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Current papers in developmental biology and gene function
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| ARCHIVE | Friday, March 29th, 2019 - Behavior |
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What's hot today
July 2022 June 2022 May 2022 April 2022 March 2022 February 2022 January 2022 December 2021 November 2021 October 2021 September 2021 August 2021 July 2021 June 2021 May 2021 April 2021 March 2021 February 2021 January 2021 December 2020 November 2020 October 2020 September 2020 March 2020 April 2017 March 2017 | Mossman, J. A., Mabeza, R. M. S., Blake, E., Mehta, N. and Rand, D. M. (2019). Age of both parents influences reproduction and egg dumping behavior in Drosophila melanogaster. J Hered. PubMed ID: 30753690
Summary: Trans-generational maternal effects have been shown to influence a broad range of offspring phenotypes. However, very little is known about paternal trans-generational effects. This study tested the trans-generational effects of maternal and paternal age, and their interaction, on daughter and son reproductive fitness in Drosophila melanogaster. Significant effects were found of parent ages on offspring reproductive fitness over 10 days post-fertilization. In daughters, older (45 days old) mothers conferred lower reproductive fitness compared to younger mothers (3 days old). In sons, father's age significantly affected reproductive fitness. The effects of two old parents were additive in both sexes and reproductive fitness was lowest when the focal individual had two old parents. Interestingly, daughter fertility was sensitive to father's age but son fertility was insensitive to mother's age, suggesting a sexual asymmetry in trans-generational effects. The egg-laying dynamics in daughters dramatically shaped this relationship. Daughters with two old parents demonstrated an extreme egg dumping behavior on day one and laid >2.35 X the number of eggs than the other three age class treatments. This study reveals significant trans-generational maternal and paternal age effects on fertility and an association with a novel egg laying behavioral phenotype in Drosophila. | Chen, W., Xue, Y., Scarfe, L., Wang, D. and Zhang, Y. (2019). Loss of Prune in circadian cells decreases the amplitude of the circadian locomotor rhythm in Drosophila. Front Cell Neurosci 13: 76. PubMed ID: 30881291
Summary: The circadian system, which has a period of about 24 h, is import for organismal health and fitness. The molecular circadian clock consists of feedback loops involving both transcription and translation, and proper function of the circadian system also requires communication among intracellular organelles. As important hubs for signaling in the cell, mitochondria integrate a variety of signals. Mitochondrial dysfunction and disruption of circadian rhythms are observed in neurodegenerative diseases and during aging. However, how mitochondrial dysfunction influences circadian rhythm is largely unknown. This study reports that Drosophila prune (pn), which localizes to the mitochondrial matrix, most likely affects the function of certain clock neurons. Deletion of pn in flies caused decreased expression of mitochondrial transcription factor TFAM and reductions in levels of mitochondrial DNA, which resulted in mitochondrial dysfunction. Loss of pn decreased the amplitude of circadian rhythms. In addition, depletion of mtDNA by overexpression of a mitochondrially targeted restriction enzyme mitoXhoI also decreased the robustness of circadian rhythms. This work demonstrates that pn is important for mitochondrial function thus involved in the regulation of circadian rhythms. |
| Toda, H., Williams, J. A., Gulledge, M. and Sehgal, A. (2019). A sleep-inducing gene, nemuri, links sleep and immune function in Drosophila. Science 363(6426): 509-515. PubMed ID: 30705188
Summary: Sleep remains a major mystery of biology. In particular, little is known about the mechanisms that account for the drive to sleep. In an unbiased screen of more than 12,000 Drosophila lines, identified a single gene, nemuri (CG31813), that induces sleep. The NEMURI protein is an antimicrobial peptide that can be secreted ectopically to drive prolonged sleep (with resistance to arousal) and to promote survival after infection. Loss of nemuri increased arousability during daily sleep and attenuated the acute increase in sleep induced by sleep deprivation or bacterial infection. Conditions that increase sleep drive induced expression of nemuri in a small number of fly brain neurons and targeted it to the sleep-promoting, dorsal fan-shaped body. It is proposed that NEMURI is a bona fide sleep homeostasis factor that is particularly important under conditions of high sleep need; because these conditions include sickness, these findings provide a link between sleep and immune function. | Giraldo, D., Adden, A., Kuhlemann, I., Gras, H. and Geurten, B. R. H. (2019). Correcting locomotion dependent observation biases in thermal preference of Drosophila. Sci Rep 9(1): 3974. PubMed ID: 30850647
Summary: Sensing environmental temperatures is essential for the survival of ectothermic organisms. One drawback of gradients is that small ectothermic animals are susceptible to cold-trapping: a physiological inability to move at the cold area of the gradient. Often cold-trapping cannot be avoided, biasing the resulting temperature preference (TP) to lower temperatures. Two mathematical models were previously developed to correct for cold-trapping. These models, however, focus on group behaviour which can lead to overestimation of cold-trapping due to group aggregation. This study presents a mathematical model that simulates the behaviour of individual Drosophila in temperature gradients. The model takes the spatial dimension and temperature difference of the gradient into account, as well as the rearing temperature of the flies. Furthermore, it allows the quantification of cold-trapping and reveals unbiased TP. Additionally, the model reveals that flies have a range of tolerable temperatures, and this measure is more informative about the behaviour than commonly used TP. |
| Geissmann, Q., Beckwith, E. J. and Gilestro, G. F. (2019). Most sleep does not serve a vital function: Evidence from Drosophila melanogaster. Sci Adv 5(2): eaau9253. PubMed ID: 30801012
Summary: Sleep appears to be a universally conserved phenomenon among the animal kingdom, but whether this notable evolutionary conservation underlies a basic vital function is still an open question. Using a machine learning-based video-tracking technology, a detailed high-throughput analysis of sleep was conducted in the fruit fly Drosophila melanogaster, coupled with a lifelong chronic and specific sleep restriction. These results show that some wild-type flies are virtually sleepless in baseline conditions and that complete, forced sleep restriction is not necessarily a lethal treatment in wild-type D. melanogaster. It was also shown that circadian drive, and not homeostatic regulation, is the main contributor to sleep pressure in flies. These results offer a new perspective on the biological role of sleep in Drosophila and, potentially, in other species. | Chen, K. F., Lowe, S., Lamaze, A., Kratschmer, P. and Jepson, J. (2019). Neurocalcin regulates nighttime sleep and arousal in Drosophila. Elife 8. PubMed ID: 30865587
Summary: Sleep-like states in diverse organisms can be separated into distinct stages, each with a characteristic arousal threshold. However, the molecular pathways underlying different sleep stages remain unclear. The fruit fly, Drosophila melanogaster, exhibits consolidated sleep during both day and night, with night sleep associated with higher arousal thresholds compared to day sleep. This study identified a role for the neuronal calcium sensor protein Neurocalcin (NCA) in promoting sleep during the night but not the day by suppressing nocturnal arousal and hyperactivity. Both circadian and light-sensing pathways define the temporal window in which NCA promotes sleep. Furthermore, NCA promotes sleep by suppressing synaptic release from a dispersed wake-promoting neural network and the mushroom bodies, a sleep-regulatory center, are a module within this network. These results advance the understanding of how sleep stages are genetically defined. |
Thursday March 28th - RNA and Transposons |
| Oberhofer, G., Ivy, T. and Hay, B. A. (2019). Cleave and Rescue, a novel selfish genetic element and general strategy for gene drive. Proc Natl Acad Sci U S A. PubMed ID: 30760597
Summary: There is great interest in being able to spread beneficial traits throughout wild populations in ways that are self-sustaining. This study describes a chromosomal selfish genetic element, CleaveR [Cleave and Rescue (ClvR)], able to achieve this goal. ClvR comprises two linked chromosomal components. One, germline-expressed Cas9 and guide RNAs (gRNAs)-the Cleaver-cleaves and thereby disrupts endogenous copies of a gene whose product is essential. The other, a recoded version of the essential gene resistant to cleavage and gene conversion with cleaved copies-the Rescue-provides essential gene function. ClvR enhances its transmission, and that of linked genes, by creating conditions in which progeny lacking ClvR die because they have no functional copies of the essential gene. In contrast, those who inherit ClvR survive, resulting in an increase in ClvR frequency. ClvR is predicted to spread to fixation under diverse conditions. To test these predictions, a ClvR element was generated in Drosophila melanogaster. ClvR (tko) is located on chromosome 3 and uses Cas9 and four gRNAs to disrupt melanogaster technical knockout (tko), an X-linked essential gene. Rescue activity is provided by tko from Drosophila virilis ClvR (tko) results in germline and maternal carryover-dependent inactivation of melanogaster tko (>99% per generation); lethality caused by this loss is rescued by the virilis transgene; ClvR (tko) activities are robust to genetic diversity in strains from five continents; and uncleavable but functional melanogaster tko alleles were not observed. Finally, ClvR (tko) spreads to transgene fixation. The simplicity of ClvR suggests it may be useful for altering populations in diverse species. | Rech, G. E., Bogaerts-Marquez, M., Barron, M. G., Merenciano, M., Villanueva-Canas, J. L., Horvath, V., Fiston-Lavier, A. S., Luyten, I., Venkataram, S., Quesneville, H., Petrov, D. A. and Gonzalez, J. (2019). Stress response, behavior, and development are shaped by transposable element-induced mutations in Drosophila. PLoS Genet 15(2): e1007900. PubMed ID: 30753202
Summary: Most of the current knowledge on the genetic basis of adaptive evolution is based on the analysis of single nucleotide polymorphisms (SNPs). Despite increasing evidence for their causal role, the contribution of structural variants to adaptive evolution remains largely unexplored. This work analyzed the population frequencies of 1,615 Transposable Element (TE) insertions annotated in the reference genome of Drosophila melanogaster, in 91 samples from 60 worldwide natural populations. A set of 300 polymorphic TEs were identified that are present at high population frequencies, and located in genomic regions with high recombination rate, where the efficiency of natural selection is high. The age and the length of these 300 TEs are consistent with relatively young and long insertions reaching high frequencies due to the action of positive selection. Besides, a set of 21 fixed TEs also likely to be adaptive was identified. Indeed evidence has been found of selection for 84 of these reference TE insertions. The analysis of the genes located nearby these 84 candidate adaptive insertions suggested that the functional response to selection is related with the GO categories of response to stimulus, behavior, and development. It was further shown that a subset of the candidate adaptive TEs affects expression of nearby genes, and five of them have already been linked to an ecologically relevant phenotypic effect. These results provide a more complete understanding of the genetic variation and the fitness-related traits relevant for adaptive evolution. Similar studies should help uncover the importance of TE-induced adaptive mutations in other species as well. |
| Keskeny, C., Raisch, T., Sgromo, A., Igreja, C., Bhandari, D., Weichenrieder, O. and Izaurralde, E. (2019). A conserved CAF40-binding motif in metazoan NOT4 mediates association with the CCR4-NOT complex. Genes Dev 33(3-4): 236-252. PubMed ID: 30692204
Summary: The multisubunit CCR4-NOT mRNA deadenylase complex plays important roles in the posttranscriptional regulation of gene expression. The NOT4 E3 ubiquitin ligase is a stable component of the CCR4-NOT complex in yeast but does not copurify with the human or Drosophila melanogaster complex. This study shows that the C-terminal regions of human and D. melanogaster NOT4 contain a conserved sequence motif that directly binds the CAF40 subunit of the CCR4-NOT complex (CAF40-binding motif [CBM]). In addition, nonconserved sequences flanking the CBM also contact other subunits of the complex. Crystal structures of the CBM-CAF40 complex reveal a mutually exclusive binding surface for NOT4 and Roquin or Bag of marbles mRNA regulatory proteins. Furthermore, CAF40 depletion or structure-guided mutagenesis to disrupt the NOT4-CAF40 interaction impairs the ability of NOT4 to elicit decay of tethered reporter mRNAs in cells. Together with additional sequence analyses, these results reveal the molecular basis for the association of metazoan NOT4 with the CCR4-NOT complex and show that it deviates substantially from yeast. They mark the NOT4 ubiquitin ligase as an ancient but nonconstitutive cofactor of the CCR4-NOT deadenylase with potential recruitment and/or effector functions. | Ankush Jagtap, P. K., Muller, M., Masiewicz, P., von Bulow, S., Hollmann, N. M., Chen, P. C., Simon, B., Thomae, A. W., Becker, P. B. and Hennig, J. (2019). Structure, dynamics and roX2-lncRNA binding of tandem double-stranded RNA binding domains dsRBD1,2 of Drosophila helicase Maleless. Nucleic Acids Res. PubMed ID: 30805612
Summary: Maleless (MLE) is an evolutionary conserved member of the DExH family of helicases in Drosophila. Besides its function in RNA editing and presumably siRNA processing, MLE is best known for its role in remodelling non-coding roX RNA in the context of X chromosome dosage compensation in male flies. MLE and its human orthologue DHX9 contain two tandem double-stranded RNA binding domains (dsRBDs) located at the N-terminal region. The two dsRBDs are essential for localization of MLE at the X-territory and it is presumed that this involves binding roX secondary structures. However, for dsRBD1 roX RNA binding has so far not been described. This study determined the solution NMR structure of dsRBD1 and dsRBD2 of MLE in tandem and investigated its role in double-stranded RNA (dsRNA) binding. The NMR and SAXS data show that both dsRBDs act as independent structural modules in solution and are canonical, non-sequence-specific dsRBDs featuring non-canonical KKxAXK RNA binding motifs. NMR titrations combined with filter binding experiments and isothermal titration calorimetry (ITC) document the contribution of dsRBD1 to dsRNA binding in vitro. Curiously, dsRBD1 mutants in which dsRNA binding in vitro is strongly compromised do not affect roX2 RNA binding and MLE localization in cells. These data suggest alternative functions for dsRBD1 in vivo. |
| Weigelt, C. M., Hahn, O., Arlt, K., Gruhn, M., Jahn, A. J., Esser, J., Werner, J. A., Klein, C., Buschges, A., Gronke, S. and Partridge, L. (2019). Loss of miR-210 leads to progressive retinal degeneration in Drosophila melanogaster. Life Sci Alliance 2(1). PubMed ID: 30670478
Summary: miRNAs are small, non-coding RNAs that regulate gene expression post-transcriptionally. This study used small RNA sequencing to identify tissue-specific miRNAs in the adult brain, thorax, gut, and fat body of Drosophila melanogaster. One of the most brain-specific miRNAs that was identified was miR-210, an evolutionarily highly conserved miRNA implicated in the regulation of hypoxia in mammals. In Drosophila, miR-210 is specifically expressed in sensory organs, including photoreceptors. miR-210 knockout mutants are not sensitive toward hypoxia but show progressive degradation of photoreceptor cells, accompanied by decreased photoreceptor potential, demonstrating an important function of miR-210 in photoreceptor maintenance and survival. | Cusumano, P., Damulewicz, M., Carbognin, E., Caccin, L., Puricella, A., Specchia, V., Bozzetti, M. P., Costa, R. and Mazzotta, G. M. (2019). The RNA helicase BELLE is involved in circadian rhythmicity and in transposons regulation in Drosophila melanogaster. Front Physiol 10: 133. PubMed ID: 30842743
Summary: Circadian clocks control and synchronize biological rhythms of several behavioral and physiological phenomena in most, if not all, organisms.Rhythmic clock-gene expression is at the base of rhythmic protein accumulation, though post-transcriptional and post-translational mechanisms have evolved to adjust and consolidate the proper pace of the clock. In Drosophila, BELLE, a conserved DEAD-box RNA helicase playing important roles in reproductive capacity, is involved in the small RNA-mediated regulation associated to the piRNA pathway. This study reports that BELLE is implicated in the circadian rhythmicity and in the regulation of endogenous transposable elements (TEs) in both nervous system and gonads. It is suggested that BELLE acts as important element in the piRNA-mediated regulation of the TEs, and the hypothesis is raised that this specific regulation could represent another level of post-transcriptional control adopted by the clock to ensure the proper rhythmicity. |
Wednesday, March 27th - Disease Models |
| Belfer, S. J., Bashaw, A. G., Perlis, M. L. and Kayser, M. S. (2019). A Drosophila model of sleep restriction therapy for insomnia. Mol Psychiatry. PubMed ID: 30824866
Summary: Insomnia is the most common sleep disorder among adults, especially affecting individuals of advanced age or with neurodegenerative disease. Insomnia is also a common comorbidity across psychiatric disorders. Cognitive behavioral therapy for insomnia (CBT-I) is the first-line treatment for insomnia; a key component of this intervention is restriction of sleep opportunity, which optimizes matching of sleep ability and opportunity, leading to enhanced sleep drive. Despite the well-documented efficacy of CBT-I, little is known regarding how CBT-I works at a cellular and molecular level to improve sleep, due in large part to an absence of experimentally-tractable animals models of this intervention. Guided by human behavioral sleep therapies, this study developed a Drosophila model for sleep restriction therapy (SRT) of insomnia. It was demonstrated that restriction of sleep opportunity through manipulation of environmental cues improves sleep efficiency in multiple short-sleeping Drosophila mutants. The response to sleep opportunity restriction requires ongoing environmental inputs, but is independent of the molecular circadian clock. This sleep opportunity restriction paradigm was applied to aging and Alzheimer's disease fly models; sleep impairments in these models are reversible with sleep restriction, with associated improvement in reproductive fitness and extended lifespan. This work establishes a model to investigate the neurobiological basis of CBT-I, and provides a platform that can be exploited toward novel treatment targets for insomnia. | Campbell, N. G., et al. (2019). Structural, functional, and behavioral insights of dopamine dysfunction revealed by a deletion in SLC6A3. Proc Natl Acad Sci U S A 116(9): 3853-3862. PubMed ID: 30755521
Summary: The human dopamine (DA) transporter (hDAT) mediates clearance of DA. Genetic variants in hDAT have been associated with DA dysfunction, a complication associated with several brain disorders, including autism spectrum disorder (ASD). This study investigated the structural and behavioral bases of an ASD-associated in-frame deletion in hDAT at N336 (N336). The deletion promoted a previously unobserved conformation of the intracellular gate of the transporter, likely representing the rate-limiting step of the transport process. It is defined by a "half-open and inward-facing" state (HOIF) of the intracellular gate that is stabilized by a network of interactions conserved phylogenetically, as was demonstrated in hDAT by Rosetta molecular modeling and fine-grained simulations, as well as in its bacterial homolog leucine transporter by electron paramagnetic resonance analysis and X-ray crystallography. The stabilization of the HOIF state is associated both with DA dysfunctions demonstrated in isolated brains of Drosophila melanogaster expressing hDAT N336 and with abnormal behaviors observed at high-time resolution. These flies display increased fear, impaired social interactions, and locomotion traits that are associated with DA dysfunction and the HOIF state. Together, these results describe how a genetic variation causes DA dysfunction and abnormal behaviors by stabilizing a HOIF state of the transporter. |
| Man Anh, H., Linh, D. M., My Dung, V. and Thi Phuong Thao, D. (2019). Evaluating dose- and time-dependent effects of vitamin C treatment on a Parkinson's disease fly model. Parkinsons Dis 2019: 9720546. PubMed ID: 30719278
Summary: Parkinson's disease (PD) is a common neurodegenerative disorder and characterized by progressive locomotive defects and loss of dopaminergic neurons (DA neuron). Currently, there is no potent therapy to cure PD, and the medications merely support to control the symptoms. It is difficult to develop an effective treatment, since the PD onset mechanism of PD is still unclear. Oxidative stress is considered as a major cause of neurodegenerative diseases, and there is increasing evidence for the association between PD and oxidative stress. Therefore, antioxidant treatment may be a promising therapy for PD. Drosophila with knockdown of dUCH, a homolog of UCH-L1 which is a PD-related gene, exhibited PD-like phenotypes including progressive locomotive impairments and DA neuron degeneration. Moreover, knockdown of dUCH led to elevated level of ROS. Thus, dUCH knockdown flies can be used as a model for screening of potential antioxidants for treating PD. Previous studies demonstrated that curcumin at 1 mM and vitamin C at 0.5 mM could improve PD-like phenotypes induced by this knockdown. With the purpose of further investigating the efficiency of vitamin C in PD treatment, dUCH knockdown Drosophila model was used to examine the dose- and time-dependent effects of vitamin C on PD-like phenotypes. The results showed that although vitamin C exerted neuroprotective effects, high doses of vitamin C and long-term treatment with this antioxidant also resulted in side effects on physiology. It is suggested that dose-dependent effects of vitamin C should be considered when used for treating PD. | Adusumalli, S., Ngian, Z. K., Lin, W. Q., Benoukraf, T. and Ong, C. T. (2019). Increased intron retention is a post-transcriptional signature associated with progressive aging and Alzheimer's disease. Aging Cell: e12928. PubMed ID: 30868713
Summary: Intron retention (IR) by alternative splicing is a conserved regulatory mechanism that can affect gene expression and protein function during adult development and age-onset diseases. However, it remains unclear whether IR undergoes spatial or temporal changes during different stages of aging or neurodegeneration like Alzheimer's disease (AD). By profiling the transcriptome of Drosophila head cells at different ages, a significant increase was observed in IR events for many genes during aging. Differential IR affects distinct biological functions at different ages and occurs at several AD-associated genes in older adults. The increased nucleosome occupancy at the differentially retained introns in young animals suggests that it may regulate the level of IR during aging. Notably, an increase in the number of IR events was also observed in healthy older mouse and human brain tissues, as well as in the cerebellum and frontal cortex from independent AD cohorts. Genes with differential IR shared many common features, including shorter intron length, no perturbation in their mRNA level, and enrichment for biological functions that are associated with mRNA processing and proteostasis. The differentially retained introns identified in AD frontal cortex have higher GC content, with many of their mRNA transcripts showing an altered level of protein expression compared to control samples. Taken together, these results suggest that an increased IR is an conserved signature that is associated with aging. By affecting pathways involved in mRNA and protein homeostasis, changes of IR pattern during aging may regulate the transition from healthy to pathological state in late-onset sporadic AD. |
| Choi, H. J., Cha, S. J. and Kim, K. (2019). Glutathione transferase modulates acute ethanol-induced sedation in Drosophila neurones. Insect Mol Biol 28(2): 246-252. PubMed ID: 30347459
