What's hot today: Current papers in developmental biology and gene function

What's hot today:
Current papers in developmental biology and gene function

Wednesday, October 30th - Adult Physiology and Metabolism

Bellanda, M., Damulewicz, M., Zambelli, B., Costanzi, E., Gregoris, F., Mammi, S., Tosatto, S. C. E., Costa, R., Minervini, G., Mazzotta, G. M. (2024). A PDZ scaffolding/CaM-mediated pathway in Cryptochrome signaling. Protein science : a publication of the Protein Society, 33(3):e4914 PubMed ID: 38358255
Summary:
Cryptochromes are cardinal constituents of the circadian clock, which orchestrates daily physiological rhythms in living organisms. A growing body of evidence points to their participation in pathways that have not traditionally been associated with circadian clock regulation, implying that cryptochromes may be subject to modulation by multiple signaling mechanisms. This study demonstrates that human CRY2 (hCRY2) forms a complex with the large, modular scaffolding protein known as Multi-PDZ Domain Protein 1 (MUPP1). This interaction is facilitated by the calcium-binding protein Calmodulin (CaM) in a calcium-dependent manner. These findings suggest a novel cooperative mechanism for the regulation of mammalian cryptochromes, mediated by calcium ions (Ca(2+) ) and CaM. It us proposed that this Ca(2+) /CaM-mediated signaling pathway may be an evolutionarily conserved mechanism that has been maintained from Drosophila to mammals, most likely in relation to its potential role in the broader context of cryptochrome function and regulation. Further, the understanding of cryptochrome interactions with other proteins and signaling pathways could lead to a better definition of its role within the intricate network of molecular interactions that govern circadian rhythms.

Bhattacharya, M., Starz-Gaiano, M. (2024). Steroid hormone signaling synchronizes cell migration machinery, adhesion and polarity to direct collective movement. J Cell Sci, 137(5) PubMed ID: 38323986
Summary:
Migratory cells - either individually or in cohesive groups - are critical for spatiotemporally regulated processes such as embryonic development and wound healing. Their dysregulation is the underlying cause of formidable health problems such as congenital abnormalities and metastatic cancers. Border cell behavior during Drosophila oogenesis provides an effective model to study temporally regulated, collective cell migration in vivo. Developmental timing in flies is primarily controlled by the steroid hormone ecdysone, which acts through a well-conserved, nuclear hormone receptor complex. Ecdysone signaling determines the timing of border cell migration, but the molecular mechanisms governing this remain obscure. This study found that border cell clusters expressing a dominant-negative form of ecdysone receptor extended ineffective protrusions. Additionally, these clusters had aberrant spatial distributions of E-cadherin (E-cad), apical domain markers and activated myosin that did not overlap. Remediating their expression or activity individually in clusters mutant for ecdysone signaling did not restore proper migration. It is proposed that ecdysone signaling synchronizes the functional distribution of E-cadherin, Atypical protein kinase C (aPKC), Discs large (Dlg1) and activated myosin post-transcriptionally to coordinate adhesion, polarity and contractility and temporally control collective cell migration.

Nelson, J. O., Slicko, A., Raz, A. A., Yamashita, Y. M. (2024). Insulin signaling regulates R2 retrotransposon expression to orchestrate transgenerational rDNA copy number maintenance. bioRxiv, PubMed ID: 38464041
Summary:
Preserving a large number of essential yet highly unstable ribosomal DNA (rDNA) repeats is critical for the germline to perpetuate the genome through generations. Spontaneous rDNA loss must be countered by rDNA copy number (CN) expansion. Germline rDNA CN expansion is best understood in Drosophila melanogaster, which relies on unequal sister chromatid exchange (USCE) initiated by DNA breaks at rDNA. The rDNA-specific retrotransposon R2 responsible for USCE-inducing DNA breaks is typically expressed only when rDNA CN is low to minimize the danger of DNA breaks; however, the underlying mechanism of R2 regulation remains unclear. This study identified the insulin receptor (InR) as a major repressor of R2 expression, limiting unnecessary R2 activity. Through single-cell RNA sequencing this study found that male germline stem cells (GSCs), the major cell type that undergoes rDNA CN expansion, have reduced InR expression when rDNA CN is low. Reduced InR activity in turn leads to R2 expression and CN expansion. It was further found that dietary manipulation alters R2 expression and rDNA CN expansion activity. This work reveals that the insulin pathway integrates rDNA CN surveying with environmental sensing, revealing a potential mechanism by which diet exerts heritable changes to genomic content.

Portela, M., Mukherjee, S., Paul, S., La Marca, J. E., Parsons, L. M., Veraksa, A., Richardson, H. E. (2024). The Drosophila tumour suppressor Lgl and Vap33 activate the Hippo pathway through a dual mechanism. J Cell Sci, 137(4) PubMed ID: 38240353
Summary:
The tumour suppressor, Lethal (2) giant larvae [Lgl) also known as L(2)gl], is an evolutionarily conserved protein that was discovered in Drosophila, where its depletion results in tissue overgrowth and loss of cell polarity. Lgl links cell polarity and tissue growth through regulation of the Notch and the Hippo signalling pathways. Lgl regulates the Notch pathway by inhibiting V-ATPase activity via Vap33. How Lgl regulates the Hippo pathway was unclear. This study showed that V-ATPase activity inhibits the Hippo pathway, whereas Vap33 acts to activate Hippo signalling. Vap33 physically and genetically interacts with the actin cytoskeletal regulators RtGEF (Pix) and http://flybase.org/reports/FBgn0033539.htm">Git, which also bind to the Hippo protein (Hpo) and are involved in the activation of the Hippo pathway. Additionally, it was shown that the ADP ribosylation factor Arf79F (Arf1), which is a Hpo interactor, is involved in the inhibition of the Hippo pathway. Altogether, these data suggest that Lgl acts via Vap33 to activate the Hippo pathway by a dual mechanism: (1) through interaction with RtGEF, Git and Arf79F, and (2) through interaction and inhibition of the V-ATPase, thereby controlling epithelial tissue growth.

Lee, H. C., Oliveira, N. M. M., Hastings, C., Baillie-Benson, P., Moverley, A. A., Lu, H. C., Zheng, Y., Wilby, E. L., Weil, T. T., Page, K. M., Fu, J., Moris, N., Stern, C. D. (2024). Regulation of long-range BMP gradients and embryonic polarity by propagation of local calcium-firing activity. Nat Commun, 15(1):1463 PubMed ID: 38368410
Summary:
Many amniote vertebrate species including humans can form identical twins from a single embryo, but this only occurs rarely. It has been suggested that the primitive-streak-forming embryonic region emits signals that inhibit streak formation elsewhere but the signals involved, how they are transmitted and how they act has not been elucidated. This study shows that short tracks of calcium firing activity propagate through extraembryonic tissue via gap junctions and prevent ectopic primitive streak formation in chick embryos. Cross-regulation of calcium activity and an inhibitor of primitive streak formation (Bone Morphogenetic Protein, BMP) via NF-kappaB and NFAT establishes a long-range BMP gradient spanning the embryo. This mechanism explains how embryos of widely different sizes can maintain positional information that determines embryo polarity. Evidence is orovided for similar mechanisms in two different human embryo models and in Drosophila, suggesting an ancient evolutionary origin.

Li, Y., Liu, D., Wang, H., Zhang, X., Lu, B., Li, S. (2024). The IRE1/Xbp1 axis restores ER and tissue homeostasis perturbed by excess Notch in Drosophila. Dev Biol, 507:11-19 PubMed ID: 38142805
Summary:
Notch signaling controls numerous key cellular processes including cell fate determination and cell proliferation. Its malfunction has been linked to many developmental abnormalities and human disorders. Overactivation of Notch signaling is shown to be oncogenic. Retention of excess Notch protein in the endoplasmic reticulum (ER) can lead to altered Notch signaling and cell fate, but the mechanism is not well understood. This study shows that V5-tagged or untagged exogenous Notch is retained in the ER when overexpressed in fly tissues. Furthermore, it was shown that Notch retention in the ER leads to robust ER enlargement and elicits a rough eye phenotype. Gain-of-function of unfolded protein response (UPR) factors IRE1 or spliced Xbp1 (Xbp1-s) alleviates Notch accumulation in the ER, restores ER morphology and ameliorates the rough eye phenotype. These results uncover a pivotal role of the IRE1/Xbp1 axis in regulating the detrimental effect of ER-localized excess Notch protein during development and tissue homeostasis.

Tuesday, October 29th - Adult Physiology and Metabolism

Kovacs, Z., Bajusz, C., Szabo, A., Borkuti, P., Vedelek, B., Benke, R., Lipinszki, Z., Kristo, I., Vilmos, P. (2024). A bipartite NLS motif mediates the nuclear import of Drosophila moesin. Frontiers in cell and developmental biology, 12:1206067 PubMed ID: 38450250
Summary:
The ERM protein family, which consists of three closely related proteins in vertebrates, ezrin, radixin, and moesin (ERM), is an ancient and important group of cytoplasmic actin-binding and organizing proteins. With their FERM domain, ERMs bind various transmembrane proteins and anchor them to the actin cortex through their C-terminal F-actin binding domain, thus they are major regulators of actin dynamics in the cell. ERMs participate in many fundamental cellular processes, such as phagocytosis, microvilli formation, T-cell activation and tumor metastasis. It has been shown that, besides its cytoplasmic activities, the single ERM protein of Drosophila melanogaster, Moesin, is also present in the cell nucleus, where it participates in gene expression and mRNA export. This study investigated the mechanism by which Moesin enters the nucleus. The nuclear import of Moesin is an NLS-mediated, active process. The nuclear localization sequence of the Moesin protein is an evolutionarily highly conserved, conventional bipartite motif located on the surface of the FERM domain. These experiments also reveal that the nuclear import of Moesin does not require PIP2 binding or protein activation, and occurs in monomeric form. It is proposed, that the balance between the phosphorylated and non-phosphorylated protein pools determines the degree of nuclear import of Moesin.

Aiswarya, K, S. A., Sarkar, S., Vishnu, S., ...., Varghese, J., Srinivasula, S. M., Singh, M. S. (2024). Real-time study of spatio-temporal dynamics (4D) of physiological activities in alive biological specimens with different FOVs and resolutions simultaneously. Sci Rep, 14(1):3542 PubMed ID: 38346989
Summary:
A microscopy imaging system was developed that gives feasibility for studying spatio-temporal dynamics of physiological activities of alive biological specimens (over entire volume not only for a particular section, i.e., in 4D). The imaging technology facilitates obtaining two image frames of a section of the larger specimen with different Field of views (FOVs) at different resolutions or magnifications simultaneously in real-time (in addition to recovery of 3D (volume) information). Again, this imaging system addresses the longstanding challenges of housing multiple light sources (6 at the maximum to date) in microscopy (in general) and light sheet fluorescence microscopy (LSFM) (in particular), by using a tuneable pulsed laser source (with an operating wavelength in the range 670 nm) in contrast to the conventional Continuous-Wave (CW) laser source being adopted for inducing photo-excitation of tagged fluorophores. This study employed four wavelengths. This study also demonstrates quantitative characterization of spatio-temporal dynamics (velocity-both amplitude and direction) of organelles (mitochondria) and their mutual correlationships. Mitochondria close to the nucleus are observed to possess a lower degree of freedom in comparison to that at the cellular periphery. In addition, the study demonstrates real-time observation and recording of the development and growth of all tracheal branches during the entire period of embryonic development. The experimental results conducted in various biological specimens (Drosophila, mouse embryo, and HeLa cells), demonstrate that the study is of great biological and technological impact.

Wright, Y., Armstrong, A. R. (2024). Differential amino acid transporter expression in adult Drosophila melanogaster tissues. microPublication biology, 2024 PubMed ID: 38481556
Summary:
Organismal macronutrient intake modulates organ and tissue function. Dietary amino acids play essential roles in metabolic processes that support normal tissue growth, repair, and function. For example, in Drosophila melanogaster, protein-deficient diets lead to reduced overall organismal growth during larval development and severely decreased egg production in adult females. Multiple tissues, therefore, must sense and respond to dietary protein input. Amino acid transporter proteins facilitate the movement of amino acids across cellular membranes. Based on high-throughput expression studies, the Drosophila genome is predicted to encode 58 amino acid transporters. This study set out to determine if there are tissue-specific amino acid requirements for proper tissue function by first assessing the complement of amino acid transporters expressed in several adult tissues. Using RT-PCR to assess transcript levels, most of the 24 amino acid transporters examined were found to be expressed in the head, thorax, abdomen, gut, and ovary, while a subset shows differential transcript expression. This work will serve as the foundation for future studies addressing the impact of physiological factors, like nutrition, on amino acid sensing by individual tissues.

Ng, A. Q. E., Chan, S. N., Pek, J. W. (2024). Nutrient-dependent regulation of a stable intron modulates germline mitochondrial quality control. Nat Commun, 15(1):1252 PubMed ID: 38341415
Summary:
Mitochondria are inherited exclusively from the mothers and are required for the proper development of embryos. Hence, germline mitochondrial quality is highly regulated during oogenesis to ensure oocyte viability. How nutrient availability influences germline mitochondrial quality control is unclear. This study found that fasting leads to the accumulation of mitochondrial clumps and oogenesis arrest in Drosophila. Fasting induces the downregulation of the DIP1-Clueless pathway, leading to an increase in the expression of a stable intronic sequence RNA called sisRNA:1. Mechanistically, sisR-1 localizes to the mitochondrial clumps to inhibit the poly-ubiquitination of the outer mitochondrial protein Porin/VDAC1, thereby suppressing p62-mediated mitophagy. Alleviation of the fasting-induced high sisR-1 levels by either sisR-1 RNAi or refeeding leads to mitophagy, the resumption of oogenesis and an improvement in oocyte quality. Thus, this study provides a possible mechanism by which fasting can improve oocyte quality by modulating the mitochondrial quality control pathway. Of note, this study found that the sisR-1 response also regulates mitochondrial clumping and oogenesis during protein deprivation, heat shock and aging, suggesting a broader role for this mechanism in germline mitochondrial quality control.

Privalova, V., Sobczyk, L., Szlachcic, E., Labecka, A. M., Czarnoleski, M. (2024). Heat tolerance in Drosophila melanogaster is influenced by oxygen conditions and mutations in cell size control pathways. Philosophical transactions of the Royal Society of London Series B, Biological sciences, 379(1896):20220490 PubMed ID: 38186282
Summary:
Understanding metabolic performance limitations is key to explaining the past, present and future of life. This study investigated whether heat tolerance in actively flying Drosophila melanogaster is modified by individual differences in cell size and the amount of oxygen in the environment. Two mutants with loss-of-function mutations in cell size control were found associated with the target of rapamycin (TOR)/insulin pathways, showing reduced (mutant rictor(Δ2)) or increased (mutant Mnt(1)) cell size in different body tissues compared to controls. Flies were exposed to a steady increase in temperature under normoxia and hypoxia until they collapsed. The upper critical temperature decreased in response to each mutation type as well as under hypoxia. Females, which have larger cells than males, had lower heat tolerance than males. Altogether, mutations in cell cycle control pathways, differences in cell size and differences in oxygen availability affected heat tolerance, but existing theories on the roles of cell size and tissue oxygenation in metabolic performance can only partially explain these results. A better understanding of how the cellular composition of the body affects metabolism may depend on the development of research models that help separate various interfering physiological parameters from the exclusive influence of cell size.

Kitamura, D., Taniguchi, K., Nakamura, M., Igaki, T. (2024). In vivo evidence for homeostatic regulation of ribosomal protein levels in Drosophila. Cell structure and function, 49(1):11-20 PubMed ID: 38199250
Summary:
The ribosome is a molecular machine essential for protein synthesis, which is composed of approximately 80 different ribosomal proteins (Rps). Studies in yeast and cell culture systems have revealed that the intracellular level of Rps is finely regulated by negative feedback mechanisms or ubiquitin-proteasome system, which prevents over- or under-abundance of Rps in the cell. However, in vivo evidence for the homeostatic regulation of intracellular Rp levels has been poor. Using Drosophila genetics, this study showed that intracellular Rp levels are regulated by proteasomal degradation of excess Rps that are not incorporated into the ribosome. By establishing an EGFP-fused Rp gene system that can monitor endogenously expressed Rp levels, it was found that endogenously expressed EGFP-RpS20 or -RpL5 is eliminated from the cell when RpS20 or RpL5 is exogenously expressed. Notably, the level of endogenously expressed Hsp83, a housekeeping gene, was not affected by exogenous expression of Hsp83, suggesting that the strict negative regulation of excess protein is specific for intracellular Rps. Further analyses revealed that the maintenance of cellular Rp levels is not regulated at the transcriptional level but by proteasomal degradation of excess free Rps as a protein quality control mechanism. These observations provide not only the in vivo evidence for the homeostatic regulation of Rp levels but also a novel genetic strategy to study in vivo regulation of intracellular Rp levels and its role in tissue homeostasis via cell competition.

Wright, Y., Armstrong, A. R. (2024). Differential amino acid transporter expression in adult Drosophila melanogaster tissues. microPublication biology, 2024 PubMed ID: 38481556
Summary:
Organismal macronutrient intake modulates organ and tissue function. Dietary amino acids play essential roles in metabolic processes that support normal tissue growth, repair, and function. For example, in Drosophila melanogaster, protein-deficient diets lead to reduced overall organismal growth during larval development and severely decreased egg production in adult females. Multiple tissues, therefore, must sense and respond to dietary protein input. Amino acid transporter proteins facilitate the movement of amino acids across cellular membranes. Based on high-throughput expression studies, the Drosophila genome is predicted to encode 58 amino acid transporters. This study set out to determine if there are tissue-specific amino acid requirements for proper tissue function by first assessing the complement of amino acid transporters expressed in several adult tissues. Using RT-PCR to assess transcript levels, most of the 24 amino acid transporters examined were found to be expressed in the head, thorax, abdomen, gut, and ovary, while a subset shows differential transcript expression. This work will serve as the foundation for future studies addressing the impact of physiological factors, like nutrition, on amino acid sensing by individual tissues.

Kovaacs, Z., Bajusz, C., Szabo, A., Borkúti, P., Vedelek, B., Benke, R., Lipinszki, Z., Kristo, I., Vilmos, P. (2024). A bipartite NLS motif mediates the nuclear import of Drosophila moesin. Frontiers in cell and developmental biology, 12:1206067 PubMed ID: 38450250
Summary:
The ERM protein family, which consists of three closely related proteins in vertebrates, ezrin, radixin, and moesin (ERM), is an ancient and important group of cytoplasmic actin-binding and organizing proteins. With their FERM domain, ERMs bind various transmembrane proteins and anchor them to the actin cortex through their C-terminal F-actin binding domain, thus they are major regulators of actin dynamics in the cell. ERMs participate in many fundamental cellular processes, such as phagocytosis, microvilli formation, T-cell activation and tumor metastasis. It has been shown that, besides its cytoplasmic activities, the single ERM protein of Drosophila melanogaster, Moesin, is also present in the cell nucleus, where it participates in gene expression and mRNA export. This study investigated the mechanism by which Moesin enters the nucleus. The nuclear import of Moesin is an NLS-mediated, active process. The nuclear localization sequence of the Moesin protein is an evolutionarily highly conserved, conventional bipartite motif located on the surface of the FERM domain. These experiments also reveal that the nuclear import of Moesin does not require PIP2 binding or protein activation, and occurs in monomeric form. It is proposed, that the balance between the phosphorylated and non-phosphorylated protein pools determines the degree of nuclear import of Moesin.

