is known about the rates at which de novo transcripts are gained and lost in individuals of the same species. This study addresses this gap and estimates the de novo transcript turnover rate with an evolutionary model. DNA long reads and RNA short reads from seven geographically remote samples of inbred individuals of Drosophila melanogaster were used to detect de novo transcripts that are gained on a short evolutionary time scale. Overall, each sampled individual contains around 2500 unspliced de novo transcripts, with most of them being sample specific. It is estimated that around 0.15 transcripts are gained per year, and that each gained transcript is lost at a rate around 5× 10-5 per year. This high turnover of transcripts suggests frequent exploration of new genomic sequences within species. These rate estimates are essential to comprehend the process and timescale of de novo gene birth. |
Monday, August 13th - Embryonic Neural Development |
| Singh, B. N., Tran, H., Kramer, J., Kirishenko, E., Changela, N., Wang, F., Feng, Y., Kumar, D., Tu, M., Lan, J., Bizet, M., Fuks, F. and Steward, R. (2023). Tet-dependent 5-hydroxymethyl-Cytosine modification of mRNA regulates the axon guidance genes robo2 and slit in Drosophila. Res Sq. PubMed ID: 36824980
Summary: Modifications of mRNA, especially methylation of adenosine, have recently drawn much attention. The much rarer modification, 5-hydroxymethylation of cytosine (5hmC), is not well understood and is the subject of this study. Vertebrate Tet proteins are 5-methylcytosine (5mC) hydroxylases enzymes catalyzing the transition of 5mC to 5hmC in DNA and have recently been shown to have the same function in messenger RNAs in both vertebrates and in Drosophila. The Tet gene is essential in Drosophila because Tet knock-out animals do not reach adulthood. This study describes the identification of Tet-target genes in the embryo and larval brain by determining Tet DNA-binding sites throughout the genome and by mapping the Tet-dependent 5hmrC modifications transcriptome-wide. 5hmrC-modified sites can be found along the entire transcript and are preferentially located at the promoter where they overlap with histone 4me3 peaks. The identified mRNAs are frequently involved in neuron and axon development and Tet knock-out led to a reduction of 5hmrC marks on specific mRNAs. Among the Tet-target genes were the robo2 receptor and its slit ligand that function in axon guidance in Drosophila and in vertebrates. Tet knock-out embryos show overlapping phenotypes with robo2 and are sensitized to reduced levels of slit. Both Robo2 and Slit protein levels were markedly reduced in Tet KO larval brains. These results establish a role for Tet-dependent 5hmrC in facilitating the translation of modified mRNAs, primarily in developing nerve cells. | Hsiao, Y. L., Chen, H. W., Chen, K. H., Tan, B. C., Chen, C. H. and Pi, H. (2023). Actin-related protein 6 facilitates proneural protein-induced gene activation for rapid neural differentiation. Development 150(5). PubMed ID: 36897355
Summary: Neurogenesis is initiated by basic helix-loop-helix proneural proteins. This study shows that Actin-related protein 6 (Arp6), a core component of the H2A.Z exchange complex SWR1, interacts with proneural proteins and is crucial for efficient onset of proneural protein target gene expression. Arp6 mutants exhibit reduced transcription in sensory organ precursors (SOPs) downstream of the proneural protein patterning event. This leads to retarded differentiation and division of SOPs and smaller sensory organs. These phenotypes are also observed in proneural gene hypomorphic mutants. Proneural protein expression is not reduced in Arp6 mutants. Enhanced proneural gene expression fails to rescue retarded differentiation in Arp6 mutants, suggesting that Arp6 acts downstream of or in parallel with proneural proteins. H2A.Z mutants display Arp6-like retardation in SOPs. Transcriptomic analyses demonstrate that loss of Arp6 and H2A.Z preferentially decreases expression of proneural protein-activated genes. H2A.Z enrichment in nucleosomes around the transcription start site before neurogenesis correlates highly with greater activation of proneural protein target genes by H2A.Z. It is proposed that upon proneural protein binding to E-box sites, H2A.Z incorporation around the transcription start site allows rapid and efficient activation of target genes, promoting rapid neural differentiation. |
| Karkali, K., Vernon, S. W., Baines, R. A., Panayotou, G., Martin-Blanco, E. (2023). Puckered and JNK signaling in pioneer neurons coordinates the motor activity of the Drosophila embryo. Nat Commun, 14(1):8186 PubMed ID: 38081827
Summary: Central nervous system organogenesis is a complex process that obeys precise architectural rules. The impact that nervous system architecture may have on its functionality remains, however, relatively unexplored. To clarify this problem, the development of the Drosophila embryonic Ventral Nerve Cord (VNC) was studied. VNC morphogenesis requires the tight control of Jun kinase (JNK) signaling in a subset of pioneer neurons, exerted in part via a negative feedback loop mediated by the dual specificity phosphatase Puckered. The JNK pathway autonomously regulates neuronal electrophysiological properties without affecting synaptic vesicle transport. Manipulating JNK signaling activity in pioneer neurons during early embryogenesis directly influences their function as organizers of VNC architecture and, moreover, uncovers a role in the coordination of the embryonic motor circuitry that is required for hatching. Together, these data reveal critical links, mediated by the control of the JNK signaling cascade by Puckered, between the structural organization of the VNC and its functional optimization. | Carranza, A., Howard, L. J., Brown, H. E., Ametepe, A. S. and Evans, T. A. (2023). Slit-independent guidance of longitudinal axons by Drosophila Robo3. bioRxiv. PubMed ID: 37214810
Summary: Drosophila Robo3 is a member of the evolutionarily conserved Roundabout (Robo) receptor family and one of three Drosophila Robo paralogs. During embryonic ventral nerve cord development, Robo3 does not participate in canonical Slit-dependent midline repulsion, but instead regulates the formation of longitudinal axon pathways at specific positions along the medial-lateral axis. Longitudinal axon guidance by Robo3 is hypothesized to be Slit dependent, but this has not been directly tested. This study create a series of Robo3 variants in which the N-terminal Ig1 domain is deleted or modified, in order to characterize the functional importance of Ig1 and Slit binding for Robo3's axon guidance activity. Robo3 was shown to require its Ig1 domain for interaction with Slit and for proper axonal localization in embryonic neurons, but deleting Ig1 from Robo3 only partially disrupts longitudinal pathway formation. Robo3 variants with modified Ig1 domains that cannot bind Slit retain proper localization and fully rescue longitudinal axon guidance. These results indicate that Robo3 guides longitudinal axons independently of Slit, and that sequences both within and outside of Ig1 contribute to this Slit-independent activity. |
| Karkali, K., Saunders, T. E., Panayotou, G. and Martin-Blanco, E. (2023). JNK signaling in pioneer neurons organizes ventral nerve cord architecture in Drosophila embryos. Nat Commun 14(1): 675. PubMed ID: 36750572
Summary: Morphogenesis of the Central Nervous System (CNS) is a complex process that obeys precise architectural rules. Yet, the mechanisms dictating these rules remain unknown. Analyzing morphogenesis of the Drosophila embryo Ventral Nerve Cord (VNC), it was observed that a tight control of JNK signaling is essential for attaining the final VNC architecture. JNK signaling in a specific subset of pioneer neurons autonomously regulates the expression of Fasciclin 2 (Fas 2) and Neurexin IV (Nrx IV) adhesion molecules, probably via the transcription factor zfh1. Interfering at any step in this cascade affects fasciculation along pioneer axons, leading to secondary cumulative scaffolding defects during the structural organization of the axonal network. The global disorder of architectural landmarks ultimately influences nervous system condensation. In summary, these data point to JNK signaling in a subset of pioneer neurons as a key element underpinning VNC architecture, revealing critical milestones on the mechanism of control of its structural organization. | Sullivan, K. G., Bashaw, G. J. (2023). Commissureless acts as a substrate adapter in a conserved Nedd4 E3 ubiquitin ligase pathway to promote axon growth across the midline. bioRxiv, PubMed ID: 37905056
Summary: In both vertebrates and invertebrates, commissural neurons prevent premature responsiveness to the midline repellant Slit by downregulating surface levels of its receptor Roundabout1 (Robo1). In Drosophila, Commissureless (Comm) plays a critical role in this process; however, there is conflicting data on the underlying molecular mechanism. This study demonstrates that the conserved PY motifs in the cytoplasmic domain of Comm are required allow the ubiquitination and lysosomal degradation of Robo1. Disruption of these motifs prevents Comm from endosomes and to promote axon growth across the midline in vivo. In addition, this tudy conclusively demonstrates a role for Nedd4 in midline crossing. Genetic analysis shows that nedd4 mutations result in midline crossing defects in the Drosophila embryonic nerve cord, which can be rescued by introduction of exogenous Nedd4. Biochemical evidence shows that Nedd4 incorporates into a three-member complex with Comm and Robo in a PY motif-dependent manner. Finally, genetic evidence is presented that Nedd4 acts with Comm in the embryonic nerve cord to downregulate Robo1 levels. Taken together, these findings demonstrate that Comm promotes midline crossing in the nerve cord by facilitating Robo ubiquitination by Nedd4, ultimately leading to its degradation. |
Friday, August 10th - Enhancers and Transcriptional Regulation |
| Raicu, A. M., Suresh, M., Arnosti, D. N. (2023). A regulatory role for the unstructured C-terminal domain of the CtBP transcriptional corepressor. J Biol Chem, 300(1):105490 PubMed ID: 38000659
Summary: The C-terminal binding protein (CtBP) is a transcriptional corepressor that plays critical roles in development, tumorigenesis, and cell fate. CtBP proteins are structurally similar to alpha hydroxyacid dehydrogenases and feature a prominent intrinsically disordered region in the C terminus. In the mammalian system, CtBP proteins lacking the C-terminal domain (CTD) are able to function as transcriptional regulators and oligomerize, putting into question the significance of this unstructured domain for gene regulation. Yet, the presence of an unstructured CTD of ∼100 residues, including some short motifs, is conserved across Bilateria, indicating the importance of maintaining this domain over evolutionary time. To uncover the significance of the CtBP CTD, naturally occurring Drosophila isoforms of CtBP that possess or lack the CTD were functionally tested, namely CtBP(L) and CtBP(S). The CRISPRi system was used to recruit dCas9-CtBP(L) and dCas9-CtBP(S) to endogenous promoters to directly compare their transcriptional impacts in vivo. Interestingly, CtBP(S) was able to significantly repress transcription of the Mpp6 promoter, while CtBP(L) was much weaker, suggesting that the long CTD may modulate CtBP's repression activity. In contrast, in cell culture, the isoforms behaved similarly on a transfected Mpp6 reporter gene. The context-specific differences in activity of these two developmentally regulated isoforms suggests that the CTD may help provide a spectrum of repression activity suitable for developmental programs. | Blum, J. A., Wells, M., Huxley-Reicher, Z., Johnson, J. E., Bateman, J. R. (2023). Transvection between non-allelic genomic positions in Drosophila. G3 (Bethesda), PubMed ID: 37949840
Summary: In Drosophila, pairing of maternal and paternal homologous chromosomes can permit trans-interactions between enhancers on one homolog and promoters on another, an example of transvection. Although trans-interactions have been observed at many loci in the Drosophila genome and in other organisms, the parameters that govern enhancer action in trans remain poorly understood. Using a transgenic reporter system, this study asked whether enhancers and promoters at non-allelic, but nearby, genomic positions can communication in trans. Using one transgenic insertion carrying the synthetic enhancer GMR and another nearby insertion carrying the hsp70 promoter driving a fluorescent reporter, it was shown that transgenes separated by 2.6 kb of linear distance can support enhancer action in trans at the 53F8 locus. Furthermore, transvection between the non-allelic insertions can be augmented by a small deletion flanking one insert, likely via changes to the paired configuration of the homologs. Subsequent analyses of other insertions in 53F8 that carry different transgenic sequences demonstrate that the capacity to support transvection between non-allelic sites varies greatly, suggesting that factors beyond the linear distance between insertion sites play an important role. Finally, analysis of transvection between nearby non-allelic sites at other genomic locations shows evidence of position effects, where one locus supported GMR action in trans over a linear distance of over 10 kb, whereas another locus showed no evidence of transvection over a span less than 200 bp. Overall, these data demonstrate that transvection between non-allelic sites represents a complex interplay between genomic context, interallelic distance, and promoter identity. |
| Bishop, T. R., Onal, P., Xu, Z., Zheng, M., Gunasinghe, H., Nien, C. Y., Small, S., Datta, R. R. (2023). Multi-level regulation of even-skipped stripes by the ubiquitous factor Zelda. Development, 150(23) PubMed ID: 37934130
Summary: The zinc-finger protein Zelda (Zld) is a key activator of zygotic transcription in early Drosophila embryos. Zld-dependent regulation was studied of the seven-striped pattern of the pair-rule gene even-skipped (eve). Individual stripes are regulated by discrete enhancers that respond to broadly distributed activators; stripe boundaries are formed by localized repressors encoded by the gap genes. The strongest effects of Zld are on stripes 2, 3 and 7, which are regulated by two enhancers in a 3.8 kb genomic fragment that includes the eve basal promoter. Zld facilitates binding of the activator Bicoid and the gap repressors to this fragment, consistent with its proposed role as a pioneer protein. To test whether the effects of Zld are direct, all canonical Zld sites in the 3.8 kb fragment were mutated, which reduced expression but failed to phenocopy the abolishment of stripes caused by removing Zld in trans. Zld also indirectly regulates the eve stripes by establishing specific gap gene expression boundaries, which provides the embryonic spacing required for proper stripe activation. | Jiang, Y., Chiu, T. P., Mitra, R., Rohs, R. (2024). Probing the role of the protonation state of a minor groove-linker histidine in Exd-Hox-DNA binding. Biophys J, 123(2):248-259 PubMed ID: 38130056
Summary: DNA recognition and targeting by transcription factors (TFs) through specific binding are fundamental in biological processes. Furthermore, the histidine protonation state at the TF-DNA binding interface can significantly influence the binding mechanism of TF-DNA complexes. Nevertheless, the role of histidine in TF-DNA complexes remains underexplored. This study employed all-atom molecular dynamics simulations using AlphaFold2-modeled complexes based on previously solved co-crystal structures to probe the role of the His-12 residue in the Extradenticle (Exd)-Sex combs reduced (Scr)-DNA complex when binding to Scr and Ultrabithorax (Ubx) target sites. These results demonstrate that the protonation state of histidine notably affected the DNA minor-groove width profile and binding free energy. Examining flanking sequences of various binding affinities derived from SELEX-seq experiments, the relationship between binding affinity and specificity was studied. How histidine protonation leads to increased binding affinity but can lower specificity was uncovered. These findings provide new mechanistic insights into the role of histidine in modulating TF-DNA binding. |
| Fujiwara, R., Zhai, S. N., Liang, D., Shah, A. P., Tracey, M., Ma, X. K., Fields, C. J., Mendoza-Figueroa, M. S., Meline, M. C., Tatomer, D. C., Yang, L., Wilusz, J. E. (2023). IntS6 and the Integrator phosphatase module tune the efficiency of select premature transcription termination events. Mol Cell, 83(24):4445-4460. PubMed ID: 37995689
Summary: The metazoan-specific Integrator complex catalyzes 3' end processing of small nuclear RNAs (snRNAs) and premature termination that attenuates the transcription of many protein-coding genes. Integrator has RNA endonuclease and protein phosphatase activities, but it remains unclear if both are required for complex function. This study shows that IntS6 (Integrator subunit 6) over-expression blocks Integrator function at a subset of Drosophila protein-coding genes, although having no effect on snRNAs or attenuation of other loci. Over-expressed IntS6 titrates protein phosphatase 2A (PP2A) subunits, thereby only affecting gene loci where phosphatase activity is necessary for Integrator function. IntS6 functions analogous to a PP2A regulatory B subunit as over-expression of canonical B subunits, which do not bind Integrator, is also sufficient to inhibit Integrator activity. These results show that the phosphatase module is critical at only a subset of Integrator-regulated genes and point to PP2A recruitment as a tunable step that modulates transcription termination efficiency. | Balasubramanian, D., Borges Pinto, P., Grasso, A., Vincent, S., Tarayre, H., Lajoignie, D., Ghavi-Helm, Y. (2023). Enhancer-promoter interactions can form independently of genomic distance and be functional across TAD boundaries. Nucleic Acids Res, PubMed ID: 38084924
Summary: Topologically Associating Domains (TADs) have been suggested to facilitate and constrain enhancer-promoter interactions. However, the role of TAD boundaries in effectively restricting these interactions remains unclear. This study shows that a significant proportion of enhancer-promoter interactions are established across TAD boundaries in Drosophila embryos, but that developmental genes are strikingly enriched in intra- but not inter-TAD interactions. This observation was pursued using the twist locus, a master regulator of mesoderm development, and systematically relocated one of its enhancers to various genomic locations. While this developmental gene can establish inter-TAD interactions with its enhancer, the functionality of these interactions remains limited, highlighting the existence of topological constraints. Furthermore, contrary to intra-TAD interactions, the formation of inter-TAD enhancer-promoter interactions is not solely driven by genomic distance, with distal interactions sometimes favored over proximal ones. These observations suggest that other general mechanisms must exist to establish and maintain specific enhancer-promoter interactions across large distances. |
Thursday, August 9th - Larval and Adult Neural Development, Structure, and Function |
| Zboray, K., Toth, A. V., Miskolczi, T. D., Pesti, K., Casanova, E., Kreidl, E., Mike, A., Szenes, Á., Sagi, L., Lukacs, P. (2023). High-throughput ligand profile characterization in novel cell lines expressing seven heterologous insect olfactory receptors for the detection of volatile plant biomarkers. Sci Rep, 13(1):21757 PubMed ID: 38066004
Summary: Agriculturally important crop plants emit a multitude of volatile organic compounds (VOCs), which are excellent indicators of their health status and their interactions with pathogens and pests. This study has developed a novel cellular olfactory panel for detecting fungal pathogen-related VOCs identified in the field, as well as during controlled inoculations of several crop plants. The olfactory panel consists of seven stable HEK293 cell lines each expressing a functional Drosophila olfactory receptor as a biosensing element along with GCaMP6, a fluorescent calcium indicator protein. An automated 384-well microplate reader was used to characterize the olfactory receptor cell lines for their sensitivity to reference VOCs. Subsequently, a set of 66 VOCs were profiled on all cell lines, covering a concentration range from 1 to 100 &mi;M. Results showed that 49 VOCs (74.2%) elicited a response in at least one olfactory receptor cell line. Some VOCs activated the cell lines even at nanomolar (ppb) concentrations. The interaction profiles obtained in this study will support the development of biosensors for agricultural applications. Additionally, the olfactory receptor proteins can be purified from these cell lines with sufficient yields for further processing, such as structure determination or integration with sensor devices. | Nakato, E., Kamimura, K., Knudsen, C., Masutani, S., Takemura, M., Hayashi, Y., Akiyama, T., Nakato, H. (2023). Differential heparan sulfate dependency of the Drosophila glypicans. J Biol Chem, 300(1):105544 PubMed ID: 3807ƒ2044
Summary: Heparan sulfate proteoglycans (HSPGs) are composed of a core protein and glycosaminoglycan (GAG) chains and serve as coreceptors for many growth factors and morphogens. To understand the molecular mechanisms by which HSPGs regulate morphogen gradient formation and signaling, it is important to determine the relative contributions of the carbohydrate and protein moieties to the proteoglycan function. To address this question, ΔGAG alleles were generated for dally and dally-like protein (dlp), two Drosophila HSPGs of the glypican family, in which all GAG-attachment serine residues are substituted to alanine residues using CRISPR/Cas9 mutagenesis. In these alleles, the glypican core proteins are expressed from the endogenous loci with no GAG modification. Analyses of the dally(Δ GAG) allele defined Dally functions that do not require heparan sulfate (HS) chains and that need both core protein and HS chains. A new, dally(ΔGAG)-specific phenotype was discovered, the formation of a posterior ectopic vein, which has never been seen in the null mutants. Unlike dally(ΔGAG), dlp(ΔGAG) mutants do not show most of the dlp null mutant phenotypes, suggesting that HS chains are dispensable for these dlp functions. As an exception, HS is essentially required for Dlp's activity at the neuromuscular junction. Thus, Drosophila glypicans show strikingly different levels of HS dependency. The ΔGAG mutant alleles of the glypicans serve as new molecular genetic toolsets highly useful to address important biological questions, such as molecular mechanisms of morphogen gradient formation. |
| Zhou, F., Tichy, A. M., Imambocus, B. N., Sakharwade, S., Rodriguez Jimenez, F. J., Gonzalez Martínez, M., Jahan, I., Habib, M., Wilhelmy, N., Burre, V., Lomker, T., Sauter, K., Helfrich-Forster, C., Pielage, J., Grunwald Kadow, I. C., Janovjak, H., Soba, P. (2023). Optimized design and in vivo application of optogenetically functionalized Drosophila dopamine receptors. Nat Commun, 14(1):8434 PubMed ID: 38114457
Summary: Neuromodulatory signaling via G protein-coupled receptors (GPCRs) plays a pivotal role in regulating neural network function and animal behavior. The recent development of optogenetic tools to induce G protein-mediated signaling provides the promise of acute and cell type-specific manipulation of neuromodulatory signals. However, designing and deploying optogenetically functionalized GPCRs (optoXRs) with accurate specificity and activity to mimic endogenous signaling in vivo remains challenging. This study optimized the design of optoXRs by considering evolutionary conserved GPCR-G protein interactions and demonstrates the feasibility of this approach using two Drosophila Dopamine receptors (optoDopRs). These optoDopRs exhibit high signaling specificity and light sensitivity in vitro. In vivo, receptor and cell type-specific effects of dopaminergic signaling in various behaviors, including the ability of optoDopRs to rescue the loss of the endogenous receptors. This work demonstrates that optoXRs can enable optical control of neuromodulatory receptor-specific signaling in functional and behavioral studies. | Yang, J., Chen, X., Jin, S., Ding, J. (2023). Structure and biochemical characterization of l-2-hydroxyglutarate dehydrogenase and its role in the pathogenesis of l-2-hydroxyglutaric aciduria. J Biol Chem, 300(1):105491 PubMed ID: 37995940
Summary: l-2-hydroxyglutarate dehydrogenase (L2HGDH) is a mitochondrial membrane-associated metabolic enzyme, which catalyzes the oxidation of l-2-hydroxyglutarate (l-2-HG) to 2-oxoglutarate (2-OG). Mutations in human L2HGDH lead to abnormal accumulation of l-2-HG, which causes a neurometabolic disorder named l-2-hydroxyglutaric aciduria (l-2-HGA). This study reports the crystal structures of Drosophila melanogaster L2HGDH (dmL2HGDH) in FAD-bound form and in complex with FAD and 2-OG and show that dmL2HGDH exhibits high activity and substrate specificity for l-2-HG. dmL2HGDH consists of an FAD-binding domain and a substrate-binding domain, and the active site is located at the interface of the two domains with 2-OG binding to the re-face of the isoalloxazine moiety of FAD. Mutagenesis and activity assay confirmed the functional roles of key residues involved in the substrate binding and catalytic reaction and showed that most of the mutations of dmL2HGDH equivalent to l-2-HGA-associated mutations of human L2HGDH led to complete loss of the activity. The structural and biochemical data together reveal the molecular basis for the substrate specificity and catalytic mechanism of L2HGDH and provide insights into the functional roles of human L2HGDH mutations in the pathogeneses of l-2-HGA. |
| Pacalon, J., Audic, G., Magnat, J., Philip, M., Golebiowski, J., Moreau, C. J., Topin, J. (2023). Elucidation of the structural basis for ligand binding and translocation in conserved insect odorant receptor co-receptors. Nat Commun, 14(1):8182 PubMed ID: 38081900
Summary: In numerous insects, the olfactory receptor family forms a unique class of heteromeric cation channels. Recent progress in resolving the odorant receptor structures offers unprecedented opportunities for deciphering their molecular mechanisms of ligand recognition. Unexpectedly, these structures in apo or ligand-bound states did not reveal the pathway taken by the ligands between the extracellular space and the deep internal cavities. By combining molecular modeling with electrophysiological recordings, amino acids were identified involved in the dynamic entry pathway and the binding of VUAA1 to Drosophila melanogaster's odorant receptor co-receptor (Orco). The results provide evidence for the exact location of the agonist binding site and a detailed and original mechanism of ligand translocation controlled by a network of conserved residues. These findings would explain the particularly high selectivity of Orcos for their ligands. | Ramya, R., Shyamala, V. B. (2023). olf413, a putative octopamine biosynthesis pathway gene is required for negative geotactic motor function in Drosophila melanogaster. Journal of genetics, 102 PubMed ID: 38073169
Summary: Olf413 gene annotated as CG12673 in the genome of Drosophila melanogaster has been predicted to code for a protein with putative function in octopamine biosynthesis. The expression pattern and the biological function of the gene awaits investigation. The present study is the first report where its expression in the late pupal brain is described as depicted by enhancer-driven green fluorescent protein (GFP) expression. With experiments using loss of function olf413 mutant flies, it was demonstrated that olf413 function is essential for motor activity, gravity sensing and spatial balancing of the body against gravity during climbing. |
Wednesday, August 8th - Larval and Adult Neural Development, Structure, and Function |
| Garner, D., Kind, E., Nern, A., Houghton, L., Zhao, A., Sancer, G., Rubin, G. M., Wernet, M. F., Kim, S. S. (2023). Connectomic reconstruction predicts the functional organization of visual inputs to the navigation center of the Drosophila brain. bioRxiv, PubMed ID: 38076786
Summary: Many animals, including humans, navigate their surroundings by visual input, yet little is understood about how visual information is transformed and integrated by the navigation system. In Drosophila melanogaster, compass neurons in the donut-shaped ellipsoid body of the central complex generate a sense of direction by integrating visual input from ring neurons, a part of the anterior visual pathway (AVP). This study densely reconstruct all neurons in the AVP using FlyWire, an AI-assisted tool for analyzing electron-microscopy data. The AVP comprises four neuropils, sequentially linked by three major classes of neurons: MeTu neurons, which connect the medulla in the optic lobe to the small unit of anterior optic tubercle (AOTUsu) in the central brain; TuBu neurons, which connect the anterior optic tubercle to the bulb neuropil; and ring neurons, which connect the bulb to the ellipsoid body. Based on neuronal morphologies, connectivity between different neural classes, and the locations of synapses, this study identified non-overlapping channels originating from four types of MeTu neurons, which were further divided into ten subtypes based on the presynaptic connections in medulla and postsynaptic connections in AOTUsu. To gain an objective measure of the natural variation within the pathway, the differences were quantified between anterior visual pathways from both hemispheres and between two electron-microscopy datasets. Furthermore, potential visual features and the visual area were infered from which any given ring neuron receives input by combining the connectivity of the entire AVP, the MeTu neurons' dendritic fields, and presynaptic connectivity in the optic lobes. These results provide a strong foundation for understanding how distinct visual features are extracted and transformed across multiple processing stages to provide critical information for computing the fly's sense of direction. | Ali, M. Z., Anushree, Ahsan, A., Ola, M. S., Haque, R., Ahsan, J. (2023). Ionotropic receptors mediate olfactory learning and memory in Drosophila. Insect Sci, PubMed ID: 38114448
Summary: Phenylacetaldehyde (PAH), an aromatic compound, is present in a diverse range of fruits including overripe bananas and prickly pear cactus, the two major host fruits for Drosophila melanogaster. PAH acts as a potent ligand for the ionotropic receptor 84a (IR84a) in the adult fruit fly and it is detected by the IR84a/IR8a heterotetrameric complex. Its role in the male courtship behavior through IR84a as an environmental aphrodisiac is of additional importance. In D. melanogaster, two distinct kinds of olfactory receptors, that is, odorant receptors (ORs) and ionotropic receptors (IRs), perceive the odorant stimuli. They display unique structural, molecular, and functional characteristics in addition to having different evolutionary origins. Traditionally, olfactory cues detected by the ORs such as ethyl acetate, 1-butanol, isoamyl acetate, 1-octanol, 4-methylcyclohexanol, etc. classified as aliphatic esters and alcohols have been employed in olfactory classical conditioning using fruit flies. This underlines the participation of OR-activated olfactory pathways in learning and memory formation. This study elucidates that likewise ethyl acetate (EA) (an OR-responsive odorant), PAH (an IR-responsive aromatic compound) too can form learning and memory when associated with an appetitive gustatory reinforcer. The association of PAH with sucrose (PAH/SUC) led to learning and formation of the long-term memory (LTM). Additionally, the Orco(1) , Ir84a(MI00501) , and Ir8a(1) mutant flies were used to confirm the exclusive participation of the IR84a/IR8a complex in PAH/SUC olfactory associative conditioning. These results highlight the involvement of IRs via an IR-activated pathway in facilitating robust olfactory behavior. |
| Brown, E. B., Zhang, J., Lloyd, E., Lanzon, E., Botero, V., Tomchik, S., Keene, A. C. (2023). Neurofibromin 1 mediates sleep depth in Drosophila. PLoS Genet, 19(12):e1011049 PubMed ID: 38091360
Summary: Neural regulation of sleep and metabolic homeostasis are critical in many aspects of human health. Despite extensive epidemiological evidence linking sleep dysregulation with obesity, diabetes, and metabolic syndrome, little is known about the neural and molecular basis for the integration of sleep and metabolic function. The RAS GTPase-activating gene Neurofibromin (Nf1) has been implicated in the regulation of sleep and metabolic rate, raising the possibility that it serves to integrate these processes, but the effects on sleep consolidation and physiology remain poorly understood. A key hallmark of sleep depth in mammals and flies is a reduction in metabolic rate during sleep. This study examined multiple measures of sleep quality to determine the effects of Nf1 on sleep-dependent changes in arousal threshold and metabolic rate. Flies lacking Nf1 fail to suppress metabolic rate during sleep, raising the possibility that loss of Nf1 prevents flies from integrating sleep and metabolic state. Sleep of Nf1 mutant flies is fragmented with a reduced arousal threshold in Nf1 mutants, suggesting Nf1 flies fail to enter deep sleep. The effects of Nf1 on sleep can be localized to a subset of neurons expressing the GABAA receptor Rdl. Sleep loss has been associated with changes in gut homeostasis in flies and mammals. Selective knockdown of Nf1 in Rdl-expressing neurons within the nervous system increases gut permeability and reactive oxygen species (ROS) in the gut, raising the possibility that loss of sleep quality contributes to gut dysregulation. Together, these findings suggest Nf1 acts in GABA-sensitive neurons to modulate sleep depth in Drosophila. | Oliveira-Ferreira, C., Gaspar, M., Vasconcelos, M. L. (2023). Neuronal substrates of egg-laying behaviour at the abdominal ganglion of Drosophila melanogaster. Sci Rep, 13(1):21941 PubMed ID: 38081887