Summary: Heavy alcohol consumption leads to neuropathological damage and alcohol use disorder, which affects the health of people and results in a cost burden. However, the genes modulating sensitivity to ethanol remain largely unknown. This study identified a novel gene, Drosophila glutathione transferase omega 1 (GstO1), which plays a critical role in regulating sensitivity to ethanol sedation. GstO1 mutant flies showed highly increased ethanol sensitivity. Furthermore, the expression level of GstO1 regulates the behavioural response to ethanol, because decreasing and increasing GstO1 affects sedation sensitivity in a contrasting manner. In addition, the RNA interference-mediated knockdown of GstO1 expression reveals that GstO1 mediates sensitivity to ethanol sedation in neurones, including dopaminergic and serotonergic neurones. Altogether, these findings provide the first evidence for the involvement of glutathione transferase in the response to alcohol in Drosophila and provide a novel mechanistic insight into the toxicity and sensitivity of ethanol exposure. | Ranson, D. C., Ayoub, S. S., Corcoran, O. and Casalotti, S. O. (2019).. Pharmacological targeting of the GABAB receptor alters Drosophila's behavioural responses to alcohol. Addict Biol. PubMed ID: 30761704
Summary: When exposed to ethanol, Drosophila melanogaster display a variety of addiction-like behaviours similar to those observed in mammals. Sensitivity to ethanol can be quantified by measuring the time at which 50% of the flies are sedated by ethanol exposure (ST50); an increase of ST50 following multiple ethanol exposures is widely interpreted as development of tolerance to ethanol. Sensitivity and tolerance to ethanol were measured after administration of the gamma-aminobutyric acid receptor B (GABAB) agonist (SKF 97541) and antagonist (CGP 54626), when compared with flies treated with ethanol alone. Dose-dependent increases and decreases in sensitivity to ethanol were observed for both the agonist and antagonist respectively. Tolerance was recorded in the presence of GABAB drugs, but the rate of tolerance development was increased by SKF 97451 and unaltered in presence of CGP 54626. This indicates that the GABAB receptor contributes to both the sensitivity to ethanol and mechanisms by which tolerance develops. The data also reinforce the usefulness of Drosophila as a model for identifying the molecular components of addictive behaviours and for testing drugs that could potentially be used for the treatment of alcohol use disorder (AUD). |
Tuesday, March 26th - Signaling |
| Zhang, S., Guo, X., Wu, H., Sun, Y., Ma, X., Li, J., Xu, Q., Wu, C., Li, Q., Jiang, C., Li, W., Ho, M. S., Lv, Z. and Xue, L. (2019). Wingless modulates activator protein-1-mediated tumor invasion. Oncogene. PubMed ID: 30683884
Summary: Metastasis begins with a subset of local tumor cells acquiring the potential to invade into surrounding tissues, and remains to be a major obstacle for cancer treatments. More than 90% of cancer patients died from tumor metastasis, instead of primary tumor growth. The canonical Wnt/beta-catenin pathway plays essential roles in promoting tumor formation, yet its function in regulating tumor metastasis and the underlying mechanisms remain controversial. This study employed well-established Drosophila tumor models to investigate the regulating mechanism of Wingless (Wg) pathway in tumor invasion. The results showed that Wg signaling is necessary and sufficient for cell polarity disruption-induced cell migration and molecular changes reminiscent of epithelial-mesenchymal transition (EMT). Moreover, reducing Wg signaling suppressed lgl-/-RasV12-induced tumor invasion, and cooperation between Arm and Ras(V12) is sufficient to induce tumor invasion. Mechanistically, this study found that cell polarity disruption activates JNK signaling, which in turn upregulate wg expression through transcription factor activator protein-1 (AP-1). A consensus AP-1 binding site was identified located in the 2nd intron of wg and was confirmed to be essential for AP-1 induced wg transcription both in vitro and in vivo. Lastly, it was confirmed that the transcriptional activation of WNT by AP-1 is conserved in human cancer cells. These evidences reveal a positive role of Wnt/beta-catenin pathway in tumor invasion, and provide a conserved mechanism that connects JNK and Wnt signaling in regulating tumor progression. | Cho, E., Kwon, M., Jung, J., Hyun Kang, D., Jin, S., Choi, S. E., Kang, Y. and Kim, E. Y. (2019). AMP-activated protein kinase regulates circadian rhythm by affecting CLOCK in Drosophila. J Neurosci. PubMed ID: 30819799
Summary: The circadian clock organizes the physiology and behavior of organisms to their daily environmental rhythms. The central circadian timekeeping mechanism in eukaryotic cells is the transcriptional-translational feedback loop (TTFL). In the Drosophila TTFL, the transcription factors CLOCK (CLK) and CYCLE (CYC) play crucial roles in activating expression of core clock genes and clock-controlled genes. Many signaling pathways converge on the CLK/CYC complex and regulate its activity to fine-tune the cellular oscillator to environmental time cues. This study aimed to identify factors that regulate CLK by performing tandem affinity purification (TAP) combined with mass spectrometry (MS) using Drosophila S2 cells that stably express HA/FLAG-tagged CLK and V5-tagged CYC. SNF4Agamma, a homolog of mammalian AMP-activated protein kinase gamma (AMPKgamma), was identified as a factor that co-purified with HA/FLAG-tagged CLK. The AMPK holoenzyme composed of a catalytic subunit AMPKalpha and two regulatory subunits, AMPKbeta and AMPKgamma, directly phosphorylated purified CLK in vitro. Locomotor behavior analysis in Drosophila revealed that knockdown of each AMPK subunit in pacemaker neurons induced arrhythmicity and long periods. Knockdown of AMPKbeta reduced CLK levels in pacemaker neurons, and thereby reduced pre-mRNA and protein levels of CLK downstream core clock genes such as period and vrille. Finally, overexpression of CLK reversed the long-period phenotype that resulted from AMPKbeta knockdown. Thus, it is concluded that AMPK, a central regulator of cellular energy metabolism, regulates the Drosophila circadian clock by stabilizing CLK and activating CLK/CYC-dependent transcription. |
| Zacharioudaki, E., Falo Sanjuan, J. and Bray, S. (2019). Mi-2/NuRD complex protects stem cell progeny from mitogenic Notch signalling. Elife 8. PubMed ID: 30694174
Summary: To progress towards differentiation, progeny of stem cells need to extinguish expression of stem cell maintenance genes. Failures in such mechanisms can drive tumorigenesis. In Drosophila neural stem cell (NSC) lineages, excessive Notch signalling results in supernumerary NSCs causing hyperplasia. However, onset of hyperplasia is considerably delayed implying there are mechanisms that resist the mitogenic signal. Monitoring the live expression of a Notch target gene, E(spl)mgamma, revealed that normal attenuation is still initiated in the presence of excess Notch activity so that re-emergence of NSC properties occurs only in older progeny. Screening for factors responsible, this study found that depletion of Mi-2/NuRD ATP remodeling complex dramatically enhanced Notch-induced hyperplasia. Under these conditions, E(spl)mgamma was no longer extinguished in NSC progeny. It is proposed that Mi-2 is required for decommissioning stem cell enhancers in their progeny, enabling the switch towards more differentiated fates and rendering them insensitive to mitogenic factors such as Notch. | Wang, Y., Maier, A., Gehring, N. and Moussian, B. (2019). Inhibition of fatty acid desaturation impairs cuticle differentiation in Drosophila melanogaster. Arch Insect Biochem Physiol: e21535. PubMed ID: 30672604
Summary: Previous work has shown that inhibition of the activity of fatty acid desaturases (Desat) perturbs signalling of the developmental timing hormone ecdysone in the fruit fly Drosophila melanogaster. To understand the impact of this effect on cuticle differentiation, a process regulated by ecdysone, this study analysed the cuticle of D. melanogaster larvae fed with the Desat inhibitor CA10556. In these larvae, the expression of most of the key cuticle genes is normal or slightly elevated at day one of CA10556 feeding. As an exception, expression of twdlM coding for a yet uncharacterised cuticle protein is completely suppressed. The cuticle of these larvae appears to be normal at the morphological level. However, these animals are sensitive to desiccation, a trait that may be associated with reduced TwdlM amounts. At day two of CA10556 feeding, expression of most of the cuticle genes tested including twdlM is suppressed. Expression of cpr47Eb coding for a chitin-binding protein is, by contrast, highly elevated suggesting that Cpr47Eb participates at a specific compensation program. Overall, the cuticle of these larvae is thinner than the cuticle of control larvae. Taken together, lipid desaturation is necessary for a coordinated deployment of a normal cuticle differentiation program. |
| Venkatesan, A., Fan, J. Y., Bouyain, S. and Price, J. L. (2019). The circadian tau mutation in Casein Kinase 1 is part of a larger domain that can be mutated to shorten circadian period. Int J Mol Sci 20(4). PubMed ID: 30769795
Summary: Drosophila Double-time (DBT) phosphorylates the circadian protein Period (PER). The period-altering mutation tau, identified in hamster casein kinase I (CKIepsilon) and created in Drosophila DBT, has been shown to shorten the circadian period in flies, as it does in hamsters. Since CKI often phosphorylates downstream of previously phosphorylated residues and the tau amino acid binds a negatively charged ion in X-ray crystal structures, this amino acid has been suggested to contribute to a phosphate recognition site for the substrate. Alternatively, the tau amino acid may affect a nuclear localization signal (NLS) with which it interacts. This study mutated the residues that were close to or part of the phosphate recognition site or NLS. Flies expressing DBT with mutations of amino acids close to or part of either of these motifs produced a shortening of period, suggesting that a domain, including the phosphate recognition site or the NLS, can be mutated to produce the short period phenotype. Mutation of residues affecting internally placed residues produced a longer period, suggesting that a specific domain on the surface of the kinase might generate an interaction with a substrate or regulator, with short periods produced when the interaction is disrupted. | Wang, M. and Lemos, B. (2019). Ribosomal DNA harbors an evolutionarily conserved clock of biological aging. Genome Res. PubMed ID: 30765617
Summary: The ribosomal DNA (rDNA) is the most evolutionarily conserved segment of the genome and gives origin to the nucleolus, an energy intensive nuclear organelle and major hub influencing myriad molecular processes from cellular metabolism to epigenetic states of the genome. The rDNA/nucleolus has been directly and mechanistically implicated in aging and longevity in organisms as diverse as yeasts, Drosophila, and humans. The rDNA is also a significant target of DNA methylation that silences supernumerary rDNA units and regulates nucleolar activity. This study introduced an age clock built exclusively with CpG methylation within the rDNA. The ribosomal clock is sufficient to accurately estimate individual age within species, is responsive to genetic and environmental interventions that modulate life-span, and operates across species as distant as humans, mice, and dogs. Further analyses revealed a significant excess of age-associated hypermethylation in the rDNA relative to other segments of the genome, and which forms the basis of the rDNA clock. These observations identified an evolutionarily conserved marker of aging that is easily ascertained, grounded on nucleolar biology, and could serve as a universal marker to gauge individual age and response to interventions in humans as well as laboratory and wild organisms across a wide diversity of species. |
Monday, March 25th - Synapses and Vesicles |
| Russo, A., Goel, P., Brace, E. J., Buser, C., Dickman, D. and DiAntonio, A. (2019). The E3 ligase Highwire promotes synaptic transmission by targeting the NAD-synthesizing enzyme dNmnat. EMBO Rep. PubMed ID: 30692130
Summary: The ubiquitin ligase Highwire restrains synaptic growth and promotes evoked neurotransmission at NMJ synapses in Drosophila. Highwire regulates synaptic morphology by downregulating the MAP3K Wallenda, but excess Wallenda signaling does not account for the decreased presynaptic release observed in highwire mutants. Hence, Highwire likely has a second substrate that inhibits neurotransmission. Highwire targets the NAD(+) biosynthetic and axoprotective enzyme dNmnat to regulate axonal injury responses. dNmnat localizes to synapses and interacts with the active zone protein Bruchpilot, leading to a hypothesis that Highwire promotes evoked release by downregulating dNmnat. This study shows that excess dNmnat is necessary in highwire mutants and sufficient in wild-type larvae to reduce quantal content, likely via disruption of active zone ultrastructure. Catalytically active dNmnat is required to drive defects in evoked release, and depletion of a second NAD(+) synthesizing enzyme is sufficient to suppress these defects in highwire mutants, suggesting that excess NAD(+) biosynthesis is the mechanism inhibiting neurotransmission. Thus, Highwire downregulates dNmnat to promote evoked synaptic release, suggesting that Highwire balances the axoprotective and synapse-inhibitory functions of dNmnat. | Metwally, E., Zhao, G., Li, W., Wang, Q. and Zhang, Y. Q. (2019). Calcium activated Calpain specifically cleaves Glutamate receptor IIA but not IIB at the Drosophila neuromuscular junction. J Neurosci. PubMed ID: 30705102
Summary: Calpains are calcium dependent, cytosolic proteinases active at neutral pH. They do not degrade but cleave substrates at limited sites. Calpains are implicated in various pathologies such as ischemia, injuries, muscular dystrophy, and neurodegeneration. Despite so, the physiological function of calpains remains to be clearly defined. Using the neuromuscular junction of Drosophila of both sexes as a model, RNAi screening was performed, and is was uncovered that calpains negatively regulated protein levels of the glutamate receptor GluRIIA but not GluRIIB. Calpains enrich at the postsynaptic area, and the calcium-dependent activation of calpains induced cleavage of GluRIIA at Q788 of its C-terminus. Further genetic and biochemical experiments revealed that different calpains genetically and physically interact to form a protein complex. The protein complex was required for the proteinase activation to downregulate GluRIIA. These data provide a novel insight into the mechanisms by which different calpains act together as a complex to specifically control GluRIIA levels and consequently synaptic function. |
| Patron, L. A., Nagatomo, K., Eves, D. T., Imad, M., Young, K., Torvund, M., Guo, X., Rogers, G. C. and Zinsmaier, K. E. (2019). Cul4 ubiquitin ligase cofactor DCAF12 promotes neurotransmitter release and homeostatic plasticity. J Cell Biol. PubMed ID: 30670470
Summary: This study genetically characterized the synaptic role of the Drosophila homologue of human DCAF12, a putative cofactor of Cullin4 (Cul4) ubiquitin ligase complexes. Deletion of Drosophila DCAF12 impairs larval locomotion and arrests development. At larval neuromuscular junctions (NMJs), DCAF12 is expressed presynaptically in synaptic boutons, axons, and nuclei of motor neurons. Postsynaptically, DCAF12 is expressed in muscle nuclei and facilitates Cul4-dependent ubiquitination. Genetic experiments identified several mechanistically independent functions of DCAF12 at larval NMJs. First, presynaptic DCAF12 promotes evoked neurotransmitter release. Second, postsynaptic DCAF12 negatively controls the synaptic levels of the glutamate receptor subunits GluRIIA, GluRIIC, and GluRIID. The down-regulation of synaptic GluRIIA subunits by nuclear DCAF12 requires Cul4. Third, presynaptic DCAF12 is required for the expression of synaptic homeostatic potentiation. It is suggested that DCAF12 and Cul4 are critical for normal synaptic function and plasticity at larval NMJs. | Sanguanini, M. and Cattaneo, A. (2018). A continuous model of physiological prion aggregation suggests a role for Orb2 in gating long-term synaptic information. R Soc Open Sci 5(12): 180336. PubMed ID: 30662713
Summary: The regulation of mRNA translation at the level of the synapse is believed to be fundamental in memory and learning at the cellular level. The family of cytoplasmic polyadenylation element binding (CPEB) proteins emerged as an important RNA-binding protein family during development and in adult neurons. Drosophila Orb2 (homologue of mouse CPEB3 protein and of the neural isoform of Aplysia CPEB) has been found to be involved in the translation of plasticity-dependent mRNAs and has been associated with long-term memory. Orb2 protein presents two main isoforms, Orb2A and Orb2B, which form an activity-induced amyloid-like functional aggregate, thought to be the translation-inducing state of the RNA-binding protein. This study presents a first two-states continuous differential model for Orb2A-Orb2B aggregation. This model provides new working hypotheses for studying the role of prion-like CPEB proteins in long-term synaptic plasticity. Moreover, this model can be used as a first step to integrate translation- and protein aggregation-dependent phenomena in synaptic facilitation rules. |
| Boda, A., Lorincz, P., Takats, S., Csizmadia, T., Toth, S., Kovacs, A. L. and Juhasz, G. (2019). Drosophila Arl8 is a general positive regulator of lysosomal fusion events. Biochim Biophys Acta Mol Cell Res 1866(4): 533-544. PubMed ID: 30590083
Summary: The small GTPase Arl8 is known to be involved in the periphery-directed motility of lysosomes. However, the overall importance of moving these vesicles is still poorly understood. This study shows that Drosophila Arl8 is required not only for the proper distribution of lysosomes, but also for autophagosome-lysosome fusion in starved fat cells, endosome-lysosome fusion in garland nephrocytes, and developmentally programmed secretory granule degradation (crinophagy) in salivary gland cells. Moreover, proper Arl8 localization to lysosomes depends on the shared subunits of the BLOC-1 and BORC complexes, which also promote autophagy and crinophagy. In conclusion, this study demonstrates that Arl8 is responsible not only for positioning lysosomes but also acts as a general lysosomal fusion factor. | Tsai, J. W., Kostyleva, R., Chen, P. L., Rivas-Serna, I. M., Clandinin, M. T., Meinertzhagen, I. A. and Clandinin, T. R. (2019). Transcriptional feedback links lipid synthesis to synaptic vesicle pools in Drosophila photoreceptors. Neuron. PubMed ID: 30737130
Summary: Neurons can maintain stable synaptic connections across adult life. However, the signals that regulate expression of synaptic proteins in the mature brain are incompletely understood. This study describes a transcriptional feedback loop between the biosynthesis and repertoire of specific phospholipids and the synaptic vesicle pool in adult Drosophila photoreceptors. Mutations that disrupt biosynthesis of a subset of phospholipids cause degeneration of the axon terminal and loss of synaptic vesicles. Although degeneration of the axon terminal is dependent on neural activity, activation of sterol regulatory element binding protein (SREBP) is both necessary and sufficient to cause synaptic vesicle loss. These studies demonstrate that SREBP regulates synaptic vesicle levels by interacting with tetraspanins, critical organizers of membranous organelles. SREBP is an evolutionarily conserved regulator of lipid biosynthesis in non-neuronal cells; these studies reveal a surprising role for this feedback loop in maintaining synaptic vesicle pools in the adult brain. |
Friday, March 22nd - Evolution |
| Whittle, C. A. and Extavour, C. G. (2019). Contrasting patterns of molecular evolution in metazoan germ line genes. BMC Evol Biol 19(1): 53. PubMed ID: 30744572
Summary: Germ lines are the cell lineages that give rise to the sperm and eggs in animals. The germ lines first arise from primordial germ cells (PGCs) during embryogenesis: these form from either a presumed derived mode of preformed germ plasm (inheritance) or from an ancestral mechanism of inductive cell-cell signalling (induction). Numerous genes involved in germ line specification and development have been identified and functionally studied. However, little is known about the molecular evolutionary dynamics of germ line genes in metazoan model systems. This study examined the molecular evolution of germ line genes within three metazoan model systems. These include the genus Drosophila (N=34 genes, inheritance), the fellow insect Apis (N=30, induction), and their more distant relative Caenorhabditis (N=23, inheritance). Using multiple species and established phylogenies in each genus, this study reports that germ line genes exhibited marked variation in the constraint on protein sequence divergence (dN/dS) and codon usage bias (CUB) within each genus. Importantly, it was found that de novo lineage-specific inheritance (LSI) genes in Drosophila (osk, pgc) and in Caenorhabditis (pie-1, pgl-1), which are essential to germ plasm functions under the derived inheritance mode, displayed rapid protein sequence divergence relative to the other germ line genes within each respective genus. This may reflect the evolution of specialized germ plasm functions and/or low pleiotropy of LSI genes, features not shared with other germ line genes. In addition, it was observed that the relative ranking of dN/dS and of CUB between genera were each more strongly correlated between Drosophila and Caenorhabditis, from different phyla, than between Drosophila and its insect relative Apis, suggesting taxonomic differences in how germ line genes have evolved. Taken together, the present results advance understanding of the evolution of animal germ line genes within three well-known metazoan models. Further, the findings provide insights to the molecular evolution of germ line genes with respect to LSI status, pleiotropy, adaptive evolution as well as PGC-specification mode. | Maier, D. (2019). The evolution of transcriptional repressors in the Notch signaling pathway: a computational analysis. Hereditas 156: 5. PubMed ID: 30679936
Summary: The Notch signaling pathway governs the specification of different cell types in flies, nematodes and vertebrates alike. Principal components of the pathway that activate Notch target genes are highly conserved throughout the animal kingdom. Despite the impact on development and disease, repression mechanisms are less well studied. Repressors are known from arthropods and vertebrates that differ strikingly by mode of action: whereas Drosophila Hairless assembles repressor complexes with CSL transcription factors, competition between activator and repressors occurs in vertebrates (for example SHARP/MINT and KyoT2). This divergence raises questions on the evolution: Are there common ancestors throughout the animal kingdom. Available genome databases representing all animal clades were searched for homologues of Hairless, SHARP and KyoT2. The most distant species with convincing Hairless orthologs belong to Myriapoda, indicating its emergence after the Mandibulata-Chelicarata radiation about 500 million years ago. SHARP shares motifs with SPEN and SPENITO proteins, present throughout the animal kingdom. The CSL interacting domain of SHARP, however, is specific to vertebrates separated by roughly 600 million years of evolution. KyoT2 bears a C-terminal CSL interaction domain (CID), present only in placental mammals but highly diverged already in marsupials, suggesting introduction roughly 100 million years ago. Based on the LIM-domains that characterize KyoT2, homologues can be found in Drosophila melanogaster (Limpet) and Hydra vulgaris (Prickle 3 like). These lack the CID of KyoT2, however, contain a PET and additional LIM domains. Conservation of intron/exon boundaries underscores the phylogenetic relationship between KyoT2, Limpet and Prickle. Most strikingly, Limpet and Prickle proteins carry a tetra-peptide motif resembling that of several CSL interactors. Overall, KyoT2 may have evolved from prickle and Limpet to a Notch repressor in mammals. It is concluded that Notch repressors appear to be specific to either chordates or arthropods. |
| May, C. M., van den Heuvel, J., Doroszuk, A., Hoedjes, K. M., Flatt, T. and Zwaan, B. J. (2019). Adaptation to developmental diet influences the response to selection on age at reproduction in the fruit fly. J Evol Biol. PubMed ID: 30735275
Summary: Experimental evolution (EE) is a powerful tool for addressing how environmental factors influence life-history evolution. While in nature different selection pressures experienced across the lifespan shape life histories, EE studies typically apply selection pressures one at a time. This study assesses the consequences of adaptation to three different developmental diets in combination with classical selection for early or late reproduction in the fruit fly Drosophila melanogaster. Response to each selection pressure is similar to that observed when they are applied independently, but the overall magnitude of the response depends on the selection regime experienced in the other life stage. For example, adaptation to increased age at reproduction increased lifespan across all diets, however, the extent of the increase was dependent on the dietary selection regime. Similarly, adaptation to a lower calorie developmental diet led to faster development and decreased adult weight, but the magnitude of the response was dependent on the age-at-reproduction selection regime. Given that multiple selection pressures are prevalent in nature, these findings suggest that trade-offs should be considered not only among traits within an organism, but also among adaptive responses to different - sometimes conflicting - selection pressures, including across life stages. | Mossman, J. A., Ge, J. Y., Navarro, F. and Rand, D. M. (2019). Mitochondrial DNA fitness depends on nuclear genetic background in Drosophila. G3 (Bethesda). PubMed ID: 30745378