Friday, October 25th - Disease Models

Coleman, C. R., Pallos, J., Arreola-Bustos, A., Wang, L., Raftery, D., Promislow, D. E. L., Martin, I. (2024). Natural Variation in Age-Related Dopamine Neuron Degeneration is Glutathione-Dependent and Linked to Life Span. bioRxiv, PubMed ID: 38405950
Summary:
Aging is the biggest risk factor for Parkinson's disease (PD), suggesting that age-related changes in the brain promote dopamine neuron vulnerability. It is unclear, however, whether aging alone is sufficient to cause significant dopamine neuron loss and if so, how this intersects with PD-related neurodegeneration. In this study, through examining a large collection of naturally varying Drosophila strains, a strong relationship was found between life span and age-related dopamine neuron loss. Naturally short-lived strains exhibit a loss of dopamine neurons but not generalized neurodegeneration, while long-lived strains retain dopamine neurons across age. Metabolomic profiling reveals lower glutathione levels in short-lived strains which is associated with elevated levels of reactive oxygen species (ROS), sensitivity to oxidative stress and vulnerability to silencing the familial PD gene parkin. Strikingly, boosting neuronal glutathione levels via glutamate-cysteine ligase (GCL) overexpression is sufficient to normalize ROS levels, extend life span and block dopamine neurons loss in short-lived backgrounds, demonstrating that glutathione deficiencies are central to neurodegenerative phenotypes associated with short longevity. These findings may be relevant to human PD pathogenesis, where glutathione depletion is frequently reported in idiopathic PD patient brain. Building on this evidence, reduced levels of GCL catalytic and modulatory subunits were detected in brain from PD patients harboring the LRRK2 G2019S mutation, implicating possible glutathione deficits in familial LRRK2-linked PD. This study across Drosophila and human PD systems suggests that glutathione plays an important role in the influence of aging on PD neurodegeneration.

Fu, B., Ma, R., Liu, F., Chen, X., Wang, M., Jin, W., Zhang, S., Wang, Y., Sun, L. (2024). New insights into ginsenoside Rg1 regulating the niche to inhibit age-induced germline stem cells depletion through targeting ECR/BMP signaling pathway in Drosophila. Aging, 16(4):3612-3630 PubMed ID: 38364249
Summary:
The age-induced imbalance in ecological niches leads to the loss of GSCs, which is the main reason for ovarian germline senescence. Ginsenoside Rg1, a compound found in the root and stem of the ginseng plant, can delay ovarian senescence. This study shed light on new insights of ginsenoside Rg1 in regulating the niche to maintain GSCs self-renewal and discussed related molecular mechanisms. The differences among GSC number, reproductive capacity of naturally aging female Drosophila after ginsenoside Rg1 feeding were analyzed by immunofluorescence and behavior monitoring. The expressions of the active factors in the niche and the BMP signaling were analyzed through Western blot and RT-qPCR. The target effect was verified in the ECR mutant and combined with the molecular docking. Ginsenoside Rg1 inhibited the age-induced reduction of the GSCs number and restored offspring production and development. Ginsenoside Rg1 promoted the expression of anchor proteins E-cadherin, stemness maintenance factor Nos and differentiation promoting factor Bam, thereby GSCs niche homeostasis was regulated. In addition, ginsenoside Rg1 was bound to the LBD region of the hormone receptor ECR. Ginsenoside Rg1 promotes the regeneration of GSCs by targeting the ECR to increase Smad1/5/8 expression and thereby activating the BMP signaling pathway. In addition, ginsenoside Rg1 maintenance of niche homeostasis to promote GSCs regeneration is dependent on ECR as demonstrated in ECR mutants. It is concluded that ginsenoside Rg1 regulated the ecological niche homeostasis of GSCs and promoted the regeneration of GSCs by targeting the ECR/BMP signaling pathway in hormone-deficient states in aging ovaries. It is of great significance for prolonging fertility potential and delaying ovarian senescence.

Setzu, M. D., Mocci, I., Fabbri, D., Carta, P., Muroni, P., Diana, A., Dettori, M. A., Casu, M. A. (2024). Neuroprotective Effects of the Nutraceutical Dehydrozingerone and Its C(2)-Symmetric Dimer in a Drosophila Model of Parkinson's Disease. Biomolecules, 14(3) PubMed ID: 38540694
Summary:
Parkinson's disease (PD) is a neurodegenerative disorder characterized by the loss of dopaminergic neurons responsible for unintended or uncontrollable movements. Mutations in the leucine-rich repeat kinase 2 locus contribute to genetic forms of PD. The fruit fly Drosophila melanogaster carrying this mutation (LRRK2-Dm) is an in vivo model of PD that develops motor impairment and stands for an eligible non-mammalian paradigm to test novel therapeutic approaches. Dehydrozingerone (DHZ) is a natural phenolic compound isolated from ginger and presents anti-inflammatory, antioxidant and neuroprotective properties, making it a potential therapeutic target for PD. This study administered DHZ and its C(2)-symmetric dimer (DHZ-DIM) at 0.5 and 1 mM for 14 and 21 days in the LRRK2-Dm, with the aim of assessing changes in rescuing motor behavior, brain dopaminergic neurons, mitochondria and synapses (T-bars). The shorter treatment with both molecules revealed efficacy at the higher dose, improving climbing behavior with a prevention of dopaminergic neuronal demise. After 21 days, a recovery of the motor disability, dopaminergic neuron loss, mitochondrial damage and T-bars failure was observed with the DHZ-DIM. These data indicate that the DHZ-DIM exerts a more potent neuroprotective effect with respect to the monomer in LRRK2-Dm, prompting further investigation of these compounds in rodent models of PD.

Li, Y., Liu, D., Zhang, X., Rimal, S., Lu, B., Li, S. (2024). RACK1 and IRE1 participate in the translational quality control of amyloid precursor protein in Drosophila models of Alzheimer's disease. J Biol Chem, 300(3):105719 PubMed ID: 38311171
Summary:
Alzheimer's disease (AD) is a progressive neurodegenerative disorder characterized by dysregulation of the expression and processing of the amyloid precursor protein (APP). Protein quality control systems are dedicated to remove faulty and deleterious proteins to maintain cellular protein homeostasis (proteostasis). Identidying mechanisms underlying APP protein regulation is crucial for understanding AD pathogenesis. However, the factors and associated molecular mechanisms regulating APP protein quality control remain poorly defined. This study shows that mutant APP with its mitochondrial-targeting sequence ablated exhibited predominant endoplasmic reticulum (ER) distribution and led to aberrant ER morphology, deficits in locomotor activity, and shortened lifespan. This study searched for regulators that could counteract the toxicity caused by the ectopic expression of this mutant APP. Genetic removal of the ribosome-associated quality control (RQC) factor RACK1 resulted in reduced levels of ectopically expressed mutant APP. By contrast, gain of RACK1 function increased mutant APP level. Additionally, overexpression of the ER stress regulator (IRE1) resulted in reduced levels of ectopically expressed mutant APP. Mechanistically, the RQC related ATPase VCP/p97 and the E3 ubiquitin ligase Hrd1 were required for the reduction of mutant APP level by IRE1. These factors also regulated the expression and toxicity of ectopically expressed wild type APP, supporting their relevance to APP biology. These results reveal functions of RACK1 and IRE1 in regulating the quality control of APP homeostasis and mitigating its pathogenic effects, with implications for the understanding and treatment of AD.

Twyning, M. J., Tufi, R., Gleeson, T. P., Kolodziej, K. M., Campesan, S., Terriente-Felix, A., Collins, L., De Lazzari, F., Giorgini, F., Whitworth, A. J. (2024). Partial loss of MCU mitigates pathology in vivo across a diverse range of neurodegenerative disease models. Cell Rep, 43(2):113681 PubMed ID: 38236772
Summary:
Mitochondrial calcium (Ca(2+)) uptake augments metabolic processes and buffers cytosolic Ca(2+) levels; however, excessive mitochondrial Ca(2+) can cause cell death. Disrupted mitochondrial function and Ca(2+) homeostasis are linked to numerous neurodegenerative diseases (NDs), but the impact of mitochondrial Ca(2+) disruption is not well understood. This study show that Drosophila models of multiple NDs (Parkinson's, Huntington's, Alzheimer's, and frontotemporal dementia) reveal a consistent increase in neuronal mitochondrial Ca(2+) levels, as well as reduced mitochondrial Ca(2+) buffering capacity, associated with increased mitochondria-endoplasmic reticulum contact sites (MERCs). Importantly, loss of the mitochondrial Ca(2+) uptake channel MCU or overexpression of the efflux channel NCLX robustly suppresses key pathological phenotypes across these ND models. Thus, mitochondrial Ca(2+) imbalance is a common feature of diverse NDs in vivo and is an important contributor to the disease pathogenesis. The broad beneficial effects from partial loss of MCU across these models presents a common, druggable target for therapeutic intervention.

Shan, Z., Li, S., Gao, Y., Jian, C., Ti, X., Zuo, H., Wang, Y., Zhao, G., Wang, Y., Zhang, Q. (2024). mtDNA extramitochondrial replication mediates mitochondrial defect effects. mtDNA extramitochondrial replication mediates mitochondrial defect effects. iScience, 27(2):108970 PubMed ID: 38322987
Summary:
A high ratio of severe mitochondrial defects causes multiple human mitochondrial diseases. However, until now, the in vivo rescue signal of such mitochondrial defect effects has not been clear. This study built fly mitochondrial defect models by knocking down the essential mitochondrial genes dMterf4 and dMrps23. Following genome-wide RNAi screens, knockdown of Med8/Tfb4/mtSSB/PolG2/mtDNA-helicase rescued dMterf4/dMrps23 RNAi-mediated mitochondrial defect effects. Extremely surprisingly, they drove mtDNA replication outside mitochondria through the Med8/Tfb4-mtSSB/PolG2/mtDNA-helicase axis to amplify cytosolic mtDNA, leading to activation of the cGAS-Sting-like IMD pathway to partially mediate dMterf4/dMrps23 RNAi-triggered effects. Moreover, the Med8/Tfb4-mtSSB/PolG2/mtDNA-helicase axis also mediated other fly mitochondrial gene defect-triggered dysfunctions and Drosophila aging. Overall, this study demarcates the mtDNA-helicase axis as a candidate mechanism to mediate mitochondrial defect effects through driving mtDNA extramitochondrial replication; dysfunction of this axis might be used for potential treatments for many mitochondrial and age-related diseases.

Thursday, October 24th - Disease Models

Pfefferkorn, R. M., Mortzfeld, B. M., Fink, C., Frieling, J. V., Bossen, J., Esser, D., Kaleta, C., Rosenstiel, P., Heine, H., Roeder, T. (2024). Recurrent Phases of Strict Protein Limitation Inhibit Tumor Growth and Restore Lifespan in A Drosophila Intestinal Cancer Model. Aging and disease, 15(1):226-244 PubMed ID: 37962464
Summary:
Diets that restrict caloric or protein intake offer a variety of benefits, including decreasing the incidence of cancer. However, whether such diets pose a substantial therapeutic benefit as auxiliary cancer treatments remains unclear. This study determined the effects of severe protein depletion on tumorigenesis in a Drosophila melanogaster intestinal tumor model, using a human RAF gain-of-function allele. Severe and continuous protein restriction significantly reduced tumor growth but resulted in premature death. Therefore, a diet was developed in which short periods of severe protein restriction alternated cyclically with periods of complete feeding. This nutritional regime reduced tumor mass, restored gut functionality, and rescued the lifespan of oncogene-expressing flies to the levels observed in healthy flies on a continuous, fully nutritious diet. Furthermore, this diet reduced the chemotherapy-induced stem cell activity associated with tumor recurrence. Transcriptome analysis revealed long-lasting changes in the expression of key genes involved in multiple major developmental signaling pathways. Overall, the data suggest that recurrent severe protein depletion effectively mimics the health benefits of continuous protein restriction, without undesired nutritional shortcomings. This provides seminal insights into the mechanisms of the memory effect required to maintain the positive effects of protein restriction throughout the phases of a full diet. Finally, the repetitive form of strict protein restriction is an ideal strategy for adjuvant cancer therapy that is useful in many tumor contexts.

Cong, B., Stamou, E., Pennel, K., McKenzie, M., Matly, A., Gopinath, S., Edwards, J., Cagan, R. (2023). WNT Signalling Promotes NF-kappaB Activation and Drug Resistance in KRAS-Mutant Colorectal Cancer. bioRxiv, PubMed ID: 38187607
Summary:
Approximately 40% of colorectal cancer (CRC) cases are characterized by KRAS mutations, rendering them insensitive to most CRC therapies. While the reasons for this resistance remain incompletely understood, one key aspect is genetic complexity: in CRC, oncogenic KRAS is most commonly paired with mutations that alter WNT and P53 activities ("RAP"). This study demonstrate dthat elevated WNT activity upregulates canonical (NF-kappaB) signalling in both Drosophila and human RAS mutant tumours. This upregulation required Toll-1 and Toll-9 and resulted in reduced efficacy of RAS pathway targeted drugs such as the MEK inhibitor trametinib. Inhibiting WNT activity pharmacologically significantly suppressed trametinib resistance in RAP tumours and more genetically complex RAP-containing 'patient avatar' models. WNT/MEK drug inhibitor combinations were further improved by targeting brm, shg, ago, rhoGAPp190 and upf1, highlighting these genes as candidate biomarkers for patients sensitive to this duel approach. These findings shed light on how genetic complexity impacts drug resistance and proposes a therapeutic strategy to reverse this resistance.

Mishra, A. K., Rodriguez, M., Torres, A. Y., Smith, M., Rodriguez, A., Bond, A., Morrissey, M. A., Montell, D. J. (2023). Hyperactive Rac stimulates cannibalism of living target cells and enhances CAR-M-mediated cancer cell killing. Proc Natl Acad Sci U S A, 120(52):e2310221120 PubMed ID: 38109551
Summary:
The 21kD GTPase Rac is an evolutionarily ancient regulator of cell shape and behavior. Rac2 is predominantly expressed in hematopoietic cells where it is essential for survival and motility. The hyperactivating mutation Rac2(E62K) also causes human immunodeficiency, although the mechanism remains unexplained. This study reports that in Drosophila, hyperactivating Rac stimulates ovarian cells to cannibalize neighboring cells, destroying the tissue. Hyperactive Rac2(E62K) stimulates human HL60-derived macrophage-like cells to engulf and kill living T cell leukemia cells. Primary mouse Rac2(+/E62K) bone-marrow-derived macrophages also cannibalize primary Rac2(+/E62K) T cells due to a combination of macrophage hyperactivity and T cell hypersensitivity to engulfment. Additionally, Rac2(+/E62K) macrophages non-autonomously stimulate wild-type macrophages to engulf T cells. Rac2(E62K) also enhances engulfment of target cancer cells by chimeric antigen receptor-expressing macrophages (CAR-M) in a CAR-dependent manner. It is proposed that Rac-mediated cell cannibalism may contribute to Rac2(+/E62K) human immunodeficiency and enhance CAR-M cancer immunotherapy.

Pinal, N., Ruiz-Losada, M., Azpiazu, N., Morata, G. (2023). Size compensation in Drosophila after generalised cell death. Frontiers in cell and developmental biology, 11:1301913 PubMed ID: 38078009
Summary:
Regeneration is a response mechanism aimed to restore tissues that have been damaged. In the wing disc of Drosophila the regenerative response to a dose of Ionizing Radiation is being studied that kills over 35% of the cells distributed all over the disc. After such treatment the discs are able to restore normal size, indicating there is a mechanism that repairs generalised damage. The role is being studied of the JNK, JAK/STAT and Wg pathways, known to be required for regeneration after localised damage in the disc. After irradiation there is size compensation in the absence of function of these pathways, indicating that they are not necessary for the compensation. Furthermore, it was also found that generalised damage does not cause an increase in the proliferation rate of surviving cells. It is proposed that irradiated discs suffer a developmental delay and resume growth at normal rate until they reach the final stereotyped size. The delay appears to be associated with a developmental reversion, because discs undergo rejuvenation towards an earlier developmental stage. It is argued that the response to generalized damage is fundamentally different from that to localized damage, which requires activity of JNK and Wg.

Xu, J., Liu, Y., Yang, F., Cao, Y., Chen, W., Li, J. S. S., Zhang, S., Comjean, A., Hu, Y., Perrimon, N. (2024). Mechanistic characterization of a Drosophila model of paraneoplastic nephrotic syndrome. Nat Commun, 15(1):1241 PubMed ID: 38336808
Summary:
Paraneoplastic syndromes occur in cancer patients and originate from dysfunction of organs at a distance from the tumor or its metastasis. A wide range of organs can be affected in paraneoplastic syndromes; however, the pathological mechanisms by which tumors influence host organs are poorly understood. Recent studies in the fly uncovered that tumor secreted factors target host organs, leading to pathological effects. In this study, using a Drosophila gut tumor model, this study characterize a mechanism of tumor-induced kidney dysfunction. Specifically, it was found that Pvf1, a PDGF/VEGF signaling ligand, secreted by gut tumors activates the PvR/JNK/Jra signaling pathway in the principal cells of the kidney, leading to mis-expression of renal genes and paraneoplastic renal syndrome-like phenotypes. This study describes an important mechanism by which gut tumors perturb the function of the kidney, which might be of clinical relevance for the treatment of paraneoplastic syndromes.

Abidi, S. N. F., Hsu, F. T., Smith-Bolton, R. K. (2023). Regenerative growth is constrained by brain tumor to ensure proper patterning in Drosophila. PLoS Genet, 19(12):e1011103 PubMed ID: 38127821
Summary:
Some animals respond to injury by inducing new growth to regenerate the lost structures. This regenerative growth must be carefully controlled and constrained to prevent aberrant growth and to allow correct organization of the regenerating tissue. However, the factors that restrict regenerative growth have not been identified. Using a genetic ablation system in the Drosophila wing imaginal disc, this study has identified one mechanism that constrains regenerative growth, impairment of which also leads to erroneous patterning of the final appendage. Regenerating discs with reduced levels of the RNA-regulator Brain tumor (Brat) exhibit enhanced regeneration, but produce adult wings with disrupted margins that are missing extensive tracts of sensory bristles. In these mutants, aberrantly high expression of the pro-growth factor Myc and its downstream targets likely contributes to this loss of cell-fate specification. Thus, Brat constrains the expression of pro-regeneration genes and ensures that the regenerating tissue forms the proper final structure.

Wednesday, October 23rd - Adult Neural Development, Structure and Function

Montanari, M., Maniere, G., Berthelot-Grosjean, M., Dusabyinema, Y., Gillet, B., Grosjean, Y., Kurz, C. L., Royet, J. (2024). Larval microbiota primes the Drosophila adult gustatory response. Nat Commun. 15(1):1341. PubMed ID: 38351056
Summary:
The survival of animals depends, among other things, on their ability to identify threats in their surrounding environment. Senses such as olfaction, vision and taste play an essential role in sampling their living environment, including microorganisms, some of which are potentially pathogenic. This study focuses on the mechanisms of detection of bacteria by the Drosophila gustatory system. The peptidoglycan (PGN) that forms the cell wall of bacteria was demonstrated to trigger an immediate feeding aversive response when detected by the gustatory system of adult flies. Although this study identified ppk23+ and Gr66a+ gustatory neurons as necessary to transduce fly response to PGN, they were demonstrated to play very different roles in the process. Time-controlled functional inactivation and in vivo calcium imaging demonstrate that while ppk23+ neurons are required in the adult flies to directly transduce PGN signal, Gr66a+ neurons must be functional in larvae to allow future adults to become PGN sensitive. Furthermore, the ability of adult flies to respond to bacterial PGN is lost when they hatch from larvae reared under axenic conditions. Recolonization of germ-free larvae, but not adults, with a single bacterial species, Lactobacillus brevis, is sufficient to restore the ability of adults to respond to PGN. These data demonstrate that the genetic and environmental characteristics of the larvae are essential to make the future adults competent to respond to certain sensory stimuli such as PGN.

Wodrich, A. P. K., Harris, B. T., Giniger, E. (2024). Changes in mitochondrial distribution occur at the axon initial segment in association with neurodegeneration in Drosophila. bioRxiv, PubMed ID: 38405730
Summary:
Changes in mitochondrial distribution are a feature of numerous age-related neurodegenerative diseases. In Drosophila, reducing the activity of Cdk5 causes a neurodegenerative phenotype and is known to affect several mitochondrial properties. Therefore, this study investigated whether alterations of mitochondrial distribution are involved in Cdk5-associated neurodegeneration. Reducing Cdk5 activity does not alter the balance of mitochondrial localization to the somatodendritic vs. axonal neuronal compartments of the mushroom body, the learning and memory center of the Drosophila brain. Changes were observed in mitochondrial distribution at the axon initial segment (AIS), a neuronal compartment located in the proximal axon involved in neuronal polarization and action potential initiation. Specifically, it was observed that mitochondria are partially excluded from the AIS in wild-type neurons, but that this exclusion is lost upon reduction of Cdk5 activity, concomitant with the shrinkage of the AIS domain that is known to occur in this condition. This mitochondrial redistribution into the AIS is not likely due to the shortening of the AIS domain itself but rather due to altered Cdk5 activity. Furthermore, mitochondrial redistribution into the AIS is unlikely to be an early driver of neurodegeneration in the context of reduced Cdk5 activity.