Summary: Egg-laying in Drosophila is the product of post-mating physiological and behavioural changes that culminate in a stereotyped sequence of actions. Egg-laying harbours a great potential as a paradigm to uncover how the appropriate motor circuits are organized and activated to generate behaviour. To study this programme, this study first describes the different phases of the egg-laying programme and the specific actions associated with each phase. Using a combination of neuronal activation and silencing experiments, neurons (OvAbg) in the abdominal ganglion were identified as key players in egg-laying. To generate and functionally characterise subsets of OvAbg, an intersectional approach was used with neurotransmitter specific lines-VGlut, Cha and Gad1. OvAbg/VGlut neurons were shown to promote initiation of egg deposition in a mating status dependent way. OvAbg/Cha neurons are required in exploration and egg deposition phases, though activation leads specifically to egg expulsion. Experiments with the OvAbg/Gad1 neurons show they participate in egg deposition. A functional connection of OvAbg neurons with brain neurons was shown. This study provides insight into the organization of neuronal circuits underlying complex motor behaviour. |
| Ishida, I. G., Sethi, S., Mohren, T. L., Abbott, L. F., Maimon, G. (2023)s. Neuronal calcium spikes enable vector inversion in the Drosophila brain. bioRxiv, PubMed ID: 38077032
Summary: A typical neuron signals to downstream cells when it is depolarized and firing sodium spikes. Some neurons, however, also fire calcium spikes when hyperpolarized. The function of such bidirectional signaling remains unclear in most circuits. This study shows how a neuron class that participates in vector computation in the fly central complex employs hyperpolarization-elicited calcium spikes to invert two-dimensional mathematical vectors. When cells switch from firing sodium to calcium spikes, this leads to a ~180° realignment between the vector encoded in the neuronal population and the fly's internal heading signal, thus inverting the vector. The calcium spikes were shown to rely on the T-type calcium channel Ca-α1T, and it is argued, via analytical and experimental approaches, that these spikes enable vector computations in portions of angular space that would otherwise be inaccessible. These results reveal a seamless interaction between molecular, cellular and circuit properties for implementing vector math in the brain. | Miyashita, T., Murakami, K., Kikuchi, E., Ofusa, K., Mikami, K., Endo, K., Miyaji, T., Moriyama, S., Konno, K., Muratani, H., Moriyama, Y., Watanabe, M., Horiuchi, J., Saitoe, M. (2023). Glia transmit negative valence information during aversive learning in Drosophila. Science, 382(6677):eadf7429 PubMed ID: 38127757
Summary: During Drosophila aversive olfactory conditioning, aversive shock information needs to be transmitted to the mushroom bodies (MBs) to associate with odor information. This study reports that aversive information is transmitted by ensheathing glia (EG) that surround the MBs. Shock induces vesicular exocytosis of glutamate from EG. Blocking exocytosis impairs aversive learning, whereas activation of EG can replace aversive stimuli during conditioning. Glutamate released from EG binds to N-methyl-d-aspartate receptors in the MBs, but because of Mg(2+) block, Ca(2+) influx occurs only when flies are simultaneously exposed to an odor. Vesicular exocytosis from EG also induces shock-associated dopamine release, which plays a role in preventing formation of inappropriate associations. These results demonstrate that vesicular glutamate released from EG transmits negative valence information required for associative learning. |
Tuesday, August 7th - Cytoskeleton and Junctions |
| Bremer, K. V., Wu, C., Patel, A. A., He, K. L., Grunfeld, A. M., Chanfreau, G. F., Quinlan, M. E. (2023). Formin tails act as a switch, inhibiting or enhancing processive actin elongation. J Biol Chem, 300(1):105557 PubMed ID: 38097186
Summary: Formins are large, multidomain proteins that nucleate new actin filaments and accelerate elongation through a processive interaction with the barbed ends of filaments. Their actin assembly activity is generally attributed to their eponymous formin homology (FH) 1 and 2 domains; however, evidence is mounting that regions outside of the FH1FH2 stretch also tune actin assembly. This study explored the underlying contributions of the tail domain, which spans the sequence between the FH2 domain and the C terminus of formins. Tails vary in length from ∼0 to >200 residues and contain a number of recognizable motifs. The most common and well-studied motif is the ∼15-residue-long diaphanous autoregulatory domain. This domain mediates all or nothing regulation of actin assembly through an intramolecular interaction with the diaphanous inhibitory domain in the N-terminal half of the protein. Multiple reports demonstrate that the tail can enhance both nucleation and processivity. This study provides a high-resolution view of the alternative splicing encompassing the tail in the formin homology domain (Fhod) family of formins during development. While four distinct tails are predicted, this study found significant levels of only two of these. The biochemical effects of the different tails were characterized. Surprisingly, the two highly expressed Fhod-tails inhibit processive elongation and diminish nucleation, while a third supports activity. These findings demonstrate a new mechanism of modulating actin assembly by formins and support a model in which splice variants are specialized to build distinct actin structures during development. | Claussen, N. H., Brauns, F., Shraiman, B. I. (2023). A Geometric Tension Dynamics Model of Epithelial Convergent Extension. ArXiv, PubMed ID: 38076522
Summary: Epithelial tissue elongation by convergent extension is a key motif of animal morphogenesis. On a coarse scale, cell motion resembles laminar fluid flow; yet in contrast to a fluid, epithelial cells adhere to each other and maintain the tissue layer (Tl) under actively generated internal tension. To resolve this apparent paradox, a model was formulated in which tissue flow occurs through adiabatic remodelling of the cellular force balance causing local cell rearrangement. It is proposed that the gradual shifting of the force balance is caused by positive feedback on myosin-generated cytoskeletal tension. Shifting force balance within a tension network causes active T1s oriented by the global anisotropy of tension. Rigidity of cells against shape changes converts the oriented internal rearrangements into net tissue deformation. Strikingly, it was found that the total amount of tissue extension depends on the initial magnitude of anisotropy and on cellular packing order. T1s degrade this order so that tissue flow is self-limiting. These findings are explained by showing that coordination of T1s depends on coherence in local tension configurations, quantified by a certain order parameter in tension space. This model reproduces the salient tissue- and cell-scale features of germ band elongation during Drosophila gastrulation, in particular the slowdown of tissue flow after approximately twofold extension concomitant with a loss of order in tension configurations. This suggests local cell geometry contains morphogenetic information and yields predictions testable in future experiments. Furthermore, this focus on defining biologically controlled active tension dynamics on the manifold of force-balanced states may provide a general approach to the description of morphogenetic flow. |
| Clements, R., Smith, T., Cowart, L., Zhumi, J., Sherrod, A., Cahill, A., Hunter, G. L. (2024). Myosin XV is a negative regulator of signaling filopodia during long-range lateral inhibition. Dev Biol, 505:110-121 PubMed ID: 37956923
Summary: The self-organization of cells during development is essential for the formation of healthy tissues and requires the coordination of cell activities at local scales. Cytonemes, or signaling filopodia, are dynamic actin-based cellular protrusions that allow cells to engage in contact mediated signaling at a distance. While signaling filopodia have been shown to support several signaling paradigms during development, less is understood about how these protrusions are regulated. This study investigated the role of the plus-end directed, unconventional MyTH4-FERM myosins in regulating signaling filopodia during sensory bristle patterning on the dorsal thorax of the fruit fly Drosophila melanogaster. Myosin XV is required for regulating signaling filopodia dynamics and, as a consequence, lateral inhibition more broadly throughout the patterning epithelium. Myosin XV was found to be required for limiting the length and number of signaling filopodia generated by bristle precursor cells. Cells with additional and longer signaling filopodia due to loss of Myosin XV are not signaling competent, due to altered levels of Delta ligand and Notch receptor along their lengths. It is concluded that Myosin XV acts to negatively regulate signaling filopodia, as well as promote the ability of signaling filopodia to engage in long-range Notch signaling. Since Myosin XV isoforms are present across several vertebrate and invertebrate systems, this may have significance for other long-range signaling mechanisms. | Ermanoska, B., Rodal, A. A. (2023). Non-muscle myosin II regulates presynaptic actin assemblies and neuronal mechanobiology. bioRxiv, PubMed ID: 38014140
Summary: Neuromuscular junctions (NMJs) are evolutionarily ancient, specialized contacts between neurons and muscles. Axons and NMJs must endure mechanical strain through a lifetime of muscle contraction, making them vulnerable to aging and neurodegenerative conditions. However, cellular strategies for mitigating this mechanical stress remain unknown. In this study, Drosophila larval NMJs were used to investigate the role of actin and myosin (actomyosin)-mediated contractility in generating and responding to cellular forces at the neuron-muscle interface. A new long-lived, low-turnover presynaptic actin core traversing the NMJ was identified, that partly co-localizes with non-muscle myosin II (NMII). Neuronal RNAi of NMII induced disorganization of this core, suggesting that this structure might have contractile properties. Interestingly, neuronal RNAi of NMII also decreased NMII levels in the postsynaptic muscle proximal to neurons, suggesting that neuronal actomyosin rearrangements propagate their effects transsynaptically. Reduced Integrin levels were also observed upon NMII knockdown, indicating that neuronal actomyosin disruption triggers rearrangements of Integrin-mediated connections between neurons and surrounding muscle tissue. In summary, this study identifies a previously uncharacterized presynaptic actomyosin subpopulation that upholds the neuronal mechanical continuum, transmits signals to adjacent muscle tissue, and collaborates with Integrin receptors to govern the mechanobiology of the neuromuscular junction. |
| Collinet, C., Bailles, A., Dehapiot, B., Lecuit, T. (2024). Mechanical regulation of substrate adhesion and de-adhesion drives a cell-contractile wave during Drosophila tissue morphogenesis. Dev Cell, 59(1):156-172.e157 PubMed ID: 38103554
Summary: During morphogenesis, mechanical forces induce large-scale deformations; yet, how forces emerge from cellular contractility and adhesion is unclear. In Drosophila embryos, a tissue-scale wave of actomyosin contractility coupled with adhesion to the surrounding vitelline membrane drives polarized tissue invagination. This process emerges subcellularly from the mechanical coupling between myosin II activation and sequential adhesion/de-adhesion to the vitelline membrane. At the wavefront, integrin clusters anchor the actin cortex to the vitelline membrane and promote activation of myosin II, which in turn enhances adhesion in a positive feedback. Following cell detachment, cortex contraction and advective flow amplify myosin II. Prolonged contact with the vitelline membrane prolongs the integrin-myosin II feedback, increases integrin adhesion, and thus slows down cell detachment and wave propagation. The angle of cell detachment depends on adhesion strength and sets the tensile forces required for detachment. Thus, this study documents how the interplay between subcellular mechanochemical feedback and geometry drives tissue morphogenesis. | Stjepic, V., Nakamura, M., Hui, J., Parkhurst, S. M. (2023). Two Septin Complexes Mediate Actin Dynamics During Cell Wound Repair. bioRxiv, PubMed ID: 38014090
Summary: Cells have robust wound repair systems to prevent further damage or infection and to quickly restore cell cortex integrity when exposed to mechanical and chemical stress. Actomyosin ring formation and contraction at the wound edge are major events during closure of the plasma membrane and underlying cytoskeleton during cell wound repair. This study shows that all five Drosophila Septins are required for efficient cell wound repair. Based on their different recruitment patterns and knockdown/mutant phenotypes, two distinct Septin complexes, Sep1-Sep2-Pnut and Sep4-Sep5-Pnut, are assembled to regulate actin ring assembly, contraction, and remodeling during the repair process. Intriguingly, these two Septin complexes have different F-actin bending activities. In addition, Anillin regulates the recruitment of only one of two Septin complexes upon wounding. These results demonstrate that two functionally distinct Septin complexes work side-by-side to discretely regulate actomyosin ring dynamics during cell wound repair. |
Monday, August 6th - Behavior |
| Cellini, B., Ferrero, M., Mongeau, J. M. (2024). Drosophila flying in augmented reality reveals the vision-based control autonomy of the optomotor response. Curr Biol, 34(1):68-78.e64 PubMed ID: 38113890
Summary: For walking, swimming, and flying animals, the optomotor response is essential to stabilize gaze. How flexible is the optomotor response? Classic work in Drosophila has argued that flies adapt flight control under augmented visual feedback conditions during goal-directed bar fixation. However, whether the lower-level, reflexive optomotor response can similarly adapt to augmented visual feedback (partially autonomous) or not (autonomous) over long timescales is poorly understood. To address this question, an augmented reality paradigm was developed to study the vision-based control autonomy of the yaw optomotor response of flying fruit flies (Drosophila). Flies were placed in a flight simulator, which permitted free body rotation about the yaw axis. By feeding back body movements in real time to a visual display, visual feedback was augmented and inverted. Thus, this experimental paradigm caused a constant visual error between expected and actual visual feedback to study potential adaptive visuomotor control. By combining experiments with control theory, it was demonstrated that the optomotor response is autonomous during augmented reality flight bouts of up to 30 min, which exceeds the reported learning epoch during bar fixation. Agreement between predictions from linear systems theory and experimental data supports the notion that the optomotor response is approximately linear and time invariant within the experimental assay. Even under positive visual feedback, which revealed the stability limit of flies in augmented reality, the optomotor response was autonomous. These results support a hierarchical motor control architecture in flies with fast and autonomous reflexes at the bottom and more flexible behavior at higher levels. | Ike, K. G. O., Lamers, S. J. C., Kaim, S., de Boer, S. F., Buwalda, B., Billeter, J. C., Kas, M. J. H. (2023). The human neuropsychiatric risk gene Drd2 is necessary for social functioning across evolutionary distant species. Molecular psychiatry, PubMed ID: 38114631
Summary: The mammalian Drd2 gene, encoding the dopamine D2 receptor (D2R), was recently indicated as a potential target in the etiology of lowered sociability (i.e., social withdrawal), a symptom of several neuropsychiatric disorders such as Schizophrenia and Major Depression. Many animal species show social withdrawal in response to stimuli, including the vinegar fly Drosophila melanogaster and mice, which also share most human disease-related genes. This study tested for causality between Drd2 and sociability and for its evolutionary conserved function in these two distant species, as well as assess its mechanism as a potential therapeutic target. During behavioral observations in groups of freely interacting D. melanogaster, Drd2 homologue (Dop2R) mutant showed decreased social interactions and locomotor activity. After confirming Drd2's social effects in flies, conditional transgenic mice lacking Drd2 in dopaminergic cells (autoreceptor KO) or in serotonergic cells (heteroreceptor KO) were studied in semi-natural environments, where they could freely interact. Autoreceptor KOs showed increased sociability, but reduced activity, while no overall effect of Drd2 deletion was observed in heteroreceptor KOs. To determine acute effects of D2R signaling on sociability, it was also shown that a direct intervention with the D2R agonist Sumanirole decreased sociability in wild type mice, while the antagonist showed no effects. Using a computational ethological approach, this study demonstrates that Drd2 regulates sociability across evolutionary distant species, and that activation of the mammalian D2R autoreceptor, in particular, is necessary for social functioning. |
| Benoit, J. B., Ajayi, O. M., Webster, A., Grieshop, K., Lewis, D., Talbott, H., Bose, J., Polak, M. (2023). Shifted levels of sleep and activity under darkness as mechanisms underlying ectoparasite resistance. bioRxiv, PubMed ID: 37961082
Summary: Parasites harm host fitness and are pervasive agents of natural selection. Host defensive traits in natural populations typically show genetic variation, which may be maintained when parasite resistance imposes fitness costs on the host coupled with variability in parasite prevalence in space and/or time. Previous work demonstrated significant evolutionary responses to artificial selection for increasing behavioral immunity to Gamasodes queenslandicus mites in replicate lines of Drosophila melanogaster. This study reports transcriptional shifts in metabolic processes between selected and control fly lines based on RNA-seq analyses. Decreased starvation resistance and increased use of nutrient reserves was also noticed in flies from mite-resistant lines. Additionally, mite-resistant lines exhibited increased behavioral activity, reduced sleep and elevated oxygen consumption under conditions of darkness. The link between resistance and sleep was confirmed in an independent panel of D. melanogaster genetic lines exhibiting variable sleep durations, showing a positive correlation between mite resistance and reduced sleep. Experimentally restraining the activity of artificially selected mite-resistant flies during exposure to parasites under dark conditions reduced their resistance advantage relative to control flies. The results suggest that ectoparasite resistance in this system involves increased dark-condition activity and metabolic gene expression at the expense of nutrient reserves and starvation resistance. | Lyth, S., Betancourt, A. J., Price, T. A. R., Verspoor, R. L. (2023). The suppression of a selfish genetic element increases a male's mating success in a fly. Ecology and evolution 13(11):e10719 PubMed ID: 37964789
Summary: X chromosome meiotic drive (XCMD) kills Y-bearing sperm during spermatogenesis, leading to the biased transmission of the selfish X chromosome. Despite this strong transmission, some natural XCMD systems remain at low and stable frequencies, rather than rapidly spreading through populations. The reason may be that male carriers can have reduced fitness, as they lose half of their sperm, only produce daughters, and may carry deleterious alleles associated with XCMD. Thus, females may benefit from avoiding mating with male carriers, yielding a further reduction in fitness. Genetic suppressors of XCMD, which block the killing of Y sperm and restore fair Mendelian inheritance, are also common and could prevent the spread of XCMD. However, whether suppressed males are as fit as a wild-type male remains an open question, as the effect that genetic suppressors may have on a male's mating success is rarely considered. This study investigated the mating ability of XCMD males and suppressed XCMD males in comparison to wild-type males in the fruit fly Drosophila subobscura, where drive remains at a stable frequency of 20% in wild populations where it occurs. Both competitive and non-competitive mating trials were used to evaluate male mating success in this system. No evidence was found that unsuppressed XCMD males were discriminated against. Remarkably, however, their suppressed XCMD counterparts had a higher male mating success compared to wild-type controls. Unsuppressed XCMD males suffered 12% lower offspring production in comparison to wild-type males. This cost appears too weak to counter the transmission advantage of XCMD, and thus the factors preventing the spread of XCMD remain unclear. |
| Shakeel, M., Kaushik, S., Tanimura, T., Brockmann, A., Kain, P. (2023). Tracking Sugar-Elicited Local Searching Behavior in Drosophila. J Vis Exp, (201) PubMed ID: 38047573
Summary: Foraging behavior is essential for the survival of organisms as it enables them to locate and acquire essential food resources. In Drosophila, hunger triggers a distinct search behavior following the consumption of small quantities of a sugar solution. This report presents a simple experimental setup to study sugar-elicited search behavior with the aim of uncovering the underlying mechanisms. Minute quantities of concentrated sugar solution elicit sustained searching behavior in flies. The involvement of path integration in this behavior has been established, as flies utilize their trajectory to return to the sugar location. The most recent findings provide evidence of temporal modulation in the initiation and intensity of the search behavior after sugar intake. This setup was used for artificial activation of specific taste-receptor neurons in the pharynx, which elicits the search behavior. Understanding the neural basis of hunger-driven searching behavior in flies contributes to the field of neurobiology as a whole, offering insights into the regulatory mechanisms that govern feeding behaviors not only in other organisms but also in humans. | Villegas, G., Pereira, M. T., Love, C. R., Edery, I. (2023). DAYWAKE implicates novel roles for circulating lipid-binding proteins as extracerebral regulators of daytime wake-sleep behavior. FEBS letters, PubMed ID: 38112219
Summary: Sleep during the midday, commonly referred to as siesta, is a common trait of animals that mainly sleep during the night. Work using Drosophila led to the identification of the daywake (dyw) gene, found to have anti-siesta activity. This study shows that the DYW protein undergoes signal peptide-dependent secretion, is present in the circulatory system, and accumulates in multiple organs, but, surprisingly, it is not detected in the brain where wake-sleep centers are located. The abundance of DYW in adult flies is regulated by age, sex, temperature, and the splicing efficiency of a nearby thermosensitive intron. It is suggested that DYW regulates daytime wake-sleep balance in an indirect, extracerebral manner, via a multi-organ network that interfaces with the circulatory system. |
Friday, August 3rd - Cell Cycle |
| Yamazoe, K., Inoue, Y. H. (2023). Cyclin B Export to the Cytoplasm via the Nup62 Subcomplex and Subsequent Rapid Nuclear Import Are Required for the Initiation of Drosophila Male Meiosis. Cells, 12(22) PubMed ID: 37998346
Summary: The cyclin-dependent kinase 1 (Cdk1)-cyclin B (CycB) complex plays critical roles in cell-cycle regulation. Before Drosophila male meiosis, CycB is exported from the nucleus to the cytoplasm via the nuclear porin 62kD (Nup62) subcomplex of the nuclear pore complex. When this export is inhibited, Cdk1 is not activated, and meiosis does not initiate. This study investigated the mechanism that controls the cellular localization and activation of Cdk1. Cdk1-CycB continuously shuttled into and out of the nucleus before meiosis. Overexpression of CycB, but not that of CycB with nuclear localization signal sequences, rescued reduced cytoplasmic CycB and inhibition of meiosis in Nup62-silenced cells. Full-scale Cdk1 activation occurred in the nucleus shortly after its rapid nuclear entry. Cdk1-dependent centrosome separation did not occur in Nup62-silenced cells, whereas Cdk1 interacted with Cdk-activating kinase and Twine/Cdc25C in the nuclei of Nup62-silenced cells, suggesting the involvement of another suppression mechanism. Silencing of roughex rescued Cdk1 inhibition and initiated meiosis. Nuclear export of Cdk1 ensured its escape from inhibition by a cyclin-dependent kinase inhibitor. The complex re-entered the nucleus via importin β at the onset of meiosis. A model is proposed regarding the dynamics and activation mechanism of Cdk1-CycB to initiate male meiosis. | Fellmeth, J. E., Jang, J. K., Persaud, M., Sturm, H., Changela, N., Parikh, A., McKim, K. S. (2023). A dynamic population of prophase CENP-C is required for meiotic chromosome segregation. PLoS Genet, 19(11):e1011066 PubMed ID: 38019881
Summary: The centromere is an epigenetic mark that is a loading site for the kinetochore during meiosis and mitosis. This mark is characterized by the H3 variant CENP-A, known as CID in Drosophila. In Drosophila, CENP-C is critical for maintaining CID at the centromeres and directly recruits outer kinetochore proteins after nuclear envelope break down. These two functions, however, happen at different times in the cell cycle. Furthermore, in Drosophila and many other metazoan oocytes, centromere maintenance and kinetochore assembly are separated by an extended prophase. This study has investigated the dynamics of function of CENP-C during the extended meiotic prophase of Drosophila oocytes and found that maintaining high levels of CENP-C for metaphase I requires expression during prophase. In contrast, CID is relatively stable and does not need to be expressed during prophase to remain at high levels in metaphase I of meiosis. Expression of CID during prophase can even be deleterious, causing ectopic localization to non-centromeric chromatin, abnormal meiosis and sterility. CENP-C prophase loading is required for multiple meiotic functions. In early meiotic prophase, CENP-C loading is required for sister centromere cohesion and centromere clustering. In late meiotic prophase, CENP-C loading is required to recruit kinetochore proteins. CENP-C is one of the few proteins identified in which expression during prophase is required for meiotic chromosome segregation. An implication of these results is that the failure to maintain recruitment of CENP-C during the extended prophase in oocytes would result in chromosome segregation errors in oocytes. |
| Wong, S. S., Wainman, A., Saurya, S., Raff, J. W. (2024). Regulation of centrosome size by the cell-cycle oscillator in Drosophila embryos. The EMBO journal, PubMed ID: 38233576
Summary: Mitotic centrosomes assemble when centrioles recruit large amounts of pericentriolar material (PCM) around themselves. In early C. elegans embryos, mitotic centrosome size appears to be set by the limiting amount of a key component. In Drosophila syncytial embryos, thousands of mitotic centrosomes are assembled as the embryo proceeds through 13 rounds of rapid nuclear division, driven by a core cell cycle oscillator. These divisions slow during nuclear cycles 11-13, and centrosomes were found to respond by reciprocally decreasing their growth rate, but increasing their growth period-so that they grow to a relatively consistent size at each cycle. At the start of each cycle, moderate CCO activity initially promotes centrosome growth, in part by stimulating Polo/PLK1 recruitment to centrosomes. Later in each cycle, high CCO activity inhibits centrosome growth by suppressing the centrosomal recruitment and/or maintenance of centrosome proteins. Thus, in fly embryos, mitotic centrosome size appears to be regulated predominantly by the core cell cycle oscillator, rather than by the depletion of a limiting component. | Rollins, K. R., Blankenship, J. T. (2023). Dysregulation of the endoplasmic reticulum blocks recruitment of centrosome-associated proteins resulting in mitotic failure. Development, 150(22) PubMed ID: 37971218
Summary: The endoplasmic reticulum (ER) undergoes a remarkable transition in morphology during cell division to aid in the proper portioning of the ER. However, whether changes in ER behaviors modulate mitotic events is less clear. Like many animal embryos, the early Drosophila embryo undergoes rapid cleavage cycles in a lipid-rich environment. This study shows that mitotic spindle formation, centrosomal maturation, and ER condensation occur with similar time frames in the early syncytium. In a screen for Rab family GTPases that display dynamic function at these stages, Rab1 was identified. Rab1 disruption led to an enhanced buildup of ER at the spindle poles and produced an intriguing 'mini-spindle' phenotype. ER accumulation around the mitotic space negatively correlates with spindle length/intensity. Importantly, centrosomal maturation is defective in these embryos, as mitotic recruitment of key centrosomal proteins is weakened after Rab1 disruption. Finally, division failures and ER overaccumulation is rescued by Dynein inhibition, demonstrating that Dynein is essential for ER spindle recruitment. These results reveal that ER levels must be carefully tuned during mitotic processes to ensure proper assembly of the division machinery. |
| Pazhayam, N. M., Frazier, L. K., Sekelsky, J. (2023). Centromere-Proximal Suppression of Meiotic Crossovers in Drosophila is Robust to Changes in Centromere Number and Repetitive DNA Content. bioRxiv, PubMed ID: 37905008
Summary: Accurate segregation of homologous chromosomes during meiosis depends on both the presence and regulated placement of crossovers (COs). The centromere effect (CE), or CO exclusion in pericentromeric regions of the chromosome, is a meiotic CO patterning phenomenon that helps prevent nondisjunction (NDJ), thereby protecting against chromosomal disorders and other meiotic defects. Despite being identified nearly a century ago, the mechanisms behind this fundamental cellular process remain unknown, with most studies of the Drosophila CE focusing on local influences of the centromere and pericentric heterochromatin. This study sought to investigate whether dosage changes in centromere number and repetitive DNA content affect the strength of the CE, using phenotypic recombination mapping. Additionally, the effects of repetitive DNA function on CE strength was also studied using satellite-DNA binding protein mutants shown to have defective centromere clustering. Despite what previous studies suggest, these results show that the Drosophila CE is robust to dosage changes in centromere number and repetitive DNA content, and potentially also to repetitive DNA function. This study suggests that the CE is unlikely to be spatially controlled, providing novel insight into the mechanisms behind the Drosophila centromere effect. | Sperling, A. L., Glover, D. M. (2023). Aneuploidy during development in facultative parthenogenetic Drosophila. Heredity, PubMed ID: 38017115
Summary: From concatenated chromosomes to polyploidization, large-scale genome changes are known to occur in parthenogenetic animals. This study report mosaic aneuploidy in larval brains of facultatively parthenogenetic Drosophila. A background of aneuploidy was identified in D. mercatorum strains, and increased levels of aneuploidy were found in the larval brain tissue of animals arising parthenogenetically versus those arising from sexual reproduction. There is also intra-individual variation in germline-derived aneuploidy within the same strain. To determine if this is a general feature of facultative parthenogenesis in drosophilids, sexually reproduced and parthenogenetic offspring were compared from an engineered facultative parthenogenetic strain of D. melanogaster. In addition to germline-derived aneuploidy, this revealed somatic aneuploidy that increased by up to fourfold in parthenogens compared to sexually reproduced offspring. Therefore, the genetic combination identified in D. mercatorum that causes facultative parthenogenesis in D. melanogaster results in aneuploidy, which indicates that the loss of mitotic control resulting in parthenogenesis causes subsequent genome variation within the parthenogenetic offspring. These findings challenge the assumption that parthenogenetic offspring are near genetic replicas of their mothers. |
Wednesday, July 31st - Disease Models |
| Landis, G. N., Bell, H. S., Peng, O., Bognar, B., Tong, A., Manea, T. D., Bao, H., Han, X., Tower, J. (2023). Dhr96[1] mutation and maternal tudor[1] mutation increase life span and reduce the beneficial effects of mifepristone in mated female Drosophila. PLoS One, 18(12):e0292820 PubMed ID: 38127988