Summary: Mitochondrial DNA (mtDNA) has been one of the most extensively studied molecules in ecological, evolutionary and clinical genetics. In its early application in evolutionary genetics, mtDNA was assumed to be a selectively neutral marker conferring negligible fitness consequences for its host. However, this dogma has been overturned in recent years due to now extensive evidence for non-neutral evolutionary dynamics. Since mtDNA proteins physically interact with nuclear proteins to provide the mitochondrial machinery for aerobic ATP production, among other cell functions, co-variation of the respective genes is predicted to affect organismal fitness. To test this hypothesis, an mtDNA-nuclear DNA introgression model was used in Drosophila melanogaster to test the fitness of genotypes in perturbation-reperturbation population cages and in a non-competitive assay for female fecundity. Genotypes consisted of both conspecific and heterospecific mtDNA-nDNA constructs, with either D. melanogaster or D. simulans mtDNAs on two alternative D. melanogaster nuclear backgrounds, to investigate mitonuclear genetic interactions (G x G effects). Considerable variation was found between nuclear genetic backgrounds on the selection of mtDNA haplotypes. In addition, there was variation in the selection on mtDNAs pre- and post- reperturbation, demonstrating overall poor repeatability of selection. There was a strong influence of nuclear background on non-competitive fecundity across all the mtDNA species types. |
| Orengo, D. J., Puerma, E. and Aguade, M. (2019). The molecular characterization of fixed inversions breakpoints unveils the ancestral character of the Drosophila guanche chromosomal arrangements. Sci Rep 9(1): 1706. PubMed ID: 30737415
Summary: Cytological studies revealed that the number of chromosomes and their organization varies across species. The increasing availability of whole genome sequences of multiple species across specific phylogenies has confirmed and greatly extended these cytological observations. In the Drosophila genus, the ancestral karyotype consists of five rod-like acrocentric chromosomes (Muller elements A to E) and one dot-like chromosome (element F), each exhibiting a generally conserved gene content. Chromosomal fusions and paracentric inversions are thus the major contributors, respectively, to chromosome number variation among species and to gene order variation within chromosomal element. The subobscura cluster of Drosophila consists in three species that retain the genus ancestral karyotype and differ by a reduced number of fixed inversions. This study used cytological information and the D. guanche genome sequence to identify and molecularly characterize the breakpoints of inversions that became fixed since the D. guanche-D. subobscura split. The results have led to a proposal of a modified version of the D. guanche cytological map of its X chromosome, and to establish that (i) most inversions became fixed in the D. subobscura lineage and (ii) the order in which the four X chromosome overlapping inversions occurred and became fixed. | Shaw, K. H., Johnson, T. K., Anderson, A., de Bruyne, M. and Warr, C. G. (2019). Molecular and functional evolution at the Odorant receptor Or22 locus in Drosophila melanogaster. Mol Biol Evol. PubMed ID: 30768139
Summary: Insect odorant receptor (Or) genes determine the responses of sensory neurons that mediate critical behaviours. The Drosophila melanogaster Or22 locus represents an interesting example of molecular evolution, with high levels of sequence divergence and copy number variation between D. melanogaster and other Drosophila species, and a corresponding high level of variability in the responses of the neuron it controls, ab3A. However, the link between Or22 molecular and functional diversity has not been established. This study shows that several naturally occurring Or22 variants generate major shifts in neuronal response properties. The molecular changes were determined that underpin these response shifts, one of which represents a chimaeric gene variant previously suggested to be under natural selection. In addition it was shown that several alternative molecular genetic mechanisms have evolved for ensuring that where there is more than one gene copy at this locus, only one functional receptor is generated. These data thus provide a causal link between the striking levels of phenotypic neuronal response variation found in natural populations of D. melanogaster and genetic variation at the Or22 locus. Since neuronal responses govern animal behaviour, it is predicted that Or22 may be a key player in underlying one or more olfactory-driven behaviours of significant adaptive importance. |
Thursday, March 21st - Adult neural development and function |
| Wei, J. Y., Chung, P. C., Chu, S. Y. and Yu, H. H. (2019). FOXO regulates cell fate specification of Drosophila ventral olfactory projection neurons. J Neurogenet: 1-8. PubMed ID: 30686090
Summary: Diverse types of neurons must be specified in the developing brain to form the functional neural circuits that are necessary for the execution of daily tasks. This study describes the participation of Forkhead box class O (FOXO) in cell fate specification of a small subset of Drosophila ventral olfactory projection neurons (vPNs). Using the two-color labeling system, twin-spot MARCM, this study determined the temporal birth order of each vPN type, and this characterization served as a foundation to investigate regulators of cell fate specification. Flies deficient for chinmo, a known temporal cell fate regulator, exhibited a partial loss of vPNs, suggesting that the gene plays a complex role in specifying vPN cell fate and is not the only regulator of this process. Interestingly, loss of foxo function resulted in the precocious appearance of late-born vPNs in place of early-born vPNs, whereas overexpression of constitutively active FOXO caused late-born vPNs to take on a morphology reminiscent of earlier born vPNs. Taken together, these data suggest that FOXO temporally regulates vPN cell fate specification. The comprehensive identification of molecules that regulate neuronal fate specification promises to provide a better understanding of the mechanisms governing the formation of functional brain tissue. | Zhou, M., Chen, N., Tian, J., Zeng, J., Zhang, Y., Zhang, X., Guo, J., Sun, J., Li, Y., Guo, A. and Li, Y. (2019). Suppression of GABAergic neurons through D2-like receptor secures efficient conditioning in Drosophila aversive olfactory learning. Proc Natl Acad Sci U S A 116(11): 5118-5125. PubMed ID: 30796183
Summary: In Drosophila, the GABAergic anterior paired lateral (APL) neurons mediate a negative feedback essential for odor discrimination; however, their activity is suppressed by learning via unknown mechanisms. In aversive olfactory learning, a group of dopaminergic (DA) neurons is activated on electric shock (ES) and modulates the Kenyon cells (KCs) in the mushroom body, the center of olfactory learning. This work finds that the same group of DA neurons also form functional synaptic connections with the APL neurons, thereby emitting a suppressive signal to the latter through Drosophila dopamine 2-like receptor (DD2R). Knockdown of either DD2R or its downstream molecules in the APL neurons results in impaired olfactory learning at the behavioral level. Results obtained from in vivo functional imaging experiments indicate that this DD2R-dependent DA-to-APL suppression occurs during odor-ES conditioning and discharges the GABAergic inhibition on the KCs specific to the conditioned odor. Moreover, the decrease in odor response of the APL neurons persists to the postconditioning phase, and this change is also absent in DD2R knockdown flies. Taken together, these findings show that DA-to-GABA suppression is essential for restraining the GABAergic inhibition during conditioning, as well as for inducing synaptic modification in this learning circuit. |
| Sen, S. Q., Chanchani, S., Southall, T. D. and Doe, C. Q. (2019). Neuroblast-specific open chromatin allows the temporal transcription factor, Hunchback, to bind neuroblast-specific loci. Elife 8. PubMed ID: 30694180
Summary: Spatial and temporal cues are required to specify neuronal diversity, but how these cues are integrated in neural progenitors remains unknown. Drosophila progenitors (neuroblasts) are a good model: they are individually identifiable with relevant spatial and temporal transcription factors known. This study tested whether spatial/temporal factors act independently or sequentially in neuroblasts. Targeted DamID was used to identify genomic binding sites of the Hunchback temporal factor in two neuroblasts (NB5-6 and NB7-4) that make different progeny. Hunchback targets were different in each neuroblast, ruling out the independent specification model. Moreover, each neuroblast had distinct open chromatin domains, which correlated with differential Hb-bound loci in each neuroblast. Importantly, the Gsb/Pax3 spatial factor, expressed in NB5-6 but not NB7-4, had genomic binding sites correlated with open chromatin in NB5-6, but not NB7-4. These data support a model in which early-acting spatial factors like Gsb establish neuroblast-specific open chromatin domains, leading to neuroblast-specific temporal factor binding and the production of different neurons in each neuroblast lineage. | Sullivan, L. F., Warren, T. L. and Doe, C. Q. (2019). Temporal identity establishes columnar neuron morphology, connectivity, and function in a Drosophila navigation circuit. Elife 8. PubMed ID: 30706848
Summary: The insect central complex (CX) is a conserved brain region containing 60+ neuronal subtypes, several of which contribute to navigation. It is not known how CX neuronal diversity is generated or how developmental origin of subtypes relates to function. This study mapped the developmental origin of four key CX subtypes and found that neurons with similar origin have similar axon/dendrite targeting. Moreover, the temporal transcription factor (TTF) Eyeless/Pax6 was found to regulate the development of two recurrently-connected CX subtypes: Eyeless loss simultaneously produces ectopic P-EN neurons with normal axon/dendrite projections, and reduces the number of E-PG neurons. Furthermore, transient loss of Eyeless during development impairs adult flies' capacity to perform celestial navigation. It is concluded that neurons with similar developmental origin have similar connectivity, that Eyeless maintains equal E-PG and P-EN neuron number, and that Eyeless is required for the development of circuits that control adult navigation. |
| Spurrier, J., Shukla, A. K., Buckley, T., Smith-Trunova, S., Kuzina, I., Gu, Q. and Giniger, E. (2019). Expression of a fragment of Ankyrin 2 disrupts the structure of the axon initial segment and causes axonal degeneration in Drosophila. Mol Neurobiol. PubMed ID: 30666562
Summary: Neurodegenerative stimuli are often associated with perturbation of the axon initial segment (AIS), but it remains unclear whether AIS disruption is causative for neurodegeneration or is a downstream step in disease progression. This study demonstrates that either of two separate, genetically parallel pathways that disrupt the AIS induce axonal degeneration and loss of neurons in the central brain of Drosophila. Expression of a portion of the C-terminal tail of the Ank2-L isoform of Ankyrin severely shortens the AIS in Drosophila mushroom body (MB) neurons, and this shortening occurs through a mechanism that is genetically separate from the previously described Cdk5alpha-dependent pathway of AIS regulation. Further, either manipulation triggers morphological degeneration of MB axons and is accompanied by neuron loss. Taken together, these results are consistent with the hypothesis that disruption of the AIS is causally related to degeneration of fly central brain neurons, and it is suggested that similar mechanisms may contribute to neurodegeneration in mammals. | Venkatasubramanian, L., Guo, Z., Xu, S., Tan, L., Xiao, Q., Nagarkar-Jaiswal, S. and Mann, R. S. (2019). Stereotyped terminal axon branching of leg motor neurons mediated by IgSF proteins DIP-alpha and Dpr10. Elife 8. PubMed ID: 30714901
Summary: For animals to perform coordinated movements requires the precise organization of neural circuits controlling motor function. Motor neurons (MNs), key components of these circuits, project their axons from the central nervous system and form precise terminal branching patterns at specific muscles. Focusing on the Drosophila leg neuromuscular system this study shows that the stereotyped terminal branching of a subset of MNs is mediated by interacting transmembrane Ig superfamily proteins DIP-alpha and Dpr10, present in MNs and target muscles, respectively. The DIP-alpha/Dpr10 interaction is needed only after MN axons reach the vicinity of their muscle targets. Live imaging suggests that precise terminal branching patterns are gradually established by DIP-alpha/Dpr10-dependent interactions between fine axon filopodia and developing muscles. Further, different leg MNs depend on the DIP-alpha and Dpr10 interaction to varying degrees that correlate with the morphological complexity of the MNs and their muscle targets. |
Wednesday, March 20th - Signaling |
| Sun, Y., Zhang, D., Guo, X., Li, W., Li, C., Luo, J., Zhou, M. and Xue, L. (2019). MKK3 modulates JNK-dependent cell migration and invasion. Cell Death Dis 10(3): 149. PubMed ID: 30770795
Summary: The c-Jun N-terminal kinase (JNK) pathway plays essential roles in regulating a variety of physiological processes including cell migration and invasion. To identify critical factors that regulate JNK-dependent cell migration, a genetic screen was carried out in Drosophila based on the loss-of-cell polarity-triggered cell migration in the wing epithelia, and MKK3 licorne (lic) was identified as an essential regulator of JNK-mediated cell migration and invasion. Loss of lic suppressed ptc > scrib-IR or ptc > Egr triggered cell migration in the wing epithelia, and Ras(v12)/lgl(-/-) induced tumor invasion in the eye discs. In addition, ectopic expression of Lic is sufficient to induce JNK-mediated but p38-independent cell migration, and cooperates with oncogenic Ras to promote tumor invasion. Consistently, Lic is able to activate JNK signaling by phosphorylating JNK, which up-regulates the matrix metalloproteinase MMP1 and integrin, characteristics of epithelial-mesenchymal transition (EMT). Moreover, lic is required for physiological JNK-mediate cell migration in thorax development. Finally, expression of human MKK3 in Drosophila is able to initiate JNK-mediated cell migration, cooperates with oncogenic Ras to trigger tumor invasion, and rescue loss-of-lic induced thorax closure defect. As previous studies suggest that MKK3 specifically phosphorylates and activates p38MAPK, these data provide the first in vivo evidence that MKK3 regulates JNK-dependent cell migration and invasion, a process evolutionarily conserved from flies to human. | Santabarbara-Ruiz, P., Esteban-Collado, J., Perez, L., Viola, G., Abril, J. F., Milan, M., Corominas, M. and Serras, F. (2019). Ask1 and Akt act synergistically to promote ROS-dependent regeneration in Drosophila. PLoS Genet 15(1): e1007926. PubMed ID: 30677014
Summary: How cells communicate to initiate a regenerative response after damage has captivated scientists during the last few decades. It is known that one of the main signals emanating from injured cells is the Reactive Oxygen Species (ROS), which propagate to the surrounding tissue to trigger the replacement of the missing cells. However, the link between ROS production and the activation of regenerative signaling pathways is not yet fully understood. This study describes the non-autonomous ROS sensing mechanism by which living cells launch their regenerative program. To this aim, Drosophila imaginal discs were used as a model system due to their well-characterized regenerative ability after injury or cell death. Cell death was genetically-induced and it was found that the Apoptosis signal-regulating kinase 1 (Ask1) is essential for regenerative growth. Ask1 senses ROS both in dying and living cells, but its activation is selectively attenuated in living cells by Akt1, the core kinase component of the insulin/insulin-like growth factor pathway. Akt1 phosphorylates Ask1 in a secondary site outside the kinase domain, which attenuates its activity. This modulation of Ask1 activity results in moderate levels of JNK signaling in the living tissue, as well as in activation of p38 signaling, both pathways required to turn on the regenerative response. These findings demonstrate a non-autonomous activation of a ROS sensing mechanism by Ask1 and Akt1 to replace the missing tissue after damage. Collectively, these results provide the basis for understanding the molecular mechanism of communication between dying and living cells that triggers regeneration. |
| Misra, J. R. and Irvine, K. D. (2019). Early girl is a novel component of the Fat signaling pathway. PLoS Genet 15(1): e1007955. PubMed ID: 30699121
Summary: The Drosophila protocadherins Dachsous and Fat regulate growth and tissue polarity by modulating the levels, membrane localization and polarity of the atypical myosin Dachs. Localization to the apical junctional membrane is critical for Dachs function, and the adapter protein Vamana/Dlish and palmitoyl transferase Approximated are required for Dachs membrane localization. However, how Dachs levels are regulated is poorly understood. This study identified the early girl gene as playing an essential role in Fat signaling by limiting the levels of Dachs protein. early girl mutants display overgrowth of the wings and reduced cross vein spacing, hallmark features of mutations affecting Fat signaling. Genetic experiments reveal that it functions in parallel with Fat to regulate Dachs. early girl encodes an E3 ubiquitin ligase, physically interacts with Dachs, and regulates its protein stability. Concomitant loss of early girl and approximated results in accumulation of Dachs and Vamana in cytoplasmic punctae, suggesting that it also regulates their trafficking to the apical membrane. These findings establish a crucial role for early girl in Fat signaling, involving regulation of Dachs and Vamana, two key downstream effectors of this pathway. | Neal, S. J., Dolezal, D., Jusic, N. and Pignoni, F. (2019). Drosophila ML-DmD17-c3 cells respond robustly to Dpp and exhibit complex transcriptional feedback on BMP signaling components. BMC Dev Biol 19(1): 1. PubMed ID: 30669963
Summary: BMP signaling is involved in myriad metazoan developmental processes, and study of this pathway in Drosophila has contributed greatly to understanding of its molecular and genetic mechanisms. These studies have benefited not only from Drosophila's advanced genetic tools, but from complimentary in vitro culture systems. However, the commonly-used S2 cell line is not intrinsically sensitive to the major BMP ligand Dpp and must therefore be augmented with exogenous pathway components for most experiments. This study identified and characterized the responses of Drosophila ML-DmD17-c3 cells, which are sensitive to Dpp stimulation and exhibit characteristic regulation of BMP target genes including Dad and brk. Dpp signaling in ML-DmD17-c3 cells is primarily mediated by the receptors Put and Tkv, with additional contributions from Wit and Sax. Furthermore, this study reports complex regulatory feedback on core pathway genes in this system. It is concluded that native ML-DmD17-c3 cells exhibit robust transcriptional responses to BMP pathway induction. It is proposed that ML-DmD17-c3 cells are well-suited for future BMP pathway analyses. |
| Nan, Z., Yang, W., Lyu, J., Wang, F., Deng, Q., Xi, Y., Yang, X. and Ge, W. (2019). Drosophila Hcf regulates Hippo signaling pathway via association with the histone H3K4 methyltransferase Trr. Biochem J. PubMed ID: 30733258
Summary: Control of organ size is a fundamental aspect in biology and plays important roles in development. The Hippo pathway is a conserved signaling cascade that controls tissue and organ size through the regulation of cell proliferation and apoptosis. This study reports on the roles of Hcf, the Drosophila homolog of Host cell factor 1, in regulating the Hippo signaling pathway. Loss of Hcf function causes tissue undergrowth and the downregulation of Hippo target gene expression. Genetic analysis reveals that Hcf is required for Hippo pathway-mediated overgrowth. Mechanistically, Hcf associates with the histone H3 lysine-4 methyltransferase Trithorax-related (Trr) to maintain H3K4 mono- and tri-methylation. Thus, it is concluded that Hcf positively regulates Hippo pathway activity through forming a complex with Trr and controlling H3K4 methylation. | Strassburger, K., Kang, E. and Teleman, A. A. (2019). Drosophila ZDHHC8 palmitoylates scribble and Ras64B and controls growth and viability. PLoS One 14(2): e0198149. PubMed ID: 30735487
Summary: Palmitoylation is an important posttranslational modification regulating diverse cellular functions. Consequently, aberrant palmitoylation can lead to diseases such as neuronal disorders or cancer. In humans there are roughly one hundred times more palmitoylated proteins than enzymes catalyzing palmitoylation (palmitoyltransferases). Therefore, it is an important challenge to establish the links between palmitoyltransferases and their targets. From publicly available data, this study found that expression of human ZDHHC8 correlates significantly with cancer survival. To elucidate the organismal function of ZDHHC8, the Drosophila ortholog of hZDHHC8, CG34449/dZDHHC8 was studied. Knockdown of dZDHHC8 causes tissue overgrowth while dZDHHC8 mutants are larval lethal. A list is provided of 159 palmitoylated proteins in Drosophila and data is presented suggesting that Scribble and Ras64B are targets of dZDHHC8. |
Tuesday, March 19th - Junctions and Cytoskeleton |
| Strutt, H., Langton, P. F., Pearson, N., McMillan, K. J., Strutt, D. and Cullen, P. J. (2019). Retromer controls planar polarity protein levels and asymmetric localization at intercellular iunctions. Curr Biol 29(3): 484-491. PubMed ID: 30661800
Summary: The coordinated polarization of cells in the plane of a tissue, termed planar polarity, is a characteristic feature of epithelial tissues. In the fly wing, trichome positioning is dependent on the core planar polarity proteins adopting asymmetric subcellular localizations at apical junctions, where they form intercellular complexes that link neighboring cells. Specifically, the seven-pass transmembrane protein Frizzled and the cytoplasmic proteins Dishevelled and Diego localize to distal cell ends, the four-pass transmembrane protein Strabismus and the cytoplasmic protein Prickle localize proximally, and the seven-pass transmembrane spanning atypical cadherin Flamingo localizes both proximally and distally. To establish asymmetry, these core proteins are sorted from an initially uniform distribution; however, the mechanisms underlying this polarized trafficking remain poorly understood. This study describes the identification of retromer, a master controller of endosomal recycling, as a key component regulating core planar polarity protein localization in Drosophila. Through generation of mutants, it was verified that loss of the retromer-associated Snx27 cargo adaptor, but notably not components of the Wash complex, reduces junctional levels of the core proteins Flamingo and Strabismus in the developing wing. It was established that Snx27 directly associates with Flamingo via its C-terminal PDZ binding motif, and Snx27 was shown to be essential for normal Flamingo trafficking. it is concluded that Wash-independent retromer function and the Snx27 cargo adaptor are important components in the endosomal recycling of Flamingo and Strabismus back to the plasma membrane and thus contribute to the establishment and maintenance of planar polarization. | Raza, Q., Choi, J. Y., Li, Y., O'Dowd, R. M., Watkins, S. C., Chikina, M., Hong, Y., Clark, N. L. and Kwiatkowski, A. V. (2019). Evolutionary rate covariation analysis of E-cadherin identifies Raskol as a regulator of cell adhesion and actin dynamics in Drosophila. PLoS Genet 15(2): e1007720. PubMed ID: 30763317
Summary: The adherens junction couples the actin cytoskeletons of neighboring cells to provide the foundation for multicellular organization. The core of the adherens junction is the cadherin-catenin complex that arose early in the evolution of multicellularity to link actin to intercellular adhesions. Over time, evolutionary pressures have shaped the signaling and mechanical functions of the adherens junction to meet specific developmental and physiological demands. Evolutionary rate covariation (ERC) identifies proteins with correlated fluctuations in evolutionary rate that can reflect shared selective pressures and functions. This study used ERC to identify proteins with evolutionary histories similar to the Drosophila E-cadherin (DE-cad) ortholog. Core adherens junction components alpha-catenin and p120-catenin displayed positive ERC correlations with DE-cad, indicating that they evolved under similar selective pressures during evolution between Drosophila species. Further analysis of the DE-cad ERC profile revealed a collection of proteins not previously associated with DE-cad function or cadherin-mediated adhesion. The function of a subset of ERC-identified candidates was analyzed by RNAi during border cell (BC) migration, and novel genes were identified that function to regulate DE-cad. Among these, the gene CG42684, which encodes a putative GTPase activating protein (GAP), was found to regulate BC migration and adhesion. CG42684 was name raskol ("to split" in Russian) and it was shown to regulates DE-cad levels and actin protrusions in BCs. It is proposed that Raskol functions with DE-cad to restrict Ras/Rho signaling and help guide BC migration. These results demonstrate that a coordinated selective pressure has shaped the adherens junction and this can be leveraged to identify novel components of the complexes and signaling pathways that regulate cadherin-mediated adhesion. |