Zhuravlev, A. V., Vetrovoy, O. V., Zalomaeva, E. S., Egozova, E. S., Nikitina, E. A., Savvateeva-Popova, E. V. (2024). Overexpression of the limk1 Gene in Drosophila melanogaster Can Lead to Suppression of Courtship Memory in Males. Biochemistry (Mosc), 89(3):393-406 PubMed ID: 38648760
Summary:
Courtship suppression is a behavioral adaptation of the fruit fly. When majority of the females in a fly population are fertilized and non-receptive for mating, a male, after a series of failed attempts, decreases its courtship activity towards all females, saving its energy and reproductive resources. The time of courtship decrease depends on both duration of unsuccessful courtship and genetically determined features of the male nervous system. Thereby, courtship suppression paradigm can be used for studying molecular mechanisms of learning and memory. p-Cofilin, a component of the actin remodeling signaling cascade and product of LIM-kinase 1 (LIMK1), regulates Drosophila melanogaster forgetting in olfactory learning paradigm. Previously, it was shown that limk1 suppression in the specific types of nervous cells differently affects fly courtship memory. Gal4 :gt; UAS system was used to induce limk1 overexpression in the same types of neurons. limk1 activation in the mushroom body, glia, and fruitless neurons decreased learning index compared to the control strain or the strain with Lim1 knockdown. In cholinergic and dopaminergic/serotoninergic neurons, both overexpression and knockdown of limk1 impaired Drosophila short-term memory. Thus, proper balance of the limk1. activity is crucial for normal cognitive activity of the fruit fly.

Nelson, N., Miller, V., Baumann, N., Broadie, K. (2024). Experience-Dependent Remodeling of Juvenile Brain Olfactory Sensory Neuron Synaptic Connectivity in an Early-Life Critical Period. J Vis Exp, (205) PubMed ID: 38497653
Summary:
Early-life olfactory sensory experience induces dramatic synaptic glomeruli remodeling in the Drosophila juvenile brain, which is experientially dose-dependent, temporally restricted, and transiently reversible only in a short, well-defined critical period. The directionality of brain circuit synaptic connectivity remodeling is determined by the specific odorant acting on the respondent receptor class of olfactory sensory neurons. In general, each neuron class expresses only a single odorant receptor and innervates a single olfactory synaptic glomerulus. In the Drosophila genetic model, the full array of olfactory glomeruli has been precisely mapped by odorant responsiveness and behavioral output. Ethyl butyrate (EB) odorant activates Or42a receptor neurons innervating the VM7 glomerulus. During the early-life critical period, EB experience drives dose-dependent synapse elimination in the Or42a olfactory sensory neurons. Timed periods of dosed EB odorant exposure allow investigation of experience-dependent circuit connectivity pruning in juvenile brain. Confocal microscopy imaging of antennal lobe synaptic glomeruli is done with Or42a receptor-driven transgenic markers that provide quantification of synapse number and innervation volume. The sophisticated Drosophila genetic toolkit enables the systematic dissection of the cellular and molecular mechanisms mediating brain circuit remodeling.

Toshima, N., Schleyer, M. (2024). IR76b-expressing neurons in Drosophila melanogaster are necessary for associative reward learning of an amino acid mixture. Biology letters, 20(2):20230519 PubMed ID: 38351746
Summary:
Learning where to find nutrients while at the same time avoiding toxic food is essential for survival of any animal. Using Drosophila melanogaster larvae as a study case, this study investigate the role of gustatory sensory neurons expressing IR76b for associative learning of amino acids, the building blocks of proteins. Surprising complexity was found in the neuronal underpinnings of sensing amino acids, and a functional division of sensory neurons. The IR76b receptor was found to be dispensable for amino acid learning, whereas the neurons expressing IR76b are specifically required for the rewarding but not the punishing effect of amino acids. This unexpected dissociation in neuronal processing of amino acids for different behavioural functions provides a study case for functional divisions of labour in gustatory systems.

Cornean, J., Molina-Obando, S., Gur, B., Bast, A., Ramos-Traslosheros, G., Chojetzki, J., Lorsch, L., Ioannidou, M., Taneja, R., Schnaitmann, C., Silies, M. (2024). Heterogeneity of synaptic connectivity in the fly visual system. Nat Commun, 15(1):1570 PubMed ID: 38383614
Summary:
Visual systems are homogeneous structures, where repeating columnar units retinotopically cover the visual field. Each of these columns contain many of the same neuron types that are distinguished by anatomic, genetic and - generally - by functional properties. However, there are exceptions to this rule. In the 800 columns of the Drosophila eye, there is an anatomically and genetically identifiable cell type with variable functional properties, Tm9. Since anatomical connectivity shapes functional neuronal properties, this study identified the presynaptic inputs of several hundred Tm9s across both optic lobes using the full adult female fly brain (FAFB) electron microscopic dataset and FlyWire connectome. This work shows that Tm9 has three major and many sparsely distributed inputs. This differs from the presynaptic connectivity of other Tm neurons, which have only one major, and more stereotypic inputs than Tm9. Genetic synapse labeling showed that the heterogeneous wiring exists across individuals. Together, these data argue that the visual system uses heterogeneous, distributed circuit properties to achieve robust visual processing.

Tuesday, October 22nd - Disease Models

Granat, L., Knorr, D. Y., Ranson, D. C., Chakrabarty, R. P., Chandel, N. S., Bateman, J. M. (2024). A Drosophila model of mitochondrial disease phenotypic heterogeneity. Biol Open, 13(2) PubMed ID: 38304969
Summary:
Mutations in genes that affect mitochondrial function cause primary mitochondrial diseases. Mitochondrial diseases are highly heterogeneous and even patients with the same mitochondrial disease can exhibit broad phenotypic heterogeneity, which is poorly understood. Mutations in subunits of mitochondrial respiratory complex I cause complex I deficiency, which can result in severe neurological symptoms and death in infancy. However, some complex I deficiency patients present with much milder symptoms. The most common nuclear gene mutated in complex I deficiency is the highly conserved core subunit NDUFS1. To model the phenotypic heterogeneity in complex I deficiency, RNAi lines were used targeting the Drosophila NDUFS1 homolog ND-75 with different efficiencies. Strong knockdown of ND-75 in Drosophila neurons resulted in severe behavioural phenotypes, reduced lifespan, altered mitochondrial morphology, reduced endoplasmic reticulum (ER)-mitochondria contacts and activation of the unfolded protein response (UPR). By contrast, weak ND-75 knockdown caused much milder behavioural phenotypes and changes in mitochondrial morphology. Moreover, weak ND-75 did not alter ER-mitochondria contacts or activate the UPR. Weak and strong ND-75 knockdown resulted in overlapping but distinct transcriptional responses in the brain, with weak knockdown specifically affecting proteosome activity and immune response genes. Metabolism was also differentially affected by weak and strong ND-75 knockdown including gamma-aminobutyric acid (GABA) levels, which may contribute to neuronal dysfunction in ND-75 knockdown flies. Several metabolic processes were only affected by strong ND-75 knockdown including the pentose phosphate pathway and the metabolite 2-hydroxyglutarate (2-HG), suggesting 2-HG as a candidate biomarker of severe neurological mitochondrial disease. Thus, the Drosophila model provides the means to dissect the mechanisms underlying phenotypic heterogeneity in mitochondrial disease.

Hewson, L., Choo, A., Webber, D. L., Trim, P. J., Snel, M. F., Fedele, A. O., Hopwood, J. J., Hemsley, K. M., O'Keefe, L. V. (2024). Drosophila melanogaster models of MPS IIIC (Hgsnat-deficiency) highlight the role of glia in disease presentation. J Inherit Metab Dis, 47(2):340-354 PubMed ID: 38238109
Summary:
Sanfilippo syndrome (Mucopolysaccharidosis type III or MPS III) is a recessively inherited neurodegenerative lysosomal storage disorder. Mutations in genes encoding enzymes in the heparan sulphate degradation pathway lead to the accumulation of partially degraded heparan sulphate, resulting ultimately in the development of neurological deficits. Mutations in the gene encoding the membrane protein heparan-α-glucosaminide N-acetyltransferase (HGSNAT; EC2.3.1.78) cause MPS IIIC (OMIM#252930), typified by impaired cognition, sleep-wake cycle changes, hyperactivity and early death, often before adulthood. The precise disease mechanism that causes symptom emergence remains unknown, posing a significant challenge in the development of effective therapeutics. As HGSNAT is conserved in Drosophila melanogaster, this study describes the creation and characterisation of the first Drosophila models of MPS IIIC. Flies with either an endogenous insertion mutation or RNAi-mediated knockdown of hgsnat were confirmed to have a reduced level of HGSNAT transcripts and age-dependent accumulation of heparan sulphate leading to engorgement of the endo/lysosomal compartment. This resulted in abnormalities at the pre-synapse, defective climbing and reduced overall activity. Altered circadian rhythms (shift in peak morning activity) were seen in hgsnat neuronal knockdown lines. Further, when hgsnat was knocked down in specific glial subsets (wrapping, cortical, astrocytes or subperineural glia), impaired climbing or reduced activity was noted, implying that hgsnat function in these specific glial subtypes contributes significantly to this behaviour and targeting treatments to these cell groups may be necessary to ameliorate or prevent symptom onset. These novel models of MPS IIIC provide critical research tools for delineating the key cellular pathways causal in the onset of neurodegeneration in this presently untreatable disorder.

Pereira, J., Melo, S., Ferreira, R. M., Carneiro, P., Yang, V., Maia, A. F., Carvalho, J., Figueiredo, C., Machado, J. C., Morais-de-Sa, E., Seruca, R., Figueiredo, J. (2024). E-cadherin variants associated with oral facial clefts trigger aberrant cell motility in a REG1A-dependent manner. Cell Commun Signal, 22(1):152 PubMed ID: 38414029
Summary:
Germline mutations of E-cadherin contribute to hereditary diffuse gastric cancer (HDGC) and congenital malformations, such as oral facial clefts (OFC). However, the molecular mechanisms through which E-cadherin loss-of-function triggers distinct clinical outcomes remain unknown. It was postulated that E-cadherin-mediated disorders result from abnormal interactions with the extracellular matrix and consequent aberrant intracellular signalling, affecting the coordination of cell migration. This study developed in vivo and in vitro models of E-cadherin mutants associated with either OFC or HDGC. Using a Drosophila approach, the impact of the different variants was addressed in cell morphology and migration ability. By combining gap closure migration assays and time-lapse microscopy, the migration pattern of cells expressing OFC or HDGC variants was investigated. The adhesion profile of the variants was evaluated using high-throughput ECM arrays, whereas RNA sequencing technology was explored for identification of genes involved in aberrant cell motility. This study demonstrated that cells expressing OFC variants exhibit an excessive motility performance and irregular leading edges, which prevent the coordinated movement of the epithelial monolayer. Importantly, OFC variants promote cell adhesion to a wider variety of extracellular matrices than HDGC variants, suggesting higher plasticity in response to different microenvironments. A distinct transcriptomic profile was unveiled in the OFC setting and REG1A was pinpointed as a putative regulator of this outcome. Consistent with this, specific RNAi-mediated inhibition of REG1A shifted the migration pattern of OFC expressing cells, leading to slower wound closure with coordinated leading edges. This study provides evidence that E-cadherin variants associated with OFC activate aberrant signalling pathways that support dynamic rearrangements of cells towards improved adaptability to the microenvironment. This proficiency results in abnormal tissue shaping and movement, possibly underlying the development of orofacial malformations.

Tio, M., Wen, R., Choo, C. N., Tan, J. B., Chua, A., Xiao, B., Sundaram, J. R., Chan, C. H. S., Tan, E. K. (2024). Genetic and pharmacologic p32-inhibition rescue CHCHD2-linked Parkinson's disease phenotypes in vivo and in cell models. Journal of biomedical science, 31(1):24 PubMed ID: 38395904
Summary:
Mutations in CHCHD2 have been linked to Parkinson's disease, however, their exact pathophysiologic roles are unclear. The /p32 protein has been suggested to interact with CHCHD2, however, the physiological functions of such interaction in the context of PD have not been clarified. Interaction between CHCHD2 and p32 was confirmed by co-immunoprecipitation experiments. The effect of p32-knockdown was studied in the transgenic Drosophila and Hela cells expressing the wild type and the pathogenic variants of hCHCHD2. The rescue ability of a custom generated p32-inhibitor was assessed in these models as well as in the human fibroblast derived neural precursor cells and the dopaminergic neurons harboring hCHCHD2-Arg145Gln. These results showed that wildtype and mutant hCHCHD2 could bind to p32 in vitro, supported by in vivo interaction between human CHCHD2 and Drosophila p32. Knockdown of p32 reduced mutant hCHCHD2 levels in Drosophila and in vitro. In Drosophila hCHCHD2 models, inhibition of p32 through genetic knockdown and pharmacological treatment using a customized p32-inhibitor restored dopaminergic neuron numbers and improved mitochondrial morphology. These were correlated with improved locomotor function, reduced oxidative stress and decreased mortality. Consistently, Hela cells expressing mutant hCHCHD2 showed improved mitochondrial morphology and function after treatment with the p32-inhibitor. As compared to the isogenic control cells, large percentage of the mutant neural precursor cells and dopaminergic neurons harboring hCHCHD2-Arg145Gln contained fragmented mitochondria which was accompanied by lower ATP production and cell viability. The NPCs harboring hCHCHD2-Arg145Gln also had a marked increase in α-synuclein expression. The p32-inhibitor was able to ameliorate the mitochondrial fragmentation, restored ATP levels, increased cell viability and reduced α-synuclein level in these cells. This study identified p32 as a modulator of CHCHD2, possibly exerting its effects by reducing the toxic mutant hCHCHD2 expression and/or mitigating the downstream effects. Inhibition of the p32 pathway can be a potential therapeutic intervention for CHCHD2-linked PD and diseases involving mitochondrial dysfunction.

Sleep, M., Landaverde, S., Lacoste, A., Tan, S., Schuback, R., Reiter, L. T., Iyengar, A. (2024). Glial expression of Drosophila UBE3A causes spontaneous seizures modulated by 5-HT signaling. bioRxiv, PubMed ID: 38370819
Summary:
Misexpression of the E3 ubiquitin ligase ube3a_angelman.htm">UBE3A is thought to contribute to a range of neurological disorders. In the context of Dup15q syndrome, excess genomic copies of UBE3A is thought to contribute to the autism, muscle tone and spontaneous seizures characteristic of the disorder. In a Drosophila model of Dup 15q syndrome, it was recently shown glial-driven expression of the UBE3A ortholog dube3a led to a "bang-sensitive" phenotype, where mechanical shock triggers convulsions, suggesting glial dube3a expression contributes to hyperexcitability in flies. This study directly compares the consequences of glial- and neuronal-driven dube3a expression on motor coordination and neuronal excitability in Drosophila. IowaFLI tracker was used, and a hidden Markov Model was developed to classify seizure-related immobilization. Both glial and neuronal driven dube3a expression led to clear motor phenotypes. However, only glial-driven dube3aexpression displayed spontaneous immobilization events, that were exacerbated at high-temperature (38 °C). Using a tethered fly preparation was monitored; flight muscle activity glial-driven dube3a flies display spontaneous spike discharges which were bilaterally synchronized indicative of seizure activity. Neither control flies, nor neuronal- dube3a overexpressing flies display such firing patterns. Prior drug screen indicated bang-sensitivity in glial-driven dube3a expressing flies could be suppressed by certain 5-HT modulators. Consistent with this report, glial-driven dube3a flies fed the serotonin reuptake inhibitor vortioxetine and the 5HT (2A) antagonist ketanserin displayed reduced immobilization and spike bursting. Together these findings highlight the potential for glial pathophysiology to drive Dup15q syndrome-related seizure activity.

Al-Sabri, M. H., Ammar, N., Korzh, S., Alsehli, A. M., Hosseini, K., Fredriksson, R., Mwinyi, J., Williams, M. J., Boukhatmi, H., Schiöth, H. B. (2024). Fluvastatin-induced myofibrillar damage is associated with elevated ROS, and impaired fatty acid oxidation, and is preceded by mitochondrial morphological changes. Sci Rep, 14(1):3338 PubMed ID: 38336990
Summary:
It has been shown that fluvastatin treatment induces myofibrillar damage and mitochondrial phenotypes in the skeletal muscles of Drosophila. However, the sequential occurrence of mitochondrial phenotypes and myofibril damage remains elusive. To address this, flies were treated with fluvastatin for two and five days, and their thorax flight muscles were examined using confocal microscopy. In the two-day fluvastatin group, compared to the control, thorax flight muscles exhibited mitochondrial morphological changes, including fragmentation, rounding up and reduced content, while myofibrils remained organized in parallel. In the five-day fluvastatin treatment, not only did mitochondrial morphological changes become more pronounced, but myofibrils became severely disorganized with significantly increased thickness and spacing, along with myofilament abnormalities, suggesting myofibril damage. These findings suggest that fluvastatin-induced mitochondrial changes precede myofibril damage. Moreover, in the five-day fluvastatin group, the mitochondria demonstrated elevated H(2)O(2) and impaired fatty acid oxidation compared to the control group, indicating potential mitochondrial dysfunction. Surprisingly, knocking down Hmgcr (Drosophila homolog of HMGCR) showed normal mitochondrial respiration in all parameters compared to controls or five-day fluvastatin treatment, which suggests that fluvastatin-induced mitochondrial dysfunction might be independent of Hmgcr inhibition. These results provide insights into the sequential occurrence of mitochondria and myofibril damage in statin-induced myopathy for future studies.

Friday, October 18th - Cell Death

Li, J., Jethva, P. N., Rohrs, H. W., Chemuru, S., Miller, K., Gross, M. L., Beckingham, K. M. (2024). Hydrogen/Deuterium Exchange Mass Spectrometry Provides Insights into the Role of Drosophila Testis-Specific Myosin VI Light Chain AndroCaM.. Biochemistry, 63(5):610-624 PubMed ID: 38357882
Summary:
In Drosophila testis, myosin VI plays a special role, distinct from its motor function, by anchoring components to the unusual actin-based structures (cones) that are required for spermatid individualization. For this, the two calmodulin (CaM) light-chain molecules of myosin VI are replaced by androcam (ACaM), a related protein with 67% identity to CaM. Although ACaM has a similar bi-lobed structure to CaM, with two EF hand-type Ca(2+) binding sites per lobe, only one functional Ca(2+) binding site operates in the amino-terminus. To understand this light chain substitution, hydrogen-deuterium exchange mass spectrometry (HDX-MS) was used to examine dynamic changes in ACaM and CaM upon Ca(2+) binding and interaction with the two CaM binding motifs of myosin VI (insert2 and IQ motif). HDX-MS reveals that binding of Ca(2+) to ACaM destabilizes its N-lobe but stabilizes the entire C-lobe, whereas for CaM, Ca(2+) binding induces a pattern of alternating stabilization/destabilization throughout. The conformation of this stable holo-C-lobe of ACaM seems to be a "prefigured" version of the conformation adopted by the holo-C-lobe of CaM for binding to insert2 and the IQ motif of myosin VI. Strikingly, the interaction of holo-ACaM with either peptide converts the holo-N-lobe to its Ca(2+)-free, more stable, form. Thus, ACaM in vivo should bind the myosin VI light chain sites in an apo-N-lobe/holo-C-lobe state that cannot fulfill the Ca(2+)-related functions of holo-CaM required for myosin VI motor assembly and activity. These findings indicate that inhibition of myosin VI motor activity is a precondition for transition to an anchoring function.