Summary: Mating and receipt of male Sex Peptide hormone cause increased egg laying, increased midgut size and decreased life span in female Drosophila. Feeding mated females with the synthetic steroid mifepristone decreases egg production, reduces midgut size, and increases life span. Several gene mutations were assayed to investigate possible mechanisms for mifepristone action. Drosophila Dhr96 is a hormone receptor, and a key positive regulator of midgut lipid uptake and metabolism. Dhr961 null mutation increased female life span, and reduced the effects of mifepristone on life span, suggesting thatDhr961 mutation and mifepristone may act in part through the same mechanism. Consistent with this idea, lipidomics analysis revealed that mating increases whole-body levels of triglycerides and fatty-acids in triglycerides, and these changes are reversed by mifepristone. Maternal tudor1 mutation results in females that lack the germ-line and produce no eggs. Maternal tudor1 mutation increased mated female life span, and reduced but did not eliminate the effects of mating and mifepristone on life span. This indicates that decreased egg production may be related to the life span benefits of mifepristone, but is not essential. Mifepristone increases life span in w[1118] mutant mated females, but did not increase life span in w[1118] mutant virgin females. Mifepristone decreased egg production in w[1118] mutant virgin females, indicating that decreased egg production is not sufficient for mifepristone to increase life span. Mifepristone increases life span in virgin females of some, but not all, white[+] and mini-white[+] strains. Backcrossing of mini-white[+] transgenes into the w[1118] background was not sufficient to confer a life span response to mifepristone in virgin females. Taken together, the data support the hypothesis that mechanisms for mifepristone life span increase involve reduced lipid uptake and/or metabolism, and suggest that mifepristone may increase life span in mated females and virgin females through partly different mechanisms. | Utsuno, Y., Hamada, K., Hamanaka, K., Miyoshi, K., Tsuchimoto, K., Sunada, S., Itai, T., Sakamoto, M., Tsuchida, N., Uchiyama, Y., Koshimizu, E., Fujita, A., Miyatake, S., Misawa, K., Mizuguchi, T., Kato, Y., Saito, K., Ogata, K., Matsumoto, N. (2023). Novel missense variants cause intermediate phenotypes in the phenotypic spectrum of SLC5A6-related disorders. Journal of human genetics, PubMed ID: 38012394
Summary: Mammalian SLC5A6 encodes the sodium-dependent multivitamin transporter, a transmembrane protein that uptakes biotin, pantothenic acid, and lipoic acid. Biallelic SLC5A6 variants cause sodium-dependent multivitamin transporter deficiency (SMVTD) and childhood-onset biotin-responsive peripheral motor neuropathy (COMNB), which both respond well to replacement therapy with the above three nutrients. SMVTD usually presents with various symptoms in multiple organs, such as gastrointestinal hemorrhage, brain atrophy, and global developmental delay, at birth or in infancy. Without nutrient replacement therapy, SMVTD can be lethal in early childhood. COMNB is clinically milder and has a later onset than SMVTD, at approximately 10 years of age. COMNB symptoms are mostly limited to peripheral motor neuropathy. This study reports three patients from one Japanese family harboring novel compound heterozygous missense variants in SLC5A6, namely NM_021095.4:c.[221C>T];[642G>C] p.[(Ser74Phe)];[(Gln214His)]. Both variants were predicted to be deleterious through multiple lines of evidence, including amino acid conservation, in silico predictions of pathogenicity, and protein structure considerations. Drosophila analysis also showed c.221C>T to be pathogenic. All three patients had congenital brain cysts on neonatal cranial imaging, but no other morphological abnormalities. They also had a mild motor developmental delay that almost completely resolved despite no treatment. In terms of severity, their phenotypes were intermediate between SMVTD and COMNB. From these findings a new SLC5A6-related disorder is proposed, spontaneously remitting developmental delay with brain cysts (SRDDBC) whose phenotypic severity is between that of SMVTD and COMNB. Further clinical and genetic evidence is needed to support this suggestion. |
| Kosakamoto, H., Obata, F., Kuraishi, J., Aikawa, H., Okada, R., Johnstone, J. N., Onuma, T., Piper, M. D. W., Miura, M. (2023). Early-adult methionine restriction reduces methionine sulfoxide and extends lifespan in Drosophila. Nat Commun, 14(1):7832 PubMed ID: 38052797
Summary: Methionine restriction (MetR) extends lifespan in various organisms, but its mechanistic understanding remains incomplete. Whether MetR during a specific period of adulthood increases lifespan is not known. In Drosophila, MetR is reported to extend lifespan only when amino acid levels are low. By using an exome-matched holidic medium, this study showed that decreasing Met levels to 10% extends Drosophila lifespan with or without decreasing total amino acid levels. MetR during the first four weeks of adult life only robustly extends lifespan. MetR in young flies induces the expression of many longevity-related genes, including Methionine sulfoxide reductase A (MsrA), which reduces oxidatively-damaged Met. MsrA induction is foxo-dependent and persists for two weeks after cessation of the MetR diet. Loss of MsrA attenuates lifespan extension by early-adulthood MetR. This study highlights the age-dependency of the organismal response to specific nutrients and suggests that nutrient restriction during a particular period of life is sufficient for healthspan extension. | Pragati, Sarkar, S. (2023). Targeted upregulation of dMyc restricts JNK-mediated degeneration of dopaminergic neurons in the paraquat-induced Parkinson's disease model of Drosophila.. Neurosci Res, PubMed ID: 37913999
Summary: Parkinson's disease is the second most common neurodegenerative disease characterized by the loss of dopaminergic neurons in the brain. Parkinson's disease has both familial and sporadic cases of origin governed differentially by genetic and/or environmental factors. Different epidemiological studies have proposed an association between the pathogenesis of cancer and Parkinson's disease; however, a precise correlation between these two illnesses could not be established yet. This study examined the disease-modifying property of dmyc (a Drosophila homolog of human cmyc proto-oncogene) in the paraquat-induced sporadic Parkinson's disease model of Drosophila. Targeted upregulation of dMyc significantly restricts paraquat-mediated neurotoxicity. Paraquat feeding reduces the cellular level of dMyc. It is further noted that targeted upregulation of dMyc in paraquat-exposed flies mitigates degeneration of dopaminergic neurons by reinstating the aberrantly activated JNK pathway, and this in turn improves the motor performance and survival rate of the flies. This study provides the first evidence that improved cellular level of dMyc could efficiently minimize the neurotoxic effects of paraquat, which could be beneficial in designing novel therapeutic strategies against Parkinson's disease. |
| Cheng, J., Wu, B. T., Liu, H. P., Lin, W. Y. (2023). Tyrosine Metabolism Pathway Is Downregulated in Dopaminergic Neurons with LRRK2 Overexpression in Drosophila. Int J Mol Sci, 24(21) PubMed ID: 37958569
Summary: LRRK2 mutations are the leading cause of familial Parkinson's disease (PD) and are a significant risk factor for idiopathic PD cases. However, the molecular mechanisms underlying the degeneration of dopaminergic (DA) neurons in LRRK2 PD patients remain unclear. To determine the translatomic impact of LRRK2 expression in DA neurons, gene set enrichment analysis (GSEA) was used to analyze a translating ribosome affinity purification (TRAP) RNA-seq dataset from a DA-neuron-specific-expressing Drosophila model. The tyrosine metabolism pathway, including tyrosine hydroxylase (TH), was found to be downregulated in DA neurons with LRRK2 overexpression; in contrast, the Hippo signaling pathway is downregulated in the G2019S mutant compared to wild-type LRRK2 in the DA neurons. These results imply that the downregulation of tyrosine metabolism occurs before pronounced DA neuron loss and that LRRK2 may downregulate the tyrosine metabolism in a DA-neuron-loss-independent way. | Sujkowski, A. L., Ranxhi, B., Prifti, M. V., Alam, N., Todi, S. V., Tsou, W. L. (2023). Progressive degeneration in a new Drosophila model of Spinocerebellar Ataxia type 7. bioRxiv, PubMed ID: 37986914
Summary: Spinocerebellar ataxia type 7 (SCA7) is a progressive neurodegenerative disorder resulting from abnormal expansion of polyglutamine (polyQ) in its disease protein, ataxin-7 (ATXN7). ATXN7 is part of Spt-Ada-Gcn5 acetyltransferase (SAGA), an evolutionarily conserved transcriptional coactivation complex with critical roles in chromatin remodeling, cell signaling, neurodifferentiation, mitochondrial health and autophagy. SCA7 is dominantly inherited and characterized by genetic anticipation and high repeat-length instability. Patients with SCA7 experience progressive ataxia, atrophy, spasticity, and blindness. There is currently no cure for SCA7, and therapies are aimed at alleviating symptoms to increase quality of life. This study reports novel Drosophila lines of SCA7 with polyQ repeats in wild-type and human disease patient range. ATXN7 expression has age- and polyQ repeat length-dependent reduction in survival and retinal instability, concomitant with increased ATXN7 protein aggregation. These new lines will provide important insight on disease progression that can be used in the future to identify therapeutic targets for SCA7 patients |
Tuesday, July 30th - Larval and Adult Physiology and Metabolism |
| Evans, A., Ferrer, A. J., Fradkov, E., Shomar, J. W., Forer, J., Klein, M. (2023). Temperature sensitivity and temperature response across development in the Drosophila larva. Frontiers in molecular neuroscience, 16:1275469 PubMed ID: 37965044
Summary: This study examined how sensory neuronal and behavioral responses to temperature variation both change across the development of the larva. Looking at a wide range of non-nociceptive isotropic thermal environments, this study characterized the dependence of speed, turning rate, and other behavioral components on temperature, distinguishing the physical effects of temperature from behavior changes based on sensory processing. This study also characterized the strategies larvae use to modulate individual behavioral components to produce directed navigation along thermal gradients, and how these strategies change during physical development. Simulations based on modified random walks show where thermotaxis in each developmental stage fits into the larger context of possible navigation strategies. This study also investigate cool sensing neurons in the larva's dorsal organ ganglion, characterizing neural response to sine-wave modulation of temperature while performing single-cell-resolution 3D imaging. The sensitivity of these neurons, which produce signals in response to extremely small temperature changes, was determined. Combining thermotaxis results with neurophysiology data, this study observed, across development, sensitivity to temperature change as low as a few thousandths of a °C per second, or a few hundredths of a °C in absolute temperature change. | Miyamoto, T., Hedjazi, S., Miyamoto, C., Amrein, H. (2023). Drosophila Neuronal Glucose 6 Phosphatase is a modulator of Neuropeptide Release that regulates muscle glycogen stores via FMRFamide signaling. bioRxiv PubMed ID: 38077084
Summary: Neuropeptides (NPs) and their cognate receptors are critical molecular effectors of diverse physiological processes and behaviors. A non-canonical function has been reported of the Drosophila Glucose-6-Phosphatase (G6P) gene in a subset of neurosecretory cells in the CNS that governs systemic glucose homeostasis in food deprived flies. This study shows that G6P expressing neurons define 7 groups of neuropeptide secreting cells, 5 in the brain and 2 in the thoracic ganglia. Using the glucose homeostasis phenotype as a screening tool, one such group, located in the thoracic ganglia and expressing FMRFamide (FMRFa (G6P)) neuropeptides, was shown to be necessary and sufficient to maintain systemic glucose homeostasis in starved flies. It was further shown that the receptor for FMRFamides (FMRFaR) is one key target of G6P dependent NP signaling and essential for the build-up of glycogen stores in the jump muscle. Lastly, measurements of the Golgi apparatus of FMRFa (G6P) neurons and neuropeptide released into the hemolymph suggests that G6P enhances FMRFa signaling by increasing the capacity of the neurosecretory system. A general model is proposed in which the main role of G6P is to counteract glycolysis in peptidergic neurons for the purpose of optimizing the intracellular environment best suited for the expansion of the Golgi apparatus, boosting release of neuropeptides, which through the activation of specific neuropeptide receptors, enhances signaling in respective target tissues. |
| Qiao, S., Bernasek, S., Gallagher, K. D., O'Connell, J., Yamada, S., Bagheri, N., Amaral, L. A. N., Carthew, R. W. (2024). Energy metabolism modulates the regulatory impact of activators on gene expression. Development, 151(1) PubMed ID: 38063847
Summary: Gene expression is a regulated process fueled by ATP consumption. Therefore, regulation must be coupled to constraints imposed by the level of energy metabolism. This study explored this relationship both theoretically and experimentally. A stylized mathematical model predicts that activators of gene expression have variable impact depending on metabolic rate. Activators become less essential when metabolic rate is reduced and more essential when metabolic rate is enhanced. In the Drosophila eye, expression dynamics of the yan gene are less affected by loss of EGFR-mediated activation when metabolism is reduced, and the opposite effect is seen when metabolism is enhanced. The effects are also seen at the level of pattern regularity in the adult eye, where loss of EGFR-mediated activation is mitigated by lower metabolism. It is proposed that gene activation is tuned by energy metabolism to allow for faithful expression dynamics in the face of variable metabolic conditions. | Xu, B., Hwangbo, D. S., Saurabh, S., Rosensweig, C., Allada, R., Kath, W. L., Braun, R. (2023). Temperature-driven coordination of circadian transcriptome regulation. bioRxiv, PubMed ID: 37961403
Summary: The circadian rhythm is an evolutionarily-conserved molecular oscillator that enables species to anticipate rhythmic changes in their environment. At a molecular level, the core clock genes induce a circadian oscillation in thousands of genes in a tissue-specific manner, orchestrating myriad biological processes. While studies have investigated how the core clock circuit responds to environmental perturbations such as temperature, the downstream effects of such perturbations on circadian regulation remain poorly understood. By analyzing bulk-RNA sequencing of Drosophila fat bodies harvested from flies subjected to different environmental conditions, a highly condition-specific circadian transcriptome was generated. Further employing a reference-based gene regulatory network (Reactome), evidence was found of increased gene-gene coordination at low temperatures and synchronization of rhythmic genes that are network neighbors. These results point to the mechanisms by which the circadian clock mediates the fly's response to seasonal changes in temperature. |
| Huang, W. H., Kajal, K., Wibowo, R. H., Amartuvshin, O., Kao, S. H., Rastegari, E., Lin, C. H., Chiou, K. L., Pi, H. W., Ting, C. T., Hsu, H. J. (2024). Excess dietary sugar impairs Drosophila adult stem cells via elevated reactive oxygen species-induced JNK signaling. Development, 151(1) PubMed ID: 38063853
Summary: High-sugar diets (HSDs) often lead to obesity and type 2 diabetes, both metabolic syndromes associated with stem cell dysfunction. However, it is unclear whether excess dietary sugar affects stem cells. This study reports that HSD impairs stem cell function in the intestine and ovaries of female Drosophila prior to the onset of insulin resistance, a hallmark of type 2 diabetes. Although 1 week of HSD leads to obesity, impaired oogenesis and altered lipid metabolism, insulin resistance does not occur. HSD increases glucose uptake by germline stem cells (GSCs) and triggers reactive oxygen species-induced JNK signaling, which reduces GSC proliferation. Removal of excess sugar from the diet reverses these HSD-induced phenomena. A similar phenomenon is found in intestinal stem cells (ISCs), except that HSD disrupts ISC maintenance and differentiation. Interestingly, tumor-like GSCs and ISCs are less responsive to HSD, which may be because of their dependence on glycolytic metabolism and high energy demand, respectively. This study suggests that excess dietary sugar induces oxidative stress and damages stem cells before insulin resistance develops, a mechanism that may also occur in higher organisms. | Bordet, G., Bamgbose, G., Tulin, A. V. (2023). Poly(ADP-ribosyl)ating enzymes coordinate changes in the expression of metabolic genes with developmental progression. Sci Rep, 13(1):20320 PubMed ID: 37985852
Summary: Metabolism, known to be temporally regulated to meet evolving energy demands, plays a crucial role in shaping developmental pace. Recent studies have demonstrated that two key proteins PARP1 and PARG play a regulatory role in the transcription of both morphogenic and metabolic genes. Intriguingly, in Drosophila, the depletion of PARP1 or PARG proteins causes a developmental arrest before pupation, resulting in individuals unable to complete their development. This phenotype highlights the critical involvement of poly(ADP-ribosyl)ating enzymes in regulating the metamorphic process. This study provides compelling evidence that these enzymes intricately coordinate transcriptional changes in both developmental and metabolic pathways during metamorphosis. Specifically, they promote the expression of genes crucial for pupation, while simultaneously negatively regulating the expression of metabolic genes before the transition to the pupal stage. Additionally, these enzymes suppress the expression of genes that are no longer required during this transformative period. These findings shed light on the intricate interplay between poly(ADP-ribosyl)ating enzymes, developmental processes, and metabolic regulation before metamorphosis and highlight a new role of poly(ADP-ribosyl)ating enzymes in the global regulation of transcription. |
Monday, July 29th - Larval and Adult Development |
| Tyson, J. J., Monshizadeh, A., Shvartsman, S. Y., Shingleton, A. W. (2023). A dynamical model of growth and maturation in Drosophila. Proc Natl Acad Sci U S A, 120(49):e2313224120 PubMed ID: 38015844
Summary: The decision to stop growing and mature into an adult is a critical point in development that determines adult body size, impacting multiple aspects of an adult's biology. In many animals, growth cessation is a consequence of hormone release that appears to be tied to the attainment of a particular body size or condition. Nevertheless, the size-sensing mechanism animals use to initiate hormone synthesis is poorly understood. This study develop a simple mathematical model of growth cessation in Drosophila melanogaster, which is ostensibly triggered by the attainment of a critical weight (CW) early in the last instar. Attainment of CW is correlated with the synthesis of the steroid hormone ecdysone, which causes a larva to stop growing, pupate, and metamorphose into the adult form. A model suggests that, contrary to expectation, the size-sensing mechanism that initiates metamorphosis occurs before the larva reaches CW; that is, the critical-weight phenomenon is a downstream consequence of an earlier size-dependent developmental decision, not a decision point itself. Further, this size-sensing mechanism does not require a direct assessment of body size but emerges from the interactions between body size, ecdysone, and nutritional signaling. Because many aspects of this model are evolutionarily conserved among all animals, the model may provide a general framework for understanding how animals commit to maturing from their juvenile to adult form. | Liu, S. P., Yin, H. D., Li, W. J., Qin, Z. H., Yang, Y., Huang, Z. Z., Zong, L., Liu, X. K., Du, Z., Fan, W. L., Zhang, Y. Q., Zhang, D., Zhang, Y. E., Liu, X. Y., Yang, D., Ge, S. Q. (2023). The Morphological Transformation of the Thorax during the Eclosion of Drosophila melanogaster (Diptera: Drosophilidae).. Insects, 14(11) PubMed ID: 37999092
Summary: The model organism Drosophila melanogaster, as a species of Holometabola, undergoes a series of transformations during metamorphosis. To deeply understand its development, it is crucial to study its anatomy during the key developmental stages. This study describes the anatomical systems of the thorax, including the endoskeleton, musculature, nervous ganglion, and digestive system, from the late pupal stage to the adult stage, based on micro-CT and 3D visualizations. The development of the endoskeleton causes original and insertional changes in muscles. Several muscles change their shape during development in a non-uniform manner with respect to both absolute and relative size; some become longer and broader, while others shorten and become narrower. Muscular shape may vary during development. The number of muscular bundles also increases or decreases. Growing muscles are probably anchored by the tissues in the stroma. Some muscles and tendons are absent in the adult stage, possibly due to the hardened sclerites. Nearly all flight muscles are present by the third day of the pupal stage, which may be due to the presence of more myofibers with enough mitochondria to support flight power. There are sexual differences in the same developmental period. In contrast to the endodermal digestive system, the functions of most thoracic muscles change in the development from the larva to the adult in order to support more complex locomotion under the control of a more structured ventral nerve cord based on the serial homology proposed herein. |
| He, Q., Fan, X., Wang, S., Chen, S., Chen, J. (2023). Juvenile hormone inhibits adult cuticle formation in Drosophila melanogaster through Kr-h1/Dnmt2-mediated DNA methylation of Acp65A promoter. Insect Mol Biol, PubMed ID: 37916965
Summary: \Differentiation of imaginal epidermal cells of Drosophila melanogaster to form adult cuticles occurs at approximately 40-93 h after puparium formation. Juvenile hormone (JH) given at pupariation results in formation of a second pupal cuticle in the abdomen instead of the adult cuticle. Although the adult cuticle gene Acp65A has been reported to be down-regulated following JH treatment, the regulatory mechanism remains unclear. This study found that the JH primary response gene Kruppel homologue 1 (Kr-h1) plays a vital role in the repression of adult cuticle formation through the mediation of JH action. Overexpression of Kr-h1 mimicked-while knocking down of Kr-h1 attenuated-the inhibitory action of JH on the formation of the adult abdominal cuticle. Further, it was found that Kr-h1 inhibited the transcription of Acp65A by directly binding to the consensus Kr-h1 binding site (KBS) within the Acp65A promoter region. Moreover, the DNA methyltransferase Dnmt2 was shown to interact with Kr-h1, combined with the KBS to promote the DNA methylation of sequences around the KBS, in turn inhibiting the transcription of Acp65A. This study advances understanding of the molecular basis of the "status quo" action of JH on the Drosophila adult metamorphosis. | Piscitello-GOmez, R., Gruber, F. S., Krishna, A., Duclut, C., Modes, C. D., Popovic, M., Julicher, F., Dye, N. A., Eaton, S. (2023). Core PCP mutations affect short time mechanical properties but not tissue morphogenesis in the Drosophila pupal wing. Elife, 12 PubMed ID: 38117039
Summary: How morphogenetic movements are robustly coordinated in space and time is a fundamental open question in biology. This question was studied using the wing of Drosophila melanogaster, an epithelial tissue that undergoes large-scale tissue flows during pupal stages. Previous work showed that pupal wing morphogenesis involves both cellular behaviors that allow relaxation of mechanical tissue stress, as well as cellular behaviors that appear to be actively patterned. This study shows that these active cellular behaviors are not guided by the core planar cell polarity (PCP) pathway, a conserved signaling system that guides tissue development in many other contexts. No significant phenotype was found on the cellular dynamics underlying pupal morphogenesis in mutants of core PCP. Furthermore, using laser ablation experiments, coupled with a rheological model to describe the dynamics of the response to laser ablation, it is concluded that while core PCP mutations affect the fast timescale response to laser ablation they do not significantly affect overall tissue mechanics. In conclusion, this work shows that cellular dynamics and tissue shape changes during Drosophila pupal wing morphogenesis do not require core PCP as an orientational guiding cue. |
| Bollepogu Raja, K. K., Yeung, K., Shim, Y. K., Li, Y., Chen, R., Mardon, G. (2023). A single cell genomics atlas of the Drosophila larval eye reveals distinct photoreceptor developmental timelines. Nat Commun, 14(1):7205 PubMed ID: 37938573
Summary: The Drosophila eye is a powerful model system to study the dynamics of cell differentiation, cell state transitions, cell maturation, and pattern formation. However, a high-resolution single cell genomics resource that accurately profiles all major cell types of the larval eye disc and their spatiotemporal relationships is lacking. This study reports transcriptomic and chromatin accessibility data for all known cell types in the developing eye. Photoreceptors appear as strands of cells that represent their dynamic developmental timelines. As photoreceptor subtypes mature, they appear to assume a common transcriptomic profile that is dominated by genes involved in axon function. Cell type maturation genes, enhancers, and potential regulators, were identified as well as genes with distinct R3 or R4 photoreceptor specific expression. Finally, it was observed that the chromatin accessibility between cones and photoreceptors is distinct. These single cell genomics atlases will greatly enhance the power of the Drosophila eye as a model system. | Imura, E., Enya, S., Niwa, R. (2023). A Drosophila melanogaster ortholog of pentatricopeptide repeat domain 3 ( PTCD3) is essential for development. microPublication biology, 2023 PubMed ID: 38074476
Summary: Mitochondrial DNA (mtDNA) replication and transcription are essential for cellular energy metabolism. It has been suggested that pentatricopeptide repeat (PPR) proteins regulate various aspects of mitochondrial RNA metabolism, including transcription, processing, maturation and stability, and protein synthesis. However, an in vivo requirement of PPR proteins in RNA metabolism has not been fully examined. This paper focuses on the Drosophila melanogaster homolog of PPR domain 3 (PTCD3), encoded by the CG4679 gene. A loss-of-function mutant of PTCD3 is lethal during the second instar. In addition, mutants exhibit reduced expression of a group of genes related to mitochondrial function and ribosome biogenesis, and conversely, they show up-regulated expression of neuronal development-related genes. These results suggest that PTCD3 has important functions in relation to mtDNA and is essential for development. |
Friday, July 26th - Adult Physiology and Metabolism |
| Evans, A., Ferrer, A. J., Fradkov, E., Shomar, J. W., Forer, J., Klein, M. (2023). Temperature sensitivity and temperature response across development in the Drosophila larva. Frontiers in molecular neuroscience. 16:1275469 PubMed ID: 37965044
Summary: The surrounding thermal environment is highly important for the survival and fitness of animals, and as a result most exhibit behavioral and neural responses to temperature changes. Signals generated by thermosensory neurons were studied in Drosophila larvae, and also the physical and sensory effects of temperature variation on locomotion and navigation were studied. In particular how sensory neuronal and behavioral responses to temperature variation both change across the development of the larva were characterized. Looking at a wide range of non-nociceptive isotropic thermal environments, this study characterize the dependence of speed, turning rate, and other behavioral components on temperature, distinguishing the physical effects of temperature from behavior changes based on sensory processing. The strategies larvae use to modulate individual behavioral components to produce directed navigation along thermal gradients, and how these strategies change during physical development were characterized. Simulations based on modified random walks show where thermotaxis in each developmental stage fits into the larger context of possible navigation strategies. Cool sensing neurons were studied in the larva's dorsal organ ganglion, characterizing neural response to sine-wave modulation of temperature while performing single-cell-resolution 3D imaging. The sensitivity of these neurons, which produce signals in response to extremely small temperature changes, was characterized. Combining thermotaxis results with neurophysiology data across development, sensitivity to temperature change was observed as low as a few thousandths of a °C per second, or a few hundredths of a °C in absolute temperature change. | Kim, N., Ahn, Y., Ko, K., Kim, B., Han, K., Suh, H. J., Jung, J., Hong, K. B. (2023). Yeast Hydrolysate Inhibits Lipid Accumulation via Regulation of Lipid Accumulation-Related Genes in a Drosophila Model of High-Sugar Diet-Induced Obesity. Int J Mol Sci, 24(22) PubMed ID: 38003491
Summary: The increasing frequency of processed food consumption has led to the higher ingestion of sugar, increasing the risk of chronic diseases, such as obesity. Yeast hydrolysates (YHs) inhibit body fat accumulation. However, the action mechanism of YH in relation to high-sugar diet-induced obesity is still unclear. Therefore, this study aimed to evaluate the biological effects of YH on lipid accumulation and verify behavioral changes and carbohydrate metabolic gene regulation in high-sugar diet-fed fruit flies. Adult male flies (Drosophila melanogaster; 2-5 days old) were exposed to 20% sucrose for obesity induction. In high-sugar-fed Drosophila, the effect of YH was compared with that of yeast extract. The effects of YH on body conditions and lipid droplet size were quantified and analyzed. Behavioral factors were evaluated by analyzing circadian rhythm patterns and neurotransmitter content, and a molecular approach was used to analyze the expression of metabolism-related genes. Dietary supplementation with YH did not reduce total sugar content, but significantly decreased the triglyceride (TG) levels in Drosophila. A behavioral analysis showed that the total number of night-time activities increased significantly with YH treatment in a dose-dependent manner. In addition, YH effectively regulated the gene expression of insulin-like peptides related to carbohydrate metabolism as well as genes related to lipogenesis. The TG content was significantly reduced at a YH concentration of 0.5%, confirming that the active compound in YH effectively suppresses fat accumulation. These findings support that YH is a potential anti-obesity food material via regulating carbohydrate metabolism in Drosophila. |
| Ghosn, Z. A., Sparks, K. M., Spaulding, J. L., Vutukuri, S., Ahmed, M. J. J., VanBerkum, M. F. A. (2024). Divalent metal content in diet affects severity of manganese toxicity in Drosophila. Biol Open, 13(1) PubMed ID: 38117005
Summary: Dysregulation of manganese (Mn) homeostasis is a contributing factor in many neuro-degenerative diseases. Adult Drosophila are sensitive to excessive levels of dietary Mn, dying relatively early, and exhibiting biochemical and mobility changes reminiscent of Parkinsonian conditions. To further study Mn homeostasis in Drosophila, this study sought to test lower levels of dietary Mn (5 mM), and a striking difference in Canton-S adult survivorship on different food was noted. On a cornmeal diet, Mn-treated flies live only about half as long as untreated siblings. Yet, with the same Mn concentration in a molasses diet, adults survive about 80% as long as untreated siblings, and adults raised on a sucrose-yeast diet are completely insensitive to this low dose of dietary Mn. By manipulating metal ion content in the cornmeal diet, and measuring the metal content in each diet, the difference in lifespan were traced to the levels of calcium and magnesium in the food, suggesting that these ions are involved in Mn uptake and/or use. Based on these findings, it is recommended that the total dietary load of metal ions be considered when assessing Mn toxicity. | Bordet, G., Bamgbose, G., Tulin, A. V. (2023). Poly(ADP-ribosyl)ating enzymes coordinate changes in the expression of metabolic genes with developmental progression. Sci Rep, 13(1):20320 PubMed ID: 37985852
Summary: Metabolism, known to be temporally regulated to meet evolving energy demands, plays a crucial role in shaping developmental pace. Recent studies have demonstrated that two key proteins PARP1 and PARG play a regulatory role in the transcription of both morphogenic and metabolic genes. Intriguingly, in Drosophila, the depletion of PARP1 or PARG proteins causes a developmental arrest before pupation, resulting in individuals unable to complete their development. This phenotype highlights the critical involvement of poly(ADP-ribosyl)ating enzymes in regulating the metamorphic process. This study provides compelling evidence that these enzymes intricately coordinate transcriptional changes in both developmental and metabolic pathways during metamorphosis. Specifically, they promote the expression of genes crucial for pupation, while simultaneously negatively regulating the expression of metabolic genes before the transition to the pupal stage. Additionally, these enzymes suppress the expression of genes that are no longer required during this transformative period. These findings shed light on the intricate interplay between poly(ADP-ribosyl)ating enzymes, developmental processes, and metabolic regulation before metamorphosis and highlight a new role of poly(ADP-ribosyl)ating enzymes in the global regulation of transcription. |