| Jouette, J., Guichet, A. and Claret, S. B. (2019). Dynein-mediated transport and membrane trafficking control PAR3 polarised distribution. Elife 8. PubMed ID: 30672465
Summary: The scaffold protein PAR3 and the kinase PAR1 are essential proteins that control cell polarity. Their precise opposite localisations define plasma membrane domains with specific functions. PAR3 and PAR1 are mutually inhibited by direct or indirect phosphorylations, but their fates once phosphorylated are poorly known. Through precise spatiotemporal quantification of PAR3 localisation in the Drosophila oocyte, this study identified several mechanisms responsible for its anterior cortex accumulation and its posterior exclusion. PAR3 posterior plasma membrane exclusion depends on PAR1 and an endocytic mechanism relying on RAB5 and PI(4,5)P2. In a second phase, microtubules and the dynein motor, in connection with vesicular trafficking involving RAB11 and IKK-related kinase, IKKepsilon, are required for PAR3 transport towards the anterior cortex. Altogether, these results point to a connection between membrane trafficking and dynein-mediated transport to sustain PAR3 asymmetry. | Petri, J., Syed, M. H., Rey, S. and Klambt, C. (2019). Non-cell-autonomous function of the GPI-anchored protein Undicht during septate junction assembly. Cell Rep 26(6): 1641-1653. PubMed ID: 30726744
Summary: Occluding cell-cell junctions are pivotal during the development of many organs. One example is septate junction (SJ) strands, which are found in vertebrates and invertebrates. Although several proteins have been identified that are responsible for septate junction formation in Drosophila, it is presently unclear how these structures are formed or how they are positioned in a coordinated manner between two neighboring cells and within the tissue. This study identified a GPI-anchored protein called Undicht required for septate junction formation. Clonal analysis and rescue experiments show that Undicht acts in a non-cell-autonomous manner. It can be released from the plasma membrane by the proteolytic activity of two related ADAM10-like proteases, Kuzbanian and Kuzbanian-like. It is proposed that juxtacrine function of Undicht coordinates the formation of septate junction strands on two directly neighboring cells, whereas paracrine activity of Undicht controls the formation of occluding junctions within a tissue. |
| Xu, C., Tang, H. W., Hung, R. J., Hu, Y., Ni, X., Housden, B. E. and Perrimon, N. (2019). The septate junction protein Tsp2A restricts intestinal stem cell activity via endocytic regulation of aPKC and Hippo signaling. Cell Rep 26(3): 670-688.e676. PubMed ID: 30650359
Summary: Hippo signaling and the activity of its transcriptional coactivator, Yorkie (Yki), are conserved and crucial regulators of tissue homeostasis. In the Drosophila midgut, after tissue damage, Yki activity increases to stimulate stem cell proliferation, but how Yki activity is turned off once the tissue is repaired is unknown. From an RNAi screen, the septate junction (SJ) protein tetraspanin 2A (Tsp2A) was identified as a tumor suppressor. Tsp2A undergoes internalization to facilitate the endocytic degradation of atypical protein kinase C (aPKC), a negative regulator of Hippo signaling. In the Drosophila midgut epithelium, adherens junctions (AJs) and SJs are prominent in intestinal stem cells or enteroblasts (ISCs or EBs) and enterocytes (ECs), respectively. When ISCs differentiate toward ECs, Tsp2A is produced, participates in SJ assembly, and turns off aPKC and Yki-JAK-Stat activity. Altogether, this study uncovers a mechanism allowing the midgut to restore Hippo signaling and restrict proliferation once tissue repair is accomplished (Xu, 2019a). | Davidson, A. J., Millard, T. H., Evans, I. R. and Wood, W. (2019). Ena orchestrates remodelling within the actin cytoskeleton to drive robust Drosophila macrophage chemotaxis. J Cell Sci. PubMed ID: 30718364
Summary: The actin cytoskeleton is the engine that powers the inflammatory chemotaxis of immune cells to sites of tissue damage or infection. This study combined genetics with live, in vivo imaging to investigate how cytoskeletal rearrangements drive macrophage recruitment to wounds in Drosophila. The actin-regulatory protein Ena is a master regulator of lamellipodial dynamics in migrating macrophages where it remodels the cytoskeleton to form linear filaments that can then be bundled together by the cross-linker Fascin. In contrast, the formin Dia generates rare, probing filopods for specialised functions that are not required for migration. Ena's role in lamellipodial bundling is so fundamental that its over-expression increases bundling even in the absence of Fascin by marshalling the remaining cross-linking proteins to compensate. This reorganisation of the lamellipod generates cytoskeletal struts that push against the membrane to drive leading edge advancement and boost cell speed. Thus, Ena-mediated remodeling extracts the most from the cytoskeleton to power robust macrophage chemotaxis during their inflammatory recruitment to wounds. |
Monday, March 18th - Oogenesis and Spermatogenesis |
| Gartner, S. M. K., Hundertmark, T., Nolte, H., Theofel, I., Eren-Ghiani, Z., Tetzner, C., Duchow, T. B., Rathke, C., Kruger, M. and Renkawitz-Pohl, R. (2019). Stage-specific testes proteomics of Drosophila melanogaster identifies essential proteins for male fertility. Eur J Cell Biol. PubMed ID: 30679029
Summary: Spermiogenesis in Drosophila melanogaster is a highly conserved process and essential for male fertility. In this haploid phase of spermatogenesis, motile sperm are assembled from round cells, and flagella and needle-shaped nuclei with highly compacted genomes are formed. As transcription takes place mainly in spermatocytes and transcripts relevant for post-meiotic sperm development are translationally repressed for days, this study comparatively analysed the proteome of larval testes (only germ cell stages before meiotic divisions), testes of 1-2-day-old pupae (germ cell stages before meiotic divisions, meiotic and early spermatid stages) and adult flies (germ cell stages before meiotic divisions, meiotic and early spermatid stages, late spermatids and sperm). 6,171 proteins were identified; 61 proteins were detected solely in one stage and are thus enriched, namely 34 in larval testes, 77 in pupal testes and 214 in adult testes. To substantiate mass spectrometric data, the stage-specific synthesis and importance for male fertility were analyzed of a number of uncharacterized proteins. For example, Mst84B (gene CG1988), a very basic cysteine- and lysine-rich nuclear protein and was present in the transition phase from a histone-based to a protamine-based chromatin structure. CG6332 encodes d-Theg, which is related to the mouse tHEG and human THEG proteins. Mutants of d-Theg were sterile due to the lack of sperm in the seminal vesicles. This catalogue of proteins of the different stages of testis development in D. melanogaster will pave the road for future analyses of spermatogenesis. | Gruntenko, N. E., Karpova, E. K., Adonyeva, N. V., Andreenkova, O. V., Burdina, E. V., Ilinsky, Y. Y., Bykov, R. A., Menshanov, P. N. and Rauschenbach, I. Y. (2019). Drosophila female fertility and juvenile hormone metabolism depends on the type of Wolbachia infection. J Exp Biol. PubMed ID: 30679245
Summary: Maternally inherited intracellular bacteria Wolbachia cause both parasitic and mutualistic effects on their numerous insect hosts that include manipulating the host reproductive system in order to increase the bacteria spreading in a host population, and increasing the host fitness. This study demonstrates that the type of Wolbachia infection determines the effect on Drosophila melanogaster egg production as a proxy for fecundity and metabolism of juvenile hormone (JH), which acts as gonadotropin in adult insects. This study used six D. melanogaster lineages carrying the nuclear background of interbred Bi90 lineage and cytoplasmic backgrounds with Wolbachia of different genotype variants or without it. wMelCS genotype of Wolbachia decreases the egg production in the infected D. melanogaster females in the beginning of oviposion and increases it later (since the sixth day after eclosion), wMelPop Wolbachia strain causes the opposite effect, while wMel, wMel2 and wMel4 genotypes of Wolbachia do not show any effect on these traits compared to uninfected Bi90 D. melanogaster females. The intensity of JH catabolism negatively correlates with the fecundity level in the flies carrying both wMelCS and wMelPop Wolbachia The JH catabolism in females infected with genotypes of wMel group does not differ from that in uninfected females. The effects of wMelCS and wMelPop infection on egg production can be leveled by the modulation of JH titre (via precocene/JH treatment of the flies). Thus, at least one of the mechanisms, promoting the effect of Wolbachia on D. melanogaster female fecundity, is mediated by JH. |
| Yu, J., Chen, B., Zheng, B., Qiao, C., Chen, X., Yan, Y., Luan, X., Xie, B., Liu, J., Shen, C., He, Z., Hu, X., Liu, M., Li, H., Shao, Q. and Fang, J. (2019). ATP synthase is required for male fertility and germ cell maturation in Drosophila testes. Mol Med Rep. PubMed ID: 30628672
Summary: Germ cell maturation is essential for spermatogenesis and testis homeostasis. ATP synthase serves significant roles in energy storage in germ cell survival and is catalyzed by alterations in the mitochondrial membrane proton concentration. The intrinsic cellular mechanisms governing stem cell maturation remain largely unknown. In the present study, in vivo RNA interference (RNAi) screening of major ATP synthase subunits was performed, and the function of ATP synthase for male fertility and spermatogenesis in Drosophila was explored. A UAS/Gal4 transcription factor system was used to knock down gene expression in specific cell types, and immunofluorescence staining was conducted to assess the roles of ATP synthase subunits in Drosophila testes. It was knockdown of ATP synthase was shown to result in male infertility and abnormal spermatogenesis in Drosophila testes. In addition, knockdown of the ATP synthase beta subunit in germ cells resulted in defects in male infertility and germ cell maturation, while the hub and cyst cell populations were maintained. Other major ATP synthase subunits were also examined and similar phenotypes in Drosophila testes were identified. Taken together, the data from the present study revealed that ATP synthase serves important roles for male fertility during spermatogenesis by regulating germ cell maturation in Drosophila testes. | Sreejith, P., Jang, W., To, V., Hun Jo, Y., Biteau, B. and Kim, C. (2019). Lin28 is a critical factor in the function and aging of Drosophila testis stem cell niche. Aging (Albany NY). PubMed ID: 30713156
Summary: Age-related decline in stem cell function is observed in many tissues from invertebrates to humans. While cell intrinsic alterations impair stem cells, aging of the stem cell niche also significantly contributes to the loss of tissue homeostasis associated with reduced regenerative capacity. Hub cells, which constitute the stem cell niche in the Drosophila testis, exhibit age-associated decline in number and activities, yet underlying mechanisms are not fully understood. This study shows that Lin28, a highly conserved RNA binding protein, is expressed in hub cells and its expression dramatically declines in old testis. lin28 mutant testes exhibit hub cell loss and defective hub architecture, recapitulating the normal aging process. Importantly, maintained expression of Lin28 prolongs hub integrity and function in aged testes, suggesting that Lin28 decline is a driver of hub cell aging. Mechanistically, the level of unpaired (upd), a stem cell self-renewal factor, is reduced in lin28 mutant testis and Lin28 protein directly binds and stabilizes upd transcripts, in a let-7 independent manner. Altogether, these results suggest that Lin28 acts to protect upd transcripts in hub cells, and reduction of Lin28 in old testis leads to decreased upd levels, hub cell aging and loss of the stem cell niche. |
| Gao, Y., Mao, Y., Xu, R. G., Zhu, R., Zhang, M., Sun, J., Shen, D., Peng, P., Xie, T. and Ni, J. Q. (2019). Defining gene networks controlling the maintenance and function of the differentiation niche by an in vivo systematic RNAi screen. J Genet Genomics. PubMed ID: 30745214
Summary: In the Drosophila ovary, escort cells (ECs) extrinsically control germline stem cell (GSC) maintenance and progeny differentiation. However, the underlying mechanisms remain poorly understood. This study identified 173 EC genes for their roles in controlling GSC maintenance and progeny differentiation by using an in vivo systematic RNAi approach. Of the identified genes, 10 and 163 are required in ECs to promote GSC maintenance and progeny differentiation, respectively. The genes required for progeny differentiation fall into different functional categories, including transcription, mRNA splicing, protein degradation, signal transduction and cytoskeleton regulation. In addition, the GSC progeny differentiation defects caused by defective ECs are often associated with BMP signaling elevation, indicating that preventing BMP signaling is a general functional feature of the differentiation niche. Lastly, exon junction complex (EJC) components, which are essential for mRNA splicing, are required in ECs to promote GSC progeny differentiation by maintaining ECs and preventing BMP signaling. Therefore, this study has identified the major regulators of the differentiation niche, which provides important insights into how stem cell progeny differentiation is extrinsically controlled. | Lenhart, K. F., Capozzoli, B., Warrick, G. S. D. and DiNardo, S. (2018). Diminished Jak/STAT signaling causes early-onset aging defects in stem cell cytokinesis. Curr Biol. PubMed ID: 30612906 Summary: Tissue renewal becomes compromised with age. Although defects in niche and stem cell behavior have been implicated in promoting age-related decline, the causes of early-onset aging defects are unknown. This study has identified an early consequence of aging in germline stem cells (GSCs) in the Drosophila testis. Aging disrupts the unique program of GSC cytokinesis, with GSCs failing to abscise from their daughter cells. Abscission failure significantly disrupts both self-renewal and the generation of differentiating germ cells. Extensive live imaging and genetic analyses show that abscission failure is due to inappropriate retention of F-actin at the intercellular bridges between GSC-daughter cells. Furthermore, F-actin is regulated by the Jak/STAT pathway-increasing or decreasing pathway activity can rescue or exacerbate the age-induced abscission defect, respectively. Even subtle decreases to STAT activity are sufficient to precociously age young GSCs and induce abscission failure. Thus, this work has identified the earliest age-related defect in GSCs and has revealed a unique role for an established niche signaling pathway in controlling stem cell cytokinesis and in regulating stem cell behavior with age. |
Friday, March 15th |
| Gartner, S. M. K., Hundertmark, T., Nolte, H., Theofel, I., Eren-Ghiani, Z., Tetzner, C., Duchow, T. B., Rathke, C., Kruger, M. and Renkawitz-Pohl, R. (2019). Stage-specific testes proteomics of Drosophila melanogaster identifies essential proteins for male fertility. Eur J Cell Biol. PubMed ID: 30679029
Summary: Spermiogenesis in Drosophila melanogaster is a highly conserved process and essential for male fertility. In this haploid phase of spermatogenesis, motile sperm are assembled from round cells, and flagella and needle-shaped nuclei with highly compacted genomes are formed. As transcription takes place mainly in spermatocytes and transcripts relevant for post-meiotic sperm development are translationally repressed for days, this study comparatively analysed the proteome of larval testes (only germ cell stages before meiotic divisions), testes of 1-2-day-old pupae (germ cell stages before meiotic divisions, meiotic and early spermatid stages) and adult flies (germ cell stages before meiotic divisions, meiotic and early spermatid stages, late spermatids and sperm). 6,171 proteins were identified; 61 proteins were detected solely in one stage and are thus enriched, namely 34 in larval testes, 77 in pupal testes and 214 in adult testes. To substantiate mass spectrometric data, the stage-specific synthesis and importance for male fertility were analyzed of a number of uncharacterized proteins. For example, Mst84B (gene CG1988), a very basic cysteine- and lysine-rich nuclear protein and was present in the transition phase from a histone-based to a protamine-based chromatin structure. CG6332 encodes d-Theg, which is related to the mouse tHEG and human THEG proteins. Mutants of d-Theg were sterile due to the lack of sperm in the seminal vesicles. This catalogue of proteins of the different stages of testis development in D. melanogaster will pave the road for future analyses of spermatogenesis. | Gruntenko, N. E., Karpova, E. K., Adonyeva, N. V., Andreenkova, O. V., Burdina, E. V., Ilinsky, Y. Y., Bykov, R. A., Menshanov, P. N. and Rauschenbach, I. Y. (2019). Drosophila female fertility and juvenile hormone metabolism depends on the type of Wolbachia infection. J Exp Biol. PubMed ID: 30679245
Summary: Maternally inherited intracellular bacteria Wolbachia cause both parasitic and mutualistic effects on their numerous insect hosts that include manipulating the host reproductive system in order to increase the bacteria spreading in a host population, and increasing the host fitness. This study demonstrates that the type of Wolbachia infection determines the effect on Drosophila melanogaster egg production as a proxy for fecundity and metabolism of juvenile hormone (JH), which acts as gonadotropin in adult insects. This study used six D. melanogaster lineages carrying the nuclear background of interbred Bi90 lineage and cytoplasmic backgrounds with Wolbachia of different genotype variants or without it. wMelCS genotype of Wolbachia decreases the egg production in the infected D. melanogaster females in the beginning of oviposion and increases it later (since the sixth day after eclosion), wMelPop Wolbachia strain causes the opposite effect, while wMel, wMel2 and wMel4 genotypes of Wolbachia do not show any effect on these traits compared to uninfected Bi90 D. melanogaster females. The intensity of JH catabolism negatively correlates with the fecundity level in the flies carrying both wMelCS and wMelPop Wolbachia The JH catabolism in females infected with genotypes of wMel group does not differ from that in uninfected females. The effects of wMelCS and wMelPop infection on egg production can be leveled by the modulation of JH titre (via precocene/JH treatment of the flies). Thus, at least one of the mechanisms, promoting the effect of Wolbachia on D. melanogaster female fecundity, is mediated by JH. |
| Yu, J., Chen, B., Zheng, B., Qiao, C., Chen, X., Yan, Y., Luan, X., Xie, B., Liu, J., Shen, C., He, Z., Hu, X., Liu, M., Li, H., Shao, Q. and Fang, J. (2019). ATP synthase is required for male fertility and germ cell maturation in Drosophila testes. Mol Med Rep. PubMed ID: 30628672
Summary: Germ cell maturation is essential for spermatogenesis and testis homeostasis. ATP synthase serves significant roles in energy storage in germ cell survival and is catalyzed by alterations in the mitochondrial membrane proton concentration. The intrinsic cellular mechanisms governing stem cell maturation remain largely unknown. In the present study, in vivo RNA interference (RNAi) screening of major ATP synthase subunits was performed, and the function of ATP synthase for male fertility and spermatogenesis in Drosophila was explored. A UAS/Gal4 transcription factor system was used to knock down gene expression in specific cell types, and immunofluorescence staining was conducted to assess the roles of ATP synthase subunits in Drosophila testes. It was knockdown of ATP synthase was shown to result in male infertility and abnormal spermatogenesis in Drosophila testes. In addition, knockdown of the ATP synthase beta subunit in germ cells resulted in defects in male infertility and germ cell maturation, while the hub and cyst cell populations were maintained. Other major ATP synthase subunits were also examined and similar phenotypes in Drosophila testes were identified. Taken together, the data from the present study revealed that ATP synthase serves important roles for male fertility during spermatogenesis by regulating germ cell maturation in Drosophila testes. | Sreejith, P., Jang, W., To, V., Hun Jo, Y., Biteau, B. and Kim, C. (2019). Lin28 is a critical factor in the function and aging of Drosophila testis stem cell niche. Aging (Albany NY). PubMed ID: 30713156
Summary: Age-related decline in stem cell function is observed in many tissues from invertebrates to humans. While cell intrinsic alterations impair stem cells, aging of the stem cell niche also significantly contributes to the loss of tissue homeostasis associated with reduced regenerative capacity. Hub cells, which constitute the stem cell niche in the Drosophila testis, exhibit age-associated decline in number and activities, yet underlying mechanisms are not fully understood. This study shows that Lin28, a highly conserved RNA binding protein, is expressed in hub cells and its expression dramatically declines in old testis. lin28 mutant testes exhibit hub cell loss and defective hub architecture, recapitulating the normal aging process. Importantly, maintained expression of Lin28 prolongs hub integrity and function in aged testes, suggesting that Lin28 decline is a driver of hub cell aging. Mechanistically, the level of unpaired (upd), a stem cell self-renewal factor, is reduced in lin28 mutant testis and Lin28 protein directly binds and stabilizes upd transcripts, in a let-7 independent manner. Altogether, these results suggest that Lin28 acts to protect upd transcripts in hub cells, and reduction of Lin28 in old testis leads to decreased upd levels, hub cell aging and loss of the stem cell niche. |
| Gao, Y., Mao, Y., Xu, R. G., Zhu, R., Zhang, M., Sun, J., Shen, D., Peng, P., Xie, T. and Ni, J. Q. (2019). Defining gene networks controlling the maintenance and function of the differentiation niche by an in vivo systematic RNAi screen. J Genet Genomics. PubMed ID: 30745214
Summary: In the Drosophila ovary, escort cells (ECs) extrinsically control germline stem cell (GSC) maintenance and progeny differentiation. However, the underlying mechanisms remain poorly understood. This study identified 173 EC genes for their roles in controlling GSC maintenance and progeny differentiation by using an in vivo systematic RNAi approach. Of the identified genes, 10 and 163 are required in ECs to promote GSC maintenance and progeny differentiation, respectively. The genes required for progeny differentiation fall into different functional categories, including transcription, mRNA splicing, protein degradation, signal transduction and cytoskeleton regulation. In addition, the GSC progeny differentiation defects caused by defective ECs are often associated with BMP signaling elevation, indicating that preventing BMP signaling is a general functional feature of the differentiation niche. Lastly, exon junction complex (EJC) components, which are essential for mRNA splicing, are required in ECs to promote GSC progeny differentiation by maintaining ECs and preventing BMP signaling. Therefore, this study has identified the major regulators of the differentiation niche, which provides important insights into how stem cell progeny differentiation is extrinsically controlled. | Lenhart, K. F., Capozzoli, B., Warrick, G. S. D. and DiNardo, S. (2018). Diminished Jak/STAT signaling causes early-onset aging defects in stem cell cytokinesis. Curr Biol. PubMed ID: 30612906 Summary: Tissue renewal becomes compromised with age. Although defects in niche and stem cell behavior have been implicated in promoting age-related decline, the causes of early-onset aging defects are unknown. This study has identified an early consequence of aging in germline stem cells (GSCs) in the Drosophila testis. Aging disrupts the unique program of GSC cytokinesis, with GSCs failing to abscise from their daughter cells. Abscission failure significantly disrupts both self-renewal and the generation of differentiating germ cells. Extensive live imaging and genetic analyses show that abscission failure is due to inappropriate retention of F-actin at the intercellular bridges between GSC-daughter cells. Furthermore, F-actin is regulated by the Jak/STAT pathway-increasing or decreasing pathway activity can rescue or exacerbate the age-induced abscission defect, respectively. Even subtle decreases to STAT activity are sufficient to precociously age young GSCs and induce abscission failure. Thus, this work has identified the earliest age-related defect in GSCs and has revealed a unique role for an established niche signaling pathway in controlling stem cell cytokinesis and in regulating stem cell behavior with age. |