Yu, Y., Chen, D., Farmer, S. M., Xu, S., Rios, B., Solbach, A., Ye, X., Ye, L., Zhang, S. (2024). Endolysosomal trafficking controls yolk granule biogenesis in vitellogenic Drosophila oocytes. PLoS Genet, 20(2):e1011152 PubMed ID: 38315726
Summary:
Endocytosis and endolysosomal trafficking are essential for almost all aspects of physiological functions of eukaryotic cells. As understanding on these membrane trafficking events are mostly from studies in yeast and cultured mammalian cells, one challenge is to systematically evaluate the findings from these cell-based studies in multicellular organisms under physiological settings. One potentially valuable in vivo system to address this challenge is the vitellogenic oocyte in Drosophila, which undergoes extensive endocytosis by Yolkless (Yl), a low-density lipoprotein receptor (LDLR), to uptake extracellular lipoproteins into oocytes and package them into a specialized lysosome, the yolk granule, for storage and usage during later development. However, by now there is still a lack of sufficient understanding on the molecular and cellular processes that control yolk granule biogenesis. By creating genome-tagging lines for Yl receptor and analyzing its distribution in vitellogenic oocytes, close association was observed of different endosomal structures with distinct phosphoinositides and actin cytoskeleton dynamics. Rab5 and Rab11, but surprisingly not Rab4 and Rab7, are essential for yolk granules biogenesis. Instead, evidence was uncovered for a potential role of Rab7 in actin regulation and observed a notable overlap of Rab4 and Rab7, two Rab GTPases that have long been proposed to have distinct spatial distribution and functional roles during endolysosomal trafficking. Through a small-scale RNA interference (RNAi) screen on a set of reported Rab5 effectors, yolk granule biogenesis was shown to largely follow the canonical endolysosomal trafficking and maturation processes. Further, the data suggest that the RAVE/V-ATPase complexes function upstream of or in parallel with Rab7, and are involved in earlier stages of endosomal trafficking events. Together, this study provides s novel insights into endolysosomal pathways and establishes vitellogenic oocyte in Drosophila as an excellent in vivo model for dissecting the highly complex membrane trafficking events in metazoan.

Zeng, B., Knapp, E. M., Skaritanov, E., Oramas, R., Sun, J. (2024). ETS transcription factors regulate precise matrix metalloproteinase expression and follicle rupture in Drosophila. Development, 151(5). PubMed ID: 38345299
Summary:
Drosophila matrix metalloproteinase 2 (MMP2) is specifically expressed in posterior follicle cells of stage-14 egg chambers (mature follicles) and is crucial for the breakdown of the follicular wall during ovulation, a process that is highly conserved from flies to mammals. The factors that regulate spatiotemporal expression of MMP2 in follicle cells remain unknown. This study demonstrates crucial roles for the ETS-family transcriptional activator Pointed (Pnt) and its endogenous repressor Yan in the regulation of MMP2 expression. Pnt was found to be expressed in posterior follicle cells and overlaps with MMP2 expression in mature follicles. Genetic analysis demonstrated that pnt is both required and sufficient for MMP2 expression in follicle cells. In addition, Yan was temporally upregulated in stage-13 follicle cells to fine-tune Pnt activity and MMP2 expression. Furthermore, a 1.1 kb core enhancer was discovered that is responsible for the spatiotemporal expression of MMP2 and contains multiple pnt/yan binding motifs. Mutation of Pnt/Yan binding sites significantly impaired the Mmp2 enhancer activity. These data reveal a mechanism of transcriptional regulation of Mmp2 expression in Drosophila ovulation, which could be conserved in other biological systems.

Dennis, C., Pouchin, P., Richard, G., Mirouse, V. (2024). Basement membrane diversification relies on two competitive secretory routes defined by Rab10 and Rab8 and modulated by dystrophin and the exocyst complex. PLoS Genet, 20(3):e1011169 PubMed ID: 38437244
Summary:
The basement membrane (BM) is an essential structural element of tissues, and its diversification participates in organ morphogenesis. However, the traffic routes associated with BM formation and the mechanistic modulations explaining its diversification are still poorly understood. Drosophila melanogaster follicular epithelium relies on a BM composed of oriented BM fibrils and a more homogenous matrix. This study determined the specific molecular identity and cell exit sites of BM protein secretory routes. First, Rab10 and Rab8 were found to define two parallel routes for BM protein secretion. When both routes were abolished, BM production was fully blocked; however, genetic interactions revealed that these two routes competed. Rab10 promoted lateral and planar-polarized secretion, whereas Rab8 promoted basal secretion, leading to the formation of BM fibrils and homogenous BM, respectively. The dystrophin-associated protein complex (DAPC) and Rab10 were both present in a planar-polarized tubular compartment containing BM proteins. DAPC was essential for fibril formation and sufficient to reorient secretion towards the Rab10 route. Moreover, this study identified a dual function for the exocyst complex in this context. First, the Exo70 subunit directly interacted with dystrophin to limit its planar polarization. Second, the exocyst complex was also required for the Rab8 route. Altogether, these results highlight important mechanistic aspects of BM protein secretion and illustrate how BM diversity can emerge from the spatial control of distinct traffic routes.

Zhang, S. Q., Liu, J. L. (2024)s. Dynamic Cytoophidia during Late-Stage Drosophila Oogenesis. Int J Mol Sci, 25(5) PubMed ID: 38473824
Summary:
CTP synthase (CTPS) catalyzes the final step of de novo synthesis of CTP. CTPS was first discovered to form filamentous structures termed cytoophidia in Drosophila ovarian cells. Subsequent studies have shown that cytoophidia are widely present in cells of three life domains. In the Drosophila ovary model, previous studies mainly focused on the early and middle stages, with less involvement in the later stages. This work focused on the later stages of female germline cells in Drosophila. Live-cell imaging was used to capture the continuous dynamics of cytoophidia in Stages 10-12. The heterogeneity of cytoophidia was noticed in the two types of germline cells (nurse cells and oocytes), manifested in significant differences in morphology, distribution, and dynamics. Surprisingly, it was also found that neighboring nurse cells in the same egg chamber exhibit multiple dynamic patterns of cytoophidia over time. Although the described dynamics may be influenced by the in vitro incubation conditions, this observation provides an initial understanding of the dynamics of cytoophidia during late-stage Drosophila oogenesis.

Mallart, C., Netter, S., Chalvet, F., Claret, S., Guichet, A., Montagne, J., Pret, A. M., Malartre, M. (2024). JAK-STAT-dependent contact between follicle cells and the oocyte controls Drosophila anterior-posterior polarity and germline development. Nat Commun, 15(1):1627 PubMed ID: 38388656
Summary:
The number of embryonic primordial germ cells in Drosophila is determined by the quantity of germ plasm, whose assembly starts in the posterior region of the oocyte during oogenesis. This study reports that extending JAK-STAT activity in the posterior somatic follicular epithelium leads to an excess of primordial germ cells in the future embryo. It was shown that JAK-STAT signaling is necessary for the differentiation of approximately 20 specialized follicle cells maintaining tight contact with the oocyte. These cells define, in the underlying posterior oocyte cortex, the anchoring of the germ cell determinant oskar mRNA. It was revealed that the apical surface of these posterior anchoring cells extends long filopodia penetrating the oocyte. WTwo JAK-STAT targets were identified in these cells that are each sufficient to extend the zone of contact with the oocyte, thereby leading to production of extra primordial germ cells. JAK-STAT signaling thus determines a fixed number of posterior anchoring cells required for anterior-posterior oocyte polarity and for the development of the future germline.

Thursday, October 17th - Cell death

Matamoro-Vidal, A., Cumming, T., Davidovic, A., Levillayer, F., Levayer, R. (2024). Patterned apoptosis has an instructive role for local growth and tissue shape regulation in a fast-growing epithelium. Development, 151(5). PubMed ID: 38215743
Summary:
What regulates organ size and shape remains one fundamental mystery of modern biology. Research in this area has primarily focused on deciphering the regulation in time and space of growth and cell division, while the contribution of cell death has been overall neglected. This includes studies of the Drosophila wing, one of the best-characterized systems for the study of growth and patterning, undergoing massive growth during larval stage and important morphogenetic remodeling during pupal stage. So far, it has been assumed that cell death was relatively neglectable in this tissue both during larval stage and pupal stage, and as a result, the pattern of growth was usually attributed to the distribution of cell division. Using systematic mapping and registration combined with quantitative assessment of clone size and disappearance as well as live imaging, this paper outline a persistent pattern of cell death and clone elimination emerging in the larval wing disc and persisting during pupal wing morphogenesis. Local variation of cell death is associated with local variation of clone size, pointing to an impact of cell death on local growth that is not fully compensated by proliferation. Using morphometric analyses of adult wing shape and genetic perturbations, evidence is provided that patterned death locally and globally affects adult wing shape and size. This study describes a roadmap for precise assessment of the contribution of cell death to tissue shape and outlines an important instructive role of cell death in modulating quantitatively local growth and morphogenesis of a fast-growing tissue.

Tan, J. Y. K., Chew, L. Y., Juhasz, G., Yu, F. (2024). Interplay between autophagy and CncC regulates dendrite pruning in Drosophila. Proc Natl Acad Sci U S A, 121(10):e2310740121 PubMed ID: 38408233
Summary:
Autophagy is essential for the turnover of damaged organelles and long-lived proteins. It is responsible for many biological processes such as maintaining brain functions and aging. Impaired autophagy is often linked to neurodevelopmental and neurodegenerative diseases in humans. However, the role of autophagy in neuronal pruning during development remains poorly understood. This study reports that autophagy regulates dendrite-specific pruning of ddaC sensory neurons in parallel to local caspase activation. Impaired autophagy causes the formation of ubiquitinated protein aggregates in ddaC neurons, dependent on the autophagic receptor Ref(2)P. Furthermore, the metabolic regulator AMP-activated protein kinase and the insulin-target of rapamycin pathway act upstream to regulate autophagy during dendrite pruning. Importantly, autophagy is required to activate the transcription factor CncC (Cap "n" collar isoform C), thereby promoting dendrite pruning. Conversely, CncC also indirectly affects autophagic activity via proteasomal degradation, as impaired CncC results in the inhibition of autophagy through sequestration of Atg8a into ubiquitinated protein aggregates. Thus, this study demonstrates the important role of autophagy in activating CncC prior to dendrite pruning, and further reveals an interplay between autophagy and CncC in neuronal pruning.

Shinno, K., Miura, Y., Iijima, K. M., Suzuki, E., Ando, K. (2024). Axonal distribution of mitochondria maintains neuronal autophagy during aging via eIF2beta. bioRxiv, PubMed ID: 38293064
Summary:
Neuronal aging and neurodegenerative diseases are accompanied by proteostasis collapse, while cellular factors that trigger it are not identified. Impaired mitochondrial transport in the axon is another feature of aging and neurodegenerative diseases. Using Drosophila, this study found that genetic depletion of axonal mitochondria causes dysregulation of translation and protein degradation. Axons with mitochondrial depletion showed abnormal protein accumulation, and autophagic defects. Lowering neuronal ATP levels by blocking glycolysis did not reduce autophagy, suggesting that autophagic defects are associated with mitochondrial distribution. eIF2β was found to be upregulated by depletion of axonal mitochondria via proteome analysis. Phosphorylation of eIF2α, another subunit of eIF2, was lowered, and global translation was suppressed. Neuronal overexpression of eIF2β phenocopied the autophagic defects and neuronal dysfunctions, and lowering eIF2β expression rescued those perturbations caused by depletion of axonal mitochondria. These results indicate the mitochondria-eIF2β axis maintains proteostasis in the axon, of which disruption may underly the onset and progression of age-related neurodegenerative diseases.

Nicolson, S., Manning, J. A., Lim, Y., Jiang, X., Kolze, E., Dayan, S., Umargamwala, R., Xu, T., Sandow, J. J., Webb, A. I., Kumar, S., Denton, D. (2024). The Drosophila ZNRF1/2 homologue, detour, interacts with HOPS complex and regulates autophagy. Communications biology, 7(1):183 PubMed ID: 38360932
Summary:
Autophagy, the process of elimination of cellular components by lysosomal degradation, is essential for animal development and homeostasis. Using the autophagy-dependent Drosophila larval midgut degradation model this study identified an autophagy regulator, the RING domain ubiquitin ligase CG14435 (detour). Depletion of detour resulted in increased early-stage autophagic vesicles, premature tissue contraction, and overexpression of detour or mammalian homologues, ZNRF1 and ZNRF2, increased autophagic vesicle size. The ablation of ZNRF1 or ZNRF2 in mammalian cells increased basal autophagy. WDetour interacting proteins were identified including HOPS subunits, deep orange (dor/VPS18), Vacuolar protein sorting 16A (VPS16A), and light (lt/VPS41); detour promotes their ubiquitination. The detour mutant accumulated autophagy-related proteins in young adults, displayed premature ageing, impaired motor function, and activation of innate immunity. Collectively, these findings suggest a role for detour in autophagy, likely through regulation of HOPS complex, with implications for healthy aging.

Tian, L., Li, Y., Shi, Y. (2024). Dark and Dronc activation in Drosophila melanogaster. Proc Natl Acad Sci U S A, 121(9):e2312784121 PubMed ID: 38381783
Summary:
The onset of apoptosis is characterized by a cascade of caspase activation, where initiator caspases are activated by a multimeric adaptor complex known as the apoptosome. In Drosophila melanogaster, the initiator caspase Dronc undergoes autocatalytic activation in the presence of the Dark apoptosome. Despite rigorous investigations, the activation mechanism for Dronc remains elusive. This study reports the cryo-EM structures of an auto-inhibited monomer and a single-layered, multimeric Dark/Dronc complex. Biochemical analysis suggests that the auto-inhibited Dark oligomerizes upon binding to Dronc, which is sufficient for the activation of both Dark and Dronc. In contrast, the previously observed double-ring Dark apoptosome may represent a non-functional or "off-pathway" conformation. These findings expand understanding on the molecular mechanism of apoptosis in Drosophila.

Pak, T. F., Pitt-Francis, J., Baker, R. E. (2024). A mathematical framework for the emergence of winners and losers in cell competition. Journal of theoretical biology, 577:111666 PubMed ID: 37956955
Summary:
Cell competition is a process in multicellular organisms where cells interact with their neighbours to determine a "winner" or "oser" status. The loser cells are eliminated through programmed cell death, leaving only the winner cells to populate the tissue. Cell competition is context-dependent; the same cell type can win or lose depending on the cell type it is competing against. Hence, winner/loser status is an emergent property. A key question in cell competition is: how do cells acquire their winner/loser status? In this paper, a mathematical framework is proposed for studying the emergence of winner/loser status based on a set of quantitative criteria that distinguishes competitive from non-competitive outcomes. This framework is applied in a cell-based modelling context to both highlight the crucial role of active cell death in cell competition and identify the factors that drive cell competition.

Wednesday, October 16th - Disease Models

Comparative exploration of mammalian deafness gene homologues in the Drosophila auditory organ shows genetic correlation between insect and vertebrate hearing Sutton, D. C., Andrews, J. C., Dolezal, D. M., Park, Y. J., Li, H., Eberl, D. F., Yamamoto, S., Groves, A. K. (2024). . PLoS One, 19(2):e0297846 PubMed ID: 38412189
Summary:
Johnston's organ, the Drosophila auditory organ, is anatomically very different from the mammalian organ of Corti. However, recent evidence indicates significant cellular and molecular similarities exist between vertebrate and invertebrate hearing, suggesting that Drosophila may be a useful platform to determine the function of the many mammalian deafness genes whose underlying biological mechanisms are poorly characterized. The goal of this study was a comprehensive screen of all known orthologues of mammalian deafness genes in the fruit fly to better understand conservation of hearing mechanisms between the insect and the fly and ultimately gain insight into human hereditary deafness. Bioinformatic comparisons were used to screen previously reported human and mouse deafness genes; 156 of them have orthologues in Drosophila melanogaster. Fluorescent imaging of T2A-GAL4 gene trap and GFP or YFP fluorescent protein trap lines were examined for 54 of the Drosophila genes; 38 were found to be expressed in different cell types in Johnston's organ. The function of strong loss-of-function mutants in three genes expressed in Johnston's organ (Cad99C, Msp-300, and Koi) was phenotypically characterized using a courtship assay and electrophysiological recordings of sound-evoked potentials. Cad99C and klaroid (koi) were found to have significant courtship defects. However, when these genes were tested for electrophysiological defects in hearing response, no significant difference was observed suggesting the courtship defects were not caused by hearing deficiencies. Furthermore, a UAS/RNAi approach was used to test the function of seven genes and found two additional genes, CG5921 and Myo10a, that gave a statistically significant delay in courtship but not in sound-evoked potentials. These results suggest that many mammalian deafness genes have Drosophila homologues expressed in the Johnston's organ, but that their requirement for hearing may not necessarily be the same as in mammals.

Imomnazarov, K., Lopez-Scarim, J., Bagheri, I., Joers, V., Tansey, M. G., Martín-Peña, A. (2024). Biochemical fractionation of human α-Synuclein in a Drosophila model of synucleinopathies. bioRxiv, PubMed ID: 38370694
Summary:
Synucleinopathies are a group of central nervous system pathologies that are characterized by neuronal accumulation of misfolded and aggregated α-synuclein in proteinaceous depositions known as Lewy Bodies (LBs). The transition of α-synuclein from its physiological to pathological form has been associated with several post-translational modifications such as phosphorylation and an increasing degree of insolubility, which also correlate with disease progression in postmortem specimens from human patients. Neuronal expression of α-synuclein in model organisms, including Drosophila melanogaster, has been an approach to study its physiological effects. Biochemical analysis of α-synuclein solubility via high-speed ultracentrifugation with buffers of increasing detergent strength offers a potent method for identification of α-synuclein biochemical properties and the associated pathology stage. This study tested different detergents for their ability to solubilize human α-synuclein carrying the pathological mutation A53T from brains of aged flies. The effect of sonication on solubility of human α-synuclein was also tested and was optimized to discriminate relative amounts of soluble/insoluble human α-synuclein from dopaminergic neurons of the Drosophila brain. The data established that, using a 5% SDS buffer, the 3-step protocol distinguishes between cytosolic soluble proteins in fraction 1, detergent-soluble proteins in fraction 2 and insoluble proteins in fraction 3. This protocol shows that sonication breaks down α-synuclein insoluble complexes from the fly brain, making them soluble in the SDS buffer and enriching fraction 2 of the protocol.

Perlegos, A. E., Byrns, C. N., Bonini, N. M. (2024). Cell type-specific regulation of m(6) A modified RNAs in the aging Drosophila brain. Aging Cell, 23(3):e14076 PubMed ID: 38205931
Summary:
The aging brain is highly vulnerable to cellular stress, and neurons employ numerous mechanisms to combat neurotoxic proteins and promote healthy brain aging. The RNA modification m(6) A is highly enriched in the Drosophila brain and is critical for the acute heat stress response of the brain. This study examined m(6) A in the fly brain with the chronic stresses of aging and degenerative disease. m(6) A levels dynamically increased with both age and disease in the brain, marking integral neuronal identity and signaling pathway transcripts that decline in level with age and disease. Unexpectedly, there is opposing impact of m(6) A transcripts in neurons versus glia, which conferred different outcomes on animal health span upon Mettl3 knockdown to reduce m(6) A: whereas Mettl3 function is normally beneficial to neurons, it is deleterious to glia. Moreover, knockdown of Mettl3 in glial tauopathy reduced tau pathology and increased animal survival. These findings provide mechanistic insight into regulation of m(6) A modified transcripts with age and disease, highlighting an overall beneficial function of Mettl3 in neurons in response to chronic stresses, versus a deleterious impact in glia.