| Colombo, M., Grauso, L., Lanzotti, V., Incerti, G., Adamo, A., Storlazzi, A., Gigliotti, S., Mazzoleni, S. (2023). Self-DNA Inhibition in Drosophila melanogaster Development: Metabolomic Evidence of the Molecular Determinants. Biology, 12(11) PubMed ID: 37997977
Summary: This study investigated the effects of dietary delivered self-DNA in the model insect Drosophila melanogaster. Self-DNA administration resulted in low but significant lethality in Drosophila larvae and considerably extended the fly developmental time. This was characterized by the abnormal persistence of the larvae in the L2 and L3 stages, which largely accounted for the average 72 h delay observed in pupariation, as compared to controls. In addition, self-DNA exposure affected adult reproduction by markedly reducing both female fecundity and fertility, further demonstrating its impact on Drosophila developmental processes. The effects on the metabolites of D. melanogaster larvae after exposure to self-DNA were studied by NMR, LC-MS, and molecular networking. The results showed that self-DNA feeding reduces the amounts of all metabolites, particularly amino acids and N-acyl amino acids, which are known to act as lipid signal mediators. An increasing amount of phloroglucinol was found after self-DNA exposure and correlated to developmental delay and egg-laying suppression. Pidolate, a known intermediate in the γ-glutamyl cycle, also increased after exposure to self-DNA and correlated to the block of insect oogenesis. | Xu, B., Hwangbo, D. S., Saurabh, S., Rosensweig, C., Allada, R., Kath, W. L., Braun, R. (2023). Temperature-driven coordination of circadian transcriptome regulation. bioRxiv, PubMed ID: 37961403
Summary: The circadian rhythm is an evolutionarily-conserved molecular oscillator that enables species to anticipate rhythmic changes in their environment. At a molecular level, the core clock genes induce a circadian oscillation in thousands of genes in a tissue-specific manner, orchestrating myriad biological processes. While studies have investigated how the core clock circuit responds to environmental perturbations such as temperature, the downstream effects of such perturbations on circadian regulation remain poorly understood. By analyzing bulk-RNA sequencing of Drosophila fat bodies harvested from flies subjected to different environmental conditions, a highly condition-specific circadian transcriptome was characterized. Further employing a reference-based gene regulatory network (Reactome), evidence was found of increased gene-gene coordination at low temperatures and synchronization of rhythmic genes that are network neighbors. The results point to the mechanisms by which the circadian clock mediates the fly's response to seasonal changes in temperature. |
Thursday, July 25th - Chromatin |
| Chavan, A., Isenhart, R., Nguyen, S. C., Kotb, N., Harke, J., Sintsova, A., Ulukaya, G., Uliana, F., Ashiono, C., Kutay, U., Pegoraro, G., Rangan, P., Joyce, E. F., Jagannathan, M. (2023). A nuclear architecture screen in Drosophila identifies Stonewall as a link between chromatin position at the nuclear periphery and germline stem cell fate. bioRxiv, PubMed ID: 38014085
Summary: The association of genomic loci to the nuclear periphery is proposed to facilitate cell-type specific gene repression and influence cell fate decisions. However, the interplay between gene position and expression remains incompletely understood, in part because the proteins that position genomic loci at the nuclear periphery remain unidentified. This study used an Oligopaint-based HiDRO screen targeting ~1000 genes to discover novel regulators of nuclear architecture in Drosophila cells. The heterochromatin-associated protein, Stonewall (Stwl), was identified as a factor promoting perinuclear chromatin positioning. In female germline stem cells (GSCs), Stwl binds and positions chromatin loci, including GSC differentiation genes, at the nuclear periphery. Strikingly, Stwl-dependent perinuclear positioning is associated with transcriptional repression, highlighting a likely mechanism for Stwl's known role in GSC maintenance and ovary homeostasis. Thus, this study identifies perinuclear anchors in Drosophila and demonstrates the importance of gene repression at the nuclear periphery for cell fate. | Niederhuber, M. J., Leatham-Jensen, M., McKay, D. J. (2023). The SWI/SNF nucleosome remodeler constrains enhancer activity during Drosophila wing development. Genetics, PubMed ID: 37949841
Summary: It is currently unclear how the short stretches of DNA that are individually unmask by transcription factor binding yield the kilobase-sized accessible regions characteristic of active enhancers. A genetic screen was performed to investigate the role of nucleosome remodelers in control of dynamic enhancer activity. The Drosophila SWI/SNF complex, BAP, was shown to be required for repression of a temporally dynamic enhancer, brdisc. Contrary to expectations, it was found that the BAP-specific subunit Osa is dispensable for mediating changes in chromatin accessibility between early and late stages of wing development. Instead, it was found that Osa is required to constrain the levels of brdisc activity when the enhancer is normally active. Genome-wide profiling reveals that Osa directly binds brdisc as well as thousands of other developmentally dynamic regulatory sites. Transgenic reporter analyses demonstrate that Osa is required for activation and for constraint of different sets of target enhancers in the same cells. Moreover, Osa loss results in hyperactivation of the Notch ligand Delta and development of ectopic sensory structures patterned by Notch signaling early in development. Together, these findings indicate that proper constraint of enhancer activity is necessary for regulation of dose-dependent developmental events. |
| Zhao, X., Yang, X., Lv, P., Xu, Y., Wang, X., Zhao, Z., Du, J. (2024). Polycomb regulates circadian rhythms in Drosophila in clock neurons. Life science alliance, 7(1) PubMed ID: 37914396
Summary: Circadian rhythms are essential physiological feature for most living organisms. Previous studies have shown that epigenetic regulation plays a crucial role. There is a knowledge gap in the chromatin state of some key clock neuron clusters. This study shows that circadian rhythm is affected by the epigenetic regulator Polycomb (Pc) within the Drosophila clock neurons. To investigate the molecular mechanisms underlying the roles of Pc in these clock neuron clusters, targeted DamID (TaDa) was used to identify genes significantly bound by Pc in the neurons marked by C929-Gal4 (including l-LNvs cluster), R6-Gal4 (including s-LNvs cluster), R18H11-Gal4 (including DN1 cluster), and DVpdf-Gal4, pdf-Gal80 (including LNds cluster). It shows that Pc binds to the genes involved in the circadian rhythm pathways, arguing a direct role for Pc in regulating circadian rhythms through specific clock genes. This study shows the identification of Pc targets in the clock neuron clusters, providing potential resource for understanding the regulatory mechanisms of circadian rhythms by the PcG complex. Thus, this study provided an example for epigenetic regulation of adult behavior. | Josserand, M., Rubanova, N., Stefanutti, M., Roumeliotis, S., Espenel, M., Marshall, O. J., Servant, N., Gervais, L., Bardin, A. J. (2023). Chromatin state transitions in the Drosophila intestinal lineage identify principles of cell-type specification. Dev Cell, 58(24):3048-3063. PubMed ID: 38056452
Summary: Tissue homeostasis relies on rewiring of stem cell transcriptional programs into those of differentiated cells. This study investigated changes in chromatin occurring in a bipotent adult stem cells. Combining mapping of chromatin-associated factors with statistical modeling, genome-wide transitions during differentiation were identified in the adult Drosophila intestinal stem cell (ISC) lineage. Active, stem-cell-enriched genes transition to a repressive heterochromatin protein-1-enriched state more prominently in enteroendocrine cells (EEs) than in enterocytes (ECs), in which the histone H1-enriched Black state is preeminent. In contrast, terminal differentiation genes associated with metabolic functions follow a common path from a repressive, primed, histone H1-enriched Black state in ISCs to active chromatin states in EE and EC cells. Furthermore, it was found that lineage priming has an important function in adult ISCs, and histone H1 was identfied as a mediator of this process. These data define underlying principles of chromatin changes during adult multipotent stem cell differentiation. |
| Zhimulev, I. F., Vatolina, T. Y., Pokholkova, G. V., Antonenko, O. V., Maltseva, M. V. (2023). Different Protein Groups Involved in Transcription Regulation in Development and Housekeeping Genes in Drosophila. Doklady Biochemistry and biophysics, 512(1):261-265 PubMed ID: 38093127
Summary: Antibodies to histone modifications and an insulator protein involved in the processes of transcription initiation and elongation are mapped in Drosophila polytene chromosomes. The CHRIZ protein (chromatin insulator) and H3K36me3 histone modification (RNA elongation) are detected only in the localization of housekeeping genes (interbands and gray bands of polytene chromosomes) and never in the regions of developmental genes (black bands and large puffs arising from them). Antibodies to H3S10P histone modification, which is associated with the initial elongation of the RNA strand during transcription, are found exclusively in small puffs, but not in housekeeping gene localization sites or large ecdysone-induced puffs, where housekeeping genes are localized. Antibodies to H4R3me2 histone modification (a co-repressor of the ecdysone receptor) are detected only in large ecdysone-induced puffs. | Lundkvist, M. J., Lizana, L., Schwartz, Y. B. (2023). Forecasting histone methylation by Polycomb complexes with minute-scale precision. Sci Adv, 9(51):eadj8198 PubMed ID: 38134278
Summary: Animals use the Polycomb system to epigenetically repress developmental genes. The repression requires trimethylation of lysine 27 of histone H3 (H3K27me3) by Polycomb Repressive Complex 2 (PRC2), but the dynamics of this process is poorly understood. To bridge the gap, a computational model was developed that forecasts H3K27 methylation in Drosophila with high temporal resolution and spatial accuracy of contemporary experimental techniques. Using this model, it was shown that pools of methylated H3K27 in dividing cells are defined by the effective concentration of PRC2 and the replication frequency. The allosteric stimulation by preexisting H3K27me3 makes PRC2 better in methylating developmental genes as opposed to indiscriminate methylation throughout the genome. Applied to Drosophila development, this model argues that, in this organism, the intergenerationally inherited H3K27me3 does not "survive" rapid cycles of embryonic chromatin replication and is unlikely to transmit the memory of epigenetic repression to the offspring. This model is adaptable to other organisms, including mice and humans. |
Wednesday, July 24th - RNAs and Transposons |
| Khanal, S., de Cruz, M., Strickland, B., Mansfield, K., Lai, E. C., Flynt, A. (2023). A tailed mirtron promotes longevity in Drosophila. Nucleic Acids Res, PubMed ID: 38048325
Summary: Thousands of atypical microRNAs (miRNAs) have been described in the genomes of animals; however, it is unclear if many of these non-canonical miRNAs can measurably influence phenotypes. Mirtrons are the largest class of non-canonical miRNAs that are produced from hairpins excised by splicing, which after debranching become substrates for Dicer and load into RISC. Most mirtrons require additional processing after splicing to remove 'tail' residues interposed between one of the host intron splice sites and base of the hairpin precursor structure. Despite most mirtrons requiring tail removal no function has been elucidated for a tailed species, indeed for all mirtrons identified function has only been assigned to a single species. This study examined miR-1017, a mirtron with a 3' tail, which is well expressed and conserved in Drosophila species. miR-1017 can extend lifespan when ectopically expressed in the neurons, which seems partly due to this miRNA targeting its host transcript, acetylcholine receptor Dα2. Unexpectedly it was found that not only did miR-1017 function in trans but also in cis by affecting splicing of Dα2. This suggests a mechanism for mirtron evolution where initial roles of structural elements in splicing lead to secondary acquisition of trans-regulatory function. | Shi, J., Xu, J., Ma, J., He, F. (2023). tRNA-derived small RNAs are embedded in the gene regulatory program instructing Drosophila metamorphosis. Genome research, 33(12):2119-2132 PubMed ID: 37973194
Summary: A class of noncoding RNAs, referred to as tsRNAs, is emerging with a potential to exert a new layer in gene regulation. These RNAs are breakdown products of tRNAs, either through active processing or passive cleavage or both. Since tRNAs are part of the general machinery for translation, their expression levels and activities are tightly controlled, raising the possibility that their breakdown products, tsRNAs, may provide a link between the overall translational status of a cell to specific changes in gene regulatory network. It was hypothesized that Drosophila pupation, being a special developmental stage during which there is a global limitation of nutrients, represents a system in which such a link may readily reveal itself. This study shows that specific tsRNAs indeed exhibit a dynamic accumulation upon entering the pupal stage. Experiments are described to characterize the mode of tsRNA action and, through the use of such gained knowledge, conduct a genome-wide analysis to assess the functions of dynamically expressed tsRNAs. The results show that the predicted target genes are highly enriched in biological processes specific to this stage of development including metamorphosis. It is further shown that tsRNA action is required for successful pupation, providing direct support to the hypothesis that tsRNAs accumulated during this stage are critical to the gene expression program at this stage of development. |
| Wharton, T. H., Marhabaie, M., Wharton, R. P. (2023). Significant roles in RNA-binding for the amino-terminal domains of Drosophila Pumilio and Nanos. bioRxiv, PubMed ID: 37961211
Summary: The Drosophila Pumilio (Pum) and Nanos (Nos) RNA-binding proteins govern abdominal segmentation in the early embryo, as well as a variety of other events during development. They bind together to a compound Nanos Response Element (NRE) present in thousands of maternal mRNAs in the ovary and embryo, including hunchback (hb) mRNA, thereby regulating poly-adenylation, translation, and stability. Many studies support a model in which mRNA recognition and effector recruitment are achieved by distinct regions of each protein. The well-ordered Pum and Nos RNA-binding domains (RBDs) are sufficient to specifically recognize NREs; the relatively larger low-complexity N-terminal domains (NTDs) of each protein have been thought to act by recruiting mRNA regulators. This study used yeast interaction assays to show that the NTDs also play a significant role in recognition of the NRE, acting via two mechanisms. First, the Pum and Nos NTDs interact in trans to promote assembly of the Pum/Nos/NRE ternary complex. Second, the Pum NTD acts via an unknown mechanism in cis, modifying base recognition by its RBD. These activities of the Pum NTD are important for its regulation of maternal hb mRNA in vivo. | Zhang, S., Wang, R., Zhu, X., Zhang, L., Liu, X., Sun, L. (2023). Characteristics and expression of lncRNA and transposable elements in Drosophila aneuploidy. iScience, 26(12):108494 PubMed ID: 38125016
Summary: Aneuploidy can globally affect the expression of the whole genome, which is detrimental to organisms. Dosage-sensitive regulators usually have multiple intermolecular interactions, and changes in their stoichiometry are responsible for the dysregulation of the regulatory network. Currently, studies on noncoding genes in aneuploidy are relatively rare. This study examined the characteristics and expression profiles of long noncoding RNAs (lncRNAs) and transposable elements (TEs) in aneuploid Drosophila. It was found that lncRNAs and TEs are affected by genomic imbalance and appear to be more sensitive to an inverse dosage effect than mRNAs. Several dosage-sensitive lncRNAs and TEs were detected for their expression patterns during embryogenesis, and their biological functions in the ovary and testes were investigated using tissue-specific RNAi. This study advances understanding of the noncoding sequences in imbalanced genomes and provides a novel perspective for the study of aneuploidy-related human diseases such as cancer. |
| Jagtap, P. K. A., Muller, M., Kiss, A. E., Thomae, A. W., Lapouge, K., Beck, M., Becker, P. B., Hennig, J. (2023). Structural basis of RNA-induced autoregulation of the DExH-type RNA helicase maleless. Mol Cell, 83(23):4318-4333.e4310 PubMed ID: 37989319
Summary: RNA unwinding by DExH-type helicases underlies most RNA metabolism and function. It remains unresolved if and how the basic unwinding reaction of helicases is regulated by auxiliary domains. This study explored the interplay between the RecA and auxiliary domains of the RNA helicase maleless (MLE) from Drosophila using structural and functional studies. MLE was shown to exist in a dsRNA-bound open conformation and that the auxiliary dsRBD2 domain aligns the substrate RNA with the accessible helicase tunnel. In an ATP-dependent manner, dsRBD2 associates with the helicase module, leading to tunnel closure around ssRNA. Furthermore, these structures provide a rationale for blunt-ended dsRNA unwinding and 3'-5' translocation by MLE. Structure-based MLE mutations confirm the functional relevance of this model for RNA unwinding. These findings contribute to understanding of the fundamental mechanics of auxiliary domains in DExH helicase MLE, which serves as a model for its human ortholog and potential therapeutic target, DHX9/RHA. | Crane, A. B., Jetti, S. K., Littleton, J. T. (2024). A stochastic RNA editing process targets a limited number of sites in individual Drosophila glutamatergic motoneurons. bioRxiv, PubMed ID: 38798345
Summary: RNA editing is a post-transcriptional source of protein diversity and occurs across the animal kingdom. Given the complete profile of mRNA targets and their editing rate in individual cells is unclear, this study analyzed single cell RNA transcriptomes from Drosophila larval tonic and phasic glutamatergic motoneuron subtypes to determine the most highly edited targets and identify cell-type specific editing. From approximately 15,000 genes encoded in the genome, 316 high confidence A-to-I canonical RNA edit sites were identified, with 102 causing missense amino acid changes in proteins regulating membrane excitability, synaptic transmission, and cellular function. Some sites showed 100% editing in single neurons as observed with mRNAs encoding mammalian AMPA receptors. However, most sites were edited at lower levels and generated variable expression of edited and unedited mRNAs within individual neurons. Together, these data provide insights into how the RNA editing landscape alters protein function to modulate the properties of two well-characterized neuronal populations in Drosophila . |
Tuesday, July 23rd - Synapse and Vesicles |
| Zhang, Y., Wang, T., Cai, Y., Cui, T., Kuah, M., Vicini, S., Wang, T. (2023). Role of alpha2delta-3 in regulating calcium channel localization at presynaptic active zones during homeostatic plasticity. Frontiers in molecular neuroscience, 16:1253669 PubMed ID: 38025261
Summary: The homeostatic modulation of synaptic transmission is an evolutionarily conserved mechanism that is critical for stabilizing the nervous system. At the Drosophila neuromuscular junction (NMJ), presynaptic homeostatic potentiation (PHP) compensates for impairments in postsynaptic glutamate receptors due to pharmacological blockade or genetic deletion. During PHP, there is an increase in presynaptic neurotransmitter release, counteracting postsynaptic changes and restoring excitation to baseline levels. Previous studies have shown that α2δ-3 (straightjacket), an auxiliary subunit of voltage-gated calcium channels (VGCCs), is essential for both the rapid induction and sustained expression of PHP at the Drosophila NMJ. However, the molecular mechanisms by which α2δ-3 regulates neurotransmitter release during PHP remain to be elucidated. This study utilized electrophysiological, confocal imaging, and super-resolution imaging approaches to explore how α2δ-3 regulates synaptic transmission during PHP. These findings suggest that α2δ-3 governs PHP by controlling the localization of the calcium channel pore-forming α1 subunit at presynaptic release sites, or active zones. Moreover, the role of two structural domains within α2δ-3 in regulating neurotransmitter release and calcium channel localization was examined. The results highlight that these domains in α2δ-3 serve distinct functions in controlling synaptic transmission and presynaptic calcium channel abundance, at baseline in the absence of perturbations and during PHP. In summary, this research offers compelling evidence that α2δ-3 is an indispensable signaling component for controlling calcium channel trafficking and stabilization in homeostatic plasticity. | Kang, C. J., Guzmán-Clavel, L. E., Lei, K., Koo, M., To, S., Roche, J. P. (2023). The exocyst subunit Sec15 is critical for proper synaptic development and function at the Drosophila NMJ. Molecular and cellular neurosciences, 128:103914 PubMed ID: 38086519
Summary: The exocyst protein complex is important for targeted vesicle fusion in a variety of cell types, however, its function in neurons is still not entirely known. This study found that presynaptic knockdown (KD) of the exocyst component sec15 by transgenic RNAi expression caused a number of unexpected morphological and physiological defects in the synapse. These include the development of active zones (AZ) devoid of essential presynaptic proteins, an increase in the branching of the presynaptic arbor, the appearance of satellite boutons, and a decrease in the amplitude of stimulated postsynaptic currents as well as a decrease in the frequency of spontaneous synaptic vesicle release. The release of extracellular vesicles from the presynaptic neuron was greatly diminished in the Sec15 KDs. These effects were mimicked by presynaptic knockdown of Rab11, a protein known to interact with the exocyst. sec15 RNAi expression caused an increase in phosphorylated Mothers against decapentaplegic (pMad) in the presynaptic terminal, an indication of enhanced bone morphogenic protein (BMP) signaling. Some morphological phenotypes caused by Sec15 knockdown were reduced by attenuation of BMP signaling through knockdown of wishful thinking (Wit), while other phenotypes were unaffected. Individual knockdown of multiple proteins of the exocyst complex also displayed a morphological phenotype similar to Sec15 KD. It is concluded that Sec15, functioning as part of the exocyst complex, is critically important for proper formation and function of neuronal synapses. A model is proposed in which Sec15 is involved in the trafficking of vesicles from the recycling endosome to the cell membrane as well as possibly trafficking extracellular vesicles for presynaptic release and these processes are necessary for the correct structure and function of the synapse. |
| DePew, A. T., Bruckner, J. J., O'Connor-Giles, K. M., Mosca, T. J. (2023). Neuronal LRP4 directs the development, maturation, and cytoskeletal organization of peripheral synapses. bioRxiv, PubMed ID: 37961323
Summary: Synapse development requires multiple signaling pathways to accomplish the myriad of steps needed to ensure a successful connection. Transmembrane receptors on the cell surface are optimally positioned to facilitate communication between the synapse and the rest of the neuron and often function as synaptic organizers to synchronize downstream signaling events. One such organizer, the LDL receptor-related protein LRP4, is a cell surface receptor most well-studied postsynaptically at mammalian neuromuscular junctions. Recent work, however, has identified emerging roles for LRP4 as a presynaptic molecule, but how LRP4 acts as a presynaptic organizer, what roles LRP4 plays in organizing presynaptic biology, and the downstream mechanisms of LRP4 are not well understood. This study shows that LRP4 functions presynaptically at Drosophila neuromuscular synapses, acting in motor neurons to instruct multiple aspects of pre- and postsynaptic development. Loss of presynaptic LRP4 results in a range of developmental defects, impairing active zone organization, synapse growth, physiological function, microtubule organization, synaptic ultrastructure, and synapse maturation. It was further demonstrated that LRP4 promotes most aspects of presynaptic development via a downstream SR-protein kinase, SRPK79D. SRPK79D overexpression suppresses synaptic defects associated with loss of lrp4. These data demonstrate a function for LRP4 as a peripheral synaptic organizer acting presynaptically, highlight a downstream mechanism conserved with its CNS function, and indicate previously unappreciated roles for LRP4 in cytoskeletal organization, synapse maturation, and active zone organization, underscoring its developmental importance. | Waller, T. J., Collins, C. A. (2023). Opposing roles of Fos, Raw, and SARM1 in the regulation of axonal degeneration and synaptic structure. Frontiers in cellular neuroscience, 17:1283995 PubMed ID: 38099151
Summary: The degeneration of injured axons is driven by conserved molecules, including the sterile armadillo TIR domain-containing protein SARM1, the cJun N-terminal kinase JNK, and regulators of these proteins. These molecules are also implicated in the regulation of synapse development though the mechanistic relationship of their functions in degeneration vs. development is poorly understood. This study uncovered disparate functional relationships between SARM1 and the transmembrane protein Raw in the regulation of Wallerian degeneration and synaptic growth in motoneurons of Drosophila melanogaster. Genetic data suggest that Raw antagonizes the downstream output MAP kinase signaling mediated by Drosophila SARM1 (dSarm). This relationship is revealed by dramatic synaptic overgrowth phenotypes at the larval neuromuscular junction when motoneurons are depleted for Raw or overexpress dSarm. While Raw antagonizes the downstream output of dSarm to regulate synaptic growth, it shows an opposite functional relationship with dSarm for axonal degeneration. Loss of Raw leads to decreased levels of dSarm in axons and delayed axonal degeneration that is rescued by overexpression of dSarm, supporting a model that Raw promotes the activation of dSarm in axons. However, inhibiting Fos also decreases dSarm levels in axons but has the opposite outcome of enabling Wallerian degeneration. The combined genetic data suggest that Raw, dSarm, and Fos influence each other's functions through multiple points of regulation to control the structure of synaptic terminals and the resilience of axons to degeneration. |
| Ermanoska, B., Rodal, A. A. (2023). Non-muscle myosin II regulates presynaptic actin assemblies and neuronal mechanobiology. bioRxiv, PubMed ID: 38014140
Summary: Neuromuscular junctions (NMJs) are evolutionarily ancient, specialized contacts between neurons and muscles. Axons and NMJs must endure mechanical strain through a lifetime of muscle contraction, making them vulnerable to aging and neurodegenerative conditions. However, cellular strategies for mitigating this mechanical stress remain unknown. In this study, Drosophila larval NMJs were used to investigate the role of actin and myosin (actomyosin)-mediated contractility in generating and responding to cellular forces at the neuron-muscle interface. A new long-lived, low-turnover presynaptic actin core traversing the NMJ, which partly co-localizes with non-muscle myosin II (NMII). Neuronal RNAi of NMII induced disorganization of this core, suggesting that this structure might have contractile properties. Interestingly, neuronal RNAi of NMII also decreased NMII levels in the postsynaptic muscle proximal to neurons, suggesting that neuronal actomyosin rearrangements propagate their effects transsynaptically. Reduced Integrin levels were observed upon NMII knockdown, indicating that neuronal actomyosin disruption triggers rearrangements of Integrin-mediated connections between neurons and surrounding muscle tissue. In summary, this study identifies a previously uncharacterized presynaptic actomyosin subpopulation that upholds the neuronal mechanical continuum, transmits signals to adjacent muscle tissue, and collaborates with Integrin receptors to govern the mechanobiology of the neuromuscular junction. | Tsarouhas, V., Liu, D., Tsikala, G., Engstrom, Y., Strigini, M., Samakovlis, C. (2023). A surfactant lipid layer of endosomal membranes facilitates airway gas filling in Drosophila.. Curr Biol, 33(23):5132-5146. PubMed ID: 37992718
Summary: The mechanisms underlying the construction of an air-liquid interface in respiratory organs remain elusive. This study used live imaging and genetic analysis to describe the morphogenetic events generating an extracellular lipid lining of the Drosophila airways required for their gas filing and animal survival. Sequential Rab39/Syx1A/Syt1-mediated secretion of lysosomal acid sphingomyelinase (Drosophila ASM [dASM]) and Rab11/Rab35/Syx1A/Rop-dependent exosomal secretion provides distinct components for lipid film assembly. Tracheal inactivation of Rab11 or Rab35 or loss of Rop results in intracellular accumulation of exosomal, multi-vesicular body (MVB)-derived vesicles. On the other hand, loss of dASM or Rab39 causes luminal bubble-like accumulations of exosomal membranes and liquid retention in the airways. Inactivation of the exosomal secretion in dASM mutants counteracts this phenotype, arguing that the exosomal secretion provides the lipid vesicles and that secreted lysosomal dASM organizes them into a continuous film. These results reveal the coordinated functions of extracellular vesicle and lysosomal secretions in generating a lipid layer crucial for airway gas filling and survival. |
Friday, July 19th - Larval and Adult Neural Structure, Development and Function |
| Ahmed, O. M., Crocker, A., Murthy, M. (2023). Transcriptional profiling of Drosophila male-specific P1 (pC1) neurons. bioRxiv, PubMed ID: 37986870
Summary: In Drosophila melanogaster, the P1 (pC1) cluster of male-specific neurons, consisting of 20 interneurons per hemibrain, both integrates sensory cues and drives or modulates behavioral programs such as courtship, in addition to contributing to a social arousal state. The behavioral function of these neurons is linked to the genes they express, which underpin their capacity for synaptic signaling, neuromodulation, and physiology. Yet, P1 (pC1) neurons have not been fully characterized at the transcriptome level. Moreover, it is unknown how the molecular landscape of P1 (pC1) neurons acutely changes after flies engage in social behaviors, where baseline P1 (pC1) neural activity is expected to increase. To address these two gaps, single cell-type RNA sequencing was used to profile and compare the transcriptomes of P1 (pC1) neurons harvested from socially paired versus solitary male flies. Compared to control transcriptome datasets, it was found that P1 (pC1) neurons are enriched in 2,665 genes, including those encoding receptors, neuropeptides, and cell-adhesion molecules (dprs/DIPs). Furthermore, courtship is characterized by changes in ~300 genes, including those previously implicated in regulating behavior (e.g. DopEcR, Octβ3R, Fife, kairos, radish). Finally, a suite of genes was identified that link conspecific courtship with the innate immune system. Together, these data serve as a molecular map for future studies of an important set of higher-order and sexually-dimorphic neurons. | Xiao, N., Xu, S., Li, Z. K., Tang, M., Mao, R., Yang, T., Ma, S. X., Wang, P. H., Li, M. T., Sunilkumar, A., Rouyer, F., Cao, L. H., Luo, D. G. (2023). A single photoreceptor splits perception and entrainment by cotransmission. Nature, 623(7987):562-570 PubMed ID: 37880372
Summary: Vision enables both image-forming perception, driven by a contrast-based pathway, and unconscious non-image-forming circadian photoentrainment, driven by an irradiance-based pathway. Although two distinct photoreceptor populations are specialized for each visual task, image-forming photoreceptors can additionally contribute to photoentrainment of the circadian clock in different species. However, it is unknown how the image-forming photoreceptor pathway can functionally implement the segregation of irradiance signals required for circadian photoentrainment from contrast signals required for image perception. This study reports that the Drosophila R8 photoreceptor separates image-forming and irradiance signals by co-transmitting two neurotransmitters, histamine and acetylcholine. This segregation is further established postsynaptically by histamine-receptor-expressing unicolumnar retinotopic neurons and acetylcholine-receptor-expressing multicolumnar integration neurons. The acetylcholine transmission from R8 photoreceptors is sustained by an autocrine negative feedback of the cotransmitted histamine during the light phase of light-dark cycles. At the behavioural level, elimination of histamine and acetylcholine transmission impairs R8-driven motion detection and circadian photoentrainment, respectively. Thus, a single type of photoreceptor can achieve the dichotomy of visual perception and circadian photoentrainment as early as the first visual synapses, revealing a simple yet robust mechanism to segregate and translate distinct sensory features into different animal behaviours. |