Friday, March 15th |
| Kottler, B., Faville, R., Bridi, J. C. and Hirth, F. (2019). Inverse control of turning behavior by Dopamine D1 receptor signaling in columnar and ring neurons of the central complex in Drosophila. Curr Biol. PubMed ID: 30713106
Summary: Action selection is a prerequisite for decision-making and a fundamental aspect to any goal-directed locomotion; it requires integration of sensory signals and internal states to translate them into action sequences. This paper introduces a novel behavioral analysis to study neural circuits and mechanisms underlying action selection and decision-making in freely moving Drosophila. Preferred patterns of motor activity and turning behavior were discovered. These patterns are impaired in FoxP mutant flies, which present an altered temporal organization of motor actions and turning behavior, reminiscent of indecisiveness. Then, focusing on central complex (CX) circuits known to integrate different sensory modalities and controlling premotor regions, action sequences and turning behavior were shown to be regulated by dopamine D1-like receptor (Dop1R1) signaling. Dop1R1 inputs onto CX columnar ellipsoid body-protocerebral bridge gall (E-PG) neuron and ellipsoid body (EB) R2/R4m ring neuron circuits both negatively gate motor activity but inversely control turning behavior. Although flies deficient of D1 receptor signaling present normal turning behavior despite decreased activity, restoring Dop1R1 level in R2/R4m-specific circuitry affects the temporal organization of motor actions and turning. EB R2/R4m neurons are in contact with E-PG neurons that are thought to encode body orientation and heading direction of the fly. These findings suggest that Dop1R1 signaling in E-PG and EB R2/4 m circuits are compared against each other, thereby modulating patterns of activity and turning behavior for goal-directed locomotion. | Rohith, B. N. and Shyamala, B. V. (2019). Developmental deformity due to scalloped non-function in Drosophila brain leads to cognitive impairment. Dev Neurobiol. PubMed ID: 30676700
Summary: Neural identity and wiring specificity are fundamental to brain function. Factors affecting proliferation of the progenitor cells leading to an expansion or regression of specific neuronal clusters are expected to challenge the process of formation of precise synaptic connections with their partners and their further integration to result in proper functional neural circuitry. This study has investigated the role of scalloped, a Hippo pathway gene in Drosophila brain development and has shown that its function is critical to regulate proliferation of Mushroom Body Neuroblasts and to limit the neuronal cluster size to normal in the fly brain. The consequent effect of the anatomical phenotype of mutant flies on the brain function was tested, as exemplified by their cognitive performance. The neural expansion in important neural clusters of the olfactory pathway, caused by Scalloped inactivation, imparts severe disabilities in learning, short-term memory and long-term memory. Scalloped knockdown in alphabeta Kenyon Cell clusters drastically reduces long-term memory performance. Scalloped deficiency induced neural expansion in antennal lobe and ellipsoid body neurons, bring down short-term memory performance significantly. It was also demonstrate that the cognitive impairments observed in this stdy are not due to a problem in memory formation or execution in the adult, but are due to the developmental deformities caused in the respective class of neurons. These results strongly indicate that the additional neurons generated by Scalloped inactivation are not synergistically integrated into, but rather perturb the formation of precise functional circuitry. |
| Jang, W., Lee, S., Choi, S. I., Chae, H. S., Han, J., Jo, H., Hwang, S. W., Park, C. S. and Kim, C. (2019). Impairment of proprioceptive movement and mechanical mociception in Drosophila melanogaster larvae lacking Ppk30, a Drosophila member of the DEG/ENaC family. Genes Brain Behav: e12545. PubMed ID: 30675754
Summary: The mechanosensory neurons of Drosophila larvae are demonstrably activated by diverse mechanical stimuli, but the mechanisms underlying this function are not completely understood. This study reports a genetic, immunohistochemical, and electrophysiological analysis of the Ppk30 ion channel, a member of the Drosophila pickpocket (ppk) family, counterpart of the mammalian Degenerin/Epithelial Na(+) Channel family. Ppk30 mutant larvae displayed deficits in proprioceptive movement and mechanical nociception, which are detected by class IV sensory (mdIV) neurons. The same neurons also detect heat nociception, which was not impaired in ppk30 mutant larvae. Similarly, Ppk30 mutation did not alter gentle touch mechanosensation, a distinct mechanosensation detected by other neurons, suggesting that Ppk30 has a functional role in mechanosensation in mdIV neurons. Consistently, Ppk30 was expressed in class IV neurons, but was not detectable in other larval skin sensory neurons. Mutant phenotypes were rescued by expressing Ppk30 in mdIV neurons. Electrophysiological analysis of heterologous cells expressing Ppk30 did not detect mechanosensitive channel activities, but did detect acid-induced currents. These data demonstrate that Ppk30 has a role in mechanosensation, but not in thermosensation, in class IV neurons, and possibly has other functions related to acid response. | Sears, J. C., Choi, W. J. and Broadie, K. (2019). Fragile X mental retardation protein positively regulates PKA anchor Rugose and PKA activity to control actin assembly in learning/memory circuitry. Neurobiol Dis. PubMed ID: 30771457
Summary: Recent work shows Fragile X Mental Retardation Protein (FMRP) drives the translation of very large proteins (>2000 aa) mediating neurodevelopment. Loss of function results in Fragile X syndrome (FXS), the leading heritable cause of intellectual disability (ID) and autism spectrum disorder (ASD). Using the Drosophila FXS disease model, FMRP was found to positively regulate the translation of the very large A-Kinase Anchor Protein (AKAP) Rugose (>3000 aa), homolog of ASD-associated human Neurobeachin (NBEA). In the central brain Mushroom Body (MB) circuit, where Protein Kinase A (PKA) signaling is necessary for learning/memory, FMRP loss reduces Rugose levels and targeted FMRP over-expression elevates Rugose levels. Using a new in vivo transgenic PKA activity reporter (PKA-SPARK), FMRP loss was found to reduces PKA activity in MB Kenyon Cells whereas FMRP overexpression elevates PKA activity. Consistently, loss of Rugose reduces PKA activity, but Rugose overexpression has no independent effect. A well-established PKA output is regulation of F-actin cytoskeleton dynamics. In the FXS disease model, F-actin is aberrantly accumulated in MB lobes and single MB Kenyon cells. Consistently, Rugose loss results in similar F-actin accumulation. Moreover, targeted FMRP, Rugose and PKA overexpression all result in increased F-actin accumulation in the MB circuit. These findings uncover a FMRP-Rugose-PKA mechanism regulating actin cytoskeleton. |
| Si, G., Kanwal, J. K., Hu, Y., Tabone, C. J., Baron, J., Berck, M., Vignoud, G. and Samuel, A. D. T. (2019). Structured odorant response patterns across a complete olfactory receptor neuron population. Neuron. PubMed ID: 30683545
Summary: Odor perception allows animals to distinguish odors, recognize the same odor across concentrations, and determine concentration changes. How the activity patterns of primary olfactory receptor neurons (ORNs), at the individual and population levels, facilitate distinguishing these functions remains poorly understood. This study interrogated the complete ORN population of the Drosophila larva across a broadly sampled panel of odorants at varying concentrations. The activity of each ORN was found to scale with the concentration of any odorant via a fixed dose-response function with a variable sensitivity. Sensitivities across odorants and ORNs follow a power-law distribution. Much of receptor sensitivity to odorants is accounted for by a single geometrical property of molecular structure. Similarity in the shape of temporal response filters across odorants and ORNs extend these relationships to fluctuating environments. These results uncover shared individual- and population-level patterns that together lend structure to support odor perceptions. | Sapiro, A. L., Shmueli, A., Henry, G. L., Li, Q., Shalit, T., Yaron, O., Paas, Y., Billy Li, J. and Shohat-Ophir, G. (2019). Illuminating spatial A-to-I RNA editing signatures within the Drosophila brain. Proc Natl Acad Sci U S A 116(6): 2318-2327. PubMed ID: 30659150
Summary: Adenosine-to-inosine (A-to-I) RNA editing, catalyzed by ADAR enzymes, is a ubiquitous mechanism that generates transcriptomic diversity. This process is particularly important for proper neuronal function; however, little is known about how RNA editing is dynamically regulated between the many functionally distinct neuronal populations of the brain. This study presents a spatial RNA editing map in the Drosophila brain and shows that different neuronal populations possess distinct RNA editing signatures. After purifying and sequencing RNA from genetically marked groups of neuronal nuclei, a large number of editing sites were identified, and editing levels were compared in hundreds of transcripts across nine functionally different neuronal populations. Distinct editing repertoires were found for each population, including sites in repeat regions of the transcriptome and differential editing in highly conserved and likely functional regions of transcripts that encode essential neuronal genes. These changes are site-specific and not driven by changes in Adar expression, suggesting a complex, targeted regulation of editing levels in key transcripts. This fine-tuning of the transcriptome between different neurons by RNA editing may account for functional differences between distinct populations in the brain. |
Thursday, March 14th - Adult Development |
| Narbonne-Reveau, K. and Maurange, C. (2019). Developmental regulation of regenerative potential in Drosophila by ecdysone through a bistable loop of ZBTB transcription factors. PLoS Biol 17(2): e3000149. PubMed ID: 30742616
Summary: In many organisms, the regenerative capacity of tissues progressively decreases as development progresses. However, the developmental mechanisms that restrict regenerative potential remain unclear. In Drosophila, wing imaginal discs become unable to regenerate upon damage during the third larval stage (L3). This study shows that production of ecdysone after larvae reach their critical weight (CW) terminates the window of regenerative potential by acting on a bistable loop composed of 2 antagonistic Broad-complex/Tramtrack/Bric-a-brac Zinc-finger (ZBTB) genes: chinmo and broad (br). Around mid L3, ecdysone signaling silences chinmo and activates br to switch wing epithelial progenitors from a default self-renewing to a differentiation-prone state. Before mid L3, Chinmo promotes a strong regenerative response upon tissue damage. After mid L3, Br installs a nonpermissive state that represses regeneration. Transient down-regulation of ecdysone signaling or Br in late L3 larvae enhances chinmo expression in damaged cells that regain the capacity to regenerate. This work unveils a mechanism that ties the self-renewing and regenerative potential of epithelial progenitors to developmental progression. | Schultheis, D., Schwirz, J. and Frasch, M. (2019). RNAi screen in Tribolium reveals involvement of F-BAR proteins in myoblast fusion and visceral muscle morphogenesis in insects. G3 (Bethesda). PubMed ID: 30733382
Summary: In a large-scale RNAi screen in Tribolium castaneum for genes with knock-down phenotypes in the larval somatic musculature, one recurring phenotype was the appearance of larval muscle fibers that were significantly thinner than those in control animals. Several of the genes producing this knock-down phenotype corresponded to orthologs of Drosophila genes that are known to participate in myoblast fusion, particularly via their effects on actin polymerization. A new gene previously not implicated in myoblast fusion but displaying a similar thin-muscle knock-down phenotype was the Tribolium ortholog of Nostrin, which encodes an F-BAR and SH3 domain protein. Genetic studies of Nostrin and Cip4, a gene encoding a structurally related protein, in Drosophila show that the encoded F-BAR proteins jointly contribute to efficient myoblast fusion during larval muscle development. Together with the F-Bar protein Syndapin they are also required for normal embryonic midgut morphogenesis. In addition, Cip4 is required together with Nostrin during the profound remodeling of the midgut visceral musculature during metamorphosis. It is proposed that these F-Bar proteins help govern proper morphogenesis particularly of the longitudinal midgut muscles during metamorphosis. |
| Gui, J., Huang, Y., Montanari, M., Toddie-Moore, D., Kikushima, K., Nix, S., Ishimoto, Y. and Shimmi, O. (2019). Coupling between dynamic 3D tissue architecture and BMP morphogen signaling during Drosophila wing morphogenesis. Proc Natl Acad Sci U S A. PubMed ID: 30760594
Summary: At the level of organ formation, tissue morphogenesis drives developmental processes in animals, often involving the rearrangement of two-dimensional (2D) structures into more complex three-dimensional (3D) tissues. These processes can be directed by growth factor signaling pathways. However, little is known about how such morphological changes affect the spatiotemporal distribution of growth factor signaling. Using the Drosophila pupal wing, this study addresses how Decapentaplegic (Dpp)/bone morphogenetic protein (BMP) signaling and 3D wing morphogenesis are coordinated. Dpp, expressed in the longitudinal veins (LVs) of the pupal wing, initially diffuses laterally within both dorsal and ventral wing epithelia during the inflation stage to regulate cell proliferation. Dpp localization is then refined to the LVs within each epithelial plane, but with active interplanar signaling for vein patterning/differentiation, as the two epithelia appose. These data further suggest that the 3D architecture of the wing epithelia and the spatial distribution of BMP signaling are tightly coupled, revealing that 3D morphogenesis is an emergent property of the interactions between extracellular signaling and tissue shape changes. | Iyer, K. V., Piscitello-Gomez, R., Paijmans, J., Julicher, F. and Eaton, S. (2019). Epithelial viscoelasticity is regulated by mechanosensitive E-cadherin turnover. Curr Biol. PubMed ID: 30744966
Summary: Studying how epithelia respond to mechanical stresses is key to understanding tissue shape changes during morphogenesis. This study examined the viscoelastic properties of the Drosophila wing epithelium during pupal morphogenesis by quantifying mechanical stress and cell shape as a function of time. A delay of 8 h was found between maximal tissue stress and maximal cell elongation, indicating a viscoelastic deformation of the tissue. This viscoelastic behavior emerges from the mechanosensitivity of endocytic E-cadherin turnover. The increase in E-cadherin turnover in response to stress is mediated by mechanosensitive relocalization of the E-cadherin binding protein p120-catenin (p120) from cell junctions to cytoplasm. Mechanosensitivity of E-cadherin turnover is lost in p120 mutant wings, where E-cadherin turnover is constitutively high. In this mutant, the relationship between mechanical stress and stress-dependent cell dynamics is altered. Cells in p120 mutant deform and undergo cell rearrangements oriented along the stress axis more rapidly in response to mechanical stress. These changes imply a lower viscosity of wing epithelium. Taken together, these findings reveal that p120-dependent mechanosensitive E-cadherin turnover regulates viscoelastic behavior of epithelial tissues. |
Wednesday, March 13th - Behavior |
| Nojima, T., Chauvel, I., Houot, B., Bousquet, F., Farine, J. P., Everaerts, C., Yamamoto, D. and Ferveur, J. F. (2019). The desaturase1 gene affects reproduction before, during and after copulation in Drosophila melanogaster. J Neurogenet: 1-20. PubMed ID: 30724684
Summary: Desaturase1 (desat1) is one of the few genes known to be involved in the two complementary aspects of sensory communication - signal emission and signal reception - in Drosophila melanogaster. In particular, desat1 is necessary for the biosynthesis of major cuticular pheromones in both males and females. It is also involved in the male ability to discriminate sex pheromones. Each of these two sensory communication aspects depends on distinct desat1 putative regulatory regions. This study used (i) mutant alleles resulting from the insertion/excision of a transposable genomic element inserted in a desat1 regulatory region, and (ii) transgenics made with desat1 regulatory regions used to target desat1 RNAi. These genetic variants were used to study several reproduction-related phenotypes. In particular, this study investigated the fecundity of various mutant and transgenic desat1 females with regard to the developmental fate of their progeny. The mating performance in pairs of flies was compared with altered desat1 expression in various desat1-expressing tissues together with their inability to disengage at the end of copulation. Moreover, the developmental origin of altered sex pheromone discrimination was investigated in male flies. Attempts were made to map some of the tissues involved in these reproduction-related phenotypes. Given that desat1 is expressed in many brain neurons and in non-neuronal tissues required for varied aspects of reproduction, the data suggest that the regulation of this gene has evolved to allow the optimal reproduction and a successful adaptation to varied environments in this cosmopolitan species. | Qiao, H., Keesey, I. W., Hansson, B. S. and Knaden, M. (2019). Gut microbiota affects development and olfactory behavior in Drosophila melanogaster. J Exp Biol. PubMed ID: 30679242
Summary: It has been shown that gut microbes are very important for the behavior and development of Drosophila, as the beneficial microbes are involved in the identification of suitable feeding and oviposition places. However, in what way these associated gut microbes influence the fitness-related behaviors of Drosophila melanogaster remains unclear. This study shows that D. melanogaster exhibits different behavioral preferences towards gut microbes. Both adults and larvae were attracted by the headspace of Saccharomyces cerevisiae and Lactobacillus plantarum, but were repelled by Acetobacter malorum in behavioral assays, indicating an olfactory mechanism involved in these preference behaviors. While the attraction to yeast was governed by olfactory sensory neurons expressing the odorant co-receptor Orco, the observed behaviors towards the other microbes still remained in flies lacking this co-receptor. By experimentally manipulating the microbiota of the flies, flies were found to not strive for a diverse microbiome by e.g. increasing their preference towards gut microbes that they had not experienced previously. Instead, in some cases the flies even increased preference for the microbes they were reared on. Furthermore, exposing Drosophila larvae to all three microbes promoted Drosophila's development while only exposure to S. cerevisiae and A. malorum resulted in the development of larger ovaries and in increased egg numbers the flies laid in an oviposition assay. Thus this study provides a better understanding of how gut microbes affect insect behavior and development, and offers an ecological rationale for preferences of flies for different microbes in their natural environment. |
| Monier, M., Nobel, S., Danchin, E. and Isabel, G. (2018). Dopamine and serotonin are both required for mate-copying in Drosophila melanogaster. Front Behav Neurosci 12: 334. PubMed ID: 30687036
Summary: Mate-copying is a form of social learning in which the mate-choice decision of an individual (often a female) is influenced by the mate-choice of conspecifics. Drosophila melanogaster females are known to perform such social learning, and in particular, to mate-copy after a single observation of one conspecific female mating with a male of one phenotype, while the other male phenotype is rejected. This study shows that this form of social learning is dependent on serotonin and dopamine. Using a pharmacological approach, dopamine or serotonin synthesis in adult virgin females with 3-iodotyrosine (3-IY) and DL-para-chlorophenylalanine (PCPA), respectively, and then their mate-copying performance was tested. While control females without drug treatment copied the choice of the demonstrator, drug-treated females with reduced dopamine or serotonin chose randomly. To ensure the specificity of the drugs, the direct precursors of the neurotransmitters, either the dopamine precursor L-3,4-dihydroxyphenylalanine (L-DOPA) or the serotonin precursor 5-L-hydroxytryptophan (5-HTP) were given together with the drug, (respectively 3-IY and PCPA) resulting in a full rescue of the mate-copying defects. This indicates that dopamine and serotonin are both required for mate-copying. These results give a first insight into the mechanistic pathway underlying this form of social learning in D. melanogaster. | Ni, J. D., Gurav, A. S., Liu, W., Ogunmowo, T. H., Hackbart, H., Elsheikh, A., Verdegaal, A. A. and Montell, C. (2019). Differential regulation of the Drosophila sleep homeostat by circadian and arousal inputs. Elife 8. PubMed ID: 30719975
Summary: One output arm of the sleep homeostat in Drosophila appears to be a group of neurons with projections to the dorsal fan-shaped body (dFB neurons) of the central complex in the brain. However, neurons that regulate the sleep homeostat remain poorly understood. Using neurogenetic approaches combined with Ca(2+) imaging, synaptic connections were characterized between dFB neurons and distinct sets of upstream sleep-regulatory neurons. One group of the sleep-promoting upstream neurons is a set of circadian pacemaker neurons that activates dFB neurons via direct glutaminergic excitatory synaptic connections. Opposing this population, a group of arousal-promoting neurons downregulates dFB axonal output with dopamine. Co-activating these two inputs leads to frequent shifts between sleep and wake states. dFB neurons were shown to release the neurotransmitter GABA and inhibit octopaminergic arousal neurons. It is proposed that dFB neurons integrate synaptic inputs from distinct sets of upstream sleep-promoting circadian clock neurons, and arousal neurons. |
| Rimal, S., Sang, J., Poudel, S., Thakur, D., Montell, C. and Lee, Y. (2019). Mechanism of acetic acid gustatory repulsion in Drosophila. Cell Rep 26(6): 1432-1442.e1434. PubMed ID: 30726729
Summary: The decision to consume or reject a food based on the degree of acidity is critical for animal survival. However, the gustatory receptors that detect sour compounds and influence feeding behavior have been elusive. Using the fly, Drosophila melanogaster, it was revealed that a member of the ionotropic receptor family, IR7a, is essential for rejecting foods laced with high levels of acetic acid. IR7a is dispensable for repulsion of other acidic compounds, indicating that the gustatory sensation of acids occurs through a repertoire rather than a single receptor. The fly's main taste organ, the labellum, is decorated with bristles that house dendrites of gustatory receptor neurons (GRNs). IR7a is expressed in a subset of bitter GRNs rather than GRNs dedicated to sour taste. These findings indicate that flies taste acids through a repertoire of receptors, enabling them to discriminate foods on the basis of acid composition rather than just pH. | McKellar, C. E., Lillvis, J. L., Bath, D. E., Fitzgerald, J. E., Cannon, J. G., Simpson, J. H. and Dickson, B. J. (2019). Threshold-based ordering of sequential actions during Drosophila courtship. Curr Biol 29(3): 426-434.e426. PubMed ID: 30661796
Summary: Goal-directed animal behaviors are typically composed of sequences of motor actions whose order and timing are critical for a successful outcome. Although numerous theoretical models for sequential action generation have been proposed, few have been supported by the identification of control neurons sufficient to elicit a sequence. This study identified a pair of descending neurons that coordinate a stereotyped sequence of engagement actions during Drosophila melanogaster male courtship behavior. These actions are initiated sequentially but persist cumulatively, a feature not explained by existing models of sequential behaviors. Evidence was found consistent with a ramp-to-threshold mechanism, in which increasing neuronal activity elicits each action independently at successively higher activity thresholds. |
Tuesday, March 12th - Enhancers and Transcriptonal Regulation |
| Surkova, S., Sokolkova, A., Kozlov, K., Nuzhdin, S. V. and Samsonova, M. (2019). Quantitative analysis reveals genotype- and domain- specific differences between mRNA and protein expression of segmentation genes in Drosophila. Dev Biol. PubMed ID: 30629954