Reynolds, C. J., Gillen, C. M., Burke, R., Tsering, Y., Loucks, E., Judd-Mole, S., Dow, J. A. T., Romero, M. F. (2024). Drosophila ClC-c Is a Homolog of Human CLC-5 and a New Model for Dent Disease Type 1. Kidney360, 5(3):414-426 PubMed ID: 38233994
Summary:
Drosophila can be a model for Dent Disease type 1. Drosophila Chloride channel-c (Clc-C) mutations function similar to human CLC-5 Dent 1 mutations. The Drosophila ClC-c (CG5284) has sequence homology with human ClC-5. Ion transport function and activity of Drosophila ClC-c and homologous DD1 variants were assessed by voltage clamp electrophysiology. Membrane localization was demonstrated in Drosophila expressing a GFP-labeled construct of ClC-c. Genetic expression of an RNAi against ClC-c mRNA was used to generate a knockdown fly that serves as a DD1 disease model. Tubule secretion of cations and protein were assessed, as well as the crystal formation in the Malpighian tubules. Voltage clamp experiments demonstrate that ClC-c is voltage-gated with Cl(−)-dependent and pH-sensitive currents. Inclusion of homologous DD1 mutations pathogenic variants (S393L, R494W, and Q777X) impairs ClC-c ion transport activity. In vivo expression of ClC-c-eGFP in Malpighian tubules reveals that the membrane transporter localizes to the apical membrane and nearby cytosolic regions. RNAi knockdown of ClC-c (48% decreased mRNA expression) causes increased secretion of both urinary protein and Ca(2+) and increased occurrence of spontaneous tubule crystals. It is concluded that Drosophila ClC-c shows orthologous function and localization to human ClC-5. Thus, Drosophila and ClC-c regulation may be useful for future investigations of Cl(−) transport, Ca(2+) homeostasis, and urinary protein loss in DD1.

Dubey, S. K., Lloyd, T. E., Tapadia, M. G. (2024). Disrupted nuclear import of cell cycle proteins in Huntington's/PolyQ disease causes neurodevelopment defects in cellular and Drosophila model. Heliyon, 10(4):e26393 PubMed ID: 38434042
Summary:
Huntington's disease is caused by an expansion of CAG repeats in exon 1 of the huntingtin gene encoding an extended PolyQ tract within the Huntingtin protein (mHtt). This expansion results in selective degeneration of striatal medium spiny projection neurons in the basal ganglia. The mutation causes abnormalities during neurodevelopment in human and mouse models. This study reports that mHtt/PolyQ aggregates inhibit the cell cycle in the Drosophila brain during development. PolyQ aggregates disrupt the nuclear pore complexes of the cells preventing the translocation of cell cycle proteins such as Cks30A, Cyclin E, E2F and PCNA from cytoplasm to the nucleus, thus affecting cell cycle progression. PolyQ aggregates also disrupt the nuclear pore complex and nuclear import in mHtt expressing mammalian CAD neurons. PolyQ toxicity and cell cycle defects can be restored by enhancing RanGAP-mediated nuclear import, suggesting a potential therapeutic approach for this disease.

Hussain, R., Lim, C. X., Shaukat, Z., Islam, A., Caseley, E. A., Lippiat, J. D., Rychkov, G. Y., Ricos, M. G., Dibbens, L. M. (2024). Drosophila expressing mutant human KCNT1 transgenes make an effective tool for targeted drug screening in a whole animal model of KCNT1-epilepsy. Sci Rep, 14(1):3357 PubMed ID: 38336906
Summary:
Mutations in the KCNT1 potassium channel cause severe forms of epilepsy which are poorly controlled with current treatments. In vitro studies have shown that KCNT1-epilepsy mutations are gain of function, significantly increasing K(+) current amplitudes. To investigate if Drosophila can be used to model human KCNT1 epilepsy, Drosophila melanogaster lines carrying human KCNT1 were generated with the patient mutation G288S, R398Q or R928C. Expression of each mutant channel in GABAergic neurons gave a seizure phenotype which responded either positively or negatively to 5 frontline epilepsy drugs most commonly administered to patients with KCNT1-epilepsy, often with little or no improvement of seizures. Cannabidiol showed the greatest reduction of the seizure phenotype while some drugs increased the seizure phenotype. This study shows that Drosophila has the potential to model human KCNT1- epilepsy and can be used as a tool to assess new treatments for KCNT1- epilepsy.

Tuesday, October 15th - Behavior

Noer, N. K., Rohde, P. D., Sorensen, P., Bahrndorff, S., Kristensen, T. N. (2024). Diurnal variation in genetic parameters for locomotor activity in Drosophila melanogaster assessed under natural thermal conditions. J Evol Biol, 37(3):336-345 PubMed ID: 38320319
Summary:
In nature, organisms are exposed to variable and occasionally stressful environmental conditions. Responses to diurnal and seasonal fluctuations, such as temperature and food accessibility, involve adaptive behavioural and physiological changes. While much work has been done on understanding the genetic architecture and evolutionary potential of stress tolerance traits under constant thermal conditions, there has been less focus on the quantitative genetic background in variable environments. In this study, the Drosophila Genetic Reference Panel (DGRP) was used to investigate the locomotor activity, a key behavioural trait, under variable natural thermal conditions during the summer in a temperate environment. Male flies from 100 DGRP lines were exposed to natural thermal and light conditions in Drosophila activity monitors across three experimental days. Activity was found to be highly temperature and time dependent and varied between lines both within and between days. Furthermore, variation was observed in genetic and environmental variance components, with low to moderate estimates of the heritability for locomotor activity, consistently peaking in the afternoons. Moreover, the estimated genetic correlations of locomotor activity between two time points was shown to decrease, as the absolute differences in ambient temperature increased. In conclusion, this study found that the genetic background for locomotor activity is environment specific, and it is concluded that more variable and unpredictable future temperatures will likely have a strong impact on the evolutionary trajectories of behavioural traits in ectotherms.

Ryvkin, J., Omesi, L., Kim, Y. K., ..., Nassel, D. R., Heberlein, U., Shohat-Ophir, G. (2024). Failure to mate enhances investment in behaviors that may promote mating reward and impairs the ability to cope with stressors via a subpopulation of Neuropeptide F receptor neurons. PLoS Genet, 20(1):e1011054 PubMed ID: 38236837
Summary:
Living in dynamic environments such as the social domain, where interaction with others determines the reproductive success of individuals, requires the ability to recognize opportunities to obtain natural rewards and cope with challenges that are associated with achieving them. As such, actions that promote survival and reproduction are reinforced by the brain reward system, whereas coping with the challenges associated with obtaining these rewards is mediated by stress-response pathways, the activation of which can impair health and shorten lifespan. As a model system to study the impact of failure to obtain a natural reward, the well-established courtship suppression paradigm in Drosophila melanogaster was used as means to induce repeated failures to obtain sexual reward in male flies. It was discovered that beyond the known reduction in courtship actions caused by interaction with non-receptive females, repeated failures to mate induce a stress response characterized by persistent motivation to obtain the sexual reward, reduced male-male social interaction, and enhanced aggression. This frustrative-like state caused by the conflict between high motivation to obtain sexual reward and the inability to fulfill their mating drive impairs the capacity of rejected males to tolerate stressors such as starvation and oxidative stress. It was further shown that sensitivity to starvation and enhanced social arousal is mediated by the disinhibition of a small population of neurons that express receptors for the fly homologue of neuropeptide Y. These findings demonstrate for the first time the existence of social stress in flies and offers a framework to study mechanisms underlying the crosstalk between reward, stress, and reproduction in a simple nervous system that is highly amenable to genetic manipulation.

Wu, Y., Wang, Q., Yang, W., Zhang, S., Mao, C. X., He, N., Zhou, S., Zhou, C., Liu, W. (2023). The cluster digging behavior of larvae confers trophic benefits to fitness in insects. Insect Sci, PubMed ID: 38161191
Summary:
Collective behaviors efficiently impart benefits to a diversity of species ranging from bacteria to humans. Fly larvae tend to cluster and form coordinated digging groups under crowded conditions, yet understanding the rules governing this behavior is in its infancy. This study primarily took advantage of the Drosophila model to investigate cooperative foraging behavior. Drosophila-related species and the black soldier fly have evolved a conserved strategy of cluster digging in food foraging. Subsequently, relative factors were investigated, including larval stage, population density, and food stiffness and quality, that affect the cluster digging behavior. Remarkably, oxygen supply through the posterior breathing spiracles is necessary for the organization of digging clusters. More importantly, a mathematical model was devised to accurately calculate how the cluster digging behavior expands food resources by diving depth, cross-section area, and food volume. Cluster digging behavior was found to approximately increases 2.2 fold depth, 1.7-fold cross-section area, and 2.4 fold volume than control groups, respectively. Amplification of food sources significantly facilitates survival, larval development, and reproductive success of Drosophila challenged with competition for limited food resources, thereby conferring trophic benefits to fitness in insects. Overall, these findings highlight that the cluster digging behavior is a pivotal behavior for their adaptation to food scarcity, advancing a better understanding of how this cooperative behavior confers fitness benefits in the animal kingdom.

Westeinde, E. A., Kellogg, E., Dawson, P. M., Lu, J., Hamburg, L., Midler, B., Druckmann, S., Wilson, R. I. (2024). Transforming a head direction signal into a goal-oriented steering command. Nature, 626(8000):819-826 PubMed ID: 38326621
Summary:
To navigate, heading direction must be continuously estimated, and deviations from the goal must be extimated. Direction estimation is accomplished by ring attractor networks in the head direction system. However, it is not fully understood how the sense of direction is used to guide action. Drosophila connectome analyses( reveals three cell populations (PFL3R, PFL3L and PFL2) that connect the head direction system to the locomotor system. This study used imaging, electrophysiology and chemogenetic stimulation during navigation to show how these populations function. Each population receives a shifted copy of the head direction vector, such that their three reference frames are shifted approximately 120%deg; relative to each other. Each cell type then compares its own head direction vector with a common goal vector; specifically, it evaluates the congruence of these vectors via a nonlinear transformation. The output of all three cell populations is then combined to generate locomotor commands. PFL3R cells are recruited when the fly is oriented to the left of its goal, and their activity drives rightward turning; the reverse is true for PFL3L. Meanwhile, PFL2 cells increase steering speed, and are recruited when the fly is oriented far from its goal. PFL2 cells adaptively increase the strength of steering as directional error increases, effectively managing the tradeoff between speed and accuracy. Together, these results show how a map of space in the brain can be combined with an internal goal to generate action commands, via a transformation from world-centric coordinates to body-centric coordinates.

Gautham, A. K., Miner, L. E., Franco, M. N., Thornquist, S. C., Crickmore, M. A. (2024). Molecular control of temporal integration matches decision-making to motivational state. BioRxiv, PubMed ID: 38496671
Summary:
Motivations bias responses to stimuli, producing behavioral outcomes that match ones needs and goals. This study describes a mechanism behind this phenomenon: adjusting the time over which stimulus-derived information is permitted to accumulate toward a decision. As a Drosophila copulation progresses, the male becomes less likely to continue mating through challenges. This study shows that a set of Copulation Decision Neurons (CDNs) flexibly integrates information about competing drives to mediate this decision. Early in mating, dopamine signaling restricts CDN integration time by potentiating CaMKII activation in response to stimulatory inputs, imposing a high threshold for changing behaviors. Later into mating, the timescale over which the CDNs integrate termination-promoting information expands, increasing the likelihood of switching behaviors. Scalable windows of temporal integration at dedicated circuit nodes are suggested as a key but underappreciated variable in state-based decision-making.

Eiman, M. N., Kumar, S., Serrano Negron, Y. L., Tansey, T. R., Harbison, S. T. (2024). Genome-wide association in Drosophila identifies a role for Piezo and Proc-R in sleep latency. Sci Rep, 14(1):260 PubMed ID: 38168575
Summary:
Sleep latency, the amount of time that it takes an individual to fall asleep, is a key indicator of sleep need. Sleep latency varies considerably both among and within species and is heritable, but lacks a comprehensive description of its underlying genetic network. This study conductd a genome-wide association study of sleep latency. Using previously collected sleep and activity data on a wild-derived population of flies, sleep latency was calculated, confirming significant, heritable genetic variation for this complex trait. 520 polymorphisms were identified in 248 genes contributing to variability in sleep latency. Tests of mutations in 23 candidate genes and additional putative pan-neuronal knockdown of 9 of them implicated CG44153, Piezo, Proc-R and Rbp6 in sleep latency. Two large-effect mutations in the genes Proc-R and Piezo were further confirmed via genetic rescue. This work greatly enhances understanding of the genetic factors that influence variation in sleep latency.

Monday, October 14th - Adult Physiology and Metabolism/h3>

Mudunuri, A., Chandrakanth, M., Khan, S., Sura, C., Kumar, N., Tung, S. (2024). Diet-induced plasticity of life-history traits and gene expression in outbred Drosophila melanogaster population. Ecology and evolution, 14(2):e10976 PubMed ID: 38343564
Summary:
Food is fundamental for the survival of organisms, governing growth, maintenance, and reproduction through the provision of essential macronutrients. However, access to food with optimum macronutrient composition, which will maximize the evolutionary fitness of an organism, is not always guaranteed. This leads to dietary mismatches with potential impacts on organismal performance. To understand the consequences of such dietary mismatches, this study examined the effects of isocaloric diets varying in macronutrient composition on eight key organismal traits spanning across the lifespan of a large outbred Drosophila melanogaster population (n ~ 2500). The findings reveal that carbohydrate-reduced isocaloric diets correlates to accelerated pre-adult development and boosts reproductive output without impacting pre-adult viability and body size. Conversely, an elevated dietary carbohydrate content correlated to reduced lifespan in flies, evidenced by accelerated functional senescence including compromised locomotor activity and deteriorating gut integrity. Furthermore, transcriptomic analysis indicated a substantial difference in gene regulatory landscapes between flies subject to high-carbohydrate versus high-protein diet, with elevated protein levels indicating transcriptomes primed for reduced synthesis of fatty acids. Taken together, this study helps advance understanding of the effect of macronutrient composition on life history traits and their interrelations, offering critical insights into potential adaptive strategies that organisms might adopt against the continual dietary imbalances prevalent in the rapidly evolving environment.

Privalova, V., Sobczyk, L., Szlachcic, E., Labecka, A. M., Czarnoleski, M. (2024). Heat tolerance in Drosophila melanogaster is influenced by oxygen conditions and mutations in cell size control pathways. Philosophical transactions of the Royal Society of London Series B, Biological sciences, 379(1896):20220490 PubMed ID: 38186282
Summary:
Understanding metabolic performance limitations is key to explaining the past, present and future of life. This study investigated whether heat tolerance in actively flying Drosophila melanogaster is modified by individual differences in cell size and the amount of oxygen in the environment. Two mutants were used with loss-of-function mutations in cell size control associated with the target of rapamycin (TOR)/insulin pathways, showing reduced (mutant rictor(Δ2)) or increased (mutant Mnt(1)) cell size in different body tissues compared to controls. Flies were exposed to a steady increase in temperature under normoxia and hypoxia until they collapsed. The upper critical temperature decreased in response to each mutation type as well as under hypoxia. Females, which have larger cells than males, had lower heat tolerance than males. Altogether, mutations in cell cycle control pathways, differences in cell size and differences in oxygen availability affected heat tolerance, but existing theories on the roles of cell size and tissue oxygenation in metabolic performance can only partially explain the results. A better understanding of how the cellular composition of the body affects metabolism may depend on the development of research models that help separate various interfering physiological parameters from the exclusive influence of cell size. This article is part of the theme issue 'The evolutionary significance of variation in metabolic rates'.

Chiang, A. C. Y., Jezek, J., Mu, P., Di, Y., Klucnika, A., Jaburek, M., Jezek, P., Ma, H. (2024). Two mitochondrial DNA polymorphisms modulate cardiolipin binding and lead to synthetic lethality. Nat Commun, 15(1):611 PubMed ID: 38242869
Summary:
Genetic screens have been used extensively to probe interactions between nuclear genes and their impact on phenotypes. Probing interactions between mitochondrial genes and their phenotypic outcome, however, has not been possible due to a lack of tools to map the responsible polymorphisms. Using a toolkit previous work established in Drosophila, over 300 recombinant mitochondrial genomes were isolated and a naturally occurring polymorphism was mapped at the cytochrome c oxidase III residue 109 (CoIII109) that fully rescues the lethality and other defects associated with a point mutation in cytochrome c oxidase I (CoIT300I). Cardiolipin is an essential phospholipid that makes up to 15%–20% of the inner mitochondrial membrane. Through lipidomics profiling, biochemical assays and phenotypic analyses, this study showed that the CoIII109 polymorphism modulates binding of cardiolipin, an essential phospholipid that makes up to 15%–20% of the inner mitochondrial membrane, binding to prevent complex IV instability caused by the CoIT300I mutation. This study demonstrates the feasibility of genetic interaction screens in animal mitochondrial DNA. It unwraps the complex intra-genomic interplays underlying disorders linked to mitochondrial DNA and how they influence disease expression.

Mendoza-Grimau, V., Perez-Galvez, A., Busturia, A., Fontecha, J. (2024). Lipidomic profiling of Drosophila strains Canton-S and white(1118) reveals intraspecific lipid variations in basal metabolic rate. rostaglandins, leukotrienes, and essential fatty acids, 201:102618 PubMed ID: 38795635
Summary:
Drosophila melanogaster is a well-established model system for studies on lipid metabolism and energy homeostasis. In this study, the main components of the lipid profile of two widely utilized Drosophila strains were identified and quantified, namely Canton-S and white(1118), under identical experimental conditions. Differences observed between the strains can be attributed to inherent metabolic divergences, thus limiting the influence of confounding factors. Using the comprehensive lipid data acquired, cluster analysis and PLS-DA techniques were appled to ascertain whether the lipidome could effectively differentiate between the strains. Certain lipid features, such as triacylglycerols, polar lipids, and specific sterol components, could be distinguished between flies of both strains regardless of sex. These results suggest that although Canton-S and white(1118) have similar lipid profiles and distributions, a selected subset of lipids demonstrates clear discriminatory potential between strains, thereby bearing significant implications for planning biological studies using these strains as control references.

Mure, A., Sugiura, Y., Maeda, R., Honda, K., Sakurai, N., Takahashi, Y., Watada, M., Katoh, T., Gotoh, A., Gotoh, Y., Taniguchi, I., Nakamura, K., Hayashi, T., Katayama, T., Uemura, T., Hattori, Y. (2023). Identification of key yeast species and microbe-microbe interactions impacting larval growth of Drosophila in the wild. Elife, 12 PubMed ID: 38150375
Summary:
Microbiota consisting of various fungi and bacteria have a significant impact on the physiological functions of the host. However, it is unclear which species are essential to this impact and how they affect the host. This study analyzed and isolated microbes from natural food sources of Drosophila larvae, and investigated their functions. Hanseniaspora uvarum is the predominant yeast responsible for larval growth in the earlier stage of fermentation. As fermentation progresses, Acetobacter orientalis emerges as the key bacterium responsible for larval growth, although yeasts and lactic acid bacteria must coexist along with the bacterium to stabilize this host-bacterial association. By providing nutrients to the larvae in an accessible form, the microbiota contributes to the upregulation of various genes that function in larval cell growth and metabolism. Thus, this study elucidates the key microbial species that support animal growth under microbial transition.

Betz, L. S., DiAngelo, J. R. (2024). The regulation of triglyceride and glycogen storage by Glucose transporter 1 ( Glut1 ) in Drosophila fat tissue. microPublication biology, 2024 PubMed ID: 38495587
Summary:
Obesity reflects an imbalance in nutrient storage resulting in excess fat accumulation. The molecules that tissues use to regulate nutrient storage are not well understood. A previously published genetic screen using Drosophila melanogaster larvae identified Glut1 , a transmembrane glucose transporter, as a potential obesity gene. To identify the adipose-specific functions of this gene, Glut1 levels were decreased using RNAi targeted to fly fat tissue. Adult Glut1 RNAi flies have lower glycogen and triglyceride levels, as well as decreased FASN1 RNA expression. This suggests that Glut1 functions to promote glycogen and triglyceride storage and fatty acid synthesis in Drosophila adipose tissue.