| Dillon, N. R., Manning, L., Hirono, K., Doe, C. Q. (2023). Seven-up acts in neuroblasts to specify adult central complex neuron identity and initiate neuroblast decommissioning. bioRxiv, PubMed ID: 37961302
Summary: An open question in neurobiology is how diverse neuron cell types are generated from a small number of neural stem cells. In the Drosophila larval central brain, there are eight bilateral Type 2 neuroblast (T2NB) lineages that express a suite of early temporal factors followed by a different set of late temporal factors and generate the majority of the central complex (CX) neurons. The early-to-late switch is triggered by the orphan nuclear hormone receptor Seven-up (Svp), yet little is known about this Svp-dependent switch in specifying CX neuron identities. This study shows (i) birthdate the CX neurons P-EN and P-FN (early and late, respectively); (ii) show that Svp is transiently expressed in all early T2NBs; and (iii) show that loss of Svp expands the population of early born P-EN neurons at the expense of late born P-FN neurons. Furthermore, in the absence of Svp, T2NBs fail decommissioning and abnormally extend their lineage into week-old adults. It is concludes that Svp is required to specify CX neuron identity, as well as to initiate T2NB decommissioning. | Dallmann, C. J., Agrawal, S., Cook, A., Brunton, B. W., Tuthill, J. C. (2023). Presynaptic inhibition selectively suppresses leg proprioception in behaving Drosophila. bioRxiv, PubMed ID: 37961558
Summary: The sense of proprioception is mediated by internal mechanosensory neurons that detect joint position and movement. To support a diverse range of functions, from stabilizing posture to coordinating movements, proprioceptive feedback to limb motor control circuits must be tuned in a context-dependent manner. How proprioceptive feedback signals are tuned to match behavioral demands remains poorly understood. Using calcium imaging in behaving Drosophila, this study found that the axons of position-encoding leg proprioceptors are active across behaviors, whereas the axons of movement-encoding leg proprioceptors are suppressed during walking and grooming. Using connectomics, a specific class of interneurons was identified that provide GABAergic presynaptic inhibition to the axons of movement-encoding proprioceptors. These interneurons are active during self-generated but not passive leg movements and receive input from descending neurons, suggesting they are driven by predictions of leg movement originating in the brain. Predictively suppressing expected proprioceptive feedback provides a mechanism to attenuate reflexes that would otherwise interfere with voluntary movement. |
| Tao, L., Wechsler, S. P., Bhandawat, V. (2023). Sensorimotor transformation underlying odor-modulated locomotion in walking Drosophila. Nat Commun, 14(1):6818 PubMed ID: 37884581
Summary: Most real-world behaviors - such as odor-guided locomotion - are performed with incomplete information. Activity in olfactory receptor neuron (ORN) classes provides information about odor identity but not the location of its source. This study investigated the sensorimotor transformation that relates ORN activation to locomotion changes in Drosophila by optogenetically activating different combinations of ORN classes and measuring the resulting changes in locomotion. Three features describe this sensorimotor transformation: First, locomotion depends on both the instantaneous firing frequency (f) and its change (df); the two together serve as a short-term memory that allows the fly to adapt its motor program to sensory context automatically. Second, the mapping between (f, df) and locomotor parameters such as speed or curvature is distinct for each pattern of activated ORNs. Finally, the sensorimotor mapping changes with time after odor exposure, allowing information integration over a longer timescale. | Sang, J., Dhakal, S., Shrestha, B., Nath, D. K., Kim, Y., Ganguly, A., Montell, C., Lee, Y. (2023). A single pair of pharyngeal neurons functions as a commander to reject high salt in Drosophila melanogaster. bioRxiv, PubMed ID: 37904986
Summary: Salt is a crucial for survival, while excessive NaCl can be detrimental. In the fruit fly, Drosophila melanogaster, an internal taste organ, the pharynx, is a critical gatekeeper impacting the decision to accept or reject a food. Currently, understanding of the mechanism through which pharyngeal gustatory receptor neurons (GRNs) sense high salt are rudimentary. This study found that a member of the ionotropic receptor family, IR60b, is exclusively expressed in a pair of GRNs activated by high salt. Using a two-way choice assay (DrosoX) to measure ingestion, it was demonstrated that IR60b and two coreceptors IR25a and IR76b, are required to prevent high salt consumption. Mutants lacking external taste organs but retaining the pharynx exhibit much higher salt avoidance than flies with all taste organs but missing the three IRs. These findings highlight the critical role for IRs in a pair of pharyngeal GRNs to control ingestion of high salt. |
Thursday, July 18th - Stress |
| Santos, M. A., Antunes, M. A., Grandela, A., Carromeu-Santos, A., Quina, A. S., Santos, M., Matos, M., Simoes, P. (2023). Heat-induced female biased sex ratio during development is not mitigated after prolonged thermal selection. BMC ecology and evolution, 23(1):64 PubMed ID: 37919666
Summary: The negative impacts of climate change on biodiversity are consistently increasing. Developmental stages are particularly sensitive in many ectotherms. Moreover, sex-specific differences in how organisms cope with thermal stress can produce biased sex ratios upon emergence, with potentially major impacts on population persistence. This is an issue that needs investigation, particularly testing whether thermal selection can alleviate sex ratio distortions in the long-term is a critical but neglected issue. This study reports an experiment analyzing the sex ratio patterns at different developmental temperatures in Drosophila subobscura populations subjected to long-term experimental evolution (~ 30 generations) under a warming environment. Exposure to high developmental temperatures consistently promotes sex ratio imbalance upon emergence, with a higher number of female than male offspring. Furthermore, it was found that thermal selection resulting from evolution in a warming environment did not alleviate such sex ratio distortions generated by heat stress. This study has demonstrated that heat stress during development can lead to clear sex ratio deviations upon emergence likely because of differential survival between sexes. In face of these findings, it is likely that sex ratio deviations of this sort occur in natural populations when facing environmental perturbation. The inability of many insects to avoid thermal shifts during their (more) sessile developmental stages makes this finding particularly troublesome for population subsistence in face of climate warming events. | Olufs, Z. P. G., Wassarman, D. A., Perouansky, M. (2023). Stress pathways induced by volatile anesthetics and failure of preconditioning in a mitochondrial Complex I mutant. Anesthesiology, PubMed ID: 38118175
Summary: Carriers of mutations in the mitochondrial electron transport chain (mETC) are at increased risk of anesthetic-induced neurotoxicity. To investigate the neurotoxicity mechanism, a Drosophila model of Leigh syndrome was used. Model flies carried a mutation in ND23 that encodes an mETC Complex I subunit. Why ND2360114 mutants become susceptible to lethal, oxygen-modulated neurotoxicity within 24 h of exposure to isoflurane but not sevoflurane was investigated. Transcriptomics and qRT-PCR were used to identify genes that are differentially expressed in mutants but not wild type fly heads at 30 min after exposure to high versus low toxicity conditions. The mutation had a greater effect on isoflurane- than sevoflurane-mediated changes in gene expression. Isoflurane and sevoflurane did not affect expression of heat shock protein (Hsp) genes (Hsp22, Hsp27, and Hsp68) in wild type flies, but isoflurane substantially increased expression of these genes in mutant flies. Furthermore, isoflurane and sevoflurane induced expression of oxidative (GstD1and GstD2) and xenobiotic (Cyp6a8 and Cyp6a14) stress genes to a similar extent in wild type flies, but the effect of isoflurane was largely reduced in ND2360114 flies. It is concluded that mutation of an mETC Complex I subunit generates differential effects of isoflurane and sevoflurane on gene expression that may underlie their differential effects on neurotoxicity. Additionally, the mutation produces resistance to preconditioning by stresses that protect the brain in other contexts. Therefore, Complex I activity modifies molecular and physiological effects of anesthetics in an anesthetic-specific manner. |
| Aguilera, J., Duan, J., Lee, S. M., Ray, M., Larschan, E. (2023). The CLAMP GA-binding transcription factor regulates heat stress-induced transcriptional repression by associating with 3D loop anchors. bioRxiv, PubMed ID: 37873306
Summary: In order to survive when exposed to heat heat stress (HS), organisms activate stress response genes and repress constitutive gene expression to prevent the accumulation of potentially toxic RNA and protein products. Although many studies have elucidated the mechanisms that drive HS-induced activation of stress response genes across species, little is known about repression mechanisms or how genes are targeted for activation versus repression context-specifically. The mechanisms of heat stress-regulated activation have been well-studied in Drosophila, in which the GA-binding transcription factor GAF is important for activating genes upon heat stress. A functionally distinct GA-binding transcription factor (TF) protein, CLAMP (Chromatin-linked adaptor for MSL complex proteins), is essential for repressing constitutive genes upon heat stress but not activation of the canonical heat stress pathway. HS induces loss of CLAMP-associated 3D chromatin loop anchors associated with different combinations of GA-binding TFs prior to HS if a gene becomes repressed versus activated. Overall, this study demonstrates that CLAMP promotes repression of constitutive genes upon HS, and repression and activation are associated with the loss of CLAMP-associated 3D chromatin loops bound by different combinations of GA-binding TFs. | Juul-Kristensen, T., Keller, J. G., Borg, K. N., Hansen, N. Y., Foldager, A., Ladegaard, R., Ho, Y. P., Loeschcke, V., Knudsen, B. R. (2023). Topoisomerase 1 Activity Is Reduced in Response to Thermal Stress in Fruit Flies and in Human HeLa Cells. Biosensors, 13(11) PubMed ID: 37998125
Summary: In the modern world with climate changes and increasing pollution, different types of stress are becoming an increasing challenge. Hence, the identification of reliable biomarkers of stress and accessible sensors to measure such biomarkers are attracting increasing attention. This study demonstrated that the activity, but not the expression, of the ubiquitous enzyme topoisomerase 1 (TOP1), as measured in crude cell extracts by the REEAD sensor system, is markedly reduced in response to thermal stress in both fruit flies (Drosophila melanogaster) and cultivated human cells. This effect was observed in response to both mild-to-moderate long-term heat stress and more severe short-term heat stress in D. melanogaster. In cultivated HeLa cells a reduced TOP1 activity was observed in response to both cold and heat stress. The reduced TOP1 activity appeared dependent on one or more cellular pathways since the activity of purified TOP1 was unaffected by the utilized stress temperatures. Successful quantitative measurement of TOP1 activity was demonstrated using an easily accessible chemiluminescence readout for REEAD pointing towards a sensor system suitable for point-of-care assessment of stress responses based on TOP1 as a biomarker. |
| Torre, M., Bukhari, H., Nithianandam, V., Zanella, C. A., Mata, D. A., Feany, M. B. (2023). A Drosophila model relevant to chemotherapy-related cognitive impairment. Sci Rep, 13(1):19290 PubMed ID: 37935827
Summary: Chemotherapy-related cognitive impairment (CRCI) is a common adverse effect of treatment and is characterized by deficits involving multiple cognitive domains including memory. Despite the significant morbidity of CRCI and the expected increase in cancer survivors over the coming decades, the pathophysiology of CRCI remains incompletely understood, highlighting the need for new model systems to study CRCI. Given the powerful array of genetic approaches and facile high throughput screening ability in Drosophila, the goal of this study was to validate a Drosophila model relevant to CRCI. The chemotherapeutic agents cisplatin, cyclophosphamide, and doxorubicin were administered to adult Drosophila. Neurologic deficits were observed with all tested chemotherapies, with doxorubicin and in particular cisplatin also resulting in memory deficits. Then histologic and immunohistochemical analysis of cisplatin-treated Drosophila tissue was performed, demonstrating neuropathologic evidence of increased neurodegeneration, DNA damage, and oxidative stress. Thus, the Drosophila model relevant to CRCI recapitulates clinical, radiologic, and histologic alterations reported in chemotherapy patients. The new Drosophila model can be used for mechanistic dissection of pathways contributing to CRCI (and chemotherapy-induced neurotoxicity more generally) and pharmacologic screens to identify disease-modifying therapies. | Glineburg, M. R., Yildirim, E., Gomez, N., Li, X., Pak, J., Altheim, C., Waksmacki, J., McInerney, G., Barmada, S. J., Todd, P. K. (2023). Stress granule formation helps to mitigate neurodegeneration. bioRxiv. PubMed ID: 37986813
Summary: Cellular stress pathways that inhibit translation initiation lead to transient formation of cytoplasmic RNA/protein complexes known as stress granules. Many of the proteins found within stress granules and the dynamics of stress granule formation and dissolution are implicated in neurodegenerative disease. Whether stress granule formation is protective or harmful in neurodegenerative conditions is not known. To address this, advantage was taken of the alphavirus protein nsP3, which selectively binds dimers of the central stress granule nucleator protein G3BP (rin in Drosophila) and markedly reduces stress granule formation without directly impacting the protein translational inhibitory pathways that trigger stress granule formation. In Drosophila and rodent neurons, reducing stress granule formation with nsP3 had modest impacts on lifespan even in the setting of serial stress pathway induction. In contrast, reducing stress granule formation in models of ataxia, amyotrophic lateral sclerosis and frontotemporal dementia largely exacerbated disease phenotypes. These data support a model whereby stress granules mitigate, rather than promote, neurodegenerative cascades. |
Wednesday, July 17th - RNA and Transposons |
| Huang, Y., Pang, Y., Xu, Y., Liu, L., Zhou, H. (2024). The identification of regulatory ceRNA network involved in Drosophila Toll immune responses. Dev Comp Immunol, 151:105105 PubMed ID: 38013113
Summary: Non-coding RNAs play important roles in the innate immunity of Drosophila, with various lncRNAs and miRNAs identified to maintain Drosophila innate immune homeostasis by regulating protein functions. However, it remains unclear whether interactions between lncRNAs and miRNAs give rise to a competing endogenous RNAs (ceRNA) network. In a previous study, it was observed the highest differential expression levels of lncRNA-CR11538, lncRNA-CR33942, and lncRNA-CR46018 in wild-type flies after Gram-positive bacterial infection, prompting an investigation of their role in the regulation of Drosophila Toll immune response through RNA-seq analysis. A comprehensive bioinformatics analysis revealed that lncRNA-CR11538, lncRNA-CR33942, and lncRNA-CR46018 are involved in defense mechanisms and stimulus response. Moreover, lncRNA-CR11538 and lncRNA-CR46018 can also participate in the metabolic recovery processes following Gram-positive bacterial infection. Subsequently, GSEA screening and RT-qPCR were employed to identify seven miRNAs (miR-957, miR-1015, miR-982, miR-993, miR-1007, miR-193, and miR-978) that may be regulated by these three lncRNAs. Furthermore, the potential target genes are predicted in the Toll signaling pathway for these miRNAs and their interaction with the three lncRNAs using TargetScan and miRanda software and preliminary verification. As a result, a potential ceRNA regulatory network was established for Toll immune responses in Drosophila, comprising three lncRNAs and seven miRNAs. This study provides evidence of a ceRNA regulatory network in Drosophila Toll immune responses and offers novel insights into understanding the regulatory networks involved in the innate immunity of other animals. | Palumbo, R. J., Yang, Y., Feigon, J., Hanes, S. D. (2024). Catalytic activity of the Bin3/MePCE methyltransferase domain is dispensable for 7SK snRNP function in Drosophila melanogaster. Genetics, 226(1) PubMed ID: 37982586
Summary: Methylphosphate Capping Enzyme (MePCE) monomethylates the gamma phosphate at the 5' end of the 7SK noncoding RNA, a modification thought to protect 7SK from degradation. 7SK serves as a scaffold for assembly of a snRNP complex that inhibits transcription by sequestering the positive elongation factor P-TEFb. While much is known about the biochemical activity of MePCE in vitro, little is known about its functions in vivo, or what roles-if any-there are for regions outside the conserved methyltransferase domain. This study investigated the role of Bin3, the Drosophila ortholog of MePCE, and its conserved functional domains in Drosophila development. bin3 mutant females had strongly reduced rates of egg-laying, which was rescued by genetic reduction of P-TEFb activity, suggesting that Bin3 promotes fecundity by repressing P-TEFb. bin3 mutants also exhibited neuromuscular defects, analogous to a patient with MePCE haploinsufficiency. These defects were also rescued by genetic reduction of P-TEFb activity, suggesting that Bin3 and MePCE have conserved roles in promoting neuromuscular function by repressing P-TEFb. Unexpectedly, it was found that a Bin3 catalytic mutant (Bin3Y795A) could still bind and stabilize 7SK and rescue all bin3 mutant phenotypes, indicating that Bin3 catalytic activity is dispensable for 7SK stability and snRNP function in vivo. Finally, a metazoan-specific motif (MSM) was found outside of the methyltransferase domain, and mutant flies were generated lacking this motif (Bin3ΔMSM). Bin3ΔMSM mutant flies exhibited some-but not all-bin3 mutant phenotypes, suggesting that the MSM is required for a 7SK-independent, tissue-specific function of Bin3. |
| Milyaeva, P. A., Kukushkina, I. V., Kim, A. I., Nefedova, L. N. (2023). Stress Induced Activation of LTR Retrotransposons in the Drosophila melanogaster. Genome. Life (Basel, Switzerland), 13(12) PubMed ID: 38137873
Summary: Retrotransposons with long terminal repeats (LTR retrotransposons) are widespread in all groups of eukaryotes and are often both the cause of new mutations and the source of new sequences. Apart from their high activity in generative and differentiation-stage tissues, LTR retrotransposons also become more active in response to different stressors. The precise causes of LTR retrotransposons' activation in response to stress, however, have not yet been thoroughly investigated. This study used RT-PCR to investigate the transcriptional profile of LTR retrotransposons and piRNA clusters in response to oxidative and chronic heat stresses. Oxford Nanopore sequencing was used to investigate the genomic environment of new insertions of the retrotransposons. Bioinformatics methods were used to find the stress-induced transcription factor binding sites in LTR retrotransposons. The transposition activity and transcription level of LTR retrotransposons in response to oxidative and chronic heat stress was studied and the contribution of various factors that can affect the increase in their expression under stress conditions was assessed: the state of the piRNA-interference system, the influence of the genomic environment on individual copies, and the presence of the stress-induced transcription factor binding sites in retrotransposon sequences. The main reason for the activation of LTR retrotransposons under stress conditions is the presence of transcription factor binding sites in their regulatory sequences, which are triggered in response to stress and are necessary for tissue regeneration processes. Stress-induced transposable element activation can function as a trigger mechanism, triggering multiple signal pathways and resulting in a polyvariant cell response. | Zhang, T., Xue, Y., Su, S., Altouma, V., Ho, K., Martindale, J. L., Lee, S. K., Shen, W., Park, A., Zhang, Y., De, S., Gorospe, M., Wang, W. (2023). RNA-binding protein Nocte regulates Drosophila development by promoting translation reinitiation on mRNAs with long upstream open reading frames. Nucleic Acids Res, PubMed ID: 38000373
Summary: RNA-binding proteins (RBPs) with intrinsically disordered regions (IDRs) are linked to multiple human disorders, but their mechanisms of action remain unclear. This study reports that one such protein, Nocte, is essential for Drosophila eye development by regulating a critical gene expression cascade at translational level. Knockout of nocte in flies leads to lethality, and its eye-specific depletion impairs eye size and morphology. Nocte preferentially enhances translation of mRNAs with long upstream open reading frames (uORFs). One of the key Nocte targets, glass mRNA, encodes a transcription factor critical for differentiation of photoreceptor neurons and accessory cells, and re-expression of Glass largely rescued the eye defects caused by Nocte depletion. Mechanistically, Nocte counteracts long uORF-mediated translational suppression by promoting translation reinitiation downstream of the uORF. Nocte interacts with translation factors eIF3 and Rack1 through its BAT2 domain, and a Nocte mutant lacking this domain fails to promote translation of glass mRNA. Notably, de novo mutations of human orthologs of Nocte have been detected in schizophrenia patients. These data suggest that Nocte family of proteins can promote translation reinitiation to overcome long uORFs-mediated translational suppression, and disruption of this function can lead to developmental defects and neurological disorders. |
| Hernandez, G., Garcia, A., Weingarten-Gabbay, S., Mishra, R. K., Hussain, T., Amiri, M., Moreno-Hagelsieb, G., Montiel-Davalos, A., Lasko, P., Sonenberg, N. (2023). Functional analysis of the AUG initiator codon context reveals novel conserved sequences that disfavor mRNA translation in eukaryotes. Nucleic Acids Res, PubMed ID: 38038264
Summary: mRNA translation is a fundamental process for life. Selection of the translation initiation site (TIS) is crucial, as it establishes the correct open reading frame for mRNA decoding. Studies in vertebrate mRNAs discovered that a purine at -3 and a G at +4 (where A of the AUG initiator codon is numbered + 1), promote TIS recognition. However, the TIS context in other eukaryotes has been poorly experimentally analyzed. This study analyzed in vitro the influence of the -3, -2, -1 and + 4 positions of the TIS context in rabbit, Drosophila, wheat, and yeast. It was observed that -3A conferred the best translational efficiency across these species. However, variability was found at the + 4 position for optimal translation. In addition, the Kozak motif that was defined from mammalian cells was only weakly predictive for wheat and essentially non-predictive for yeast. Eight conserved sequences were discovered that significantly disfavored translation. Due to the big differences in translational efficiency observed among weak TIS context sequences, a novel category was defined that was termed 'barren AUG context sequences (BACS)', which represent sequences disfavoring translation. Analysis of mRNA-ribosomal complexes structures provided insights into the function of BACS. The gene ontology of the BACS-containing mRNAs is presented. | van Lopik, J., Alizada, A., Trapotsi, M. A., Hannon, G. J., Bornelov, S., Nicholson, B.C. (2023). Unistrand piRNA clusters are an evolutionarily conserved mechanism to suppress endogenous retroviruses across the Drosophila genus. Nat Commun, 14(1):7337 PubMed ID: 37957172
Summary: The PIWI-interacting RNA (piRNA) pathway prevents endogenous genomic parasites, i.e. transposable elements, from damaging the genetic material of animal gonadal cells. Specific regions in the genome, called piRNA clusters, are thought to define each species' piRNA repertoire and therefore its capacity to recognize and silence specific transposon families. The unistrand cluster flamenco (flam) is essential in the somatic compartment of the Drosophila ovary to restrict Gypsy-family transposons from infecting the neighbouring germ cells. Disruption of flam results in transposon de-repression and sterility, yet it remains unknown whether this silencing mechanism is present more widely. This study systematically characterised 119 Drosophila species and identified five additional flam-like clusters separated by up to 45 million years of evolution. Small RNA-sequencing validated these as bona-fide unistrand piRNA clusters expressed in somatic cells of the ovary, where they selectively target transposons of the Gypsy family. Together, this study provides compelling evidence of a widely conserved transposon silencing mechanism that co-evolved with virus-like Gypsy-family transposons. |
Tuesday, July 16th - Stem Cells |
| Khanbabei, A., Segura, L., Petrossian, C., Lemus, A., Cano, I., Frazier, C., Halajyan, A., Ca, D., Loza-Coll, M. (2024). Experimental validation and characterization of putative targets of Escargot and STAT, two master regulators of the intestinal stem cells in Drosophila melanogaster. Dev Biol, 505:148-163 PubMed ID: 37952851
Summary: Many organs contain adult stem cells (ASCs) to replace cells due to damage, disease, or normal tissue turnover. ASCs can divide asymmetrically, giving rise to a new copy of themselves (self-renewal) and a sister that commits to a specific cell type (differentiation). Decades of research have led to the identification of pleiotropic genes whose loss or gain of function affect diverse aspects of normal ASC biology. Genome-wide screens of these so-called genetic "master regulator" (MR) genes, have pointed to hundreds of putative targets that could serve as their downstream effectors. This study experimentally validate and characterize the regulation of several putative targets of Escargot (Esg) and the Signal Transducer and Activator of Transcription (Stat92E, a.k.a. STAT), two known MRs in Drosophila intestinal stem cells (ISCs). The results indicate that regardless of bioinformatic predictions, most experimentally validated targets show a profile of gene expression that is consistent with co-regulation by both Esg and STAT, fitting a rather limited set of co-regulatory modalities. A bioinformatic analysis of proximal regulatory sequences in specific subsets of co-regulated targets identified additional transcription factors that might cooperate with Esg and STAT in modulating their transcription. Lastly, in vivo manipulations of validated targets rarely phenocopied the effects of manipulating Esg and STAT, suggesting the existence of complex genetic interactions among downstream targets of these two MR genes during ISC homeostasis. | Banach-Latapy, A., Rincheval, V., Briand, D., Guenal, I., Speder, P. (2023). Differential adhesion during development establishes individual neural stem cell niches and shapes adult behaviour in Drosophila. PLoS Biol, 21(11):e3002352 PubMed ID: 37943883
Summary: Neural stem cells (NSCs) reside in a defined cellular microenvironment, the niche, which supports the generation and integration of newborn neurons. The mechanisms building a sophisticated niche structure around NSCs and their functional relevance for neurogenesis are yet to be understood. In the Drosophila larval brain, the cortex glia (CG) encase individual NSC lineages in membranous chambers, organising the stem cell population and newborn neurons into a stereotypic structure. It was first found that CG wrap around lineage-related cells regardless of their identity, showing that lineage information builds CG architecture. It was then discovered that a mechanism of temporally controlled differential adhesion using conserved complexes supports the individual encasing of NSC lineages. An intralineage adhesion through homophilic Neurexin-IV and Wrapper exists between NSC lineages and CG. Loss of Neuroglian results in NSC lineages clumped together and in an altered CG network, while loss of Neurexin-IV/Wrapper generates larger yet defined CG chamber grouping several lineages together. Axonal projections of newborn neurons are also altered in these conditions. Further, the loss of these 2 adhesion complexes specifically during development were linked to locomotor hyperactivity in the resulting adults. Altogether, these findings identify a belt of adhesions building a neurogenic niche at the scale of individual stem cell and provide the proof of concept that niche properties during development shape adult behaviour. |
| Russell, S. L., Castillo, J. R., Sullivan, W. T. (2023). Wolbachia endosymbionts manipulate the self-renewal and differentiation of germline stem cells to reinforce fertility of their fruit fly host. PLoS Biol, 21(10):e3002335 PubMed ID: 37874788
Summary: The alphaproteobacterium Wolbachia pipientis infects arthropod and nematode species worldwide, making it a key target for host biological control. Wolbachia-driven host reproductive manipulations, such as cytoplasmic incompatibility (CI), are credited for catapulting these intracellular bacteria to high frequencies in host populations. Positive, perhaps mutualistic, reproductive manipulations also increase infection frequencies, but are not well understood. This study identified molecular and cellular mechanisms by which Wolbachia influences the molecularly distinct processes of germline stem cell (GSC) self-renewal and differentiation. wMel infection rescues the fertility of flies lacking the translational regulator mei-P26 and is sufficient to sustain infertile homozygous mei-P26-knockdown stocks indefinitely. Cytology revealed that wMel mitigates the impact of mei-P26 loss through restoring proper pMad, Bam, Sxl, and Orb expression. In Oregon R files with wild-type fertility, wMel infection elevates lifetime egg hatch rates. Exploring these phenotypes through dual-RNAseq quantification of eukaryotic and bacterial transcripts revealed that wMel infection rescues and offsets many gene expression changes induced by mei-P26 loss at the mRNA level. Overall, this study shows that wMel infection beneficially reinforces host fertility at mRNA, protein, and phenotypic levels, and these mechanisms may promote the emergence of mutualism and the breakdown of host reproductive manipulations. | Nelson, J. O., Kumon, T., Yamashita, Y. M. (2023). rDNA magnification is a unique feature of germline stem cells. Proc Natl Acad Sci U S A, 120(47):e2314440120 PubMed ID: 37967216
Summary: Ribosomal DNA (rDNA) encodes ribosomal RNA and exists as tandem repeats of hundreds of copies in the eukaryotic genome to meet the high demand of ribosome biogenesis. Tandemly repeated DNA elements are inherently unstable; thus, mechanisms must exist to maintain rDNA copy number (CN), in particular in the germline that continues through generations. A phenomenon called rDNA magnification was discovered over 50 y ago in Drosophila as a process that recovers the rDNA CN on chromosomes that harbor minimal CN. Recent studies indicated that rDNA magnification is the mechanism to maintain rDNA CN under physiological conditions to counteract spontaneous CN loss that occurs during aging. A previous study that explored the mechanism of rDNA magnification implied that asymmetric division of germline stem cells (GSCs) may be particularly suited to achieve rDNA magnification. However, it remains elusive whether GSCs are the unique cell type that undergoes rDNA magnification or differentiating germ cells are also capable of magnification. This study provides empirical evidence that suggests that rDNA magnification operates uniquely in GSCs, but not in differentiating germ cells. Computer simulation is provided that suggests that rDNA magnification is only achievable through asymmetric GSC divisions. Despite known plasticity and transcriptomic similarity between GSCs and differentiating germ cells, GSCs' unique ability to divide asymmetrically serves a critical role of maintaining rDNA CN through generations, supporting germline immortality. |
| Pang, L. Y., DeLuca, S., Zhu, H., Urban, J. M., Spradling, A. C. (2023). Chromatin and gene expression changes during female Drosophila germline stem cell development illuminate the biology of highly potent stem cells. Elife, 12 PubMed ID: 37831064