Summary: This study has characterized differences between mRNA and protein expression of Drosophila segmentation genes at the level of individual gene expression domains. Quantitative imaging data was obtained on expression of gap genes gt and hb and pair-rule gene eve for Drosophila wild type embryos, Kr null mutants and Kr+/Kr- heterozygotes. To compare mRNA and protein expression several criteria were used including difference in amplitude and positions of expression domains, pattern shape and positional variability. For a number of gene expression domains examples are shown where protein expression does not repeat mRNA expression even after a temporal delay. Time delays were calculated between eve pattern formation at the level of mRNA and protein for wild type embryos, Kr mutants and Kr+/Kr- heterozygotes. In wild type embryos, the amplitudes of eve stripes 3 and 7 do not differ significantly at the level of mRNA, however, stripe 3 is higher than stripe 7 at the protein level. It was further shown that hb mRNA and protein expression in both anterior and posterior domains significantly differs at specific time points. The formation of hb PS4 stripe at the mRNA level proceeds five times faster than at the level of protein. With regard to spatial expression, the offset between posterior gt mRNA and protein domains is much larger in Kr mutants than in wild type embryos and heterozygotes. Finally, differences were analyzed in positional variability of eve stripe 7 expression in Kr mutants and Kr+/Kr- heterozygotes at the level of mRNA and protein. These results enable further perspectives to uncover mechanisms underlying discrepancies between mRNA and protein expression in early embryo. | Immarigeon, C., Bernat-Fabre, S., Auge, B., Faucher, C., Gobert, V., Haenlin, M., Waltzer, L., Payet, A., Cribbs, D. L., Bourbon, H. G. and Boube, M. (2019). Drosophila Mediator subunit Med1 is required for GATA-dependent developmental processes: divergent binding interfaces for conserved coactivator functions. Mol Cell Biol. PubMed ID: 30670567
Summary: DNA-bound transcription factors (TFs) governing developmental gene regulation have been proposed to recruit Polymerase II machinery at gene promoters through specific interactions with dedicated subunits of the evolutionarily-conserved Mediator complex (MED). However, whether such MED subunit specific functions and partnerships have been conserved during evolution has been poorly investigated. To address this issue, the first Drosophila loss-of-function mutants were generated for Med1, known as a specific cofactor for GATA TFs and hormone nuclear receptors in mammals. Med1 was shown to be required for cell proliferation, and hematopoietic differentiation depending on the GATA TF Serpent (Srp). Med1 binds Srp in cultured cells and in vitro through its conserved GATA Zinc Finger DNA-binding domain and the divergent Med1 C-terminal. Interestingly, GATA/Srp interaction occurs through the longest Med1 isoform, suggesting a functional diversity of MED complex populations. Furthermore, it was shown that Med1 acts as a coactivator for the GATA factor Pannier during thoracic development. In conclusion, the Med1 requirement for GATA-dependent regulatory processes is a common feature in insects and mammals, although binding interfaces have diverged. Further work in Drosophila should bring valuable insights to fully understand GATA-MED functional partnerships, which probably involve other MED subunits depending on the cellular context. |
| Ramnarine, T. J. S., Glaser-Schmitt, A., Catalan, A. and Parsch, J. (2019). Population genetic and functional analysis of a cis-regulatory polymorphism in the Drosophila melanogaster Metallothionein A gene. Genes (Basel) 10(2). PubMed ID: 30769915
Summary: Although gene expression can vary extensively within and among populations, the genetic basis of this variation and the evolutionary forces that maintain it are largely unknown. In Drosophila melanogaster, a 49-bp insertion/deletion (indel) polymorphism in the Metallothionein A (MtnA) gene is associated with variation in MtnA expression and oxidative stress tolerance. To better understand the functional and evolutionary significance of this polymorphism, it was investigated in several worldwide populations. In a German population, the deletion was present at a high and stable frequency over multiple seasons and years, and was associated with increased MtnA expression. There was, however, no evidence that the polymorphism was maintained by overdominant, seasonally fluctuating, or sexually antagonistic selection. The deletion was rare in a population from the species' ancestral range in sub-Saharan Africa and is likely the result of non-African admixture, suggesting that it spread to high frequency following the species' out-of-Africa expansion. Using data from a North American population, it was found that the deletion was associated with MtnA expression and tolerance to oxidative stress induced by menadione sodium bisulfite. These results are consistent with the deletion being selectively favored in temperate populations due to the increased MtnA expression and oxidative stress tolerance that it confers. | Lin, W. H. and Baines, R. A. (2019). Myocyte enhancer factor-2 and p300 interact to regulate expression of the homeostatic regulator Pumilio in Drosophila. Eur J Neurosci. PubMed ID: 30687963
Summary: Pumilio (Pum), an RNA-binding protein, is a key component of neuron firing-rate homeostasis that likely maintains stability of neural circuit activity in all animals, from flies to mammals. Whilst Pum is ubiquitously expressed, little is understood about how synaptic excitation regulates its expression in the CNS. This study characterized the Drosophila dpum promoter and identified multiple Myocyte enhancer factor-2 (Mef2)-binding elements. Twelve dmef2 splice variants were identifiedf and a luciferase-based assay was used to monitor dpum promoter activity. Whilst all 12 dMef2 splice variants enhance dpum promoter activity, exon 10-containing variants induce greater transactivation. Previous work shows dPum expression increases with synaptic excitation. However, no change was observed in dMef2 transcript in larval CNS, of both sexes, exposed to the proconvulsant picrotoxin. The lack of activity-dependence is indicative of additional regulation. p300 was identified as a potential candidate. By binding to dMef2, p300 represses dpum transactivation. Significantly, p300 transcript is down-regulated by enhanced synaptic excitation (picrotoxin) which, in turn, increases transcription of dpum through de-repression of dMef2. These results advance understanding of dpum by showing activity-dependent expression is regulated by an interaction between p300 and dMef2. |
| Li, K. and Baker, N. E. (2019). Transcriptional and post-transcriptional regulation of extra macrochaetae during Drosophila adult peripheral neurogenesis. Dev Biol. PubMed ID: 30771303
Summary: Regulation of the Drosophila ID protein Extra macrochaetae (Emc) is important because reduced Emc levels have been proposed to favor proneural gene activity and thereby define a prepattern for neurogenesis. Recent studies suggest a major role for post-translational control of Emc levels. To further define the mechanisms of Emc regulation, two redundant cis-regulatory regions were identified by germline transformation-rescue experiments that make use of new molecularly-defined emc mutants. The mechanisms by which Daughterless (Da) regulated Emc expression were distinguished, finding post-translational regulation in most tissues, and additional transcriptional regulation in the eye imaginal disc posterior to the morphogenetic furrow. Dpp and Hh signaling pathways repressed Emc transcriptionally and post-translationally within the morphogenetic furrow of the eye disc, whereas Wg signaling repressed Emc expression at the anterior margin of the wing imaginal disc. Although the emc 3' UTR is potentially regulatory, no effect of miRNA pathways on Emc protein levels was discernible. This work supports recent evidence that post-transcriptional mechanisms contribute more to regulation of Emc protein levels than transcriptional mechanisms do. | Osman, N. M., Kitapci, T. H., Vlaho, S., Wunderlich, Z. and Nuzhdin, S. V. (2018). Inference of transcription factor regulation patterns using gene expression covariation in natural populations of Drosophila melanogaster. Biophysics (Oxf) 63(1): 43-51. PubMed ID: 30739944
Summary: Gene regulatory networks control the complex programs that drive development. Deciphering the connections between transcription factors (TFs) and target genes is challenging, in part because TFs bind to thousands of places in the genome but control expression through a subset of these binding events. It is hypothesized that natural variation of expression levels and predictions of TF binding sites can be combined to identify TF targets. RNA-seq data was combined from 71 genetically distinct F1 Drosophila melanogaster embryos, and the correlations were calculated between TF and potential target gene expression levels, which was called "regulatory strength." To separate direct and indirect TF targets, it was hypothesize that direct TF targets will have a preponderance of binding sites in their upstream regions. Using 14 TFs active during embryogenesis, it was found that 12 TFs showed a significant correlation between their binding strength and regulatory strength on downstream targets, and 10 TFs showed a significant correlation between the number of binding sites and the regulatory effect on target genes. The general roles, e.g. Bicoid's role as an activator, and the particular interactions that were observed between these TFs, e.g. Twist's role as a repressor of sloppy paired and odd paired, generally coincide with the literature. |
Monday, March 11th - Immune Response |
| Hao, Y., Yu, S., Luo, F. and Jin, L. H. (2018). Jumu is required for circulating hemocyte differentiation and phagocytosis in Drosophila. Cell Commun Signal 16(1): 95. PubMed ID: 30518379
Summary: The regulatory mechanisms of hematopoiesis and cellular immunity show a high degree of similarity between insects and mammals, and Drosophila has become a good model for investigating cellular immune responses. Jumeau (Jumu) is a member of the winged-helix/forkhead (FKH) transcription factor family and is required for Drosophila development. Adult junu mutant flies show defective hemocyte phagocytosis and a weaker defense capability against pathogen infection. This study investigated the role of junu in the regulation of larval hemocyte development and phagocytosis. In vivo phagocytosis assays, immunohistochemistry, Real-time quantitative PCR and immunoblotting were performed to investigate the effect of junu on hemocyte phagocytosis. 5-Bromo-2-deoxyUridine (BrdU) labeling, phospho-histone H3 (PH3) and TdT-mediated dUTP Nick-End Labeling (TUNEL) staining were performed to analyze the proliferation and apoptosis of hemocyte; immunohistochemistry and Mosaic analysis with a repressible cell marker (MARCM) clone analysis were performed to investigate the role of Jumu in the activation of Toll pathway. Jumu was found to indirectly control hemocyte phagocytosis by regulating the expression of NimC1 and cytoskeleton reorganization. The loss of junu also causes abnormal proliferation and differentiation in circulating hemocytes. Thesee results suggest that a severe deficiency of junu leads to the generation of enlarged multinucleate hemocytes by affecting the normal cell mitosis process and induces numerous lamellocytes by activating the Toll pathway. It is concluded that Jumu regulates circulating hemocyte differentiation and phagocytosis in Drosophila. These findings provide new insight into the mechanistic roles of cytoskeleton regulatory proteins in phagocytosis and establish a basis for further analyses of the regulatory mechanism of the mammalian ortholog of Jumu in mammalian innate immunity. | Chowdhury, M., Zhang, J., Xu, X. X., He, Z., Lu, Y., Liu, X. S., Wang, Y. F. and Yu, X. Q. (2019). An in vitro study of NF-kappaB factors cooperatively in regulation of Drosophila melanogaster antimicrobial peptide genes. Dev Comp Immunol 95: 50-58. PubMed ID: 30735676
Summary: An important innate immune response in Drosophila melanogaster is the production of antimicrobial peptides (AMPs). Expression of AMP genes is mediated by the Toll and immune deficiency (IMD) pathways via NF-kappaB transcription factors Dorsal, DIF and Relish. Dorsal and DIF act downstream of the Toll pathway, whereas Relish acts in the IMD pathway. Dorsal and DIF are held inactive in the cytoplasm by the IkappaB protein Cactus, while Relish contains an IkappaB-like inhibitory domain at the C-terminus. NF-kappaB factors normally form homodimers and heterodimers to regulate gene expression, but formation of heterodimers between Relish and DIF or Dorsal and the specificity and activity of the three NF-kappaB homodimers and heterodimers are not well understood. This study compared the activity of Rel homology domains (RHDs) of Dorsal, DIF and Relish in activation of Drosophila AMP gene promoters, demonstrated that Relish-RHD (Rel-RHD) interacted with both Dorsal-RHD and DIF-RHD, Relish-N interacted with DIF and Dorsal, and overexpression of individual RHD and co-expression of any two RHDs activated the activity of AMP gene promoters to various levels, suggesting formation of homodimers and heterodimers among Dorsal, DIF and Relish. Rel-RHD homodimers were stronger activators than heterodimers of Rel-RHD with either DIF-RHD or Dorsal-RHD, while DIF-RHD-Dorsal-RHD heterodimers were stronger activators than either DIF-RHD or Dorsal-RHD homodimers in activation of AMP gene promoters. The nucleotides at the 6th and 8th positions of the 3' half-sites of the kappaB motifs were identified that are important for the specificity and activity of NF-kappaB transcription factors. |
| Myers, A. L., Harris, C. M., Choe, K. M. and Brennan, C. A. (2018). Inflammatory production of reactive oxygen species by Drosophila hemocytes activates cellular immune defenses. Biochem Biophys Res Commun. PubMed ID: 30292413
Summary: The production of reactive oxygen species (ROS) is a prominent response to infection among innate immune cells such as macrophages and neutrophils. To better understand the relationship between antimicrobial and regulatory functions of blood cell ROS, this study has characterized the ROS response to infection in Drosophila hemocytes. Using fluorescent probes, a biphasic hemocyte ROS response was found to bacterial infection. In the first hour, virtually all hemocytes generate a transient ROS signal, with nonphagocytic cells including prohemocytes and crystal cells displaying exceptionally strong responses. A distinct, and more delayed ROS response starting at 90min is primarily within cells that have engulfed bacteria, and is sustained for several hours. The early response has a clear regulatory function, as dampening or intensifying the intracellular ROS level has profound effects on plasmatocyte activation. In addition, ROS are necessary and sufficient to activate JNK signalling in crystal cells, and to promote JNK-dependent crystal cell rupture. These findings indicate that Drosophila will be a promising model in which to dissect the mechanisms of ROS stimulation of immune activation. | Li, R., Huang, Y., Zhang, Q., Zhou, H., Jin, P. and Ma, F. (2019). The miR-317 functions as a negative regulator of Toll immune response and influences Drosophila survival. Dev Comp Immunol 95: 19-27. PubMed ID: 30708026
Summary: The miR-317 has been revealed to involve in the reproductive response and the larval ovary morphogenesis of Drosophila. However, whether the miR-317 can also regulate Drosophila innate immune responses, which remains unclear to date. This study verified that miR-317 can directly target the 3'UTR of Dif-Rc to down-regulate the expression levels of AMP Drs to negatively control Drosophila Toll immune response in vivo and vitro. Specially, the Dif is an important transcription factor of Toll pathway with four transcripts (Dif-Ra, Dif-Rb, Dif-Rc and Dif-Rd). The results show that miR-317 only targets to Dif-Rc, but not Dif-Ra/b/d. Furthermore, it was demonstrated that the miR-317 sponge can restore the expression levels of Drs and Dif-Rc at mRNA and protein levels. Remarkably, during Gram-positive bacterial infection, the overexpressed miR-317 flies have poor survival outcome, whereas the knockout miR-317 flies have favorable survival compared to the control group, respectively, suggesting that the miR-317 might play a key role in Drosophila survival. Taken together, this work not only reveal an innate immune function and a novel regulation pattern of miR-317, but also provide a new insight into the underlying molecular mechanisms of immunity disorder influencing on Drosophila survival. |
| Shukla, A. K., Spurrier, J., Kuzina, I. and Giniger, E. (2019). Hyperactive innate immunity causes degeneration of dopamine neurons upon altering activity of Cdk5. Cell Rep 26(1): 131-144. PubMed ID: 30605670
Summary: Innate immunity is central to the pathophysiology of neurodegenerative disorders, but it remains unclear why immunity is altered in the disease state and whether changes in immunity are a cause or a consequence of neuronal dysfunction. This study identified a molecular pathway that links innate immunity to age-dependent loss of dopaminergic neurons in Drosophila. Altering the expression of the activating subunit of the Cdk5 protein kinase (Cdk5alpha) causes severe disruption of autophagy. Second, this disruption of autophagy is both necessary and sufficient to cause the hyperactivation of innate immunity, particularly expression of anti-microbial peptides. Finally, it is the upregulation of immunity that induces the age-dependent death of dopaminergic neurons. Given the dysregulation of Cdk5 and innate immunity in human neurodegeneration and the conserved role of the kinase in the regulation of autophagy, this sequence is likely to have direct application to the chain of events in human neurodegenerative disease. | Merkling, S. H., Riahi, H., Overheul, G. J., Schenck, A. and van Rij, R. P. (2019). Peroxisome-associated Sgroppino links fat metabolism with survival after RNA virus infection in Drosophila. Sci Rep 9(1): 2065. PubMed ID: 30765784
Summary: The fruit fly Drosophila melanogaster is a valuable model organism for the discovery and characterization of innate immune pathways, but host responses to virus infection remain incompletely understood. This study describes a novel player in host defense, Sgroppino (Sgp; CG13091). Genetic depletion of Sgroppino causes hypersensitivity of adult flies to infections with the RNA viruses Drosophila C virus, cricket paralysis virus, and Flock House virus. Canonical antiviral immune pathways are functional in Sgroppino mutants, suggesting that Sgroppino exerts its activity via an as yet uncharacterized process. Sgroppino localizes to peroxisomes, organelles involved in lipid metabolism. In accordance, Sgroppino-deficient flies show a defect in lipid metabolism, reflected by higher triglyceride levels, higher body mass, and thicker abdominal fat tissue. In addition, knock-down of Pex3, an essential peroxisome biogenesis factor, increases sensitivity to virus infection. Together, these results establish a genetic link between the peroxisomal protein Sgroppino, fat metabolism, and resistance to virus infection. |
Friday, March 8th - Behavior |
| Isaac, R. E. (2019). The effect of mating and the male Sex peptide on group behaviour of post-mated female Drosophila melanogaster. Neurochem Res. PubMed ID: 30661229
Summary: Sleep is a highly conserved state in animals, but its regulation and physiological function is poorly understood. Drosophila melanogaster is an excellent model for studying sleep regulation and has been used to investigate how sex and social interactions can influence wake-sleep profiles. Previous work has shown that copulation has a profound effect on day time activity and quiescence (siesta sleep) of individual post-mated females. The effect of mating and the transfer of the 36 amino acid sex peptide in the seminal fluid was studied on the behavior of mated female Drosophila populations, where there will be on-going social interactions. The locomotor activity and sleep patterns of virgin and post-mated female D. melanogaster from three laboratory strains (Oregon-R, Canton-S and Dahomey) were recorded in social groups of 20 individuals in a 12-12 h light-dark cycle. Virgin female populations from all three fly strains displayed consolidated periods of low activity in between two sharp peaks of activity, corresponding to lights-on and lights-off. Similar light-correlated peaks were recorded for the mated female populations, however, the low afternoon activity and siesta seen in virgin populations was abolished after mating in all three strains. In contrast, night activity appeared unaffected. This post-mating effect was sustained for several days and was dependent on the male SP acting as a pheromone. Evidence from mixed populations of virgin and mated females suggests that the siesta of non-mated females is not easily disturbed by the presence of highly active post-mated females. | Jovanic, T., Winding, M., Cardona, A., Truman, J. W., Gershow, M. and Zlatic, M. (2019). Neural substrates of Drosophila larval anemotaxis. Curr Biol. PubMed ID: 30744969
Summary: Animals use sensory information to move toward more favorable conditions. Drosophila larvae can move up or down gradients of odors (chemotax), light (phototax), and temperature (thermotax) by modulating the probability, direction, and size of turns based on sensory input. Whether larvae can anemotax in gradients of mechanosensory cues is unknown. Further, although many of the sensory neurons that mediate taxis have been described, the central circuits are not well understood. This study used high-throughput, quantitative behavioral assays to demonstrate Drosophila larvae anemotax in gradients of wind speeds and to characterize the behavioral strategies involved. Larvae modulate the probability, direction, and size of turns to move away from higher wind speeds. This suggests that similar central decision-making mechanisms underlie taxis in somatosensory and other sensory modalities. By silencing the activity of single or very few neuron types in a behavioral screen, two sensory (chordotonal and multidendritic class III) and six nerve cord neuron types were found to be involved in anemotaxis. The identified neurons were reconstructed in an electron microscopy volume that spans the entire larval nervous system and it was found they received direct input from the mechanosensory neurons or from each other. In this way, local interneurons and first- and second-order subesophageal zone (SEZ) and brain projection neurons were identified. Finally, silencing a dopaminergic brain neuron type impairs anemotaxis. These findings suggest that anemotaxis involves both nerve cord and brain circuits. The candidate neurons and circuitry identified in this study provide a basis for future detailed mechanistic understanding of the circuit principles of anemotaxis. |
| Goda, T., Umezaki, Y., Alwattari, F., Seo, H. W. and Hamada, F. N. (2019). Neuropeptides PDF and DH31 hierarchically regulate free-running rhythmicity in Drosophila circadian locomotor activity. Sci Rep 9(1): 838. PubMed ID: 30696873
Summary: Neuropeptides play pivotal roles in modulating circadian rhythms. Pigment-dispersing factor (PDF) is critical to the circadian rhythms in Drosophila locomotor activity. This study demonstrates that diuretic hormone 31 (DH31) complements PDF function in regulating free-running rhythmicity using male flies. It was determined that Dh31 loss-of-function mutants (Dh31#51)) showed normal rhythmicity, whereas Dh31(#51);Pdf01 double mutants exhibited a severe arrhythmic phenotype compared to Pdf-null mutants (Pdf01). The expression of tethered-PDF or tethered-DH31 in clock cells, posterior dorsal neurons 1 (DN1ps), overcomes the severe arrhythmicity of Dh31(#51);PdfOPdf011 double mutants, suggesting that DH31 and PDF may act on DN1ps to regulate free-running rhythmicity in a hierarchical manner. Unexpectedly, the molecular oscillations in Dh31(#51);Pdf01 mutants were similar to those in Pdf01 mutants in DN1ps, indicating that DH31 does not contribute to molecular oscillations. Furthermore, a reduction in Dh31 receptor (Dh31r) expression resulted in normal locomotor activity and did not enhance the arrhythmic phenotype caused by the Pdf receptor (Pdfr) mutation, suggesting that PDFR, but not DH31R, in DN1ps mainly regulates free-running rhythmicity. Taken together, this study identifies a novel role of DH31, in which DH31 and PDF hierarchically regulate free-running rhythmicity through DN1ps. | Iftikhar, H., Johnson, N. L., Marlatt, M. L. and Carney, G. E. (2019). The role of miRNAs in Drosophila melanogaster male courtship behavior. Genetics. PubMed ID: 30683757
Summary: Drosophila melanogaster courtship, although stereotypical, continually changes based on cues received from the courtship subject. Such adaptive responses are mediated via rapid and widespread transcriptomic reprogramming, a characteristic now widely attributed to microRNAs (miRNAs) along with other players. This study conducted a large-scale miRNA knockout screen to identify miRNAs that affect various parameters of male courtship behavior. Apart from identifying miRNAs that impact male-female courtship, it was observed that miR-957 mutants performed significantly increased male-male courtship and chaining behavior whereby groups of males court one another. The effect of miR-957 reduction in specific neuronal cell clusters was tested, identifying miR-957 activity in Doublesex (DSX)-expressing and mushroom body clusters as an important regulator of male-male courtship interactions. The behavior of miR-957 mutants was further characterized, and these males were found to court male subjects vigorously but do not elicit courtship. Moreover, they fail to lower courtship efforts towards females with higher levels of anti-aphrodisiac pheromones. At the level of individual pheromones, miR-957 males show a reduced inhibitory response to both 7-Tricosene (7-T) and cis-vaccenyl acetate (cVA), with the effect being more pronounced in case of 7-T. Overall these results indicate that a single miRNA can contribute to regulation of complex behaviors, including detection or processing of chemicals that control important survival strategies such as chemical mate-guarding and maintenance of sex and species-specific courtship barriers. |