Thursday, October 10th - Stem Cells

Li, W. X. (2023). Computational simulation of JAK/STAT signaling in somatic versus germline stem cells. Dev Dyn, PubMed ID: 38126664
Summary:
The Janus kinase/signal transducer and activator of transcription (JAK/STAT) signaling pathway regulates a variety of cellular processes. A major activation event in this pathway involves the phosphorylation of a tyrosine of STAT, converting unphosphorylated STAT (uSTAT) to phosphorylated STAT (pSTAT), an active transcription factor. In a noncanonical role, uSTAT contributes to the maintenance of heterochromatin stability. As such, an increase in pSTAT concurrently reduces uSTAT, resulting in heterochromatin loss, as observed in Drosophila somatic tissues. Paradoxically, an opposing phenomenon occurs in Drosophila male germline stem cells (GSCs), where the JAK/STAT pathway remains persistently active due to a continuous supply of ligands. For this study, computational simulations were employed to dissect JAK/STAT pathway activation under different cellular contexts, mimicking somatic and germline cells. In these simulations, ordinary differential equations were leveraged to replicate the chemical reactions governing JAK/STAT signaling under different conditions. The outcomes indicate that transient ligand stimulation, typical in somatic tissues, led to a momentary reduction in uSTAT levels. Conversely, sustained ligand stimulation, a characteristic feature of the GSC niche, resulted in elevated uSTAT levels at equilibrium. The simulation suggests that the duration of ligand exposure could explain the observed opposite effects of JAK/STAT activation on heterochromatin in somatic versus GSCs.

Sood, C., Nahid, M. A., Branham, K. R., Pahl, M., Doyle, S. E., Siegrist, S. E. (2024). Delta-dependent Notch activation closes the early neuroblast temporal program to promote lineage progression and neurogenesis termination in Drosophila. Elife, 12 PubMed ID: 38391176
Summary:
Neuroblasts in Drosophila divide asymmetrically, sequentially expressing a series of intrinsic factors to generate a diversity of neuron types. These intrinsic factors known as temporal factors dictate timing of neuroblast transitions in response to steroid hormone signaling and specify early versus late temporal fates in neuroblast neuron progeny. After completing their temporal programs, neuroblasts differentiate or die, finalizing both neuron number and type within each neuroblast lineage. From a screen aimed at identifying genes required to terminate neuroblast divisions, this study identified Notch and Notch pathway components. When Notch is knocked down, neuroblasts maintain early temporal factor expression longer, delay late temporal factor expression, and continue dividing into adulthood. Delta, expressed in cortex glia, neuroblasts, and after division, their GMC progeny, regulates neuroblast Notch activity. Delta in neuroblasts is expressed high early, low late, and is controlled by the intrinsic temporal program: early factor Imp promotes Delta, late factors Syp/E93 reduce Delta. Thus, in addition to systemic steroid hormone cues, forward lineage progression is controlled by local cell-cell signaling between neuroblasts and their cortex glia/GMC neighbors: Delta transactivates Notch in neuroblasts bringing the early temporal program and early temporal factor expression to a close.

Hof-Michel, S., Cigoja, L., Huhn, S., Bokel, C. (2023). Rel governs loser elimination during stem cell competition in the Drosophila testis.European journal of cell biology, 103(1):151375 PubMed ID: 37995529
Summary:
In the Drosophila testis, a group of stromal cells termed hub provides multiple niche signals for the surrounding germline and somatic stem cells. Stem cells of both populations compete for physical retention in the niche, and clones unable to transduce any one niche signal are rapidly eliminated from the stem cell pool by differentiation. This study mapped the transcriptomes of isolated somatic cyst stem cells and differentiated cyst cells; the stem cells but not their differentiated progeny exhibit the signature of an innate immune response including the NF-κB transcription factor Relish (Rel). Related signalling pathways had previously implicated in cell competition in larval epithelia, prompting the question of whether NF-κB signalling was, despite the clear differences between the two competition scenarios, also involved in stem cell competition in the testis. This study shows 1) that in the testis Rel is dispensable for stemness, 2) that loss of Rel or the upstream receptor Toll suppresses loser elimination following a variety of different triggers used to induce loser fate, and 3) that clonal Rel activation is sufficient for the displacement of neutral or winner cells from the niche, even if these cells otherwise retain stem cell properties.

Veneti, Z., Fasoulaki, V., Kalavros, N., Vlachos, I. S., Delidakis, C., Eliopoulos, A. G. (2024). Polycomb-mediated silencing of miR-8 is required for maintenance of intestinal stemness in Drosophila melanogaster. Nat Commun, 15(1):1924 PubMed ID: 38429303
Summary:
Balancing maintenance of self-renewal and differentiation is a key property of adult stem cells. The epigenetic mechanisms controlling this balance remain largely unknown. This study reports that the Polycomb Repressive Complex 2 (PRC2) is required for maintenance of the intestinal stem cell (ISC) pool in the adult female Drosophila melanogaster. Lloss of PRC2 activity in ISCs by RNAi-mediated knockdown or genetic ablation of the enzymatic subunit Enhancer of zeste, E(z), results in loss of stemness and precocious differentiation of enteroblasts to enterocytes. Mechanistically, he microRNA miR-8 was identified as a critical target of E(z)/PRC2-mediated tri-methylation of histone H3 at Lys27 (H3K27me3) and uncovered a dynamic relationship between E(z), miR-8 and Notch signaling in controlling stemness versus differentiation of ISCs. Collectively, these findings uncover a hitherto unrecognized epigenetic layer in the regulation of stem cell specification that safeguards intestinal homeostasis.

Ridwan, S. M., Twillie, A., Poursaeid, S., Beard, E. K., Bener, M. B., Antel, M., Cowan, A. E., Matsuda, S., Inaba, M. (2024). Diffusible fraction of niche BMP ligand safeguards stem-cell differentiation. Nat Commun, 15(1):1166 PubMed ID: 38326318
Summary:
Drosophila male germline stem cells (GSCs) reside at the tip of the testis and surround a cluster of niche cells. Decapentaplegic (Dpp) is one of the well-established ligands and has a major role in maintaining stem cells located in close proximity. However, the existence and the role of the diffusible fraction of Dpp outside of the niche have been unclear. using genetically-encoded nanobodies called Morphotraps, this study physically blocked Dpp diffusion without interfering with niche-stem cell signaling; a diffusible fraction of Dpp was found to be required to ensure differentiation of GSC daughter cells, opposite of its role in maintenance of GSC in the niche. This work provides an example in which a soluble niche ligand induces opposed cellular responses in stem cells versus in differentiating descendants to ensure spatial control of the niche. This may be a common mechanism to regulate tissue homeostasis.

Sadanandappa, M. K., Bosco, G. (2024). Parasitoid cues modulate Drosophila germline development and stem cell proliferation. Cell Rep, 43(1):113657 PubMed ID: 38175752
Summary:
Environmental factors influence an organism's reproductive ability by regulating germline development and physiology. While the reproductive adaptations in response to extrinsic stress cues offer fitness and survival advantages to individuals, the mechanistic understanding of these modifications remains unclear. This study found that parasitoid wasps' stress signaling regulates Drosophila melanogaster oogenesis. Vruit flies dwelling in the wasp-infested area elevate their fecundity, and the observed reproductive response is specific to Pachycrepoideus sp., a pupal parasitoid wasp. Pachycrepoideus-specific olfactory and visual cues recruit the signaling pathways that promote germline stem cell proliferation and accelerate follicle development, increasing egg production in Drosophila females. Downregulation of signaling engaged in oocyte development by shifting flies to a non-wasp-infested environment increases apoptosis of the developing follicles. Thus, this study establishes host germline responsiveness to parasitoid-specific signals and supports a predator strategy to increase hosts for infection.

Wednesday, October 9th - Disease Models

Adedara, A. O., Bressan, G. N., Dos Santos, M. M., Fachinetto, R., Abolaji, A. O., Barbosa, N. V. (2024). Antioxidant responses driven by Hesperetin and Hesperidin counteract Parkinson's disease-like phenotypes in Drosophila melanogaster. Neurotoxicology, 101:117-127 PubMed ID: 38423185
Summary:
The study investigated the protective effects of Hesperetin (HSP) and Hesperidin (HSD) on 1 methyl, 4 phenyl, 1,2,3,6 tetrahydropyridine hydrochloride (MPTP)-induced Parkinsonism in Drosophila melanogaster. After a lifespan study to select exposure time and concentrations, flies were co-exposed to MPTP (0.4 mg/g diet), Hesperetin (0.2 and 0.4 mg/g diet), and Hesperidin (0.1 and 0.4 mg/g) for 7 days. In addition to in vivo parameters, some markers were assayed of oxidative stress and antioxidant status (lipid peroxidation, protein carbonylation, thiol content, hydrogen peroxide, and nitrate/nitrite levels, mRNA expression of Keap-1 (Kelch-like ECH associated protein 1), /Nrf2 (Nuclear factor erythroid 2 related factor 2), catalase, and glutathione-S-transferase (GST) activities), and cholinergic (acetyl cholinesterase activity (AChE) and dopaminergic signaling content and the mRNA expression of tyrosine hydroxylase (TH), monoamine oxidase (MAO-like) activity). In addition to increasing the lifespan of flies,it was found that both flavonoids counteracted the adverse effects of MPTP on survival, offspring emergence, and climbing ability of flies. Both flavonoids also reduced the oxidative damage on lipids and proteins and reestablished the basal levels of pro-oxidant species and activities of antioxidant enzymes in MPTP-exposed flies. These responses were accompanied by the normalization of the mRNA expression of Keap1/Nrf2 disrupted in flies exposed to MPTP. MPTP exposure also elicited changes in mRNA expression and content of TH as well as in MAO and AChE activity, which were reversed by HST and HSD. By efficiently hindering the oxidative stress in MPTP-exposed flies, these findings support the promising role of Hesperetin and Hesperidin as adjuvant therapy to manage Parkinsonism induced by chemicals such as MPTP.

Soldovieri, M. V., Ambrosino, P., Mosca, I., Servettini, I., Pietrunti, F., Belperio, G., Syrbe, S., Taglialatela, M., Lemke, J. R. (2024). De novo variants in KCNA3 cause developmental and epileptic encephalopathy.. Ann Neurol, 95(2):365-376 PubMed ID: 37964487
Summary:
Variants in several potassium channel genes, including KCNA1 and KCNA2, cause Developmental and Epileptic Encephalopathies (DEEs). This study investigated whether variants in KCNA3, another mammalian homologue of the Drosophila shaker family and encoding for Kv1.3 subunits, can cause DEE. Genetic analysis of study individuals was performed by routine exome or genome sequencing, usually of parent-offspring trios. Phenotyping was performed via a standard clinical questionnaire. Currents from wild-type and/or mutant Kv1.3 subunits were investigated by whole-cell patch-clamp upon their heterologous expression. Fourteen individuals, each carrying a de novo heterozygous missense variant in KCNA3, were identified. Most (12/14; 86%) had DEE with marked speech delay with or without motor delay, intellectual disability, epilepsy, and autism spectrum disorder. Functional analysis of Kv1.3 channels carrying each variant revealed heterogeneous functional changes, ranging from "pure" loss-of-function (LoF) effects due to faster inactivation kinetics, depolarized voltage-dependence of activation, slower activation kinetics, increased current inactivation, reduced or absent currents with or without dominant-negative effects, to "mixed" loss- and gain-of-function (GoF) effects. Compared to controls, Kv1.3 currents in lymphoblasts from 1 of the proband displayed functional changes similar to those observed upon heterologous expression of channels carrying the same variant. The antidepressant drug fluoxetine inhibited with similar potency the currents from wild-type and 1 of the Kv1.3 GoF variant. This study has describe a novel association of de novo missense variants in KCNA3 with a human DEE, and provide evidence that fluoxetine might represent a potential targeted treatment for individuals carrying variants with significant GoF effects.

Urena, E., Xu, B., Regan, J. C., Atilano, M. L., Minkley, L. J., Filer, D., Lu, Y. X., Bolukbasi, E., Khericha, M., Alic, N., Partridge, L. (2024). Trametinib ameliorates aging-associated gut pathology in Drosophila females by reducing Pol III activity in intestinal stem cells. Proc Natl Acad Sci U S A, 121(4):e2311313121 PubMed ID: 38241436
Summary:
Pharmacological therapies are promising interventions to slow down aging and reduce multimorbidity in the elderly. Studies in animal models are the first step toward translation of candidate molecules into human therapies, as they aim to elucidate the molecular pathways, cellular mechanisms, and tissue pathologies involved in the anti-aging effects. Trametinib, an allosteric inhibitor of MEK within the Ras/MAPK (Ras/Mitogen-Activated Protein Kinase) pathway and currently used as an anti-cancer treatment, emerged as a geroprotector candidate because it extended lifespan in the fruit fly Drosophila melanogaster. This study confirmed that trametinib consistently and robustly extends female lifespan, and reduces intestinal stem cell (ISC) proliferation, tumor formation, tissue dysplasia, and barrier disruption in guts in aged flies. In contrast, pro-longevity effects of trametinib are weak and inconsistent in males, and it does not influence gut homeostasis. Inhibition of the Ras/MAPK pathway specifically in ISCs is sufficient to partially recapitulate the effects of trametinib. Moreover, in ISCs, trametinib decreases the activity of the RNA polymerase III (Pol III), a conserved enzyme synthesizing transfer RNAs and other short, non-coding RNAs, and whose inhibition also extends lifespan and reduces gut pathology. Finally, this study showed that the pro-longevity effect of trametinib in ISCs is partially mediated by Maf1, a repressor of Pol III, suggesting a life-limiting Ras/MAPK-Maf1-Pol III axis in these cells. The mechanism of action described in this work paves the way for further studies on the anti-aging effects of trametinib in mammals and shows its potential for clinical application in humans.

Xia, Y., Wang, H., Xie, Z., Liu, Z. H., Wang, H. L. (2024). Inhibition of ferroptosis underlies EGCG mediated protection against Parkinson's disease in a Drosophila model. Free radical biology & medicine, 211:63-76 PubMed ID: 38092273
Summary:
Ferroptosis, a new type of cell death accompanied by iron accumulation and lipid peroxidation, is implicated in the pathology of Parkinson's disease (PD), which is a prevalent neurodegenerative disorder that primarily occurred in the elderly population. Epigallocatechin-3-gallate (EGCG) is the major polyphenol in green tea with known neuroprotective effects in PD patients. But whether EGCG-mediated neuroprotection against PD involves regulation of ferroptosis has not been elucidated. This study established a PD model using PINK1 mutant Drosophila. Iron accumulation, lipid peroxidation and decreased activity of GTPX, were detected in the brains of PD flies. Additionally, phenotypes of PD, including behavioral defects and dopaminergic neurons loss, were ameliorated by ferroptosis inhibitor ferrostatin-1 (Fer-1). Notably, the increased iron level, lipid peroxidation and decreased GPX activity in the brains of PD flies were relieved by EGCG. EGCG was found to exert neuroprotection mainly by restoring iron homeostasis in the PD flies. EGCG inhibited iron influx by suppressing Malvolio (Mvl) expression and simultaneously promoted the upregulation of ferritin, the intracellular iron storage protein, leading to a reduction in free iron ions. Additionally, EGCG downregulated the expression of Duox and Nox, two NADPH oxidases that produce reactive oxygen species (ROS) and increased SOD enzyme activity. Finally, modulation of intracellular iron levels or regulation of oxidative stress by genetic means exerted great influence on PD phenotypes. As such, the results demonstrated that ferroptosis has a role in the established PD model. Altogether, EGCG has therapeutic potentials for treating PD by targeting the ferroptosis pathway, providing new strategies for the prevention and treatment of PD and other neurodegenerative diseases.

Long, D. M., Cravetchi, O., Chow, E. S., Allen, C., Kretzschmar, D. (2024). The amyloid precursor protein intracellular domain induces sleep disruptions and its nuclear localization fluctuates in circadian pacemaker neurons in Drosophila and mice. Neurobiol Dis, 192:106429 PubMed ID: 38309627
Summary:
The most prominent symptom of Alzheimer's disease (AD) is cognitive decline; however, sleep and other circadian disruptions are also common in AD patients. Sleep disruptions have been connected with memory problems and therefore the changes in sleep patterns observed in AD patients may also actively contribute to cognitive decline. However, the underlying molecular mechanisms that connect sleep disruptions and AD are unclear. A characteristic feature of AD is the formation of plaques consisting of Amyloid-β (Aβ) peptides generated by cleavage of the Amyloid Precursor Protein (APP). Besides Aββ, APP cleavage generates several other fragments, including the APP intracellular domain (AICD) that has been linked to transcriptional regulation and neuronal homeostasis. This study shows that overexpression of the AICD reduces the early evening expression of two core clock genes and disrupts the sleep pattern in flies. Analyzing the subcellular localization of the AICD in pacemaker neurons, it was found that the AICD levels in the nucleus are low during daytime but increase at night. While this pattern of nuclear AICD persisted with age, the nighttime levels were higher in aged flies. Increasing the cleavage of the fly APP protein also disrupted AICD nuclear localization. Lastly, this study showed that the day/nighttime nuclear pattern of the AICD is also detectable in neurons in the suprachiasmatic nucleus of mice and that it also changes with age. Together, these data suggest that AD-associated changes in APP processing and the subsequent changes in AICD levels may cause sleep disruptions in AD.

Ueda, T., Takeuchi, T., Fujikake, N., Suzuki, M., Minakawa, E. N., Ueyama, M., Fujino, Y., Kimura, N., Nagano, S., Yokoseki, A., Onodera, O., Mochizuki, H., Mizuno, T., Wada, K., Nagai, Y. (2024). Dysregulation of stress granule dynamics by DCTN1 deficiency exacerbates TDP-43 pathology in Drosophila models of ALS/FTD. Acta neuropathologica communications, 12(1):20 PubMed ID: 38311779
Summary:
The abnormal aggregation of TDP-43 into cytoplasmic inclusions in affected neurons is a major pathological hallmark of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Although TDP-43 is aberrantly accumulated in the neurons of most patients with sporadic ALS/FTD and other TDP-43 proteinopathies, how TDP-43 forms cytoplasmic aggregates remains unknown. This study shows that a deficiency in DCTN1, a subunit of the microtubule-associated motor protein complex dynactin, perturbs the dynamics of stress granules and drives the formation of TDP-43 cytoplasmic aggregation in cultured cells, leading to the exacerbation of TDP-43 pathology and neurodegeneration in vivo. Using a Drosophila model of ALS/FTD it was demonstrated that genetic knockdown of DCTN1 accelerates the formation of ubiquitin-positive cytoplasmic inclusions of TDP-43. Knockdown of components of other microtubule-associated motor protein complexes, including dynein and kinesin, also increased the formation of TDP-43 inclusions, indicating that intracellular transport along microtubules plays a key role in TDP-43 pathology. Notably, DCTN1 knockdown delayed the disassembly of stress granules in stressed cells, leading to an increase in the formation of pathological cytoplasmic inclusions of TDP-43. These results indicate that a deficiency in DCTN1, as well as disruption of intracellular transport along microtubules, is a modifier that drives the formation of TDP-43 pathology through the dysregulation of stress granule dynamics.

Tuesday, October 8th - Signaling

Simon, N., Safyan, A., Pyrowolakis, G., Matsuda, S. (2024). Dally is not essential for Dpp spreading or internalization but for Dpp stability by antagonizing Tkv-mediated Dpp internalization. Elife, 12 PubMed ID: 38265865
Summary:
Dpp/BMP acts as a morphogen to provide positional information in the Drosophila wing disc. Key cell-surface molecules to control Dpp morphogen gradient formation and signaling are heparan sulfate proteoglycans (HSPGs). In the wing disc, two HSPGs, the glypicans Division abnormally delayed (Dally) and Dally-like (Dlp) have been suggested to act redundantly to control these processes through direct interaction of their heparan sulfate (HS) chains with Dpp. Based on this assumption, a number of models on how glypicans control Dpp gradient formation and signaling have been proposed, including facilitating or hindering Dpp spreading, stabilizing Dpp on the cell surface, or recycling Dpp. However, how distinct HSPGs act remains largely unknown. This study generated genome-engineering platforms for the two glypicans, and only Dally was foudn to be critical for Dpp gradient formation and signaling through interaction of its core protein with Dpp. This interaction was found to be in sufficient, and the HS chains of Dally were found to be essential for these functions largely without interacting with Dpp. Evidence is provided that the HS chains of Dally are not essential for spreading or recycling of Dpp but for stabilizing Dpp on the cell surface by antagonizing receptor-mediated Dpp internalization. These results provide new insights into how distinct HSPGs control morphogen gradient formation and signaling during development.