Summary: Highly potent animal stem cells either self transposons renew or launch complex differentiation programs, using mechanisms that are only partly understood. Drosophila female germline stem cells (GSCs) perpetuate without change over evolutionary time and generate cystoblast daughters that develop into nurse cells and oocytes. Cystoblasts initiate differentiation by generating a transient syncytial state, the germline cyst, and by increasing pericentromeric H3K9me3 modification, actions likely to suppress transposable element activity. Relatively open GSC chromatin is further restricted by Polycomb repression of testis or somatic cell-expressed genes briefly active in early female germ cells. Subsequently, Neijre/CBP and Myc help upregulate growth and reprogram GSC metabolism by altering mitochondrial transmembrane transport, gluconeogenesis, and other processes. In all these respects GSC differentiation resembles development of the totipotent zygote. It is proposed that the totipotent stem cell state was shaped by the need to resist transposon activity over evolutionary timescales. | Ho, K. Y. L., An, K., Carr, R. L., Dvoskin, A. D., Ou, A. Y. J., Vogl, W., Tanentzapf, G. (2023). Maintenance of hematopoietic stem cell niche homeostasis requires gap junction-mediated calcium signaling. Proc Natl Acad Sci U S A, 120(45):e2303018120 PubMed ID: 37903259
Summary: Regulation of stem cells requires coordination of the cells that make up the stem cell niche. This study describes a mechanism that allows communication between niche cells to coordinate their activity and shape the signaling environment surrounding resident stem cells. Using the Drosophila hematopoietic organ, the lymph gland, this study showed that cells of the hematopoietic niche, the posterior signaling center (PSC), communicate using gap junctions (GJs) and form a signaling network. This network allows PSC cells to exchange Ca(2+) signals repetitively which regulate the hematopoietic niche. Disruption of Ca(2+) signaling in the PSC or the GJ-mediated network connecting niche cells causes dysregulation of the PSC and blood progenitor differentiation. Analysis of PSC-derived cell signaling shows that the Hedgehog pathway acts downstream of GJ-mediated Ca(2+) signaling to modulate the niche microenvironment. These data show that GJ-mediated communication between hematopoietic niche cells maintains their homeostasis and consequently controls blood progenitor behavior. |
Monday July 15th - Signaling |
| Troost, T., Seib, E., Airich, A., Vullings, N., Necakov, A., De Renzis, S., Klein, T. (2023). The meaning of ubiquitylation of the DSL ligand Delta for the development of Drosophila. BMC Biol, 21(1):260 PubMed ID: 37974242
Summary: Ubiquitylation (ubi) of the intracellular domain of the Notch ligand Delta (Dl) by the E3 ligases Neuralized (Neur) and Mindbomb1 (Mib1) on lysines (Ks) is thought to be essential for the its signalling activity. Nevertheless, it has been previously shown that DlK2R-HA, a Dl variant where all Ks in its intracellular domain (ICD) are replaced by the structurally similar arginine (R), still possess weak activity if over-expressed. This suggests that ubi is not absolutely required for Dl signalling. However, it is not known whether the residual activity of DlK2R-HA is an effect of over-expression and, if not, whether DlK2R can provide sufficient activity for the whole development of Drosophila. To clarify these issues, this study generated and analysed Dl(attP)-DlK2R-HA, a knock-in allele into the Dl locus. This allele reveals that the sole presence of one copy of Dl(attP)-DlK2R-HA can provide sufficient activity for completion of development. It further indicates that while ubi is required for the full activity of Dl in Mib1-dependent processes, it is not essential for Neur-controlled neural development. Three modes of Dl signalling were identified that are either dependent or independent of ubi. Importantly, all modes depend on the presence of the endocytic adapter Epsin. During activation of Dl, direct binding of Epsin appears not to be an essential requirement. In addition, this analysis further reveals that the Ks are required to tune down the cis-inhibitory interaction of Dl with Notch. These results indicate that Dl can activate the Notch pathway without ubi of its ICD. It signals via three modes. Ubi is specifically required for the Mib1-dependent processes and the adjustment of cis-inhibition. In contrast to Mib1, Neur can efficiently activate Dl without ubi. Neur probably acts as an endocytic co-adapter in addition to its role as E3 ligase. Endocytosis, regulated in a ubi-dependent or ubi-independent manner is required for signalling and also suppression of cis-inhibition. The findings clarify the role of ubi of the ligands during Notch signalling. | Gutorov, R., Katz, B., Peters, M., Minke, B. (2023). Membrane lipid modulations by methyl-β-cyclodextrin uncouple the Drosophila light-activated phospholipase C from TRP and TRPL channel gating. J Biol Chem, 300(1):105484 PubMed ID: 37992804
Summary: Sterols are hydrophobic molecules, known to cluster signaling membrane-proteins in lipid rafts, while methyl-β-cyclodextrin (MβCD) has been a major tool for modulating membrane-sterol content for studying its effect on membrane proteins, including the transient receptor potential (TRP) channels. The Drosophila light-sensitive TRP channels are activated downstream of a G-protein-coupled phospholipase Cβ (PLC) cascade. In phototransduction, PLC is an enzyme that hydrolyzes phosphatidylinositol 4,5-bisphosphate (PIP2) generating diacylglycerol, inositol-tris-phosphate, and protons, leading to TRP and TRP-like (TRPL) channel openings. This study examined the effects of MβCD on Drosophila phototransduction using electrophysiology while fluorescently monitoring PIP2 hydrolysis, aiming to examine the effects of sterol modulation on PIP2 hydrolysis and the ensuing light-response in the native system. Incubation of photoreceptor cells with MβCD dramatically reduced the amplitude and kinetics of the TRP/TRPL-mediated light response. MβCD also suppressed PLC-dependent TRP/TRPL constitutive channel activity in the dark induced by mitochondrial uncouplers, but PLC-independent activation of the channels by linoleic acid was not affected. Furthermore, MβCD suppressed a constitutively active TRP mutant-channel, trpP365, suggesting that TRP channel activity is a target of MβCD action. Importantly, whole-cell voltage-clamp measurements from photoreceptors and simultaneously monitored PIP2-hydrolysis by translocation of fluorescently tagged Tubby protein domain, from the plasma membrane to the cytosol, revealed that MβCD virtually abolished the light response when having little effect on the light-activated PLC. Together, MβCD uncoupled TRP/TRPL channel gating from light-activated PLC and PIP2-hydrolysis suggesting the involvement of distinct nanoscopic lipid domains such as lipid rafts and PIP2 clusters in TRP/TRPL channel gating. |
| Strutt, H., Warrington, S., Madathil, A. C. K., Langenhan, T., Strutt, D. (2023). Molecular symmetry breaking in the Frizzled-dependent planar polarity pathway. Curr Biol, 33(24):5340-5354.e5346 PubMed ID: 37995695
Summary: The core planar polarity pathway consists of six proteins that form asymmetric intercellular complexes that segregate to opposite cell ends in developing tissues and specify polarized cell structures or behaviors. Within these complexes, the atypical cadherin Flamingo localizes on both sides of intercellular junctions, where it interacts homophilically in trans via its cadherin repeats, whereas the transmembrane proteins Frizzled and Strabismus localize to the opposite sides of apposing junctions. However, the molecular mechanisms underlying the formation of such asymmetric complexes are poorly understood. Using a novel tissue culture system, the minimum requirements were determined for asymmetric complex assembly in the absence of confounding feedback mechanisms. Complexes were shown to be intrinsically asymmetric and that an interaction of Frizzled and Flamingo in one cell with Flamingo in the neighboring cell is the key symmetry-breaking step. In contrast, Strabismus is unable to promote homophilic Flamingo trans binding and is only recruited into complexes once Frizzled has entered on the opposite side. This interaction with Strabismus requires intact intracellular loops of the seven-pass transmembrane domain of Flamingo. Once recruited, Strabismus stabilizes the intercellular complexes together with the three cytoplasmic core proteins. A model is proposed whereby Flamingo exists in a closed conformation and binding of Frizzled in one cell results in a conformational change that allows its cadherin repeats to interact with a Flamingo molecule in the neighboring cell. Flamingo in the adjacent cell then undergoes a further change in the seven-pass transmembrane region that promotes the recruitment of Strabismus. | Song, S., Cho, B., Weiner, A. T., Nissen, S. B., Ojeda Naharros, I., Sanchez Bosch, P., Suyama, K., Hu, Y., He, L., Svinkina, T., Udeshi, N. D., Carr, S. A., Perrimon, N., Axelrod, J. D. (2023). Protein phosphatase 1 regulates core PCP signaling. EMBO reports, 24(12):e56997 PubMed ID: 37975164
Summary: Planar cell polarity (PCP) signaling polarizes epithelial cells within the plane of an epithelium. Core PCP signaling components adopt asymmetric subcellular localizations within cells to both polarize and coordinate polarity between cells. Achieving subcellular asymmetry requires additional effectors, including some mediating post-translational modifications of core components. Identification of such proteins is challenging due to pleiotropy. This study used mass spectrometry-based proximity labeling proteomics to identify such regulators in the Drosophila wing. The catalytic subunit of protein phosphatase1, Pp1-87B, was identified, and it was shown to regulate core protein polarization. Pp1-87B interacts with the core protein Van Gogh and at least one serine/threonine kinase, Dco/CKIε, that is known to regulate PCP. Pp1-87B modulates Van Gogh subcellular localization and directs its dephosphorylation in vivo. PNUTS, a Pp1 regulatory subunit, also modulates PCP. While the direct substrate(s) of Pp1-87B in control of PCP is not known, these data support the model that cycling between phosphorylated and unphosphorylated forms of one or more core PCP components may regulate acquisition of asymmetry. Finally, this screen serves as a resource for identifying additional regulators of PCP signaling. |
| Wani, A. R., Chowdhury, B., Luong, J., Chaya, G. M., Patel, K., Isaacman-Beck, J., Shafer, O., Kayser, M. S., Syed, M. H. (2023). Stem cell-specific ecdysone signaling regulates the development and function of a Drosophila sleep homeostat. bioRxiv, PubMed ID: 37873323
Summary: Complex behaviors arise from neural circuits that are assembled from diverse cell types. Sleep is a conserved and essential behavior, yet little is known regarding how the nervous system generates neuron types of the sleep-wake circuit. This study focused on the specification of Drosophila sleep-promoting neurons-long-field tangential input neurons that project to the dorsal layers of the fan-shaped body neuropil in the central complex (CX). Lineage analysis and genetic birth dating were used to identify two bilateral Type II neural stem cells that generate these dorsal fan-shaped body (dFB) neurons. Adult dFB neurons express Ecdysone-induced protein E93, and loss of Ecdysone signaling or E93 in Type II NSCs results in the misspecification of the adult dFB neurons. Finally, it was shown that E93 knockdown in Type II NSCs affects adult sleep behavior. These results provide insight into how extrinsic hormonal signaling acts on NSCs to generate neuronal diversity required for adult sleep behavior. These findings suggest that some adult sleep disorders might derive from defects in stem cell-specific temporal neurodevelopmental programs. | Sanchez-Martinez, A., Martinez, A. and Whitworth, A. J. (2023). FBXO7/ntc and USP30 antagonistically set the ubiquitination threshold for basal mitophagy and provide a target for Pink1 phosphorylation in vivo. PLoS Biol 21(8): e3002244. PubMed ID: 37535686
Summary: Functional analyses of genes linked to heritable forms of Parkinson's disease (PD) have revealed fundamental insights into the biological processes underpinning pathogenic mechanisms. Mutations in PARK15/FBXO7 cause autosomal recessive PD and FBXO7 has been shown to regulate mitochondrial homeostasis. This study investigated the extent to which FBXO7 and its Drosophila orthologue, ntc, share functional homology and explored its role in mitophagy in vivo. ntc mutants partially phenocopy Pink1 and parkin mutants and ntc overexpression supresses parkin phenotypes. Furthermore, ntc can modulate basal mitophagy in a Pink1- and parkin-independent manner by promoting the ubiquitination of mitochondrial proteins, a mechanism that is opposed by the deubiquitinase USP30. This basal ubiquitination serves as the substrate for Pink1-mediated phosphorylation that triggers stress-induced mitophagy. It is proposed that FBXO7/ntc works in equilibrium with USP30 to provide a checkpoint for mitochondrial quality control in basal conditions in vivo and presents a new avenue for therapeutic approaches. |
Friday July 12th - Adult Physiology and Metabolism |
| Arch, M., Vidal, M., Fuentes, E., Abat, A. S., Cardona, P. J. (2023). The reproductive status determines tolerance and resistance to Mycobacterium marinum in Drosophila melanogaster. Evolution, medicine, and public health, 11(1):332-347 PubMed ID: 37868078
Summary: Sex and reproductive status of the host have a major impact on the immune response against infection. The aim of this study was to understand their impact on host tolerance or resistance in the systemic Mycobacterium marinum infection of Drosophila melanogaster.Host survival and bacillary load were measured at time of death, as well as expression by quantitative real-time polymerase chain reaction of immune genes (diptericin and drosomycin). This study also assessed the impact of metabolic and hormonal regulation in the protection against infection by measuring expression of upd3, impl2 and ecR. The data showed increased resistance in actively mating flies and in mated females, while reducing their tolerance to infection. Data suggests that Toll and immune deficiency (Imd) pathways determine tolerance and resistance, respectively, while higher basal levels of ecR favours the stimulation of the Imd pathway. A dual role has been found for upd3 expression, linked to increased/decreased mycobacterial load at the beginning and later in infection, respectively. Finally, impl2 expression has been related to increased resistance in non-actively mating males. These results allow further assessment on the differences between sexes and highlights the role of the reproductive status in D. melanogaster to face infections, demonstrating their importance to determine resistance and tolerance against M. marinum infection. | Hou, W. Q., Wen, D. T., Zhong, Q., Mo, L., Wang, S., Yin, X. Y., Ma, X. F. (2023). Physical exercise ameliorates age-related deterioration of skeletal muscle and mortality by activating Pten-related pathways in Drosophila on a high-salt diet. Faseb j, 37(12):e23304 PubMed ID: 37971426
Summary: The phosphatase and tensin congeners (Pten) gene affects cell growth, cell proliferation, and rearrangement of connections, and it is closely related to cellular senescence, but it remains unclear the role of muscle-Pten gene in exercise against age-related deterioration in skeletal muscle and mortality induced by a high-salt diet (HSD). In here, overexpression and knockdown of muscle Pten gene were constructed by building Mhc(GAL4) /Pten(UAS-overexpression) and Mhc(GAL4) /Pten(UAS-RNAi) system in flies, and flies were given exercise training and a HSD for 2 weeks. The results showed that muscle Pten knockdown significantly reduced the climbing speed, climbing endurance, GP(X) activity, and the expression of Pten, Sirt1, PGC-1α genes, and it significantly increased the expression of Akt and ROS level, and impaired myofibril and mitochondria of aged skeletal muscle. Pten knockdown prevented exercise from countering the HSD-induced age-related deterioration of skeletal muscle. Pten overexpression has the opposite effect on skeletal muscle aging when compared to it knockdown, and it promoted exercise against HSD-induced age-related deterioration of skeletal muscle. Pten overexpression significantly increased lifespan, but its knockdown significantly decreased lifespan of flies. Thus, current results confirmed that differential expression of muscle Pten gene played an important role in regulating skeletal muscle aging and lifespan, and it also affected the adaptability of aging skeletal muscle to physical exercise since it determined the activity of muscle Pten/Akt pathway and Pten/Sirt1/PGC-1α pathway. |
| Yan, H., Ding, M., Peng, T., Zhang, P., Tian, R., Zheng, L. (2023). Regular Exercise Modulates the dfoxo/dsrebp Pathway to Alleviate High-Fat-Diet-Induced Obesity and Cardiac Dysfunction in Drosophila. Int J Mol Sci, 24(21) PubMed ID: 37958546
Summary: Obesity is a prevalent metabolic disorder associated with various diseases, including cardiovascular conditions. While exercise is recognized as an effective approach for preventing and treating obesity, its underlying molecular mechanisms remain unclear. This study aimed to explore the impact of regular exercise on high-fat-diet-induced obesity and cardiac dysfunction in Drosophila, shedding light on its molecular mechanisms by identifying its regulation of the dfoxo and dsrebp signaling pathways. The findings demonstrated that a high-fat diet leads to weight gain, fat accumulation, reduced climbing performance, and elevated triglyceride levels in Drosophila. Additionally, cardiac microfilaments in these flies exhibited irregularities, breakages, and shortening. M-mode analysis revealed that high-fat-diet-fed Drosophila displayed increased heart rates, shortened cardiac cycles, decreased systolic intervals, heightened arrhythmia indices, reduced diastolic diameters, and diminished fractional shortening. Remarkably, regular exercise effectively ameliorated these adverse outcomes. Further analysis showed that regular exercise reduced fat synthesis, promoted lipolysis, and mitigated high-fat-diet-induced cardiac dysfunction in Drosophila. These results suggest that regular exercise may mitigate high-fat-diet-induced obesity and cardiac dysfunction in Drosophila by regulating the dfoxo and dsrebp signaling pathways, offering valuable insights into the mechanisms underlying the beneficial effects of exercise on obesity and cardiac dysfunction induced by a high-fat diet. | Bamgbose, G., Tulin, A. (2024). PARP-1 is a transcriptional rheostat of metabolic and bivalent genes during development. Life science alliance, 7(2) PubMed ID: 38012002
Summary: PARP-1 participates in various cellular processes, including gene regulation. In Drosophila, PARP-1 mutants undergo developmental arrest during larval-to-pupal transition. This study investigated PARP-1 binding and its transcriptional regulatory role at this stage. The findings revealed that PARP-1 binds and represses active metabolic genes, including glycolytic genes, whereas activating low-expression developmental genes, including a subset of "bivalent" genes in third-instar larvae. These bivalent promoters, characterized by dual enrichment of low H3K4me3 and high H3K27me3, a unimodal H3K4me1 enrichment at the transcription start site (conserved in C. elegans and zebrafish), H2Av depletion, and high accessibility, may persist throughout development. In PARP-1 mutant third-instar larvae, metabolic genes typically down-regulated during the larval-to-pupal transition in response to reduced energy needs were repressed by PARP-1. Simultaneously, developmental and bivalent genes typically active at this stage were activated by PARP-1. In addition, glucose and ATP levels were significantly reduced in PARP-1 mutants, suggesting an imbalance in metabolic regulation. It is proposed that PARP-1 is essential for maintaining the delicate balance between metabolic and developmental gene expression programs to ensure proper developmental progression. |
| Kang, P., Liu, P., Kim, J., Bolton, M., Kumar, A., Miao, T., Shimell, M., O'Connor, M. B., Powell-Coffman, J., Bai, H. (2023). Ptth regulates lifespan through innate immunity pathway in Drosophila. bioRxiv, PubMed ID: 37873203
Summary: The prothoracicotropic hormone (Ptth) is well-known for its important role in controlling insect developmental timing and body size by promoting the biosynthesis and release of ecdysone. However, the role of Ptth in adult physiology is largely unexplored. Thia study showws that Ptth null mutants (both males and females) show extended lifespan and healthspan, and exhibit increased resistance to oxidative stress. Transcriptomic analysis reveals that age-dependent upregulation of innate immunity pathway is attenuated by Ptth mutants. Intriguingly, it was found that Ptth regulates the innate immunity pathway, specifically in fly oenocytes, the homology of mammalian hepatocytes. It was further shown that oenocyte-specific overexpression of Relish shortens the lifespan, while oenocyte-specific downregulation of ecdysone signaling extends lifespan. Consistently, knocking down torso, the receptor of Ptth in the prothoracic gland also promotes longevity of the flies. Thus, these data reveal a novel function of the insect hormone Ptth in longevity regulation and innate immunity in adult Drosophila. | Yu, G., Liu, S., Yang, K., Wu, Q. (2023). Reproductive-dependent effects of B vitamin deficiency on lifespan and physiology. Frontiers in nutrition,. 10:1277715 PubMed ID: 37941770
Summary: B vitamins constitute essential micronutrients in animal organisms, executing crucial roles in numerous biological processes. B vitamin deficiency can result in severe health consequences, including the impairment of reproductive functions and increased susceptibility to age-related diseases. However, the understanding of how reproduction alters the requirements of each individual B vitamins for healthy aging and lifespan remains limited. In this study, utilizing Drosophila as a model organism, the substantial impacts of deficiencies in specific B vitamins on lifespan and diverse physiological functions were revealed, with the effects being significantly shaped by reproductive status. Notably, the dietary absence of VB(1), VB(3), VB(5), VB(6), or VB(7) significantly decreased the lifespan of wild-type females, yet demonstrated relatively little effect on ovoD1 infertile mutant females' lifespan. B vitamin deficiencies also resulted in distinct impacts on the reproduction, starvation tolerance and fat metabolism of wild-type females, though no apparent effects were observed in the infertile mutant females. Moreover, a deficiency in VB(1) reshaped the impacts of macronutrient intervention on the physiology and lifespan of fertile females in a reproductive-dependent manner. Overall, this study unravels that the reproductive status of females serves as a critical modulator of the lifespan and physiological alterations elicited by B-vitamin deficiencies. |
Thursday July 11th - Genes, Enzymes and Protein Expression. Evolution, Structure and Function |
| Varland, S., Silva, R. D., Kjosas, I., Faustino, A., Bogaert, A., Billmann, M., Boukhatmi, H., Kellen, B., Costanzo, M., Drazic, A., Osberg, C., Chan, K., Zhang, X., Tong, A. H. Y., Andreazza, S., Lee, J. J., Nedyalkova, L., Usaj, M., Whitworth, A. J., Andrews, B. J., Moffat, J., Myers, C. L., Gevaert, K., Boone, C., Martinho, R. G., Arnesen, T. (2023). N-terminal acetylation shields proteins from degradation and promotes age-dependent motility and longevity. Nat Commun, 14(1):6774 PubMed ID: 37891180
Summary: Most eukaryotic proteins are N-terminally acetylated, but the functional impact on a global scale has remained obscure. Using genome-wide CRISPR knockout screens in human cells, this study revealed a strong genetic dependency between a major N-terminal acetyltransferase and specific ubiquitin ligases. Biochemical analyses uncover that both the ubiquitin ligase complex UBR4-KCMF1 and the acetyltransferase NatC recognize proteins bearing an unacetylated N-terminal methionine followed by a hydrophobic residue. NatC KO-induced protein degradation and phenotypes are reversed by UBR knockdown, demonstrating the central cellular role of this interplay. Loss of Drosophila NatC is associated with male sterility, reduced longevity, and age-dependent loss of motility due to developmental muscle defects. Remarkably, muscle-specific overexpression of UbcE2M, one of the proteins targeted for NatC KO-mediated degradation, suppresses defects of NatC deletion. In conclusion, NatC-mediated N-terminal acetylation acts as a protective mechanism against protein degradation, which is relevant for increased longevity and motility. | Peebles, K. E., LaFever, K. S., Page-McCaw, P. S., Colon, S., Wang, D., Stricker, A. M., Ferrell, N., Bhave, G., Page-McCaw, A. (2023). Peroxidasin is required for full viability in development and for maintenance of tissue mechanics in adults. Matrix biology : journal of the International Society for Matrix Biology, PubMed ID: 38000777
Summary: Basement membranes are thin strong sheets of extracellular matrix. They provide mechanical and biochemical support to epithelia, muscles, nerves, and blood vessels, among other tissues. The mechanical properties of basement membranes are conferred in part by Collagen IV (Col4), an abundant protein of basement membranes that forms an extensive two-dimensional network through head-to-head and tail-to-tail interactions. After the Col4 network is assembled into a basement membrane, it is crosslinked by the matrix-resident enzyme Peroxidasin to form a large covalent polymer. Peroxidasin and Col4 crosslinking are highly conserved throughout the animal kingdom, indicating they are important, but homozygous mutant mice have mild phenotypes. To explore the role of Peroxidasin, mutants in Drosophila, including a new CRISPR-generated catalytic null were analyzed, and homozygotes were found to be mostly lethal with 13% viable escapers. Mouse mutants also show semi-lethality, with Mendelian analysis demonstrating ∼50% lethality and ∼50% escapers. Despite the strong mutations, the homozygous fly and mouse escapers had low but detectable levels of Col4 crosslinking, indicating the existence of inefficient alternative crosslinking mechanisms, probably responsible for the viable escapers. Fly mutant phenotypes are consistent with decreased basement membrane stiffness. Interestingly, it was found that even after basement membranes are assembled and crosslinked in wild-type animals, continuing Peroxidasin activity is required in adults to maintain tissue stiffness over time. These results suggest that Peroxidasin crosslinking may be more important than previously appreciated. |
| Markus, D., Pelletier, A., Boube, M., Port, F., Boutros, M., Payre, F., Obermayer, B., Zanet, J. (2023). The pleiotropic functions of Pri smORF peptides synchronize leg development regulators. PLoS Genet, 19(10):e1011004 PubMed ID: 37903161
Summary: The last decade witnesses the emergence of the abundant family of smORF peptides, encoded by small ORF (<100 codons), whose biological functions remain largely unexplored. Bioinformatic analyses here identify hundreds of putative smORF peptides expressed in Drosophila imaginal leg discs. Thanks to a functional screen in leg, smORF peptides involved in morphogenesis were found, including the pioneer smORF peptides Pri. Since its target Ubr3 was identified in the epidermis and pri was known to control leg development through poorly understood mechanisms, the role of Ubr3 in mediating pri function in leg was examined. pri was found to play several roles during leg development both in patterning and in cell survival. During larval stage, pri activates independently of Ubr3 tarsal transcriptional programs and Notch and EGFR signaling pathways, whereas at larval pupal transition, Pri peptides cooperate with Ubr3 to insure cell survival and leg morphogenesis. These results highlight Ubr3 dependent and independent functions of Pri peptides and their pleiotropy. Moreover, it was revealed that the smORF peptide family is a reservoir of overlooked developmental regulators, displaying distinct molecular functions and orchestrating leg development. | Bosch, J. A., Keith, N., Escobedo, F., Fisher, W. W., LaGraff, J. T., Rabasco, J., Wan, K. H., Weiszmann, R., Hu, Y., Kondo, S., Brown, J. B., Perrimon, N., Celniker, S. E. (2023). Molecular and functional characterization of the Drosophila melanogaster conserved smORFome. Cell Rep, 42(11):113311 PubMed ID: 37889754
Summary: Short polypeptides encoded by small open reading frames (smORFs) are ubiquitously found in eukaryotic genomes and are important regulators of physiology, development, and mitochondrial processes. This study focused on a subset of 298 smORFs that are evolutionarily conserved between Drosophila melanogaster and humans. Many of these smORFs are conserved broadly in the bilaterian lineage, and ∼182 are conserved in plants. This study observe remarkably heterogeneous spatial and temporal expression patterns of smORF transcripts-indicating wide-spread tissue-specific and stage-specific mitochondrial architectures. In addition, an analysis of annotated functional domains reveals a predicted enrichment of smORF polypeptides localizing to mitochondria. An embryonic ribosome profiling experiment was conducted and support was found for translation of 137 of these smORFs during embryogenesis. This study further embarked on functional characterization using CRISPR knockout/activation, RNAi knockdown, and cDNA overexpression, revealing diverse phenotypes. This study underscores the importance of identifying smORF function in disease and phenotypic diversity. |
| Guntur, A. R., Smith, J. E., Brahmandam, A., DeBauche, P., Cronmiller, C., Lundell, M. J. (2023). ZFH-2 is required for Drosophila ovarian follicle development and is expressed at the band/interband boundaries of polytene chromosomes. Dev Biol, 504:1-11 PubMed ID: 37666353
Summary: The transcription factor ZFH-2 has well-documented roles in Drosophila neurogenesis and other developmental processes. This study provides the first evidence that ZFH-2 has a role in oogenesis. ZFH-2 is expressed in the wild-type ovary and a loss of zfh-2 function produces a mutant ovary phenotype where egg chambers are reduced in number and fused. This study also showed that a loss of zfh-2 function can suppress a daughterless loss-of-function ovary phenotype suggesting a possible genetic relationship between these two genes in the ovary. It was also shown that ZFH-2 is located at the boundary between bands and interbands on polytene chromosomes and that at a subset of these sites ZFH-2 colocalizes with the insulator/promoter cofactor CP190. | Zhang, Q., Deng, K., Liu, M., Yang, S., Xu, W., Feng, T., Jie, M., Liu, Z., Sheng, X., Chen, H., Jiang, H. (2023). Phase separation of BuGZ regulates gut regeneration and aging through interaction with m(6)A regulators. Nat Commun, 14(1):6700 PubMed ID: 37872148
Summary: Exploring the role of phase separation in intracellular compartment formation is an active area of research. However, the associations of phase separation with intestinal stem cell (ISC)-dependent regeneration and aging remain unclear. This study demonstrates that BuGZ, a coacervating mitotic effector, shows age- and injury-associated condensation in Drosophila ISC nuclei during interphase. BuGZ condensation promotes ISC proliferation, affecting Drosophila gut repair and longevity. Moreover, m(6)A reader YT521-B acts as the transcriptional and functional downstream of BuGZ. The binding of YT521-B promotor or m(6)A writer Ime4/ Mettl14 to BuGZ controls its coacervation, indicating that the promotor may accelerate the phase transition of its binding transcription factor. Hence, it is proposed that phase separation and m(6)A regulators may be critical for ameliorating ISC-dependent gut regeneration and aging and requires further study. Summary: |
Wednesday, July 10th - Gonads |
| Burghardt, E., Rakijas, J., Tyagi, A., Majumder, P., Olson, B., McDonald, J. A. (2023). Transcriptome analysis reveals temporally regulated genetic networks during Drosophila border cell collective migration. BMC Genomics, 24(1):728 PubMed ID: 38041052
Summary: Collective cell migration underlies many essential processes, including sculpting organs during embryogenesis, wound healing in the adult, and metastasis of cancer cells. At mid-oogenesis, Drosophila border cells undergo collective migration. Border cells round up into a small group at the pre-migration stage, detach from the epithelium and undergo a dynamic and highly regulated migration at the mid-migration stage, and stop at the oocyte, their final destination, at the post-migration stage. While specific genes that promote cell signaling, polarization of the cluster, formation of protrusions, and cell-cell adhesion are known to regulate border cell migration, there may be additional genes that promote these distinct active phases of border cell migration. Therefore, this study sought to identify genes whose expression patterns changed during border cell migration. RNA-sequencing was performed on border cells isolated at pre-, mid-, and post-migration stages. It is reported that 1,729 transcripts, in nine co-expression gene clusters, are temporally and differentially expressed across the three migration stages. Gene ontology analyses and constructed protein-protein interaction networks identified genes expected to function in collective migration, such as regulators of the cytoskeleton, adhesion, and tissue morphogenesis, but also uncovered a notable enrichment of genes involved in immune signaling, ribosome biogenesis, and stress responses. Finally, the in vivo expression and function of a subset of identified genes in border cells was validated. Overall, these results identified differentially and temporally expressed genetic networks that may facilitate the efficient development and migration of border cells. The genes identified in this study represent a wealth of new candidates to investigate the molecular nature of dynamic collective cell migrations in developing tissues. | Chen, X., Qi, Y., Huang, Q., Sun, C., Zheng, Y., Ji, L., Shi, Y., Cheng, X., Li, Z., Zheng, S., Cao, Y., Gu, Z., Yu, J. (2023). Single-cell transcriptome characteristics of testicular terminal epithelium lineages during aging in the Drosophila. Aging Cell:e14057 PubMed ID: 38044573