| Hu, Y., Wang, Z., Liu, T. and Zhang, W. (2019). Piezo-like gene regulates locomotion in Drosophila larvae. Cell Rep 26(6): 1369-1377.e1364. PubMed ID: 30726723
Summary: To maintain proper locomotive patterns, animals constantly monitor body posture with their proprioceptive receptors. In Drosophila, the chordotonal organs (Cho) are especially important in the regulation of locomotion pattern. However, how Cho neurons that are normally activated with sound (vibration) transduce static displacement caused by body position change remains unclear. This study reports that piezo-like (pzl), a homolog for mammalian piezo1 and 2, is essential for Cho's function in locomotion. The mutant allele of pzl showed severe defects in crawling pattern and body gesture control, which were rescued by expressing Pzl specifically in Cho neurons. The ability of Cho neurons to respond to micrometer-scale body wall displacement requires pzl. Intriguingly, human or mouse Piezo1 can rescue pzl-mutant phenotypes, suggesting a conserved role of the Piezo-family proteins in locomotion. | Marie-Orleach, L., Bailey, N. W. and Ritchie, M. G. (2019). Social effects on fruit fly courtship song. Ecol Evol 9(1): 410-416. PubMed ID: 30680123
Summary: Courtship behavior in Drosophila has often been described as a classic innate behavioral repertoire. This study examined courtship song plasticity of two species in the Drosophila melanogaster subgroup. Sexual isolation between the species is influenced by two male song traits, the interpulse interval (IPI) and sinesong frequency (SSF). Neither of these showed plasticity when males had prior experience of con- and heterospecific social partners. However, males of both species produced longer bursts of song during courtship when they were exposed to social partners (either con- or heterospecific) than when they were reared in isolation. D. melanogaster carrying mutations affecting short- or medium-term memory showed a similar response to the social environment, not supporting a role for learning. These results demonstrate that the amount of song a male produces during courtship is plastic depending on the social environment, which might reflect the advantage of being able to respond to variation in intrasexual competition, but that song structure itself is relatively inflexible, perhaps due to strong selection against hybridization. |
Thursday, March 7th - Signaling |
| Le, T., Le, S. C. and Yang, H. (2019). Drosophila Subdued is a moonlighting transmembrane protein 16 (TMEM16) that transports ions and phospholipids. J Biol Chem. PubMed ID: 30700552
Summary: Transmembrane protein 16 (TMEM16) family members play numerous important physiological roles, ranging from controlling membrane excitability and secretion to mediating blood coagulation and viral infection. These diverse functions are largely due to their distinct biophysical properties. Mammalian TMEM16A and TMEM16B are Ca(2+)-activated Cl(-) channels (CaCCs), whereas mammalian TMEM16F, fungal afTMEM16 and nhTMEM16 are moonlighting (multifunctional) proteins with both Ca(2+)-activated phospholipid scramblase (CaPLSase) and Ca(2+)-activated, non-selective ion channel (CAN) activities. To further understand the biological functions of the enigmatic TMEM16 proteins in different organisms, this study, by combining an improved Annexin V-based CaPLSase-imaging assay with inside-out patch clamp technique, thoroughly characterized Subdued, a Drosophila TMEM16 ortholog. Subdued is also a moonlighting transport protein with both CAN and CaPLSase activities. Using a TMEM16F-deficient HEK293T cell line to avoid strong interference from endogenous CaPLSases, the functional characterization and mutagenesis studies revealed that Subdued is a bona fide CaPLSase. Tbe finding that Subdued is a moonlighting TMEM16 expands understanding of the molecular mechanisms of TMEM16 proteins and their evolution and physiology in both Drosophila and humans. | Duan, Y., et al. (2019). PARylation regulates stress granule dynamics, phase separation, and neurotoxicity of disease-related RNA-binding proteins. Cell Res. PubMed ID: 30728452
Summary: Mutations in RNA-binding proteins (RBPs) localized in ribonucleoprotein (RNP) granules, such as hnRNP A1 and TDP-43, promote aberrant protein aggregation, which is a pathological hallmark of various neurodegenerative diseases, such as myotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Protein posttranslational modifications (PTMs) are known to regulate RNP granules. This study investigated the function of poly(ADP-ribosyl)ation (PARylation), an important PTM involved in DNA damage repair and cell death, in RNP granule-related neurodegeneration. PARylation levels are a major regulator of the assembly-disassembly dynamics of RNP granules containing disease-related RBPs, hnRNP A1 and TDP-43. hnRNP A1 can both be PARylated and bind to PARylated proteins or poly(ADP-ribose) (PAR). It was further uncovered that PARylation of hnRNP A1 at K298 controls its nucleocytoplasmic transport, whereas PAR-binding via the PAR-binding motif (PBM) of hnRNP A1 regulates its association with stress granules. Moreover, it was revealed that PAR not only dramatically enhances the liquid-liquid phase separation of hnRNP A1, but also promotes the co-phase separation of hnRNP A1 and TDP-43 in vitro and their interaction in vivo. Finally, both genetic and pharmacological inhibition of PARP mitigates hnRNP A1- and TDP-43-mediated neurotoxicity in cell and Drosophila models of ALS. Together, these findings suggest a novel and crucial role for PARylation in regulating the dynamics of RNP granules, and that dysregulation in PARylation and PAR levels may contribute to ALS disease pathogenesis by promoting protein aggregation. |
| Kreutzberger, A. J. B., Ji, M., Aaron, J., Mihaljevic, L. and Urban, S. (2019). Rhomboid distorts lipids to break the viscosity-imposed speed limit of membrane diffusion. Science 363(6426). PubMed ID: 30705155
Summary: Enzymes that cut proteins inside membranes regulate diverse cellular events, including cell signaling, homeostasis, and host-pathogen interactions. Adaptations that enable catalysis in this exceptional environment are poorly understood. This study visualized single molecules of multiple rhomboid intramembrane proteases and unrelated proteins in living cells (human and Drosophila) and planar lipid bilayers. Notably, only rhomboid proteins were able to diffuse above the Saffman-Delbruck viscosity limit of the membrane. Hydrophobic mismatch with the irregularly shaped rhomboid fold distorted surrounding lipids and propelled rhomboid diffusion. The rate of substrate processing in living cells scaled with rhomboid diffusivity. Thus, intramembrane proteolysis is naturally diffusion-limited, but cells mitigate this constraint by using the rhomboid fold to overcome the "speed limit" of membrane diffusion. | Izore, T., Tailhades, J., Hansen, M. H., Kaczmarski, J. A., Jackson, C. J. and Cryle, M. J. (2019). Drosophila melanogaster nonribosomal peptide synthetase Ebony encodes an atypical condensation domain. Proc Natl Acad Sci U S A. PubMed ID: 30705105
Summary: Drosophila Ebony plays a central role in the regulation of histamine and dopamine in various tissues through condensation of these amines with beta-alanine. Ebony is a rare example of a nonribosomal peptide synthetase (NRPS) from a higher eukaryote and contains a C-terminal sequence that does not correspond to any previously characterized NRPS domain. This study has structurally characterized this C-terminal domain and have discovered that it adopts the aryl-alkylamine-N-acetyl transferase (AANAT) fold, which is unprecedented in NRPS biology. Through analysis of ligand-bound structures, activity assays, and binding measurements, it was determined how this atypical condensation domain is able to provide selectivity for both the carrier protein-bound amino acid and the amine substrates, a situation that remains unclear for standard condensation domains identified to date from NRPS assembly lines. These results demonstrate that the C terminus of Ebony encodes a eukaryotic example of an alternative type of NRPS condensation domain; they also illustrate how the catalytic components of such assembly lines are significantly more diverse than a minimal set of conserved functional domains. |
| Li, J. S. S. and Sean Millard, S. (2019). Deterministic splicing of Dscam2 is regulated by Muscleblind. Sci Adv 5(1): eaav1678. PubMed ID: 30746474
Summary: Alternative splicing increases the proteome diversity crucial for establishing the complex circuitry between trillions of neurons. To provide individual cells with different repertoires of protein isoforms, however, this process must be regulated. Previously, it was found that the mutually exclusive alternative splicing of Drosophila Dscam2 produces two isoforms (A and B) with unique binding properties. This splicing event is cell type specific, and the transmembrane proteins that it generates are crucial for the development of axons, dendrites, and synapses. This study shows that Muscleblind (Mbl) controls Dscam2 alternative splicing. Mbl represses isoform A and promotes the selection of isoform B. Mbl mutants exhibit phenotypes also observed in flies engineered to express a single Dscam2 isoform. Consistent with this, mbl expression is cell type specific and correlates with the splicing of isoform B. This study demonstrates how the regulated expression of a splicing factor is sufficient to provide neurons with unique protein isoforms crucial for development. | Lu, F., Portz, B. and Gilmour, D. S. (2019). The C-terminal domain of RNA polymerase II is a multivalent targeting sequence that supports Drosophila development with only consensus heptads. Mol Cell. PubMed ID: 30765194
Summary: The C-terminal domain (CTD) of RNA polymerase II (Pol II) is composed of repeats of the consensus YSPTSPS and is an essential binding scaffold for transcription-associated factors. Metazoan CTDs have well-conserved lengths and sequence compositions arising from the evolution of divergent motifs, features thought to be essential for development. On the contrary, this study shows that a truncated CTD composed solely of YSPTSPS repeats supports Drosophila viability but that a CTD with enough YSPTSPS repeats to match the length of the wild-type Drosophila CTD is defective. Furthermore, a fluorescently tagged CTD lacking the rest of Pol II dynamically enters transcription compartments, indicating that the CTD functions as a signal sequence. However, CTDs with too many YSPTSPS repeats are more prone to localize to static nuclear foci separate from the chromosomes. It is proposed that the sequence complexity of the CTD offsets aberrant behavior caused by excessive repetitive sequences without compromising its targeting function. |
Wednesday, March 6th - Disease models |
| Chen, Z. S., Wong, A. K. Y., Cheng, T. C., Koon, A. C. and Chan, H. Y. E. (2019). FipoQ/FBXO33, a Cullin-1 based ubiquitin ligase complex component modulates ubiquitination and solubility of polyglutamine disease protein. J Neurochem. PubMed ID: 30685895
Summary: Polyglutamine (polyQ) diseases describe a group of progressive neurodegenerative disorders caused by the CAG triplet repeat expansion in the coding region of the disease genes. To date, nine such diseases, including spinocerebellar ataxia type 3 (SCA3), have been reported. The formation of SDS-insoluble protein aggregates in neurons causes cellular dysfunctions, such as impairment of the ubiquitin-proteasome system (UPS), and contributes to polyQ pathologies. Recently, the E3 ubiquitin ligases, which govern substrate specificity of the UPS, have been implicated in polyQ pathogenesis. The Cullin (Cul) proteins are major components of Cullin-RING ubiquitin ligases (CRLs) complexes that are evolutionarily conserved in the Drosophila genome. This study examined the effect of individual Culs on SCA3 pathogenesis, and found that the knockdown of Cul1 expression enhances SCA3-induced neurodegeneration and reduces the solubility of expanded SCA3-polyQ proteins. The F-box proteins are substrate receptors of Cul1-based CRL. A genetic modifier screen of the 19 Drosophila F-box genes was performed, and F-box involved in polyQ pathogenesis (FipoQ) was identified as a genetic modifier of SCA3 degeneration that modulates the ubiquitination and solubility of expanded SCA3-polyQ proteins. In the human SK-N-MC cell model, F-box only protein 33 (FBXO33) exerts similar functions as FipoQ in modulating the ubiquitination and solubility of expanded SCA3-polyQ proteins. Taken together, this study demonstrates that Cul1-based CRL and its associated F-box protein, FipoQ/FBXO33, modify SCA3 protein toxicity. These findings will lead to a better understanding of the disease mechanism of SCA3, and provide insights on developing treatments against SCA3. | Hwang, S., Jeong, H., Hong, E. H., Joo, H. M., Cho, K. S. and Nam, S. Y. (2019). Low-dose ionizing radiation alleviates Abeta42-induced cell death via regulating AKT and p38 pathways in Drosophila Alzheimer's disease models. Biol Open 8(2). PubMed ID: 30670376
Summary: Ionizing radiation is widely used in medicine and is valuable in both the diagnosis and treatment of many diseases. However, its health effects are ambiguous. This paper reports that low-dose ionizing radiation has beneficial effects in human amyloid-beta42 (Abeta42)-expressing Drosophila Alzheimer's disease (AD) models. Ionizing radiation at a dose of 0.05 Gy suppressed AD-like phenotypes, including developmental defects and locomotive dysfunction, but did not alter the decreased survival rates and longevity of Abeta42-expressing flies. The same dose of gamma-irradiation reduced Abeta42-induced cell death in Drosophila AD models through downregulation of head involution defective (hid), which encodes a protein that activates caspases. However, 4 Gy of gamma-irradiation increased Abeta42-induced cell death without modulating pro-apoptotic genes grim, reaper and hid. The AKT signaling pathway, which was suppressed in Drosophila AD models, was activated by either 0.05 or 4 Gy gamma-irradiation. Interestingly, p38 mitogen-activated protein-kinase (MAPK) activity was inhibited by exposure to 0.05 Gy gamma-irradiation but enhanced by exposure to 4 Gy in Abeta42-expressing flies. In addition, overexpression of phosphatase and tensin homolog (PTEN), a negative regulator of the AKT signaling pathway, or a null mutant of AKT strongly suppressed the beneficial effects of low-dose ionizing radiation in Abeta42-expressing flies. These results indicate that low-dose ionizing radiation suppresses Abeta42-induced cell death through regulation of the AKT and p38 MAPK signaling pathways, suggesting that low-dose ionizing radiation has hormetic effects on the pathogenesis of Abeta42-associated AD. |
| Hansen, T., Thant, C., White, J. A., Banerjee, R., Thuamsang, B. and Gunawardena, S. (2019). Excess active P13K rescues huntingtin-mediated neuronal cell death but has no effect on axonal transport defects. Apoptosis. PubMed ID: 30725352
Summary: High levels of oxidative stress is detected in neurons affected by many neurodegenerative diseases, including huntington's disease. Many of these diseases also show neuronal cell death and axonal transport defects. While nuclear inclusions/accumulations likely cause cell death, previous work has shown that cytoplasmic axonal accumulations can also contribute to neuronal death. However, the cellular mechanisms responsible for activating cell death is unclear. One possibility is that perturbations in normal axonal transport alter the function of the phosphatidylinositol 3-kinase (PI3K)-protein kinase B (AKT)-pathway, a signal transduction pathway that promotes survival/growth in response to extracellular signals. To test this proposal in vivo, active PI3K was expressed in the context of pathogenic huntingtin (HTT-138Q) in Drosophila larval nerves, which show axonal transport defects and neuronal cell death. Excess expression of active P13K significantly suppressed HTT-138Q-mediated neuronal cell death, but had no effect on HTT-138Q-mediated axonal transport defects. Expression of active PI3K also rescued Paraquat-mediated cell death. Further, increased levels of pSer9 (inactive) glycogen synthase kinase 3beta was seen in HTT-138Q-mediated larval brains, and in dynein loss of function mutants, indicating the modulation of the pro-survival pathway. Intriguingly, proteins in the PI3K/AKT-pathway showed functional interactions with motor proteins. Taken together these observations suggest that proper axonal transport is likely essential for the normal function of the pro-survival PI3K/AKT-signaling pathway and for neuronal survival in vivo. These results have important implications for targeting therapeutics to early insults during neurodegeneration and death. | Hajji, K., Mteyrek, A., Sun, J., Cassar, M., Mezghani, S., Leprince, J., Vaudry, D., Masmoudi-Kouki, O. and Birman, S. (2019). Neuroprotective effects of PACAP against paraquat-induced oxidative stress in the Drosophila central nervous system. Hum Mol Genet. PubMed ID: 30715303
Summary: Parkinson's disease (PD) is a progressive neurodegenerative movement disorder that can arise after long-term exposure to environmental oxidative stressors, such as the herbicide paraquat (PQ). This study investigated the potential neuroprotective action of vertebrate pituitary adenylate cyclase-activating polypeptide (PACAP) against PQ in Drosophila. Pretreatment with this neuropeptide applied to the ventral nerve cord (VNC) at low doses markedly extended the survival of wild-type decapitated flies exposed to neurotoxic levels of PQ or dopamine (DA). In contrast and interestingly, application of a PACAP receptor antagonist, PACAP-6-38, had opposite effects, significantly decreasing the resistance of flies to PQ. PACAP also reduced PQ-induced caspase activation and reactive oxygen species (ROS) accumulation in the VNC. This study sought the endogenous neuropeptide receptor potentially involved in PACAP-mediated neuroprotection in Drosophila. Knocking down the gene encoding the receptor Han/PDFR of the neuropeptide pigment-dispersing factor (PDF) in all neurons conferred to flies higher resistance to PQ, whereas PDFR downregulation restricted to PDF or DA neurons did not increase PQ resistance, but remarkably suppressed the neuroprotective action of PACAP. Further experiments performed with Pdf and Pdfr-deficient mutant strains confirmed that PDF and its receptor are required for PACAP-mediated neuroprotection in flies. Evidence using split-GFP reconstitution is provided that PDF neurons make synaptic contacts onto DA neurons in the abdominal VNC. These results, therefore, suggest that the protective action of PACAP against PQ-induced defects in the Drosophila nervous system involves the modulation of PDFR signaling in a small number of interconnected neurons. |
| Lowe, S. A., Usowicz, M. M. and Hodge, J. J. L. (2019). Neuronal overexpression of Alzheimer's disease and Down's syndrome associated DYRK1A/minibrain gene alters motor decline. neurodegeneration and synaptic plasticity in Drosophila. Neurobiol Dis 125: 107-114. PubMed ID: 30703437
Summary: Down syndrome (DS) is characterised by abnormal cognitive and motor development, and later in life by progressive Alzheimer's disease (AD)-like dementia, neuropathology, declining motor function and shorter life expectancy. It is caused by trisomy of chromosome 21 (Hsa21), but how individual Hsa21 genes contribute to various aspects of the disorder is incompletely understood. Previous work has demonstrated a role for triplication of the Hsa21 gene DYRK1A in cognitive and motor deficits, as well as in altered neurogenesis and neurofibrillary degeneration in the DS brain, but its contribution to other DS phenotypes is unclear. This study demonstrates that overexpression of minibrain (mnb), the Drosophila ortholog of DYRK1A, in the Drosophila nervous system accelerated age-dependent decline in motor performance and shortened lifespan. Overexpression of mnb in the eye was neurotoxic and overexpression in ellipsoid body neurons in the brain caused age-dependent neurodegeneration. At the larval neuromuscular junction, an established model for mammalian central glutamatergic synapses, neuronal mnb overexpression enhanced spontaneous vesicular transmitter release. It also slowed recovery from short-term depression of evoked transmitter release induced by high-frequency nerve stimulation and increased the number of boutons in one of the two glutamatergic motor neurons innervating the muscle. These results provide further insight into the roles of DYRK1A triplication in abnormal aging and synaptic dysfunction in DS. | Campbell, N. G., Shekar, A., Aguilar, J. I., Peng, D., Navratna, V., Yang, D., Morley, A. N., Duran, A. M., Galli, G., O'Grady, B., Ramachandran, R., Sutcliffe, J. S., Sitte, H. H., Erreger, K., Meiler, J., Stockner, T., Bellan, L. M., Matthies, H. J. G., Gouaux, E., McHaourab, H. S. and Galli, A. (2019). Structural, functional, and behavioral insights of dopamine dysfunction revealed by a deletion in SLC6A3. Proc Natl Acad Sci U S A. PubMed ID: 30755521
Summary: The human dopamine (DA) transporter (hDAT) mediates clearance of DA. Genetic variants in hDAT have been associated with DA dysfunction, a complication associated with several brain disorders, including autism spectrum disorder (ASD). This study investigated the structural and behavioral bases of an ASD-associated in-frame deletion in hDAT at N336 (N336). The deletion promoted a previously unobserved conformation of the intracellular gate of the transporter, likely representing the rate-limiting step of the transport process. It is defined by a "half-open and inward-facing" state (HOIF) of the intracellular gate that is stabilized by a network of interactions conserved phylogenetically, as was demonstrated in hDAT by Rosetta molecular modeling and fine-grained simulations, as well as in its bacterial homolog leucine transporter by electron paramagnetic resonance analysis and X-ray crystallography. The stabilization of the HOIF state is associated both with DA dysfunctions demonstrated in isolated brains of Drosophila melanogaster expressing hDAT N336 and with abnormal behaviors observed at high-time resolution. These flies display increased fear, impaired social interactions, and locomotion traits were associate with DA dysfunction and the HOIF state. Together, these results describe how a genetic variation causes DA dysfunction and abnormal behaviors by stabilizing a HOIF state of the transporter. |
Tuesday, March 5th - Evolution |
| Barghi, N., Tobler, R., Nolte, V., Jaksic, A. M., Mallard, F., Otte, K. A., Dolezal, M., Taus, T., Kofler, R. and Schlotterer, C. (2019). Genetic redundancy fuels polygenic adaptation in Drosophila. PLoS Biol 17(2): e3000128. PubMed ID: 30716062
Summary: The genetic architecture of adaptive traits is of key importance to predict evolutionary responses. Most adaptive traits are polygenic-i.e., result from selection on a large number of genetic loci-but most molecularly characterized traits have a simple genetic basis. This discrepancy is best explained by the difficulty in detecting small allele frequency changes (AFCs) across many contributing loci. To resolve this, laboratory natural selection was used to detect signatures for selective sweeps and polygenic adaptation. This study exposed 10 replicates of a Drosophila simulans population to a new temperature regime and uncovered a polygenic architecture of an adaptive trait with high genetic redundancy among beneficial alleles. Convergent responses were observed for several phenotypes-e.g., fitness, metabolic rate, and fat content-and a strong polygenic response (99 selected alleles; mean s = 0.059). However, each of these selected alleles increased in frequency only in a subset of the evolving replicates. Different evolutionary paradigms were discerned based on the heterogeneous genomic patterns among replicates. Redundancy and quantitative trait (QT) paradigms fitted the experimental data better than simulations assuming independent selective sweeps. These results show that natural D. simulans populations harbor a vast reservoir of adaptive variation facilitating rapid evolutionary responses using multiple alternative genetic pathways converging at a new phenotypic optimum. This key property of beneficial alleles requires the modification of testing strategies in natural populations beyond the search for convergence on the molecular level. | Ellison, C. and Bachtrog, D. (2019). Contingency in the convergent evolution of a regulatory network: Dosage compensation in Drosophila. PLoS Biol 17(2): e3000094. PubMed ID: 30742611