Ouyang, Y., Jeong, M. Y., Cunningham, C. N., Berg, J. A., Toshniwal, A. G., Hughes, C. E., Seiler, K., Van Vranken, J. G., Cluntun, A. A., Lam, G., Winter, J. M., Akdogan, E., Dove, K. K., Nowinski, S. M., West, M., Odorizzi, G., Gygi, S. P., Dunn, C. D., Winge, D. R., Rutter, J. (2024). Phosphate starvation signaling increases mitochondrial membrane potential through respiration-independent mechanisms. Elife, 13 PubMed ID: 38251707
Summary:
Mitochondrial membrane potential directly powers many critical functions of mitochondria, including ATP production, mitochondrial protein import, and metabolite transport. Its loss is a cardinal feature of aging and mitochondrial diseases, and cells closely monitor membrane potential as an indicator of mitochondrial health. Given its central importance, it is logical that cells would modulate mitochondrial membrane potential in response to demand and environmental cues, but there has been little exploration of this question. This study reports that loss of the Sit4 protein phosphatase in yeast increases mitochondrial membrane potential, both by inducing the electron transport chain and the phosphate starvation response. Indeed, a similarly elevated mitochondrial membrane potential is also elicited simply by phosphate starvation or by abrogation of the Pho85-dependent phosphate sensing pathway. This enhanced membrane potential is primarily driven by an unexpected activity of the ADP/ATP carrier. This study also demonstrated that this connection between phosphate limitation and enhancement of mitochondrial membrane potential is observed in primary and immortalized mammalian cells as well as in Drosophila. These data suggest that mitochondrial membrane potential is subject to environmental stimuli and intracellular signaling regulation and raise the possibility for therapeutic enhancement of mitochondrial function even in defective mitochondria.

Zhang, W., Zhang, Z., Xiang, Y., ...., Hu, J., Gao, S., Liu, Q. (2024). Aurora kinase A-mediated phosphorylation triggers structural alteration of Rab1A to enhance ER complexity during mitosis. Nat Struct Mol Biol, 31(2):219-231 PubMed ID: 38177680
Summary:
Morphological rearrangement of the endoplasmic reticulum (ER) is critical for metazoan mitosis. Yet, how the ER is remodeled by the mitotic signaling remains unclear. This report shows that mitotic Aurora kinase A (AURKA) employs a small GTPase, Rab1A, to direct ER remodeling. During mitosis, AURKA phosphorylates Rab1A at Thr75. Structural analysis demonstrates that Thr75 phosphorylation renders Rab1A in a constantly active state by preventing interaction with GDP-dissociation inhibitor (GDI). Activated Rab1A is retained on the ER and induces the oligomerization of ER-shaping protein RTNs and REEPs, eventually triggering an increase of ER complexity. In various models, from Caenorhabditis elegans and Drosophila to mammals, inhibition of Rab1A(Thr75) phosphorylation by genetic modifications disrupts ER remodeling. Thus, this study reveals an evolutionarily conserved mechanism explaining how mitotic kinase controls ER remodeling and uncovers a critical function of Rab GTPases in metaphase.

Zhang, Q., Zhang, P., Yang, M., Tian, Y., Feng, C., Wei, W. (2024). Identifications of three novel alleles of Serrate in Drosophila. Cells & development, 177:203908 PubMed ID: 38403117
Summary:
The Notch signaling pathway, an evolutionarily highly conserved pathway, participates in various essential physiological processes in organisms. Activation of Notch signaling in the canonical manner requires the combination of ligand and receptor. There are two ligands of Notch in Drosophila: Delta (Dl) and Serrate (Ser). A mutation mf157 is identified for causing nicks of fly wings in genetic analysis from a mutant library (unpublished) that was established previously. Immunofluorescent staining illustrates that mf157 represses the expression of Cut and Wingless (Wg), the targets of Notch signaling. MARCM cloning analysis reveals that mf157 functions at the same level or the upstream of ligands of Notch in signaling sending cells. Sequencing demonstrates that mf157 is a novel allele of the Ser gene. Subsequently, mf553 and mf167 are also identified as new alleles of Ser from the library. Furthermore, the complementary assays and the examination of transcripts confirm the sequencing results. Besides, the repressed phenotypes of Notch signaling were reverted by transposon excision experiments of mf157. In conclusion, this study identified three fresh alleles of Ser. These works supply additional genetic resources for further study of functions of Ser and Notch signaling regulation.

Yun, H. M., Hyun, B., Song, X., Hyun, S. (2024). Piwi expressed in Drosophila adipose tissues regulates systemic IGF signaling and growth via IGF-binding protein. Biochem Biophys Res Commun, 695:149495 PubMed ID: 38211532
Summary:
Piwi and its partner, Piwi-interacting RNA (piRNA), are pivotal in suppressing the harmful effects of transposable elements (TEs) linked to genomic insertional mutagenesis. While primarily active in Drosophila's adult gonadal tissues, causing sterility in its absence, Piwi's role in post-embryonic development remains unclear. This study reveals Piwi's functional presence in the larval fat body, where it governs developmental growth through systemic insulin/insulin-like growth factor (IGF) signaling (IIS). Piwi knockdown in the fat body resulted in dysregulated TE expression, reduced developmental rate and body growth, and diminished systemic IIS activity. Notably, Piwi knockdown increased Imaginal Morphogenic Protein Late 2 (Imp-L2) expression, akin to insulin-like growth factor-binding protein 7 (IGFBP7), reducing systemic IIS and inhibiting body growth. This unveils a novel role for Piwi in larval adipose tissues, emphasizing its importance in regulating systemic IIS and overall organismal growth.

Karunaraj, P., Washington, C., Luf, M., Martino-Cortez, Y., Pfleger, C. M. (2024). Characterization of Ras Y4H mutants in Drosophila. microPublication biology, 2024 PubMed ID: 38495589
Summary:
Ras signaling plays a highly conserved role from flies to mammals in establishing proper development, and its dysregulation can lead to cancer. In Drosophila , this study demonstrated that Ras Tyrosine 4 (Y4) was required for inhibitory ubiquitination by FBgn0262937 Rabex-5. In humans, rare histidine substitution mutations at Y4 are found in HRas in cerebellar glioblastomas (cGBMs). This study reports that analogous Y4H mutations in Drosophila Ras make it less sensitive to Rabex-5-mediated ubiquitination in cells and show increased frequency of vein phenotypes per wing compared to wild-type Ras, which would be consistent with Ras gain-of-function and with their appearance in human cGBMs.

Monday, October 7th - Response to Stress

Xi, G., Lamba, S. A., Mysh, M., Poulton, J. S. (2024). Oxidative Stress Contributes to Slit Diaphragm Defects Caused by Disruption of Endocytosis. Kidney international reports, 9(2):451-463 PubMed ID: 38344712
Summary:
odocyte slit diaphragms are an important component of the glomerular filtration barrier. Podocyte injury frequently includes defects in slit diaphragms, and various mechanisms for these defects have been described, including altered endocytic trafficking of slit diaphragm proteins or oxidative stress. However, the potential relationship between endocytosis and oxidative stress in the context of slit diaphragm integrity has not been extensively considered. To examine the potential relationships between endocytosis, oxidative stress, and slit diaphragm integrity, this study induced genetic or pharmacological disruption of endocytosis in Drosophila nephrocytes (the insect orthologue of podocytes) and cultured human podocytes. Then immunofluorescence microscopy was employed to analyze protein localization and levels and to quantify signal from reactive oxygen species (ROS) dyes. Immunoprecipitation from podocyte cell lysates was used to examine effects on slit diaphragm protein complex formation (i.e., nephrin/podocin and nephrin/ZO-1). RDisruption of endocytosis in nephrocytes and podocytes led to slit diaphragm defects, elevated levels of ROS (oxidative stress), and activation of the nuclear factor erythroid 2-related factor 2 (Nrf2) antioxidant pathway. In nephrocytes with defective endocytosis, perturbation of Nrf2 signaling exacerbated slit diaphragm defects. Conversely, overexpression of Nrf2 target genes catalase or glucose-6-phosphate dehydrogenase (G6PD) significantly ameliorated slit diaphragm defects caused by disruption of endocytosis. It is concluded that xidative stress is an important consequence of defective endocytosis and contributes to the defects in slit diaphragm integrity associated with disruption of endocytic trafficking.

Srivastav, S., van der Graaf, K., Jonnalagadda, P. C., Thawani, M., McNew, J. A., Stern, M. (2024). Motor neuron activity enhances the proteomic stress caused by autophagy defects in the target muscle. PLoS One, 19(1):e0291477 PubMed ID: 38166124
Summary:
Several lines of evidence demonstrate that increased neuronal excitability can enhance proteomic stress. For example, epilepsy can enhance the proteomic stress caused by the expression of certain aggregation-prone proteins implicated in neurodegeneration. However, unanswered questions remain concerning the mechanisms by which increased neuronal excitability accomplishes this enhancement. This study tested whether increasing neuronal excitability at a particular identified glutamatergic synapse, the Drosophila larval neuromuscular junction, can enhance the proteomic stress caused by mutations in the ER fusion/GTPase gene atlastin (atl). It was previously shown that larval muscle from the atl2 null mutant is defective in autophagy and accumulates protein aggregates containing ubiquitin (poly-UB aggregates). To determine if increased neuronal excitability might enhance the increased proteomic stress caused by atl2, the TrpA1-encoded excitability channel was activated within neurons. TrpA1 activation had no effect on poly-UB aggregate accumulation in wildtype muscle, but significantly increased poly-UB aggregate number in atl2 muscle. Previous work has shown that atl loss from either neuron or muscle increases muscle poly-UB aggregate number. This study found that neuronal TrpA1 activation enhanced poly-UB aggregate number when atl2 was removed from muscle, but not from neurons. Neuronal TrpA1 activation enhanced other phenotypes conferred by muscle atl loss, such as decreased pupal size and decreased viability. Taken together, these results indicate that the proteomic stress caused by muscle atl loss is enhanced by increasing neuronal excitability.

Kim, S., Quagraine, Y., Singh, M., Kim, J. H. (2024). Rab11 suppresses neuronal stress signaling by localizing Dual leucine zipper kinase to axon terminals for protein turnover. bioRxiv, PubMed ID: 37131782
Summary:
Dual Leucine Zipper Kinase (DLK) mediates multiple neuronal stress responses, and its expression levels are constantly suppressed to prevent excessive stress signaling. Wallenda (Wnd), the Drosophila ortholog of DLK, is highly enriched in the axon terminals of Drosophila sensory neurons in vivo and that this subcellular localization is necessary for Highwire-mediated Wnd protein turnover under normal conditions. Structure-function analysis found that Wnd palmitoylation is essential for its axon terminal localization. Palmitoylation-defective Wnd accumulated in neuronal cell bodies, exhibited dramatically increased protein expression levels, and triggered excessive neuronal stress responses. Defective intracellular transport is implicated in neurodegenerative conditions. Comprehensive dominant-negative Rab protein screening identified Rab11 as an essential factor for Wnd localization in axon terminals. Consequently, Rab11 loss-of-function increased the protein levels of Wnd and induced neuronal stress responses. Inhibiting Wnd activity significantly ameliorated neuronal loss and c-Jun N-terminal kinase signaling triggered by Rab11 loss-of-function. Taken together, these suggest that DLK proteins are constantly transported to axon terminals by Rab11 for protein turnover. This study demonstrates how subcellular protein localization is coupled to protein turnover for neuronal stress signaling.

Jeong, D. J., Um, J. H., Kim, Y. Y., Shin, D. J., Im, S., Lee, K. M., Lee, Y. H., Lim, D. S., Kim, D., Yun, J. (2024). The Mst1/2-BNIP3 axis is required for mitophagy induction and neuronal viability under mitochondrial stress. Experimental & molecular medicine, 56(3):674-685 PubMed ID: 38443598
Summary:
Mitophagy induction upon mitochondrial stress is critical for maintaining mitochondrial homeostasis and cellular function. This study found that Mst1/2 (Stk3/4), key regulators of the Hippo pathway, are required for the induction of mitophagy under various mitochondrial stress conditions. Knockdown of Mst1/2 or pharmacological inhibition by XMU-MP-1 treatment led to impaired mitophagy induction upon CCCP and DFP treatment. Mechanistically, Mst1/2 induces mitophagy independently of the PINK1-Parkin pathway and the canonical Hippo pathway. Moreover, the results suggest the essential involvement of BNIP in Mst1/2-mediated mitophagy induction upon mitochondrial stress. Notably, Mst1/2 knockdown diminishes mitophagy induction, exacerbates mitochondrial dysfunction, and reduces cellular survival upon neurotoxic stress in both SH-SY5Y cells and Drosophila models. Conversely, Mst1 and Mst2 expression enhances mitophagy induction and cell survival. In addition, AAV-mediated Mst1 expression reduced the loss of TH-positive neurons, ameliorated behavioral deficits, and improved mitochondrial function in an MPTP-induced Parkinson's disease mouse model. These findings reveal the Mst1/2-BNIP3 regulatory axis as a novel mediator of mitophagy induction under conditions of mitochondrial stress and suggest that Mst1/2 play a pivotal role in maintaining mitochondrial function and neuronal viability in response to neurotoxic treatment.

Perlegos, A. E., Quan, X., Donnelly, K. M., Shen, H., Shields, E. J., Elashal, H., Fange Liu, K., Bonini, N. M. (2023). dTrmt10A impacts Hsp70 chaperone m(6)A levels and the stress response in the Drosophila brain. Sci Rep, 13(1):22999 PubMed ID: 38155219
Summary:
Chronic cellular stress has a profound impact on the brain, leading to degeneration and accelerated aging. Recent work has revealed the vital role of RNA modifications, and the proteins responsible for regulating them, in the stress response. This study defined the role of CG14618/dTrmt10A, the Drosophila counterpart of human TRMT10A a N(1)-methylguanosine methyltransferase, on m(6)A regulation and heat stress resilience in the Drosophila brain. By m(6)A-IP RNA sequencing on Drosophila head tissue, this study demonstrated that manipulating dTrmt10A levels indirectly regulates m(6)A levels on polyA + RNA. dTrmt10A exerted its influence on m(6)A levels on transcripts enriched for neuronal signaling and heat stress pathways, similar to the m(6)A methyltransferase Mettl3. Intriguingly, its impact primarily targeted 3' UTR m(6)A, setting it apart from the majority of Drosophila m(6)A-modified transcripts which display 5' UTR enrichment. Upregulation of dTrmt10A led to increased resilience to acute heat stress, decreased m(6)A modification on heat shock chaperones, and coincided with decreased decay of chaperone transcripts and increased translation of chaperone proteins. Overall, these findings establish a potential mechanism by which dTrmt10A regulates the acute brain stress response through m(6)A modification.

Perlegos, A. E., Byrns, C. N., Bonini, N. M. (2024). Cell type-specific regulation of m(6) A modified RNAs in the aging Drosophila brain. Aging Cell:e14076 PubMed ID: 38205931
Summary:
The aging brain is highly vulnerable to cellular stress, and neurons employ numerous mechanisms to combat neurotoxic proteins and promote healthy brain heat stress response of the brain. This study examined m(6) A in the fly brain with the chronic stresses of aging and degenerative disease. m(6) A levels dynamically increased with both age and disease in the brain, marking integral neuronal identity and signaling pathway transcripts that decline in level with age and disease. Unexpectedly, there is opposing impact of m(6) A transcripts in neurons versus glia, which conferred different outcomes on animal health span upon Mettl3 knockdown to reduce m(6) A: whereas Mettl3 function is normally beneficial to neurons, it is deleterious to glia. Moreover, knockdown of Mettl3 in glial tauopathy reduced tau pathology and increased animal survival. These findings provide mechanistic insight into regulation of m(6) A modified transcripts with age and disease, highlighting an overall beneficial function of Mettl3 in neurons in response to chronic stresses, versus a deleterious impact in glia.

Friday, October 4th - Cytoskeleton

Esmangart de Bournonville, T., Jaglarz, M. K., Durel, E., Le Borgne, R. (2024). ESCRT-III-dependent adhesive and mechanical changes are triggered by a mechanism detecting alteration of septate junction integrity in Drosophila epithelial cells. Elife, 13 PubMed ID: 38305711
Summary:
Barrier functions of proliferative epithelia are constantly challenged by mechanical and chemical constraints. How epithelia respond to and cope with disturbances of barrier functions to allow tissue integrity maintenance is poorly characterised. Cellular junctions play an important role in this process and intracellular traffic contribute to their homeostasis. This study revealed that, in Drosophila pupal notum, alteration of the bi- or tricellular septate junctions (SJs) triggers a mechanism with two prominent outcomes. On one hand, there is an increase in the levels of E-cadherin, F-actin, and non-muscle myosin II in the plane of adherens junctions. On the other hand, β-integrin/Vinculin-positive cell contacts are reinforced along the lateral and basal membranes. The weakening of SJ integrity, caused by the depletion of bi- or tricellular SJ components, was found to alter ESCRT-III/Vps32/Shrub distribution, reduces degradation and instead favours recycling of SJ components, an effect that extends to other recycled transmembrane protein cargoes including Crumbs, its effector β-Heavy Spectrin Karst, and β-integrin. A mechanism is proposed by which epithelial cells, upon sensing alterations of the SJ, reroute the function of Shrub to adjust the balance of degradation/recycling of junctional cargoes and thereby compensate for barrier junction defects to maintain epithelial integrity.

Wasilewicz, L. J., Gagnon, Z. E., Jung, J., Mercier, A. J. (2023). Investigating Postsynaptic Effects of a Drosophila Neuropeptide on Muscle Contraction. Journal of neurophysiology. PubMed ID: 38150542
Summary:
The Drosophila neuropeptide, DPKQDFMRFamide, was previously shown to enhance excitatory junctional potentials (EJPs) and muscle contraction in the NMJ by both presynaptic and postsynaptic actions. Since the peptide acts on both sides of the synaptic cleft, it has been difficult to examine postsynaptic modulatory mechanisms, particularly when contractions are elicited by nerve stimulation. Postsynaptic actions were examined in 3(rd) instar larvae by applying peptide and the excitatory neurotransmitter, L-glutamate, in the bathing solution to elicit contractions after silencing motor output by removing the central nervous system (CNS). DPKQDFMRFamide enhanced glutamate-evoked contractions at low concentrations (EC(50) 1.3 nM), consistent with its role as a neurohormone, and the combined effect of both substances was supra-additive. Glutamate-evoked contractions were also enhanced when transmitter release was blocked in temperature-sensitive (Shibire) mutants, confirming the peptide's postsynaptic action. The peptide increased membrane depolarization in muscle when co-applied with glutamate, and its effects were blocked by nifedipine, an L-type channel blocker, indicating effects at the plasma membrane involving calcium influx. DPKQDFMRFamide also enhanced contractions induced by caffeine in the absence of extracellular calcium, suggesting increased calcium release from the sarcoplasmic reticulum (SR) or effects downstream of calcium release from the SR. The peptide's effects do not appear to involve calcium/calmodulin-dependent protein kinase II (CaMKII), previously shown to mediate presynaptic effects. The approach used in this study might be useful for examining postsynaptic effects of neurohormones and cotransmitters in other systems.

Sekine, S., Tarama, M., Wada, H., Sami, M. M., Shibata, T., Hayashi, S. (2024). Emergence of periodic circumferential actin cables from the anisotropic fusion of actin nanoclusters during tubulogenesis. Nat Commun, 15(1):464 PubMed ID: 38267421
Summary:
The periodic circumferential cytoskeleton supports various tubular tissues. Radial expansion of the tube lumen causes anisotropic tensile stress, which can be exploited as a geometric cue. However, the molecular machinery linking anisotropy to robust circumferential patterning is poorly understood. This study aimed to reveal the emergent process of circumferential actin cable formation in a Drosophila tracheal tube. During luminal expansion, sporadic actin nanoclusters emerge and exhibit circumferentially biased motion and fusion. RNAi screening reveals the formin family protein, DAAM, as an essential component responding to tissue anisotropy, and non-muscle myosin II as a component required for nanocluster fusion. An agent-based model simulation suggests that crosslinkers play a crucial role in nanocluster formation and cluster-to-cable transition occurs in response to mechanical anisotropy. Altogether, it is proposed that an actin nanocluster is an organizational unit that responds to stress in the cortical membrane and builds a higher-order cable structure.