Summary: Aging is a complex biological process leading to impaired functions, with a variety of hallmarks. In the testis of Drosophila, the terminal epithelium region is involved in spermatid release and maturation, while its functional diversity and regulatory mechanism remain poorly understood. Single-cell RNA-sequencing analysis (scRNA-seq) was performed to characterize the transcriptomes of terminal epithelium in Drosophila testes at 2-, 10 and 40-Days. Terminal epithelium populations were defined with Metallothionein A (MtnA) and subdivided into six novel sub-cell clusters (EP0-EP5), and a series of marker genes were identified based on their expressions. The data revealed the functional characteristics of terminal epithelium populations, such as tight junction, focal adhesion, bacterial invasion, oxidative stress, mitochondrial function, proteasome, apoptosis and metabolism. Interestingly, it was also found that disrupting genes for several relevant pathways in terminal epithelium led to male fertility disorders. Moreover, a series of age-biased genes and pseudotime trajectory mediated state-biased genes were also discovered during terminal epithelium aging. Differentially expressed genes during terminal epithelium aging were mainly participated in the regulation of several common signatures, e.g. mitochondria-related events, protein synthesis and degradation, and metabolic processes. The Drosophila divergence and selection in the functional constraints of age-biased genes during aging was also explored, revealing that age-biased genes in epithelial cells of 2 Days group evolved rapidly and were endowed with greater evolutionary advantages. scRNA-seq analysis revealed the diversity of testicular terminal epithelium populations, providing a gene target resource for further systematic research of their functions during aging. |
| He, Z., Fang, Y., Zhang, F., Liu, Y., Cheng, X., Wang, J., Li, D., Chen, D., Wu, F. (2023). Adenine nucleotide translocase 2 (Ant2) is required for individualization of spermatogenesis of Drosophila melanogaster. Insect Sci, PubMed ID: 38112480
Summary: Successful completion of spermatogenesis is crucial for the perpetuation of the species. In Drosophila, spermatid individualization, a process involving changes in mitochondrial structure and function is critical to produce functional mature sperm. Ant2, encoding a mitochondrial adenine nucleotide translocase, is highly expressed in male testes and plays a role in energy metabolism in the mitochondria. However, its molecular function remains unclear. This study identified an important role of Ant2 in spermatid individualization. In Ant2 knockdown testes, spermatid individualization complexes composed of F-actin cones exhibited a diffuse distribution, and mature sperms were absent in the seminal vesicle, thus leading to male sterility. The most striking effects in Ant2-knockdown spermatids were decrease in tubulin polyglycylation and disruption of proper mitochondria derivatives function. Excessive apoptotic cells were also observed in Ant2-knockdown testes. To further investigate the phenotype of Ant2 knockdown in testes at the molecular level, complementary transcriptome and proteome analyses were performed. At the mRNA level, 868 differentially expressed genes were identified, of which 229 genes were upregulated and 639 were downregulated induced via Ant2 knockdown. iTRAQ-labeling proteome analysis revealed 350 differentially expressed proteins, of which 117 proteins were upregulated and 233 were downregulated. The expression of glutathione transferase (GstD5, GstE5, GstE8, and GstD3), proteins involved in reproduction were significantly regulated at both the mRNA and protein levels. These results indicate that Ant2 is crucial for spermatid maturation by affecting mitochondrial morphogenesis. | Herriage, H. C., Calvi, B. R. (2024). Premature endocycling of Drosophila follicle cells causes pleiotropic defects in oogenesis. bioRxiv, PubMed ID: 37873193
Summary: Endocycling cells grow and repeatedly duplicate their genome without dividing. Cells switch from mitotic cycles to endocycles in response to developmental signals during the growth of specific tissues in a wide range of organisms. The purpose of switching to endocycles, however, remains unclear in many tissues. Additionally, cells can switch to endocycles in response to conditional signals, which can have beneficial or pathological effects on tissues. However, the impact of these unscheduled endocycles on development is underexplored. This study uses Drosophila ovarian somatic follicle cells as a model to examine the impact of unscheduled endocycles on tissue growth and function. Follicle cells normally switch to endocycles at mid-oogenesis. Inducing follicle cells to prematurely switch to endocycles resulted in lethality of the resulting embryos. Analysis of ovaries with premature follicle cell endocycles revealed aberrant follicular epithelial structure and pleiotropic defects in oocyte growth, developmental gene amplification, and the migration of a special set of follicle cells known as border cells. Overall, these findings reveal how unscheduled endocycles can disrupt tissue growth and function to cause aberrant development. A premature switch to polyploid endocycles in Drosophila ovarian follicle cells caused pleiotropic defects in oogenesis and compromised female fertility, revealing new ways in which unscheduled endocycles cause developmental defects. |
| Kotb, N. M., Ulukaya, G., Chavan, A., Nguyen, S. C., Proskauer, L., Joyce, E., Hasson, D., Jagannathan, M., Rangan, P. (2023). Genome organization regulates nuclear pore complex formation and promotes differentiation during Drosophila oogenesis. bioRxiv, PubMed ID: 38014330
Summary: Genome organization can regulate gene expression and promote cell fate transitions. The differentiation of germline stem cells (GSCs) to oocytes in Drosophila involves changes in genome organization mediated by heterochromatin and the nuclear pore complex (NPC). Heterochromatin represses germ-cell genes during differentiation and NPCs anchor these silenced genes to the nuclear periphery, maintaining silencing to allow for oocyte development. Surprisingly, this study found that genome organization also contributes to NPC formation, mediated by the transcription factor Stonewall (Stwl). As GSCs differentiate, Stwl accumulates at boundaries between silenced and active gene compartments. Stwl at these boundaries plays a pivotal role in transitioning germ-cell genes into a silenced state and activating a group of oocyte genes and Nucleoporins (Nups). The upregulation of these Nups during differentiation is crucial for NPC formation and further genome organization. Thus, crosstalk between genome architecture and NPCs is essential for successful cell fate transitions. | Wenzel, M., Aquadro, C. F. (2023). Wolbachia infection at least partially rescues the fertility and ovary defects of several new Drosophila melanogaster bag of marbles protein-coding mutants. bioRxiv, PubMed ID: 37645949
Summary: The D. melanogaster protein coding gene bag of marbles (bam) plays a key role in early male and female reproduction by forming complexes with partner proteins to promote differentiation in gametogenesis. Like another germline gene, Sex lethal, bam genetically interacts with the endosymbiont Wolbachia, as Wolbachia rescues the reduced fertility of a bam hypomorphic mutant. This study explored the specificity of the bam-Wolbachia interaction by generating 22 new bam mutants, with ten mutants displaying fertility defects. Nine of these mutants trend towards rescue by the w Mel Wolbachia variant, with eight statistically significant at the fertility and/or cytological level. In some cases, fertility was increased a striking 20-fold. There is no specificity between the rescue and the known binding regions of bam, suggesting w Mel does not interact with one singular bam partner to rescue the reproductive phenotype. Whether w Mel interacts with bam in a non-specific way was tested by increasing bam transcript levels or acting upstream in germline stem cells. A fertility assessment of a bam RNAi knockdown mutant reveals that w Mel rescue is specific to functionally mutant bam alleles and no obvious evidence was found of w Mel interaction with germline stem cells in bam mutants. |
Tuesday, July 9th - Behavior |
| Abhilash, L., Shafer, O. T. (2023). A two-process model of Drosophila sleep reveals an inter-dependence between circadian clock speed and the rate of sleep pressure decay. Sleep, PubMed ID: 37930351
Summary: Sleep is controlled by two processes - a circadian clock that regulates its timing and a homeostat that regulates the drive to sleep. Drosophila has been an insightful model for understanding both processes. For four decades, Borbely and Daan's two-process model has provided a powerful framework for understanding sleep regulation. However, the field of fly sleep has not employed such a model as a framework for the investigation of sleep. To this end, this study has adapted the two-process model to the fly and establish its utility by showing that it can provide empirically testable predictions regarding the circadian and homeostatic control of fly sleep. The ultradian rhythms previously reported for loss-of-function clock mutants in the fly are shown to be robustly detectable and a predictable consequence of a functional sleep homeostat in the absence of a functioning circadian system. It was founnd that a model in which the circadian clock speed and homeostatic rates act without influencing each other provides imprecise predictions regarding how clock speed influences the strength of sleep rhythms and the amount of daily sleep. This study also found that quantitatively good fits between empirical values and model predictions were achieved only when clock speeds were positively correlated with rates of decay of sleep pressure. The results indicate that longer sleep bouts better reflect the homeostatic process than the current definition of sleep as any inactivity lasting five minutes or more. This two-process model represents a powerful framework for work on the molecular and physiological regulation of fly sleep. | O'Hara, M. K., Saul, C., Handa, A., Sehgal, A., Williams, J. A. (2023). The NFkappaB Dif is required for behavioral and molecular correlates of sleep homeostasis in Drosophila. bioRxiv, PubMed ID: 37905096
Summary: The nuclear factor binding the κ light chain in B-cells (NFκB) is involved in a wide range of cellular processes including development, growth, innate immunity, and sleep. However, efforts have been limited toward understanding how specific NFκB transcription factors function in sleep. Drosophila fruit flies carry three genes encoding NFκB transcription factors, Dorsal, Dorsal Immunity Factor (Dif), and Relish. Previous work found that loss of the Relish gene from fat body suppressed daily nighttime sleep, and abolished infection-induced sleep. This study shows that Dif regulates daily sleep and recovery sleep following prolonged wakefulness. Mutants of Dif showed reduced daily sleep and suppressed recovery in response to sleep deprivation. Pan-neuronal knockdown of Dif strongly suppressed daily sleep, indicating that in contrast to Relish, Dif functions from the central nervous system to regulate sleep. Based on the distribution of a Dif-associated GAL4 driver, it was hypothesized that its effects on sleep were mediated by the pars intercerebralis (PI). While RNAi knock-down of Dif in the PI reduced daily sleep, it had no effect on the recovery response to sleep deprivation. However, recovery sleep was suppressed when RNAi knock-down of Dif was distributed across a wider range of neurons. Induction of the nemuri (nur) antimicrobial peptide by sleep deprivation was suppressed in Dif mutants and pan-neuronal over-expression of nur also suppressed the Dif mutant phenotype. Together, these findings indicate that Dif functions from brain to target nemuri and to promote sleep. |
| Sten, T. H., Li, R., Hollunder, F., Eleazer, S., Ruta, V. (2023). Male-male interactions shape mate selection in Drosophila. bioRxiv, PubMed ID: 37961193
Summary: Males of many species have evolved behavioral traits to both attract females and repel rivals. This study explored mate selection in Drosophila from both the male and female perspective to shed light on how these key components of sexual selection - female choice and male-male competition - work in concert to guide reproductive strategies. Male flies were found to fend off competing suitors by interleaving their courtship of a female with aggressive wing flicks, which both repel competitors and generate a 'song' that obscures the female's auditory perception of other potential mates. Two higher-order circuit nodes - P1a and pC1x neurons - are coordinately recruited to allow males to flexibly interleave these agonistic actions with courtship displays, assuring they persistently pursue females until their rival falters. Together, these results suggest that female mating decisions are shaped by male-male interactions, underscoring how a male's ability to subvert his rivals is central to his reproductive success. | Safdar, M., Wessells, R. J. (2023). Octopamine Rescues Endurance and Climbing Speed in Drosophila Clk(out) Mutants with Circadian Rhythm Disruption. Cells, 12(21) PubMed ID: 37947593
Summary: Circadian rhythm disturbances are associated with various negative health outcomes, including an increasing incidence of chronic diseases with high societal costs. While exercise can protect against the negative effects of rhythm disruption, it is not available to all those impacted by sleep disruptions, in part because sleep disruption itself reduces exercise capacity. Thus, there is a need for therapeutics that bring the benefits of exercise to this population. This study investigated the relationship between exercise and circadian disturbances using a well-established Drosophila model of circadian rhythm loss, the Clkout mutant. Clkout was found to cause reduced exercise capacity, measured as post-training endurance, flight performance, and climbing speed, and these phenotypes are not rescued by chronic exercise training. However, exogenous administration of a molecule known to mediate the effects of chronic exercise, octopamine (OA), was able to effectively rescue mutant exercise performance, including the upregulation of other known exercise-mediating transcripts, without restoring the circadian rhythms of mutants. This work points the way toward the discovery of novel therapeutics that can restore exercise capacity in patients with rhythm disruption. |
| Gornostaev, N. G., Ruchin, A. B., Esin, M. N., Lazebny, O. E., Kulikov, A. M. (2023). Vertical Distribution of Fruit Flies (Diptera: Drosophilidae) in Deciduous Forests in the Center of European Russia. Insects, 14(10) PubMed ID: 37887834
Summary: Research of Diptera in temperate forests has demonstrated uneven vertical distributions of insects. In this study, the vertical distribution, seasonal fluctuations, and species diversity of Drosophilidae species in the Mordovia State Reserve were studied. This research marks the first exploration of drosophilid vertical stratification in the European part of Russia. Using traps, flies were collected in four deciduous forest sites between early June and mid-September in 2020. A total of 27,151 individuals from 10 genera and 34 drosophilid species were identified, with 6 species from 4 genera being new to the Republic of Mordovia. Drosophila obscura Fll. and Scaptodrosophila rufifrons Lw. were the most abundant species in traps. The total highest number of drosophilid flies (10,429 individuals) was captured at a height of 1.5 m, while the lowest number (5086 individuals) was recorded at 12 m. The average number of flies was 6240 and 5387 individuals at heights of 7.5 m and 3.5 m, respectively. However, the prevalence of drosophilid numbers at the 1.5-m height was not constant during the season. It was found that in the second part of July the total fly counts at heights of 7.5 m and 12 m exceeded those at 1.5 m. This study has described five different types of vertical distribution of drosophilids throughout the season, which differs markedly in mycetobionts and xylosaprobionts ecological groups. Species diversity demonstrated variations across different sites and tiers during the season, with peak diversity observed in June and September. | Yoshikawa, S., Tang, P., Simpson, J. H. (2023). Mechanosensory and command contributions to the Drosophila grooming sequence. bioRxiv, PubMed ID: 38045358
Summary: Flies groom in response to competing mechanosensory cues in an anterior to posterior order using specific legs. From behavior screens, this study identified a pair of cholinergic command-like neurons, Mago-no-Te (MGT), whose optogenetic activation elicits thoracic grooming by hind legs. Thoracic grooming is typically composed of body sweeps and leg rubs in alternation, but clonal analysis coupled with amputation experiments revealed that MGT activation only commands the body sweeps: initiation of leg rubbing requires contact between leg and thorax. With new electron microscopy (EM) connectome data for the ventral nerve cord (VNC), a circuit-based explanation was uncovered for why stimulation of posterior thoracic mechanosensory bristles initiates cleaning by the hind legs. Previous work showed that flies weigh mechanosensory inputs across the body to select which part to groom, but it was not known why the thorax was always cleaned last. The connectome for the VNC enabled identification if a pair of GABAergic inhibitory neurons, UMGT1, that receive diverse sensory inputs and synapse onto both MGT and components of its downstream pre-motor circuits. Optogenetic activation of UMGT1 suppresses thoracic cleaning, representing a mechanism by which mechanosensory stimuli on other body parts could take precedence in the grooming hierarchy. The pre-motor circuit downstream of MGT was mapped, including inhibitory feedback connections that may enable rhythmicity and coordination of limb movement during thoracic grooming. |
Monday, July 8th - Cancer, Tumors, and Growth |
| Martinez-Abarca Millan, A., Martin-Bermudo, M. D. (2023). Integrins Can Act as Suppressors of Ras-Mediated Oncogenesis in the Drosophila Wing Disc Epithelium. Cancers, 15(22) PubMed ID: 38001693
Summary: Cancer is the second leading cause of death worldwide. Key to cancer initiation and progression is the crosstalk between cancer cells and their microenvironment. The extracellular matrix (ECM) is a major component of the tumour microenvironment and integrins, main cell-ECM adhesion receptors, are involved in every step of cancer progression. However, accumulating evidence has shown that integrins can act as tumour promoters but also as tumour suppressor factors, revealing that the biological roles of integrins in cancer are complex. This incites a better understating of integrin function in cancer progression. To achieve this goal, simple model organisms, such as Drosophila, offer great potential to unravel underlying conceptual principles. This study found that in the Drosophila wing disc epithelium the βPS integrins act as suppressors of tumours induced by a gain of function of the oncogenic form of Ras, Ras(V)(12). βPS integrin depletion enhances the growth, delamination and invasive behaviour of Ras(V)(12) tumour cells, as well as their ability to affect the tumour microenvironment. These results strongly suggest that integrin function as tumour suppressors might be evolutionarily conserved. Drosophila can be used to understand the complex tumour modulating activities conferred by integrins, thus facilitating drug development. | Sanz, F. J., Martinez-Carrion, G., Solana-Manrique, C. and Paricio, N. (2023). Evaluation of type 1 diabetes mellitus as a risk factor of Parkinson's disease in a Drosophila model. J Exp Zool A Ecol Integr Physiol. PubMed ID: 37381093
Summary: Diabetes mellitus (DM) is a chronic metabolic disease characterized by high blood glucose levels, resulting from insulin dysregulation. Parkinson's disease (PD) is the most common neurodegenerative motor disorder caused by the selective loss of dopaminergic (DA) neurons in the substantia nigra pars compacta. DM and PD are both age-associated diseases that are turning into epidemics worldwide. Previous studies have indicated that type 2 DM might be a risk factor of developing PD. However, scarce information about the link between type 1 DM (T1DM) and PD does exist. This study generated a Drosophila model of T1DM based on insulin deficiency to evaluate if T1DM could be a risk factor to trigger PD onset. As expected, model flies exhibited T1DM-related phenotypes such as insulin deficiency, increased content of carbohydrates and glycogen, and reduced activity of insulin signaling. Interestingly, the results also demonstrated that T1DM model flies presented locomotor defects as well as reduced levels of tyrosine hydroxylase (a marker of DA neurons) in brains, which are typical PD-related phenotypes. In addition, T1DM model flies showed elevated oxidative stress levels, which could be causative of DA neurodegeneration. Therefore, these results indicate that T1DM might be a risk factor of developing PD, and encourage further studies to shed light into the exact link between both diseases. |
| Guo, T., Miao, C., Liu, Z., Duan, J., Ma, Y., Zhang, X., Yang, W., Xue, M., Deng, Q., Guo, P., Xi, Y., Yang, X., Huang, X., Ge, W. (2023). Impaired dNKAP function drives genome instability and tumorigenic growth in Drosophila epithelia. J Mol Cell Biol, PubMed ID: 38059855
Summary: Mutations or dysregulated expression of NF-kappaB activating protein (NKAP) family genes have been found in human cancers. How NKAP family gene mutations promote tumor initiation and progression remains to be determined. This study characterized dNKAP, the Drosophila homolog of NKAP, and showed that impaired dNKAP function causes genome instability and tumorigenic growth in a Drosophila epithelial tumor model. dNKAP-knockdown wing imaginal discs exhibit tumorigenic characteristics, including tissue overgrowth, cell invasive behavior, abnormal cell polarity, and cell adhesion defects. dNKAP knockdown causes both R-loop accumulation and DNA damage, indicating the disruption of genome integrity. Further analysis showed that dNKAP knockdown induces c-Jun N-terminal kinase (JNK)-dependent apoptosis and causes changes in cell proliferation in distinct cell populations. Activation of the Notch and JAK/STAT signaling pathways contributes to the tumorigenic growth of dNKAP-knockdown tissues. Furthermore, JNK signaling is essential for dNKAP depletion-mediated cell invasion. Transcriptome analysis of dNKAP-knockdown tissues confirmed the misregulation of signaling pathways involved in promoting tumorigenesis and revealed abnormal regulation of metabolic pathways. dNKAP knockdown and oncogenic Ras, Notch, or Yki mutations show synergies in driving tumorigenesis, further supporting the tumor-suppressive role of dNKAP. In summary, this study demonstrates that dNKAP plays a tumor-suppressive role by preventing genome instability in Drosophila epithelia and thus provides novel insights into the roles of human NKAP family genes in tumor initiation and progression. | Pfefferkorn, R. M., Mortzfeld, B. M., Fink, C., Frieling, J. V., Bossen, J., Esser, D., Kaleta, C., Rosenstiel, P., Heine, H., Roeder, T. (2024). Recurrent Phases of Strict Protein Limitation Inhibit Tumor Growth and Restore Lifespan in A Drosophila Intestinal Cancer Model. Aging and disease, 15(1):226-244 PubMed ID: 37962464
Summary: Diets that restrict caloric or protein intake offer a variety of benefits, including decreasing the incidence of cancer. However, whether such diets pose a substantial therapeutic benefit as auxiliary cancer treatments remains unclear. This study determined the effects of severe protein depletion on tumorigenesis in a Drosophila melanogaster intestinal tumor model, using a human RAF gain-of-function allele. Severe and continuous protein restriction significantly reduced tumor growth but resulted in premature death. Therefore, a diet was developed in which short periods of severe protein restriction alternated cyclically with periods of complete feeding. This nutritional regime reduced tumor mass, restored gut functionality, and rescued the lifespan of oncogene-expressing flies to the levels observed in healthy flies on a continuous, fully nutritious diet. Furthermore, this diet reduced the chemotherapy-induced stem cell activity associated with tumor recurrence. Transcriptome analysis revealed long-lasting changes in the expression of key genes involved in multiple major developmental signaling pathways. Overall, the data suggest that recurrent severe protein depletion effectively mimics the health benefits of continuous protein restriction, without undesired nutritional shortcomings. This provides seminal insights into the mechanisms of the memory effect required to maintain the positive effects of protein restriction throughout the phases of a full diet. Finally, the repetitive form of strict protein restriction is an ideal strategy for adjuvant cancer therapy that is useful in many tumor contexts. Summary: |
| Li, Y., Pan, L., Li, P., Gao, F., Wang, L., Chen, J., Li, Z., Gao, Y., Gong, Y., Jin, F. (2023). Isolation of Enterococcus faecium and determination of its mechanism for promoting the growth and development of Drosophila. Sci Rep, 13(1):18726 PubMed ID: 37907538
Summary: Intestinal symbiotic microorganisms have a strong capacity to regulate the physiological functions of their host, and Drosophila serves as a useful model. Enterococcus faecium (E. faecium) is a member of the normal intestinal flora of animals. Lactic acid bacteria (LAB) such as E. faecium can promote the growth and development of Drosophila, but the mechanism of regulation of Drosophila is poorly understood. This study found that E. faecium used a carbon source to produce probiotic acids. E. faecium is a symbiotic bacterium for Drosophila, and adult flies passed on parental flora to offspring. E. faecium promoted the growth and development of Drosophila, especially under poor nutritional conditions. E. faecium shortened the developmental process for Drosophila and accelerated the transformation from larva to pupa. Finally, E. faecium promoted the growth and development of Drosophila through TOR and insulin signalling pathways. | Hofstetter, J., Ogunleye, A., Kutschke, A., Buchholz, L. M., Wolf, E., Raabe, T., Gallant, P. (2024). Spt5 interacts genetically with Myc and is limiting for brain tumor growth in Drosophila. Life science alliance, 7(1) PubMed ID: 37935464
Summary: The transcription factor SPT5 physically interacts with MYC oncoproteins and is essential for efficient transcriptional activation of MYC targets in cultured cells. This study used Drosophila to address the relevance of this interaction in a living organism. Spt5 displays moderate synergy with Myc in fast proliferating young imaginal disc cells. During later development, Spt5-knockdown has no detectable consequences on its own, but strongly enhances eye defects caused by Myc overexpression. Similarly, Spt5-knockdown in larval type 2 neuroblasts has only mild effects on brain development and survival of control flies, but dramatically shrinks the volumes of experimentally induced neuroblast tumors and significantly extends the lifespan of tumor-bearing animals. This beneficial effect is still observed when Spt5 is knocked down systemically and after tumor initiation, highlighting SPT5 as a potential drug target in human oncology. |
Friday, July 5th - Disease Models |
| Escobedo, S. E., McGovern, S. E., Jauregui-Lozano, J. P., Stanhope, S. C., Anik, P., Singhal, K., DeBernardis, R., Weake, V. M. (2023). Targeted RNAi screen identifies transcriptional mechanisms that prevent premature degeneration of adult photoreceptors. Frontiers in epigenetics and epigenomics, 1 PubMed ID: 37901602
Summary: Aging is associated with a decline in visual function and increased prevalence of ocular disease, correlating with changes in the transcriptome and epigenome of cells in the eye. This study sought to identify the transcriptional mechanisms that are necessary to maintain photoreceptor viability and function during aging. To do this, A targeted photoreceptor-specific RNAi screen was performed in Drosophila to identify transcriptional regulators whose knockdown results in premature, age-dependent retinal degeneration. From an initial set of 155 RNAi lines each targeting a unique gene and spanning a diverse set of transcription factors, chromatin remodelers, and histone modifiers, 18 high-confidence target genes were identified whose decreased expression in adult photoreceptors leads to premature and progressive retinal degeneration. These 18 target genes were enriched for factors involved in the regulation of transcription initiation, pausing, and elongation, suggesting that these processes are essential for maintaining the health of aging photoreceptors. To identify the genes regulated by these factors, the photoreceptor transcriptome was profiled in a subset of lines. Strikingly, two of the 18 target genes, Spt5 and domino, show similar changes in gene expression to those observed in photoreceptors with advanced age. Together, these data suggest that dysregulation of factors involved in transcription initiation and elongation plays a key role in shaping the transcriptome of aging photoreceptors. Further, these findings indicate that the age-dependent changes in gene expression not only correlate but might also contribute to an increased risk of retinal degeneration | Dondi, C., Vogler, G., Gupta, A., Walls, S. M., Kervadec, A., Romero, M. R., Diop, S. B., Goode, J., Thomas, J. B., Colas, A. R., Bodmer, R., Montminy, M., Ocorr, K. (2023). The nutrient sensor CRTC & Sarcalumenin / Thinman represent a new pathway in cardiac hypertrophy. bioRxiv, PubMed ID: 37873259
Summary: Obesity and type 2 diabetes are at epidemic levels and a significant proportion of these patients are diagnosed with left ventricular hypertrophy. CREB R egulated T ranscription C o-activator (CRTC) is a key regulator of metabolism in mammalian hepatocytes, where it is activated by calcineurin (CaN) to increase expression of gluconeogenic genes. CaN is known its role in pathological cardiac hypertrophy, however, a role for CRTC in the heart has not been identified. In Drosophila, CRTC null mutants have little body fat and exhibit severe cardiac restriction, myofibrillar disorganization, cardiac fibrosis and tachycardia, all hallmarks of heart disease. Cardiac-specific knockdown of CRTC, or its coactivator CREBb, mimicked the reduced body fat and heart defects of CRTC null mutants. Comparative gene expression in CRTC loss- or gain-of-function fly hearts revealed contra-regulation of genes involved in glucose, fatty acid, and amino acid metabolism, suggesting that CRTC also acts as a metabolic switch in the heart. Among the contra-regulated genes with conserved CREB binding sites, the fly ortholog of Sarcalumenin, which is a Ca (2+) -binding protein in the sarcoplasmic reticulum, was identified. Cardiac knockdown recapitulated the loss of CRTC cardiac restriction and fibrotic phenotypes, suggesting it is a downstream effector of CRTC, that was named thinman (tmn). Importantly, cardiac overexpression of either CaN or CRTC in flies caused hypertrophy that was reversed in a CRTC mutant background, suggesting CRTC mediates hypertrophy downstream of CaN, perhaps as an alternative to NFAT. CRTC novel role in the heart is likely conserved in vertebrates as knockdown in zebrafish also caused cardiac restriction, as in flies. These data suggest that CRTC is involved in myocardial cell maintenance and that CaN-CRTC- Sarcalumenin/ tmn signaling represents a novel and conserved pathway underlying cardiac hypertrophy. |
| Buck, S. A., Rubin, S. A., Kunkhyen, T., Treiber, C. D., Xue, X., Fenno, L. E., Mabry, S. J., Sundar, V. R., Yang, Z., Shah, D., S., Awatramani, R., Watson, A. M., Waddell, S., Cheetham, C. E. J., Logan, R. W., Freyberg, Z. (2023). Sexually dimorphic mechanisms of VGLUT-mediated protection from dopaminergic neurodegeneration. bioRxiv, PubMed ID: 37873436
Summary: Parkinson's disease (PD) targets some dopamine (DA) neurons more than others. Sex differences offer insights, with females more protected from DA neurodegeneration. The mammalian vesicular glutamate transporter VGLUT2 and Drosophila ortholog dVGLUT have been implicated as modulators of DA neuron resilience. However, the mechanisms by which VGLUT2/dVGLUT protects DA neurons remain unknown. This study discovered DA neuron dVGLUT knockdown increased mitochondrial reactive oxygen species in a sexually dimorphic manner in response to depolarization or paraquat-induced stress, males being especially affected. DA neuron dVGLUT also reduced ATP biosynthetic burden during depolarization. RNA sequencing of VGLUT(+) DA neurons in mice and flies identified candidate genes that were functionally screened to further dissect VGLUT-mediated DA neuron resilience across PD models. Transcription factors modulating dVGLUT-dependent DA neuroprotection were discovered and dj-1β was identified as a regulator of sex-specific DA neuron dVGLUT expression. Overall, VGLUT protects DA neurons from PD-associated degeneration by maintaining mitochondrial health. | Cuddapah, V. A., Hsu, C. T., Li, Y., Shah, H. M., Saul, C., Killiany, S., Shon, J., Yue, Z., Gionet, G., Putt, M. E., Sehgal, A. (2023). Sleepiness, not total sleep amount, increases seizure risk. bioRxiv. PubMed ID: 37873373