Summary: The repeatability or predictability of evolution is a central question in evolutionary biology and most often addressed in experimental evolution studies. This study inferred how genetically heterogeneous natural systems acquire the same molecular changes to address how genomic background affects adaptation in natural populations. In particular, advantage was taken of independently formed neo-sex chromosomes in Drosophila species that have evolved dosage compensation by co-opting the dosage-compensation male-specific lethal (MSL) complex to study the mutational paths that have led to the acquisition of hundreds of novel binding sites for the MSL complex in different species. This complex recognizes a conserved 21-bp GA-rich sequence motif that is enriched on the X chromosome, and newly formed X chromosomes recruit the MSL complex by de novo acquisition of this binding motif. Recently formed sex chromosomes were identified in the D. melanica and D. robusta species groups by genome sequencing and generate genomic occupancy maps of the MSL complex to infer the location of novel binding sites. Diverse mutational paths were utilized in each species to evolve hundreds of de novo binding motifs along the neo-X, including expansions of microsatellites and transposable element (TE) insertions. However, the propensity to utilize a particular mutational path differs between independently formed X chromosomes and appears to be contingent on genomic properties of that species, such as simple repeat or TE density. This establishes the "genomic environment" as an important determinant in predicting the outcome of evolutionary adaptations. |
| Arguello, J. R., Laurent, S. and Clark, A. G. (2019). Demographic history of the human commensal Drosophila melanogaster. Genome Biol Evol. PubMed ID: 30715331
Summary: The cohabitation of Drosophila melanogaster with humans is nearly ubiquitous. Though it has been well-established that this fly species originated in sub-Saharan Africa, and only recently has spread globally, many details of its swift expansion remain unclear. Elucidating the demographic history of D. melanogaster provides a unique opportunity to investigate how human movement might have impacted patterns of genetic diversity in a commensal species, as well as providing neutral null models for studies aimed at identifying genomic signatures of local adaptation. This study used whole-genome data from five populations (Africa, North America, Europe, Central Asia, and the South Pacific) to carry out demographic inferences, with particular attention to the inclusion of migration and admixture. The importance of these parameters for model fitting is demonstrated, and how previous estimates of divergence times are likely to be significantly underestimated as a result of not including them is shown. Finally, how human movement along early shipping routes might have shaped the present-day population structure of D. melanogaster is discussed. | Karr, T. L., Southern, H., Rosenow, M. A., Gossmann, T. I. and Snook, R. R. (2019). The old and the new: discovery proteomics identifies putative novel seminal fluid proteins in Drosophila. Mol Cell Proteomics. PubMed ID: 30760537
Summary: Seminal fluid proteins (SFPs), the non-sperm component of male ejaculates produced by male accessory glands, are viewed as central mediators of reproductive fitness. SFPs effect both male and female post-mating functions and show molecular signatures of rapid adaptive evolution. While SFP identification was historically challenging, advances in label-free quantitative proteomics expands the scope of studying other systems to further advance the field. This study applied label-free quantitative proteomics to identify the accessory gland proteome and secretome in Drosophila pseudoobscura, a close relative of D. melanogaster, and use the dataset to identify both known and putative novel SFPs. Using this approach 163 putative SFPs were identified, 32% of which overlapped with previously identified D. melanogaster SFPs. SFPs with known extracellular annotation evolve more rapidly than other proteins produced by or contained within the accessory gland. These results will further the understanding of the evolution of SFPs and the underlying male accessory gland proteins that mediate reproductive fitness of the sexes. |
| Da Lage, J. L., Thomas, G. W. C., Bonneau, M. and Courtier-Orgogozo, V. (2019). Evolution of salivary glue genes in Drosophila species. BMC Evol Biol 19(1): 36. PubMed ID: 30696414
Summary: At the very end of the larval stage Drosophila expectorate a glue secreted by their salivary glands to attach themselves to a substrate while pupariating. The glue is a mixture of apparently unrelated proteins, some of which are highly glycosylated and possess internal repeats. Because species adhere to distinct substrates (i.e. leaves, wood, rotten fruits), glue genes are expected to evolve rapidly. Available genome sequences and PCR-sequencing of regions of interest were used to investigate the glue genes in 20 Drosophila species. A new gene in addition was discovered to the seven glue genes annotated in D. melanogaster. A phase 1 intron was identified at a conserved position present in five of the eight glue genes of D. melanogaster, suggesting a common origin for those glue genes. A slightly significant rate of gene turnover was inferred. Both the number of repeats and the repeat sequence were found to diverge rapidly, even between closely related species. High repeat number variation was also detected at the intrapopulation level in D. melanogaster. It is concluded that most conspicuous signs of accelerated evolution are found in the repeat regions of several glue genes. | Hsu, S. K., Jaksic, A. M., Nolte, V., Barghi, N., Mallard, F., Otte, K. A. and Schlotterer, C. (2019). A 24 h age difference causes twice as much gene expression divergence as 100 generations of adaptation to a novel environment. Genes (Basel) 10(2). PubMed ID: 30696109
Summary: Gene expression profiling is one of the most reliable high-throughput phenotyping methods, allowing researchers to quantify the transcript abundance of expressed genes. Because many biotic and abiotic factors influence gene expression, it is recommended to control them as tightly as possible. This study shows that a 24 h age difference of Drosophila simulans females that were subjected to RNA sequencing (RNA-Seq) five and six days after eclosure resulted in more than 2000 differentially expressed genes. This is twice the number of genes that changed expression during 100 generations of evolution in a novel hot laboratory environment. Importantly, most of the genes differing in expression due to age introduce false positives or negatives if an adaptive gene expression analysis is not controlled for age. These results indicate that tightly controlled experimental conditions, including precise developmental staging, are needed for reliable gene expression analyses, in particular in an evolutionary framework. |
Monday, March 4th - Synapse and Vesicles |
| Goel, P., Li, X. and Dickman, D. (2019). Estimation of the readily releasable synaptic vesicle pool at the Drosophila larval neuromuscular junction. Bio Protoc 9(1). PubMed ID: 30761328
Summary: Presynaptic boutons at nerve terminals are densely packed with synaptic vesicles, specialized organelles for rapid and regulated neurotransmitter secretion. Upon depolarization of the nerve terminal, synaptic vesicles fuse at specializations called active zones that are localized at discrete compartments in the plasma membrane to initiate synaptic transmission. A small proportion of synaptic vesicles are docked and primed for immediate fusion upon synaptic stimulation, which together comprise the readily releasable pool. The size of the readily releasable pool is an important property of synapses, which influences release probability and can dynamically change during various forms of plasticity. A detailed protocol is described for estimating the readily releasable pool at a model glutamatergic synapse, the Drosophila neuromuscular junction. This synapse is experimentally robust and amenable to sophisticated genetic, imaging, electrophysiological, and pharmacological approaches. The experimental design, electrophysiological recording procedure, and quantitative analysis that is necessary to determine the readily releasable pool size is described in this paper. This technique requires the use of a two-electrode voltage-clamp recording configuration in elevated external Ca(2+) with high frequency stimulation. This assay is sused to measure the readily releasable pool size and reveal that a form of homeostatic plasticity modulates this pool with synapse-specific and compartmentalized precision. This powerful approach can be utilized to illuminate the dynamics of synaptic vesicle trafficking and plasticity and determine how synaptic function adapts and deteriorates during states of altered development, stress and neuromuscular disease. | Gratz, S. J., Goel, P., Bruckner, J. J., Hernandez, R. X., Khateeb, K., Macleod, G. T., Dickman, D. and O'Connor-Giles, K. M. (2019). Endogenous tagging reveals differential regulation of Ca(2+) channels at single AZs during presynaptic homeostatic potentiation and depression. J Neurosci. PubMed ID: 30692227
Summary: Neurons communicate through Ca(2+)-dependent neurotransmitter release at presynaptic active zones (AZs). Neurotransmitter release properties play a key role in defining information flow in circuits and are tuned during multiple forms of plasticity. Despite their central role in determining neurotransmitter release properties, little is known about how Ca(2+) channel levels are modulated to calibrate synaptic function. This study used CRISPR to tag the Drosophila CaV2 Ca(2+) channel Cacophony (Cac) and, in males in which all endogenous Cac channels are tagged, investigated the regulation of endogenous Ca(2+) channels during homeostatic plasticity. Heterogeneously distributed Cac was found to be highly predictive of neurotransmitter release probability at individual AZs and differentially regulated during opposing forms of presynaptic homeostatic plasticity. Specifically, AZ Cac levels are increased during chronic and acute presynaptic homeostatic potentiation (PHP), and live imaging during acute expression of PHP reveals proportional Ca(2+) channel accumulation across heterogeneous AZs. In contrast, endogenous Cac levels do not change during presynaptic homeostatic depression (PHD), implying that the reported reduction in Ca(2+) influx during PHD is achieved through functional adaptions to pre-existing Ca(2+) channels. Thus, distinct mechanisms bi-directionally modulate presynaptic Ca(2+) levels to maintain stable synaptic strength in response to diverse challenges, with Ca(2+) channel abundance providing a rapidly tunable substrate for potentiating neurotransmitter release over both acute and chronic timescales. |
| Wasserman, S. S., Shteiman-Kotler, A., Harris, K., Iliadi, K. G., Persaud, A., Zhong, Y., Zhang, Y., Fang, X., Boulianne, G. L., Stewart, B. and Rotin, D. (2018). Regulation of SH3PX1 by dNedd4-long at the Drosophila Neuromuscular Junction. J Biol Chem. PubMed ID: 30518551
Summary: Drosophila Nedd4 (dNedd4) is a HECT E3 ubiquitin ligase present in two major isoforms: short (dNedd4S) and long (dNedd4Lo), with the latter containing two unique regions (N-terminus and Middle). While dNedd4S promotes neuromuscular synaptogenesis (NMS), dNedd4Lo inhibits it and impairs larval locomotion. To explain how dNedd4Lo inhibits NMS, mass spectrometry was performed to find its binding partners and identified SH3PX1, which binds dNedd4Lo unique Middle region. SH3PX1 contains SH3, PX and BAR domains and is present at neuromuscular junctions, where it regulates active zone ultrastructure and presynaptic neurotransmitter release. This study demonstrates direct binding of SH3PX1 to the dNedd4Lo Middle region (which contains a Pro rich sequence) in vitro and in cells, via the SH3PX1-SH3 domain. In Drosophila S2 cells, dNedd4Lo overexpression reduces SH3PX1 levels at the cell periphery. In vivo overexpression of dNedd4Lo post-synaptically, but not pre-synaptically, reduces SH3PX1 levels at the subsynaptic reticulum and impairs neurotransmitter release. Unexpectedly, larvae that overexpress dNedd4Lo post-synaptically and are heterozygous for a null mutation in SH3PX1 display increased neurotransmission compared to dNedd4Lo or SH3PX1 mutant larvae alone, suggesting a compensatory effect from the remaining SH3PX1 allele. These results suggest a postsynaptic - specific regulation of SH3PX1 by dNedd4Lo. | Jantrapirom, S., Cao, D., Wang, J., Hing, H., Tabone, C. J., Lantz, K., de Belle, J. S., Qiu, Y. T., Smid, H. M., Yamaguchi, M., Noordermeer, J. N., Fradkin, L. G. and Potikanond, S. (2019). Dystrophin is required for normal synaptic gain in the Drosophila olfactory circuit. Brain Res. PubMed ID: 30711401
Summary: The Drosophila olfactory system provides an excellent model to elucidate the neural circuits that control behaviors elicited by environmental stimuli. Despite significant progress in defining olfactory circuit components and their connectivity, little is known about the mechanisms that transfer the information from the primary antennal olfactory receptor neurons to the higher order brain centers. This study shows that the Dystrophin Dp186 isoform is required in the olfactory system circuit for olfactory functions. Using two-photon calcium imaging, this study found the reduction of calcium influx in olfactory receptor neurons (ORNs) and also the defect of GABAA mediated inhibitory input in the projection neurons (PNs) in Dp186 mutation. Moreover, the Dp186 mutant flies which display a decreased odor avoidance behavior were rescued by Dp186 restoration in the Drosophila olfactory neurons in either the presynaptic ORNs or the postsynaptic PNs. Therefore, these results revealed a role for Dystrophin, Dp 186 isoform in gain control of the olfactory synapse via the modulation of excitatory and inhibitory synaptic inputs to olfactory projection neurons. |
| Williams, J. L., Shearin, H. K. and Stowers, R. S. (2019). Conditional synaptic vesicle markers for Drosophila. G3 (Bethesda). PubMed ID: 30635441
Summary: The release of neurotransmitters from synaptic vesicles (SVs) at pre-synaptic release sites is the principle means by which information transfer between neurons occurs. Knowledge of the location of SVs within a neuron can thus provide valuable clues about the location of neurotransmitter release within a neuron and the downstream neurons to which a given neuron is connected, thus providing important information for understanding how neural circuits generate behavior. This study presents the development and characterization of four conditional tagged SV markers for Drosophila melanogaster. This characterization includes evaluation of conditionality, specificity for SV localization, and sensitivity of detection in diverse neuron subtypes. These four SV markers are genome-edited variants of the synaptic vesicle-specific protein Rab3. They depend on either the B2 or FLP recombinases for conditionality, and incorporate GFP or mCherry fluorescent proteins, or FLAG or HA epitope tags, for detection. | Perez-Moreno, J. J. and O'Kane, C. J. (2018). GAL4 drivers specific for type Ib and type Is motor neurons in Drosophila. G3 (Bethesda). PubMed ID: 30530644
Summary: The Drosophila melanogaster larval neuromuscular system is extensively used by researchers to study neuronal cell biology, and Drosophila glutamatergic motor neurons have become a major model system. There are two main types of glutamatergic motor neurons, Ib and Is, with different structural and physiological properties at synaptic level at the neuromuscular junction. To generate genetic tools to identify and manipulate motor neurons of each Type, GAL4 driver lines were screened for this purpose. This study describes GAL4 drivers specific for examples of neurons within each Type, Ib or Is. These drivers showed high expression levels and were expressed in only few motor neurons, making them amenable tools for specific studies of both axonal and synapse biology in identified Type I motor neurons. |
Friday, March 1st - Neural Development and Function |
| Golovin, R. M., Vest, J., Vita, D. J. and Broadie, K. (2019). Activity-dependent remodeling of Drosophila olfactory sensory neuron brain innervation during an early-Life critical period. J Neurosci. PubMed ID: 30755492
Summary: Critical periods are windows of development when the environment has a pronounced effect on brain circuitry. Models of neurodevelopmental disorders including autism spectrum disorders, intellectual disabilities and schizophrenia are linked to disruption of critical period remodeling. Critical periods open with the onset of sensory experience, however it remains unclear exactly how sensory input modifies brain circuits. This study examined olfactory sensory neuron (OSN) innervation of the Drosophila antennal lobe (AL) of both sexes as a genetic model of this question. Olfactory sensory experience during an early-use critical period was shown to drives loss of OSN innervation of AL glomeruli and subsequent axon retraction in a dose-dependent mechanism. This remodeling does not result from olfactory receptor loss or OSN degeneration, but rather from rapid synapse elimination and axon pruning in the target olfactory glomerulus. Removal of the odorant stimulus only during the critical period leads to OSN re-innervation, demonstrating remodeling is transiently reversible. This synaptic refinement requires the OSN-specific olfactory receptor and downstream activity. Conversely, blocking OSN synaptic output elevates glomeruli remodeling. GABAergic neurotransmission has no detectable role, but glutamatergic signaling via NMDA receptors is required for OSN synaptic refinement. Altogether, these results demonstrate that OSN inputs into the brain manifest robust, experience-dependent remodeling during an early-life critical period, which requires olfactory reception, OSN activity and NMDA receptor signaling. This work reveals a pathway linking initial olfactory sensory experience to glutamatergic neurotransmission in the activity-dependent remodeling of brain neural circuitry in an early-use critical period. | Castells-Nobau, A., Eidhof, I., Fenckova, M., Brenman-Suttner, D. B., Scheffer-de Gooyert, J. M., Christine, S., Schellevis, R. L., van der Laan, K., Quentin, C., van Ninhuijs, L., Hofmann, F., Ejsmont, R., Fisher, S. E., Kramer, J. M., Sigrist, S. J., Simon, A. F. and Schenck, A. (2019). Conserved regulation of neurodevelopmental processes and behavior by FoxP in Drosophila. PLoS One 14(2): e0211652. PubMed ID: 30753188
Summary: FOXP proteins form a subfamily of evolutionarily conserved transcription factors involved in the development and functioning of several tissues, including the central nervous system. In humans, mutations in FOXP1 and FOXP2 have been implicated in cognitive deficits including intellectual disability and speech disorders. Drosophila exhibits a single ortholog, called FoxP, but due to a lack of characterized mutants, understanding of the gene remains poor. This study shows that the dimerization property required for mammalian FOXP function is conserved in Drosophila. In flies, FoxP is enriched in the adult brain, showing strong expression in ~1000 neurons of cholinergic, glutamatergic and GABAergic nature. Drosophila loss-of-function mutants and UAS-FoxP transgenic lines were generated for ectopic expression, and they were used to characterize FoxP function in the nervous system. At the cellular level, it was demonstrated that Drosophila FoxP is required in larvae for synaptic morphogenesis at axonal terminals of the neuromuscular junction and for dendrite development of dorsal multidendritic sensory neurons. In the developing brain, FoxP plays important roles in alpha-lobe mushroom body formation. Finally, at a behavioral level, this study showed that Drosophila FoxP is important for locomotion, habituation learning and social space behavior of adult flies. This work shows that Drosophila FoxP is important for regulating several neurodevelopmental processes and behaviors that are related to human disease or vertebrate disease model phenotypes. This suggests a high degree of functional conservation with vertebrate FOXP orthologues and established flies as a model system for understanding FOXP related pathologies. |
| Cheng, S., Ashley, J., Kurleto, J. D., Lobb-Rabe, M., Park, Y. J., Carrillo, R. A. and Ozkan, E. (2019). Molecular basis of synaptic specificity by immunoglobulin superfamily receptors in Drosophila. Elife 8. PubMed ID: 30688651
Summary: In stereotyped neuronal networks, synaptic connectivity is dictated by cell surface proteins, which assign unique identities to neurons, and physically mediate axon guidance and synapse targeting. Two groups of immunoglobulin superfamily proteins in Drosophila, Dprs and DIPs, have been identified as strong candidates for synapse targeting functions. This study uncovers the molecular basis of specificity in Dpr-DIP mediated cellular adhesions and neuronal connectivity. First, five crystal structures of Dpr-DIP and DIP-DIP complexes are presented, highlighting the evolutionary and structural origins of diversification in Dpr and DIP proteins and their interactions. It was further shown that structures can be used to rationally engineer receptors with novel specificities or modified affinities, which can be used to study specific circuits that require Dpr-DIP interactions to help establish connectivity. This study investigated one pair, engineered Dpr10 and DIP-alpha, for function in the neuromuscular circuit in flies, and reveal roles for homophilic and heterophilic binding in wiring (Cheng, 2019) | Chai, P. C., Cruchet, S., Wigger, L. and Benton, R. (2019). Sensory neuron lineage mapping and manipulation in the Drosophila olfactory system. Nat Commun 10(1): 643. PubMed ID: 30733440
Summary: Nervous systems exhibit myriad cell types, but understanding how this diversity arises is hampered by the difficulty to visualize and genetically-probe specific lineages, especially at early developmental stages prior to expression of unique molecular markers. This study used a genetic immortalization method to analyze the development of sensory neuron lineages in the Drosophila olfactory system, from their origin to terminal differentiation. This approach was applied to define a fate map of nearly all olfactory lineages and refine the model of temporal patterns of lineage divisions. Taking advantage of a selective marker for the lineage that gives rise to Or67d pheromone-sensing neurons and a genome-wide transcription factor RNAi screen, the spatial and temporal requirements for Pointed, an ETS family member, was identified in this developmental pathway. Transcriptomic analysis of wild-type and Pointed-depleted olfactory tissue reveals a universal requirement for this factor as a switch-like determinant of fates in these sensory lineages. |
| Ashley, J., Sorrentino, V., Lobb-Rabe, M., Nagarkar-Jaiswal, S., Tan, L., Xu, S., Xiao, Q., Zinn, K. and Carrillo, R. A. (2019). Transsynaptic interactions between IgSF proteins DIP-alpha and Dpr10 are required for motor neuron targeting specificity. Elife 8. PubMed ID: 30714906
Summary: The Drosophila larval neuromuscular system provides an ideal context in which to study synaptic partner choice, because it contains a small number of pre- and postsynaptic cells connected in an invariant pattern. The discovery of interactions between two subfamilies of IgSF cell surface proteins, the Dprs and the DIPs, provided new candidates for cellular labels controlling synaptic specificity. This study shows that DIP-alpha is expressed by two identified motor neurons, while its binding partner Dpr10 is expressed by postsynaptic muscle targets. Removal of either DIP-alpha or Dpr10 results in loss of specific axonal branches and NMJs formed by one motor neuron, MNISN-1s, while other branches of the MNISN-1s axon develop normally. The temporal and spatial expression pattern of dpr10 correlates with muscle innervation by MNISN-1s during embryonic development. A model is presented whereby DIP-alpha and Dpr10 on opposing synaptic partners interact with each other to generate proper motor neuron connectivity (Ashley, 2019). | Komori, H., Golden, K. L., Kobayashi, T., Kageyama, R. and Lee, C. Y. (2018). Multilayered gene control drives timely exit from the stem cell state in uncommitted progenitors during Drosophila asymmetric neural stem cell division. Genes Dev 32(23-24): 1550-1561. PubMed ID: 30463902
Summary: Self-renewal genes maintain stem cells in an undifferentiated state by preventing the commitment to differentiate. Robust inactivation of self-renewal gene activity following asymmetric stem cell division allows uncommitted stem cell progeny to exit from an undifferentiated state and initiate the commitment to differentiate. Nonetheless, how self-renewal gene activity at mRNA and protein levels becomes synchronously terminated in uncommitted stem cell progeny is unclear. This study demonstrates that a multilayered gene regulation system terminates self-renewal gene activity at all levels in uncommitted stem cell progeny in the fly neural stem cell lineage. The RNA-binding protein Brain tumor (Brat) targets the transcripts of a self-renewal gene, deadpan (dpn), for decay by recruiting the deadenylation machinery to the 3' untranslated region (UTR). Furthermore, a nuclear protein, Insensible, was identified that complements Cullin-mediated proteolysis to robustly inactivate Dpn activity by limiting the level of active Dpn through protein sequestration. The synergy between post-transcriptional and transcriptional control of self-renewal genes drives timely exit from the stem cell state in uncommitted progenitors. This proposed multilayered gene regulation system could be broadly applicable to the control of exit from stemness in all stem cell lineages. |
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