Summary: Sleep loss has been associated with increased seizure risk since antiquity. Despite this observation standing the test of time, how poor sleep drives susceptibility to seizures remains unclear. To identify underlying mechanisms, sleep was restricted in Drosophila epilepsy models and a method was developed to identify spontaneous seizures using quantitative video tracking. This study found that sleep loss exacerbates seizures but only when flies experience increased sleep need, or sleepiness, and not necessarily with reduced sleep quantity. This is supported by the paradoxical finding that acute activation of sleep-promoting circuits worsens seizures, because it increases sleep need without changing sleep amount. Sleep-promoting circuits become hyperactive after sleep loss and are associated with increased whole-brain activity. During sleep restriction, optogenetic inhibition of sleep-promoting circuits to reduce sleepiness protects against seizures. Downregulation of the 5HT1A serotonin receptor in sleep-promoting cells mediates the effect of sleep need on seizures, and an FDA-approved 5HT1A agonist was identified to mitigate seizures. These findings demonstrate that while homeostatic sleep is needed to recoup lost sleep, it comes at the cost of increasing seizure susceptibility. This study provides an unexpected perspective on interactions between sleep and seizures, and surprisingly implicate sleep- promoting circuits as a therapeutic target for seizure control. |
| Ahn, J. S., Mahbub, N. U., Kim, S., Kim, H. B., Choi, J. S., Chung, H. J., Hong, S. T. (2023). Nectandrin B significantly increases the lifespan of Drosophila - Nectandrin B for longevity. Aging, 15(22):12749-12762 PubMed ID: 37983180
Summary: Phytochemicals are increasingly recognized in the field of healthy aging as potential therapeutics against various aging-related diseases. Nutmeg, derived from the Myristica fragrans tree, is an example. Nutmeg has been extensively studied and proven to possess antioxidant properties that protect against aging and alleviate serious diseases such as cancer, heart disease, and liver disease. However, the specific active ingredient in nutmeg responsible for these health benefits has not been identified thus far. In this study presents evidence that Nectandrin B (NecB), a bioactive lignan compound isolated from nutmeg, significantly extended the lifespan of the fruit fly Drosophila melanogaster by as much as 42.6% compared to the control group. NecB also improved age-related symptoms including locomotive deterioration, body weight gain, eye degeneration, and neurodegeneration in aging D. melanogaster. This result represents the most substantial improvement in lifespan observed in animal experiments to date, suggesting that NecB may hold promise as a potential therapeutic agent for promoting longevity and addressing age-related degeneration. | Lenzi, C., Piat, A., Schlich, P., Ducau, J., Bregliano, J. C., Aguilaniu, H., Laurencon, A. (2023). Parental age effect on the longevity and healthspan in Drosophila melanogaster and Caenorhabditis elegans. Aging, 15(21):11720-11739 PubMed ID: 37917003
Summary: Several studies have investigated the effect of parental age on biological parameters such as reproduction, lifespan, and health; however, the results have been inconclusive, largely due to inter-species variation and/or modest effect sizes. This studsy examined the effect of parental age on the lifespan, reproductive capacity, and locomotor activity of genetic isogenic lines of the nematode Caenorhabditis elegans and the fruit fly Drosophila melanogaster. The progeny of successive generations of old parents had significantly shorter lifespans than the progeny of young parents in both species. Moreover, this study investigated the fertility, fecundity, and locomotor activity of C. elegans. Interestingly, both the shorter lifespan and deteriorated healthspan of the progeny were significantly improved by switching to only one generation of younger parents. Collectively, these data demonstrate that the detrimental effect of older parental age on the longevity of the progeny can be reversed, suggesting the existence of a beneficial non-genetic mechanism. |
Wednesday, July 3rd - Adult Neural Structure, Development, and Function |
| Martin, M., Gutierrez-Avino, F., Shaikh, M. N., Tejedor, F. J. (2023). A novel proneural function of Asense is integrated with the sequential actions of Delta-Notch, L'sc and Su(H) to promote the neuroepithelial to neuroblast transition. PLoS Genet, 19(10):e1010991 PubMed ID: 37871020
Summary: In order for neural progenitors (NPs) to generate distinct populations of neurons at the right time and place during CNS development, they must switch from undergoing purely proliferative, self-renewing divisions to neurogenic, asymmetric divisions in a tightly regulated manner. In the developing Drosophila optic lobe, neuroepithelial (NE) cells of the outer proliferation center (OPC) are progressively transformed into neurogenic NPs called neuroblasts (NBs) in a medial to lateral proneural wave. The cells undergoing this transition express Lethal of Scute (L'sc), a proneural transcription factor (TF) of the Acheate Scute Complex (AS-C). There is also a peak of expression of Asense (Ase), another AS-C TF, in the cells neighboring those with transient L'sc expression. These peak of Ase cells help to identify a new transitional stage as they have lost NE markers and L'sc, they receive a strong Notch signal and barely exhibit NB markers. This expression of Ase is necessary and sufficient to promote the NE to NB transition in a more robust and rapid manner than that of l'sc gain of function or Notch loss of function. Thus, these data provide the first direct evidence of a proneural role for Ase in CNS neurogenesis. Strikingly, it was found that strong Delta-Notch signaling at the lateral border of the NE triggers l'sc expression, which in turn induces ase expression in the adjacent cells through the activation of Delta-Notch signaling. These results reveal two novel non-conventional actions of Notch signaling in driving the expression of proneural factors, in contrast to the repression that Notch signaling exerts on them during classical lateral inhibition. Finally, Suppressor of Hairless (Su(H)), which seems to be upregulated late in the transitioning cells and in NBs, represses l'sc and ase, ensuring their expression is transient. Thus, these data identify a key proneural role of Ase that is integrated with the sequential activities of Delta-Notch signaling, L'sc, and Su(H), driving the progressive transformation of NE cells into NBs. | Peng, Y. X., Liu, Z. Y., Lin, P. X., Su, S. C., Gao, C. F., Wu, S. F. (2023). Reverse genetic study reveals the molecular targets of chordotonal organ TRPV channel modulators. Pesticide biochemistry and physiology, 196:105584 PubMed ID: 37945222
Summary: Insecticides have been widely used for the control of insect pests that have a significant impact on agriculture and human health. A better understanding of insecticide targets is needed for effective insecticide design and resistance management. Pymetrozine, afidopyropen and flonicamid are reported to target on proteins that located on insect chordotonal organs, resulting in the disruption of insect coordination and the inhibition of feeding. This study systematically examined the susceptibility of six Drosophila melanogaster mutants (five transient receptor potential channels and one mechanoreceptor) to three commercially used insecticides, in order to identify the receptor subunits critical to the insect's response to insecticides. The results showed that iav1, nan36a and wtrw1/i< mutants exhibited significantly reduced susceptibility to pymetrozine and afidopyropen, but not to flonicamid. The number of eggs produced by the three mutant females were significantly less than that of the w(1118) strain. Meanwhile, the longevity of all male mutants and females of nan36a and wtrw1 mutants was significantly shorter than that of the w(1118) strain as the control. However, no gravitaxis defects were observed in wtrw1 mutants and the anti-gravitaxis ofwtrw1 mutants was abolished by pymetrozine. Behavioral assays using thermogenetic tools further confirmed the bioassay results and supported the idea that Nan as a TRPV subfamily member located in Drosophila chordotonal neurons, acting as a target of pymetrozine, which interferes with Drosophila and causes motor deficits with gravitaxis defects. Taken together, this study elucidates the interactions of pymetrozine and afidopyropen with TRPV channels, Nan and Iav, and TRPA channel, Wtrw. This research provides another evidence that pymetrozine and afidopyropen might target on nan, iav and wtrw channels and provides insights into the development of sustainable pest management strategies. |
| Gonzalez, Y. R., Kamkar, F., Jafar-Nejad, P., Wang, S., Qu, D., Alvarez, L. S., Hawari, D., Sonnenfeld, M., Slack, R. S., Albert, P. R., Park, D. S., Joselin, A. (2023). PFTK1 kinase regulates axogenesis during development via RhoA activation. BMC Biol, 21(1):240 PubMed ID: 37907898
Summary: BAPFTK1/Eip63E is a member of the cyclin-dependent kinases (CDKs) family and plays an important role in normal cell cycle progression. Eip63E expresses primarily in postnatal and adult nervous system in Drosophila melanogaster but its role in CNS development remains unknown. This study sought to understand the function of Eip63E in the CNS by studying the fly ventral nerve cord during development. The results demonstrate that Eip63E regulates axogenesis in neurons and its deficiency leads to neuronal defects. Functional interaction studies performed using the same system identify an interaction between Eip63E and the small GTPase Rho1. Furthermore, deficiency of Eip63E homolog in mice, PFTK1, in a newly generated PFTK1 knockout mice results in increased axonal outgrowth confirming that the developmental defects observed in the fly model are due to defects in axogenesis. Importantly, RhoA phosphorylation and activity are affected by PFTK1 in primary neuronal cultures. GDP-bound inactive RhoA is a substrate of PFTK1 and PFTK1 phosphorylation is required for RhoA activity. In conclusion, this work establishes an unreported neuronal role of PFTK1 in axon development mediated by phosphorylation and activation of GDP-bound RhoA. The results presented add to understanding of the role of Cdks in the maintenance of RhoA-mediated axon growth and its impact on CNS development and axonal regeneration. | Ahn, J. E., Amrein, H. (2023). Opposing chemosensory functions of closely related gustatory receptors. bioRxiv, PubMed ID: 37905057
Summary: Most animals have functionally distinct populations of taste cells, expressing receptors that are tuned to compounds of different valence. In D. melanogaster, primary sensory neurons express taste receptors that are tuned to distinct groups of chemicals, thereby activating neural ensembles that elicit either feeding or avoidance behavior. Members of a family of ligand gated receptor channels, the Gustatory receptors (Grs), play a central role in these behaviors. In general, closely related, evolutionarily conserved Gr proteins are co-expressed in the same type of taste neurons, tuned to chemically related compounds, and therefore triggering the same behavioral response. This study reports that members of the Gr28 subfamily are expressed in largely non-overlapping sets of taste neurons in Drosophila larvae, detect chemicals of different valence and trigger opposing feeding behaviors. The intrinsic properties of Gr28 neurons by were determined by expressing the mammalian Vanilloid Receptor (VR1), which is activated by capsaicin, a chemical to which wildtype Drosophila larvae do not respond. When VR1 is expressed in Gr28a neurons, larvae become attracted to capsaicin, consistent with reports showing that Gr28a itself encodes a receptor for nutritious RNA. In contrast, expression of VR1 in two pairs of Gr28b.c neurons triggers avoidance to capsaicin. Moreover, neuronal inactivation experiments show that the Gr28b.c neurons are necessary for avoidance of several bitter compounds. Lastly, behavioral experiments of Gr28 deficient larvae and live Ca (2+) imaging studies of Gr28b.c neurons revealed that denatonium benzoate, a synthetic bitter compound that shares structural similarities with natural bitter chemicals, is a ligand for a receptor complex containing a Gr28b.c or Gr28b.a subunit. Thus, the Gr28 proteins, which have been evolutionarily conserved over 260 million years in insects, represent the first taste receptor subfamily in which specific members mediate behavior with opposite valence. |
| Dweck, H. K. M., Carlson, J. R. (2023). Diverse mechanisms of taste coding in Drosophila. Sci Adv, 9(46):eadj7032 PubMed ID: 37976361
Summary: Taste systems encode chemical cues that drive vital behaviors. This study has elucidated noncanonical features of taste coding using an unconventional kind of electrophysiological analysis. Taste neurons of Drosophila are much more sensitive than previously thought. They have a low spontaneous firing frequency that depends on taste receptors. Taste neurons have a dual function as olfactory neurons: They are activated by most tested odorants, including N,N-diethyl-meta-toluamide (DEET), at a distance. DEET can also inhibit certain taste neurons, revealing that there are two modes of taste response: activation and inhibition. Electrophysiological OFF responses were characterized and it was found that the tastants that elicit them are related in structure. OFF responses link tastant identity to behavior: the magnitude of the OFF response elicited by a tastant correlated with the egg laying behavior it elicited. In summary, the sensitivity and coding capacity of the taste system are much greater than previously known. | Jouandet, G. C., Alpert, M. H., Simoes, J. M., Suhendra, R., Frank, D. D., Levy, J. I., Para, A., Kath, W. L., Gallio, M. (2023). Rapid threat assessment in the Drosophila thermosensory system. Nat Commun, 14(1):7067 PubMed ID: 37923719
Summary: Neurons that participate in sensory processing often display "ON" responses, i.e., fire transiently at the onset of a stimulus. ON transients are widespread, perhaps universal to sensory coding, yet their function is not always well-understood. This study shows that ON responses in the Drosophila thermosensory system extrapolate the trajectory of temperature change, priming escape behavior if unsafe thermal conditions are imminent. First, it was shown that second-order thermosensory projection neurons (TPN-IIIs) and their Lateral Horn targets (TLHONs), display ON responses to thermal stimuli, independent of direction of change (heating or cooling) and of absolute temperature. Instead, they track the rate of temperature change, with TLHONs firing exclusively to rapid changes (>0.2 °C/s). Next, connectomics were used to track TLHONs' output to descending neurons that control walking and escape, and modeling and genetic silencing to demonstrate how ON transients can flexibly amplify aversive responses to small thermal change. These results suggest that, across sensory systems, ON transients may represent a general mechanism to systematically anticipate and respond to salient or dangerous conditions. |
Tuesday, July 2nd - Adult Physiology and Metabolism |
| Malik, D. M., Sengupta, A., Sehgal, A., Weljie, A. M. (2023). Altered Metabolism During the Dark Period in Drosophila Short Sleep Mutants. bioRxiv, PubMed ID: 37961245
Summary: Sleep is an almost universally required state in biology. Disrupted sleep has been associated with adverse health risks including metabolic perturbations. Sleep is in part regulated via circadian mechanisms, however, metabolic dysfunction at different times of day arising from sleep disruption is unclear. This study used targeted liquid chromatography-mass spectrometry to probe metabolic alterations using high-resolution temporal sampling of two Drosophila short sleep mutants, fumin and sleepless, across a circadian day. Discriminant analyses revealed overall distinct metabolic profiles for mutants when compared to a wild type dataset. Altered levels of metabolites involved in nicotinate/nicotinamide, alanine, aspartate, and glutamate, glyoxylate and dicarboxylate metabolism, and the TCA cycle were observed in mutants suggesting increased energetic demands. Furthermore, rhythmicity analyses revealed fewer 24 hr rhythmic metabolites in both mutants. Interestingly, mutants displayed two major peaks in phases while wild type displayed phases that were less concerted. In contrast to 24 hr rhythmic metabolites, an increase in the number of 12 hr rhythmic metabolites was observed in fumin while sleepless displayed a decrease. These results support that decreased sleep alters the overall metabolic profile with short sleep mutants displaying altered metabolite levels associated with a number of pathways in addition to altered neurotransmitter levels. | Tang, C., Li, Q., Wang, X., Yu, Z., Ping, X., Qin, Y., Liu, Y., Zheng, L. (2024). Cardiac Timeless Trans-Organically Regulated by miR-276 in Adipose Tissue Modulates Cardiac Function. Function (Oxford, England), 5(1):zqad064 PubMed ID: 38058384
Summary: The interconnection between cardiac function and circadian rhythms is of great importance. While the role of the biological clock gene Timeless (Tim) in circadian rhythm has been extensively studied, its impact on cardiac function remains largely been unexplored. Previous research has provided experimental evidence for the regulation of the heart by adipose tissue and the targeting of miR-276a/b on Timeless. However, the extent to which adipose tissue regulates cardiac Timeless genes trans-organically through miR-276a/b, and subsequently affects cardiac function, remains uncertain. Therefore, the objective of this study was to investigate the potential trans-organ modulation of the Timeless gene in the heart by adipose tissue through miR-276a/b. Cardiac-specific Timeless knockdown and overexpression was shown to result in a significant increase in heart rate (HR) and a significant decrease in Heart period (HP), diastolic intervals (DI), systolic intervals (SI), diastolic diameter (DD), and systolic diameter (SD). miR-276b systemic knockdown resulted in a significant increase in DI, arrhythmia index (AI), and fractional shortening (FS) significantly increased and SI, DD and SD significantly decreased. Adipose tissue-specific miR-276a/b knockdown and miR-276a overexpression resulted in a significant increase in HR and a significant decrease in DI and SI, which were improved by exercise intervention. This study presents a novel finding that highlights the significance of the heart circadian clock gene Timeless in heart function. Additionally, it demonstrates that adipose tissue exerts trans-organ modulation on the expression of the heart Timeless gene via miR-276a/b. |
| Li, X., Karpac, J. (2023). A distinct Acyl-CoA binding protein (ACBP6) shapes tissue plasticity during nutrient adaptation in Drosophila. Nat Commun, 14(1):7599 PubMed ID: 37989752
Summary: Nutrient availability is a major selective force in the evolution of metazoa, and thus plasticity in tissue function and morphology is shaped by adaptive responses to nutrient changes. Utilizing Drosophila, this study revealed that distinct calibration of acyl-CoA metabolism, mediated by Acbp6 (Acyl-CoA binding-protein 6), is critical for nutrient-dependent tissue plasticity. Drosophila Acbp6, which arose by evolutionary duplication and binds acyl-CoA to tune acetyl-CoA metabolism, is required for intestinal resizing after nutrient deprivation through activating intestinal stem cell proliferation from quiescence. Disruption of acyl-CoA metabolism by Acbp6 attenuation drives aberrant 'switching' of metabolic networks in intestinal enterocytes during nutrient adaptation, impairing acetyl-CoA metabolism and acetylation amid intestinal resizing. STAT92e, whose function is influenced by acetyl-CoA levels, was also identified as a key regulator of acyl-CoA and nutrient-dependent changes in stem cell activation. These findings define a regulatory mechanism, shaped by acyl-CoA metabolism, that adjusts proliferative homeostasis to coordinately regulate tissue plasticity during nutrient adaptation. | Mappin, F., Bellantuono, A. J., Ebrahimi, B., DeGennaro, M. (2023). Odor-evoked transcriptomics of Aedes aegypti mosquitoes. PLoS One, 18(10):e0293018 PubMed ID: 37874813
Summary: Modulation of odorant receptors mRNA induced by prolonged odor exposure is highly correlated with ligand-receptor interactions in Drosophila as well as mammals of the Muridae family. If this response feature is conserved in other organisms, this presents an intriguing initial screening tool when searching for novel receptor-ligand interactions in species with predominantly orphan olfactory receptors. This study demonstrates that mRNA modulation in response to 1-octen-3-ol odor exposure occurs in a time- and concentration-dependent manner in Aedes aegypti mosquitoes. To investigate gene expression patterns at a global level, an odor-evoked transcriptome was generated associated with 1-octen-3-ol odor exposure. Transcriptomic data revealed that ORs and OBPs were transcriptionally responsive whereas other chemosensory gene families showed little to no differential expression. Alongside chemosensory gene expression changes, transcriptomic analysis found that prolonged exposure to 1-octen-3-ol modulated xenobiotic response genes, primarily members of the cytochrome P450, insect cuticle proteins, and glucuronosyltransferases families. Together, these findings suggest that mRNA transcriptional modulation of olfactory receptors caused by prolonged odor exposure is pervasive across taxa and can be accompanied by the activation of xenobiotic responses. |
| Malik, D. M., Rhoades, S. D., Zhang, S. L., Sengupta, A., Barber, A., Haynes, P., Arnadottir, E. S., Pack, A., Kibbey, R. G., Sehgal, A., Weljie, A. M. (2023). Glucose Challenge Uncovers Temporal Fungibility of Metabolic Homeostasis Throughout the Day. bioRxiv, PubMed ID: 37961230
Summary: Rhythmicity is a central feature of behavioral and biological processes including metabolism, however, the mechanisms of metabolite cycling are poorly understood. A robust oscillation in a network of key metabolite pathways downstream of glucose is described in humans, then these pathways mechanistically probed through purpose-built (13)C(6)-glucose isotope tracing in Drosophila every 4h. A temporal peak in biosynthesis was noted by broad labelling of pathways downstream of glucose in wild-type flies shortly following lights on. Krebs cycle labelling was generally increased in a hyperactive mutant (fumin) along with glycolysis labelling primarily observed at dawn. Surprisingly, neither underlying feeding rhythms nor the presence of food explains the rhythmicity of glucose processing across genotypes. These results are consistent with clinical data demonstrating detrimental effects of mis-timed energy intake. This approach provides a window into the dynamic range of metabolic processing ability through the day and mechanistic basis for exploring circadian metabolic homeostasis in disease states. | Qiao, S., Bernasek, S., Gallagher, K. D., Yamada, S., Bagheri, N., Amaral, L. A. N., Carthew, R. W. (2023). Energy metabolism modulates the regulatory impact of activators on gene expression. bioRxiv, PubMed ID: 37961620
Summary: Gene expression is a regulated process fueled by ATP consumption. Therefore, regulation must be coupled to constraints imposed by the level of energy metabolism. This study explored this relationship both theoretically and experimentally. A stylized mathematical model predicts that activators of gene expression have variable impact depending on metabolic rate. Activators become less essential when metabolic rate is reduced and more essential when metabolic rate is enhanced. In the Drosophila eye, expression dynamics of the yan gene are less affected by loss of EGFR-mediated activation when metabolism is reduced, and the opposite effect is seen when metabolism is enhanced. The effects are also seen at the level of pattern regularity in the adult eye, where loss of EGFR-mediated activation is mitigated by lower metabolism. It is proposed that gene activation is tuned by energy metabolism to allow for faithful expression dynamics in the face of variable metabolic conditions. |
Monday, July 1st - Monday - Chromatin, DNA, and Chromosome Dynamics |
| Gemeinhardt, T. M., Regy, R. M., Mendiola, A. J., Ledterman, H. J., Henrickson, A., Phan, T. M., Kim, Y. C., Demeler, B., Kim, C. A., Mittal, J., Francis, N. J. (2023). How a disordered linker in the Polycomb protein Polyhomeotic tunes phase separation and oligomerization. bioRxiv, PubMed ID: 37961422
Summary: The Polycomb Group (PcG) complex PRC1 represses transcription, forms condensates in cells, and modifies chromatin architecture. These processes are connected through the essential, polymerizing Sterile Alpha Motif (SAM) present in the PRC1 subunit Polyhomeotic (Ph). In vitro, Ph SAM drives formation of short oligomers and phase separation with DNA or chromatin in the context of a Ph truncation ("mini-Ph"). Oligomer length is controlled by the long disordered linker (L) that connects the SAM to the rest of Ph--replacing Drosophila Ph linker (PhL) with the evolutionarily diverged human PHC3L strongly increases oligomerization. How the linker controls SAM polymerization, and how polymerization and the linker affect condensate formation are not know. This analyzed PhL and PHC3L using biochemical assays and molecular dynamics (MD) simulations. PHC3L promotes mini-Ph phase separation and makes it relatively independent of DNA. In MD simulations, basic amino acids in PHC3L form contacts with acidic amino acids in the SAM. Engineering the SAM to make analogous charge-based contacts with PhL increased polymerization and phase separation, partially recapitulating the effects of the PHC3L. Ph to PHC3 linker swaps and SAM surface mutations alter Ph condensate formation in cells, and Ph function in Drosophila imaginal discs. Thus, SAM-driven phase separation and polymerization are conserved between flies and mammals, but the underlying mechanisms have diverged through changes to the disordered linker. | Shaukat, A., Bakhtiari, M. H., Chaudhry, D. S., Khan, M. H. F., Akhtar, J., Abro, A. H., Haseeb, M. A., Sarwar, A., Mazhar, K., Umer, Z., Tariq, M. (2024). Mask exhibits trxG-like behavior and associates with H3K27ac marked chromatin. Dev Biol, 505:130-140 PubMed ID: 37981061
Summary: The Trithorax group (trxG) proteins counteract the repressive effect of Polycomb group (PcG) complexes and maintain transcriptional memory of active states of key developmental genes. Although chromatin structure and modifications appear to play a fundamental role in this process, it is not clear how trxG prevents PcG-silencing and heritably maintains an active gene expression state. This study reports a hitherto unknown role of Drosophila Multiple ankyrin repeats single KH domain (Mask), which emerged as one of the candidate trxG genes in a reverse genetic screen. The genome-wide binding profile of Mask correlates with known trxG binding sites across the Drosophila genome. In particular, the association of Mask at chromatin overlaps with CBP and H3K27ac, which are known hallmarks of actively transcribed genes by trxG. Importantly, Mask predominantly associates with actively transcribed genes in Drosophila. Depletion of Mask not only results in the downregulation of trxG targets but also correlates with diminished levels of H3K27ac. The fact that Mask positively regulates H3K27ac levels in flies was also found to be conserved in human cells. Strong suppression of Pc mutant phenotype by mutation in mask provides physiological relevance that Mask contributes to the anti-silencing effect of trxG, maintaining expression of key developmental genes. Since Mask is a downstream effector of multiple cell signaling pathways, it is proposed that Mask may connect cell signaling with chromatin mediated epigenetic cell memory governed by trxG. |
| Godneeva, B., Fejes Toth, K., Quan, B., Chou, T. F., Aravin, A. A. (2023). Impact of Germline Depletion of Bonus on Chromatin State in Drosophila Ovaries. Cells, 12(22) PubMed ID: 37998364
Summary: Gene expression is controlled via complex regulatory mechanisms involving transcription factors, chromatin modifications, and chromatin regulatory factors. Histone modifications, such as H3K27me3, H3K9ac, and H3K27ac, play an important role in controlling chromatin accessibility and transcriptional output. In vertebrates, the Transcriptional Intermediary Factor 1 (TIF1) family of proteins play essential roles in transcription, cell differentiation, DNA repair, and mitosis. This study focused on Bonus, the sole member of the TIF1 family in Drosophila, to investigate its role in organizing epigenetic modifications. The findings demonstrated that depleting Bonus in ovaries leads to a mild reduction in the H3K27me3 level over transposon regions and alters the distribution of active H3K9ac marks on specific protein-coding genes. Additionally, through mass spectrometry analysis, novel interacting partners of Bonus in ovaries were identifed, such as PolQ, providing a comprehensive understanding of the associated molecular pathways. Furthermore, this research revealed Bonus's interactions with the Polycomb Repressive Complex 2 and its co-purification with select histone acetyltransferases, shedding light on the underlying mechanisms behind these changes in chromatin modifications. | Golovnin, A., Melnikova, L., Babosha, V., Pokholkova, G. V., Slovohotov, I., Umnova, A., Maksimenko, O., Zhimulev, I. F., Georgiev, P. (2023). The N-Terminal Part of Drosophila CP190 Is a Platform for Interaction with Multiple Architectural Proteins. Int J Mol Sci, 24(21) PubMed ID: 37958900
Summary: CP190 is a co-factor in many Drosophila architectural proteins, being involved in the formation of active promoters and insulators. CP190 contains the N-terminal BTB/POZ (Broad-Complex, Tramtrack and Bric a brac/POxvirus and Zinc finger) domain and adjacent conserved regions involved in protein interactions. This study examined the functional roles of these domains of CP190 in vivo. The best-characterized architectural proteins with insulator functions, Pita, Su(Hw), and M1BP, Opbp, and ZIPIC, interacted with one or both of the highly conserved regions in the N-terminal part of CP190. Transgenic lines of D. melanogaster expressing CP190 mutants with a deletion of each of these domains were obtained. The results showed that these mutant proteins only partially compensated for the functions of CP190, weakly binding to selective chromatin sites. Further analysis confirmed the essential role of these domains in recruitment to regulatory regions associated with architectural proteins. The N-terminal of CP190 was found to be sufficient for recruiting Z4 and Chromator proteins and successfully achieving chromatin opening. Taken together, these results and the results of previous studies showed that the N-terminal region of CP190 is a platform for simultaneous interaction with various DNA-binding architectural proteins and transcription complexes. |
| Al Zouabi, L., Stefanutti, M., Roumeliotis, S., Le Meur, G., Boumard, B., Riddiford, N., Rubanova, N., Bohec, M., Gervais, L., Servant, N., Bardin, A. J. (2023). Chromatin state transitions in the Drosophila intestinal lineage identify principles of cell-type specification. Dev Cell, 58(24):3048-3063 PubMed ID: 38056452
Summary: Tissue homeostasis relies on rewiring of stem cell transcriptional programs into those of differentiated cells. This study investigate changes in chromatin occurring in a bipotent adult stem cells. Combining mapping of chromatin-associated factors with statistical modeling, genome-wide transitions during differentiation were identified in the adult Drosophila intestinal stem cell (ISC) lineage. Active, stem-cell-enriched genes transition to a repressive heterochromatin protein-1-enriched state more prominently in enteroendocrine cells (EEs) than in enterocytes (ECs), in which the histone H1-enriched Black state is preeminent. In contrast, terminal differentiation genes associated with metabolic functions follow a common path from a repressive, primed, histone H1-enriched Black state in ISCs to active chromatin states in EE and EC cells. Furthermore, Lineage priming was found to have an important function in adult ISCs, and histone H1 as a mediator of this process. These data define underlying principles of chromatin changes during adult multipotent stem cell differentiation. | Al Zouabi, L., Stefanutti, M., Roumeliotis, S., Le Meur, G., Boumard, B., Riddiford, N., Rubanova, N., Bohec, M., Gervais, L., Servant, N., Bardin, A. J. (2023). Molecular underpinnings and environmental drivers of loss of heterozygosity in Drosophila intestinal stem cells . Cell Rep 38032794
Summary: During development and aging, genome mutation leading to loss of heterozygosity (LOH) can uncover recessive phenotypes within tissue compartments. This phenomenon occurs in normal human tissues and is prevalent in pathological genetic conditions and cancers. While studies in yeast have defined DNA repair mechanisms that can promote LOH, the predominant pathways and environmental triggers in somatic tissues of multicellular organisms are not well understood. This study investigated mechanisms underlying LOH in intestinal stem cell in Drosophila. Infection with the pathogenic bacteria, Erwinia carotovora carotovora 15, but not Pseudomonas entomophila, increases LOH frequency. Using whole genome sequencing of somatic LOH events, it was demonstrated that they arise primarily via mitotic recombination. Molecular features and genetic evidence argue against a break-induced replication mechanism and instead support cross-over via double Holliday junction-based repair. This study provides a mechanistic understanding of mitotic recombination, an important mediator of LOH, and its effects on stem cells in vivo. |
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