Wu, Z., Chen, H., Zhang, Y., Wang, Y., Wang, Q., Augiere, C., Hou, Y., Fu, Y., Peng, Y., Durand, B., Wei, Q. (2024). Cep131-Cep162 and Cby-Fam92 complexes cooperatively maintain Cep290 at the basal body and contribute to ciliogenesis initiation. PLoS Biol, 22(3):e3002330 PubMed ID: 38442096
Summary:
Cilia play critical roles in cell signal transduction and organ development. Defects in cilia function result in a variety of genetic disorders. Cep290 is an evolutionarily conserved ciliopathy protein that bridges the ciliary membrane and axoneme at the basal body (BB) and plays critical roles in the initiation of ciliogenesis and TZ assembly. How Cep290 is maintained at BB and whether axonemal and ciliary membrane localized cues converge to determine the localization of Cep290 remain unknown. This study reports that the Cep131-Cep162 module near the axoneme and the Cby-Fam92 module close to the membrane synergistically control the BB localization of Cep290 and the subsequent initiation of ciliogenesis in Drosophila. Concurrent deletion of any protein of the Cep131-Cep162 module and of the Cby-Fam92 module leads to a complete loss of Cep290 from BB and blocks ciliogenesis at its initiation stage. These results reveal that the first step of ciliogenesis strictly depends on cooperative and retroactive interactions between Cep131-Cep162, Cby-Fam92 and Cep290, which may contribute to the complex pathogenesis of Cep290-related ciliopathies.

Nelson, A. C., Rolls, M. M., Ciocanel, M. V., McKinley, S. A. (2024). Minimal Mechanisms of Microtubule Length Regulation in Living Cells. ArXiv, PubMed ID: 37904745
Summary:
The microtubule cytoskeleton is responsible for sustained, long-range intracellular transport of mRNAs, proteins, and organelles in neurons. Neuronal microtubules must be stable enough to ensure reliable transport, but they also undergo dynamic instability, as their plus and minus ends continuously switch between growth and shrinking. This process allows for continuous rebuilding of the cytoskeleton and for flexibility in injury settings. Motivated by in vivo experimental data on microtubule behavior in Drosophila neurons, a mathematical model is proposed of dendritic microtubule dynamics, with a focus on understanding microtubule length, velocity, and state-duration distributions. Limitations on microtubule growth phases were found to be needed for realistic dynamics, but the type of limiting mechanism leads to qualitatively different responses to plausible experimental perturbations. It is therefore propose and investigate two minimally-complex length-limiting factors: limitation due to resource (tubulin) constraints and limitation due to catastrophe of large-length microtubules. Simulations of a detailed stochastic model were combined with steady-state analysis of a mean-field ordinary differential equations model to map out qualitatively distinct parameter regimes. This provides a basis for predicting changes in microtubule dynamics, tubulin allocation, and the turnover rate of tubulin within microtubules in different experimental environments.

Gao, Z., Huang, E., Wang, W., Xu, L., Xu, W., Zheng, T., Rui, M. (2024). Patronin regulates presynaptic microtubule organization and neuromuscular junction development in Drosophila.. iScience, 27(2):108944 PubMed ID: 38318379
Summary:
Synapses are fundamental components of the animal nervous system. Synaptic cytoskeleton is essential for maintaining proper neuronal development and wiring. Perturbations in neuronal microtubules (MTs) are correlated with numerous neuropsychiatric disorders. Despite discovering multiple synaptic MT regulators, the importance of MT stability, and particularly the polarity of MT in synaptic function, is still under investigation. This study identified Patronin , an MT minus-end-binding protein, for its essential role in presynaptic regulation of MT organization and neuromuscular junction (NMJ) development. Analyses indicate that Patronin regulates synaptic development independent of Klp10A. Subsequent research elucidates that it is Short stop (Shot), a member of the Spectraplakin family of large cytoskeletal linker molecules, works synergistically with Patronin to govern NMJ development. The possibility is raised that normal synaptic MT polarity contributes to proper NMJ morphology. Overall, this study demonstrates an unprecedented role of Patronin, and a potential involvement of MT polarity in synaptic development.

Wednesday, October 2nd - Immune Response

Ono, M., Matsumura, T., Sung, E. J., Koyama, T., Ochiai, M., Shears, S. B., Hayakawa, Y. (2024). Drosophila cytokine GBP2 exerts immune responses and regulates GBP1 expression through GPCR receptor Mthl10. Insect biochemistry and molecular biology. 167:104086 PubMed ID: 38295885
Summary:
Growth-blocking peptide (GBP), an insect cytokine, was first found in armyworm Mythimna separata. A functional analogue of GBP, stress-responsive peptide (SRP), was also identified in the same species. SRP gene expression has been demonstrated to be enhanced by GBP, indicating that both cytokines are organized within a hierarchical regulatory network. Although GBP1 (CG15917) and GBP2 (CG11395) have been identified in Drosophila melanogaster, immunological functions have only been characterized for GBP1. It is expected that the biological responses of two structurally similar peptides should be coordinated, but there is little information on this topic. This study demonstrate that GBP2 replicates the GBP1-mediated cellular immune response from Drosophila S2 cells. Moreover, the GBP2-induced response was silenced by pre-treatment with dsRNA targeting the GBP receptor gene, Mthl10. Furthermore, treatment of S2 cells with GBP2 enhanced GBP1 expression levels, but GBP1 did not affect GBP2 expression. GBP2 derived enhancement of GBP1 expression was not observed in the presence of GBP1, indicating that GBP2 is an upstream expressional regulator of a GBP1/GBP2 cytokine network. GBP2-induced enhancement of GBP1 expression was not observed in Mthl10 knockdown cells. Enhancement of GBP2 expression was observed in both Drosophila larvae and S2 cells under heat stress conditions; expressional enhancement of both GBP1 and GBP2 was eliminated in Mthl10 knockdown cells and larvae. Finally, Ca(2+) mobilization assay in GCaMP3-expressing S2 cells demonstrated that GBP2 mobilizes Ca(2+) upstream of Mthl10. This finding revealed that Drosophila GBP1 and GBP2 control immune responses as well as their own expression levels through a hierarchical cytokine network, indicating that Drosophila GBP1/GBP2 system can be a simple model that is useful to investigate the detailed regulatory mechanism of related cytokine complexes.

Hedelin, L., Thie baut, A., Huang, J., Li, X., Lemoine, A., Haas, G., Meignin, C., Cai, H., Waterhouse, R. M., Martins, N., Imler, J. L. (2024). Investigating the Evolution of Drosophila STING-Dependent Antiviral Innate Immunity by Multispecies Comparison of 2'3'-cGAMP Responses. Mol Biol Evol, 41(3) PubMed ID: 38377349
Summary:
Viruses represent a major threat to all animals, which defend themselves through induction of a large set of virus-stimulated genes that collectively control the infection. In vertebrates, these genes include interferons that play a critical role in the amplification of the response to infection. Virus- and interferon-stimulated genes include restriction factors targeting the different steps of the viral replication cycle, in addition to molecules associated with inflammation and adaptive immunity. Predictably, antiviral genes evolve dynamically in response to viral pressure. As a result, each animal has a unique arsenal of antiviral genes. This study exploited the capacity to experimentally activate the evolutionarily conserved stimulator of IFN genes (STING) signaling pathway by injection of the cyclic dinucleotide 2'3'-cyclic guanosine monophosphate-adenosine monophosphate into flies to define the repertoire of STING-regulated genes in 10 Drosophila species, spanning 40 million years of evolution. These data reveal a set of conserved STING-regulated factors, including STING itself, a cGAS-like-receptor, the restriction factor pastel, and the antiviral protein Vago, but also 2 key components of the antiviral RNA interference pathway, Dicer-2, and Argonaute2. In addition, unknown species- or lineage-specific genes were identified that have not been previously associated with resistance to viruses. These data provide insight into the core antiviral response in Drosophila flies and pave the way for the characterization of previously unknown antiviral effectors.

Vincow, E. S., Thomas, R. E., Milstein, G., Pareek, G., Bammler, T., MacDonald, J., Pallanck, L. (2023). Glucocerebrosidase deficiency leads to neuropathology via cellular immune activation. bioRxiv, PubMed ID: 38168223
Summary:
Mutations in GBA (glucosylceramidase beta), which encodes the lysosomal enzyme glucocerebrosidase (GCase), are the strongest genetic risk factor for the neurodegenerative disorders Parkinson's disease (PD) and Lewy body dementia. Recent work has suggested that neuroinflammation may be an important factor in the risk conferred by GBA mutations. This study therefore systematically tested the contributions of immune-related genes to neuropathology in a Drosophila model of GCase deficiency.Target immune factors were identified via RNA-Seq and proteomics on heads from GCase-deficient flies, revealing both increased abundance of humoral factors and increased macrophage activation. The identified immune factors were then manipulated and their effect was measured on head protein aggregates, a hallmark of neurodegenerative disease. Genetic ablation of humoral (secreted) immune factors did not suppress the development of protein aggregation. By contrast, re-expressing Gba1b in activated macrophages suppressed head protein aggregation in Gba1b mutants and rescued their lifespan and behavioral deficits. Moreover, reducing the GCase substrate glucosylceramide in activated macrophages also ameliorated Gba1b mutant phenotypes. Taken together, these findings show that glucosylceramide accumulation due to GCase deficiency leads to macrophage activation, which in turn promotes the development of neuropathology.

Huang, Y., Wang, T., Jiang, C., Li, S., Zhou, H., Li, R. (2024). Relish-facilitated lncRNA-CR11538 suppresses Drosophila Imd immune response and maintains immune homeostasis via decoying Relish away from antimicrobial peptide promoters. Dev Comp Immunol, 151:105098 PubMed ID: 37956726
Summary:
Innate immunity plays a crucial role in host defense against pathogen invasion and its strength and duration requires precise control. Long non-coding RNAs (lncRNAs) have become important regulators of innate immunity, yet their roles in Drosophila immune responses remain largely unknown. This study identified that the overexpression of lncRNA-CR11538 inhibits the expression of antimicrobial peptides (AMPs) Dpt and AttA in Drosophila upon Escherichia coli (E. coli) infection, and influences the survival rate of flies after E. cloacae infection. Mechanically, lncRNA-CR11538 decoys Relish away from AMPs promoter region. It was further revealed that Relish can promote the transcription of lncRNA-CR11538. After analyzing the dynamic expression profile of lncRNA-CR11538 during Imd immune response, a hypothesis was put forward that in the late stage of Imd immune response, lncRNA-CR11538 can be activated by Relish and further decoy Relish away from the AMPs promoter to suppress excessive immune signal and maintain immune homeostasis. It is proposed that this mechanism provides insights into the complex regulatory networks controlling immune responses in Drosophila and suggests potential targets for therapeutic intervention in diseases involving dysregulated immune responses.

Lu, M., Wei, D., Shang, J., Li, S., Song, S., Luo, Y., Tang, G., Wang, C. (2024). Suppression of Drosophila antifungal immunity by a parasite effector via blocking GNBP3 and GNBP-like 3, the dual receptors for beta-glucans. Cell Rep, 43(1):113642 PubMed ID: 38175756
Summary:
The tactics used by animal pathogens to combat host immunity are largely unclear. This study reports the depiction of the virulence-required effector Tge1 deployed by the entomopathogen Metarhizium robertsii to suppress Drosophila antifungal immunity. Tge1 can target both GNBP3 and GNBP-like 3 (GL3), and the latter can bind to β-glucans like GNBP3, whereas the glucan binding by both receptors can be attenuated by Tge1. As opposed to the surveillance GNBP3, GL3 is inducible in Drosophila depending on the Toll pathway via a positive feedback loop mechanism. Losses of GNBP3 and GL3 genes result in the deregulations of protease cascade, Spstzle maturation, and antimicrobial gene expressions in Drosophila upon fungal challenges. Fly survival assays confirm that GL3 plays a more essential role than GNBP3 in combating fungal infections. In addition to evidencing the gene-for-gene interactions between fungi and insects, these data advance insights into Drosophila antifungal immunity.

Wei, Z., Zhang, M., Chen, Y., Hu, H., Zhao, X., Zheng, Y., Tran, N. T., Feng, H., Zeng, C., Li, S. (2024). Spatzle maintains homeostasis of hemolymph microbiota in Scylla paramamosain through Toll2. Fish & shellfish immunology:109385 PubMed ID: 38242262
Summary:
The Toll pathway is crucial for innate immune responses in organisms (including Drosophila and mammals). The Spatzle protein outside of cells acts as a ligand for Toll receptors, enabling the transfer of signals from outside the cell to the inside. However, the function of Spatzle in the immune system of mud crab (Scylla paramamosain) remains unclear. This research discovered a novel Spatzle gene (Sp-Spz) in mud crab, which showed extensive expression in all the tissues that were examined. The RNA interference exhibited the correlation between Sp-Spz and the anti-lipopolysaccharide factors (ALFs). Knockdown of Sp-Spz decreased the expression of Sp-Toll2 but not Sp-Toll1. In Drosophila Schneider 2 cells, Sp-Spz was found interacted with Sp-Toll2. Moreover, the depletion of Sp-Spz caused the separation of hepatic lobules from the basement membrane, resulting in the disruption of the structural coherence of hepatopancreatic cells. Additionally, the knockdown of Sp-Spz resulted in changes to the composition of the hemolymph microbiota, specifically affecting the proportions of different phylum and family levels. The findings indicated that Sp-Spz may promote the synthesis of ALFs via Sp-Toll2, thereby influencing the homeostasis of microbiota in the hemolymph. In this study, novel insights into mud crab immunity are provided.

Tuesday, October 1st - Gonads

Carpinteyro-Ponce, J., Machado, C. A. (2024). The Complex Landscape of Structural Divergence Between the Drosophila pseudoobscura and D. persimilis Genomes. Genome biology and evolution, 16(3) PubMed ID: 38482945
Summary:
Structural genomic variants are key drivers of phenotypic evolution. They can span hundreds to millions of base pairs and can thus affect large numbers of genetic elements. Although structural variation is quite common within and between species, its characterization depends upon the quality of genome assemblies and the proportion of repetitive elements. Using new high-quality genome assemblies, this study reports a complex and previously hidden landscape of structural divergence between the genomes of Drosophila persimilis and D. pseudoobscura, two classic species in speciation research, and study the relationships among structural variants, transposable elements, and gene expression divergence. The new assemblies confirm the already known fixed inversion differences between these species. Consistent with previous studies showing higher levels of nucleotide divergence between fixed inversions relative to collinear regions of the genome, a significant overrepresentation of INDELs was found inside the inversions. It was found that transposable elements accumulate in regions with low levels of recombination, and spatial correlation analyses reveal a strong association between transposable elements and structural variants. A strong association is reported between differentially expressed (DE) genes and structural variants and an overrepresentation of DE genes inside the fixed chromosomal inversions that separate this species pair. Interestingly, species-specific structural variants are overrepresented in DE genes involved in neural development, spermatogenesis, and oocyte-to-embryo transition. Overall, these results highlight the association of transposable elements with structural variants and their importance in driving evolutionary divergence.

Sabaris, G., Ortiz, D. M., Laiker, I., Mayansky, I., Naik, S., Cavalli, G., Stern, D. L., Preger-Ben Noon, E., Frankel, N. (2024). The Density of Regulatory Information Is a Major Determinant of Evolutionary Constraint on Noncoding DNA in Drosophila. Mol Biol Evol, 41(2) PubMed ID: 38364113
Summary:
Evolutionary analyses have estimated that ~60% of nucleotides in intergenic regions of the Drosophila melanogaster genome are functionally relevant, suggesting that regulatory information may be encoded more densely in intergenic regions than has been revealed by most functional dissections of regulatory DNA. This issue was approached through a functional dissection of the regulatory region of the gene shavenbaby (svb). Most of the ~90 kb of this large regulatory region is highly conserved in the genus Drosophila, though characterized enhancers occupy a small fraction of this region. By analyzing the regulation of svb in different contexts of Drosophila development, it was found that the regulatory information that drives svb expression in the abdominal pupal epidermis is organized in a different way than the elements that drive svb expression in the embryonic epidermis. While in the embryonic epidermis svb is activated by compact enhancers separated by large inactive DNA regions, svb expression in the pupal epidermis is driven by regulatory information distributed over broader regions of svb cis-regulatory DNA. In the same vein, other developmental genes also display a dense distribution of putative regulatory elements in their regulatory regions. Furthermore, it was found that a large percentage of conserved noncoding DNA of the Drosophila genome is contained within regions of open chromatin. These results suggest that part of the evolutionary constraint on noncoding DNA of Drosophila is explained by the density of regulatory information, which may be greater than previously appreciated.

Cavey, M., Charroux, B., Travaillard, S., Manière, G., Berthelot-Grosjean, M., Quitard, S., Minervino, C., Detailleur, B., Grosjean, Y., Prud'homme, B. (2023). Increased sugar valuation contributes to the evolutionary shift in egg-laying behavior of the fruit pest Drosophila suzukii. PLoS Biol, 21(12):e3002432 PubMed ID: 38079457
Summary:
Behavior evolution can promote the emergence of agricultural pests by changing their ecological niche. For example, the insect pest Drosophila suzukii has shifted its oviposition (egg-laying) niche from fermented fruits to ripe, non-fermented fruits, causing significant damage to a wide range of fruit crops worldwide. This study investigated the chemosensory changes underlying this evolutionary shift and ask whether fruit sugars, which are depleted during fermentation, are important gustatory cues that direct D. suzukii oviposition to sweet, ripe fruits. D. suzukii was shown to have expanded its range of oviposition responses to lower sugar concentrations than the model D. melanogaster, which prefers to lay eggs on fermented fruit. The increased response of D. suzukii to sugar correlates with an increase in the value of sugar relative to a fermented strawberry substrate in oviposition decisions. In addition, it was shown by genetic manipulation of sugar-gustatory receptor neurons (GRNs) that sugar perception is required for D. suzukii to prefer a ripe substrate over a fermented substrate, but not for D. melanogaster to prefer the fermented substrate. Thus, sugar is a major determinant of D. suzukii's choice of complex substrates. Calcium imaging experiments in the brain's primary gustatory center (ssuboesophageal zone) show that D. suzukii GRNs are not more sensitive to sugar than their D. melanogaster counterparts, suggesting that increased sugar valuation is encoded in downstream circuits of the central nervous system (CNS). Taken together, these data suggest that evolutionary changes in central brain sugar valuation computations are involved in driving D. suzukii's oviposition preference for sweet, ripe fruit.

Flynn, J. M., Ahmed-Braimah, Y. H., Long, M., Wing, R. A., Clark, A. G. (2024). High-Quality Genome Assemblies Reveal Evolutionary Dynamics of Repetitive DNA and Structural Rearrangements in the Drosophila virilis Subgroup. Genome biology and evolution, 16(1) PubMed ID: 38159044
Summary:
High-quality genome assemblies across a range of nontraditional model organisms can accelerate the discovery of novel aspects of genome evolution. The Drosophila virilis group has several attributes that distinguish it from more highly studied species in the Drosophila genus, such as an unusual abundance of repetitive elements and extensive karyotype evolution, in addition to being an attractive model for speciation genetics. This study used long-read sequencing to assemble five genomes of three virilis group species and characterized sequence and structural divergence and repetitive DNA evolution. The contiguous genome assemblies allow characterization of chromosomal arrangements with ease and can facilitate analysis of inversion breakpoints. A small panel of resequenced strains was leveraged to explore the genomic pattern of divergence and polymorphism in this species; known demographic histories were shown to largely predict the extent of genome-wide segregating polymorphism. It was further found that a neo-X chromosome in Drosophila americana displays X-like levels of nucleotide diversity. Unusual repetitive elements were found to be responsible for much of the divergence in genome composition among species. Helitron-derived tandem repeats tripled in abundance on the Y chromosome in D. americana compared to Drosophila novamexicana, accounting for most of the difference in repeat content between these sister species. Repeats with characteristics of both transposable elements and satellite DNAs expanded by 3-fold, mostly in euchromatin, in both D. americana and D. novamexicana compared to D. virilis. These results represent a major advance in understanding of genome biology in this emerging model clade.

Talagala, S., Rakosy, E., Long, T. A. F. (2024). Sexual selection and the nonrandom union of gametes: retesting for assortative mating by fitness in Drosophila melanogaster. Evolution, 78(1):26-38 PubMed ID: 37875133
Summary:
While numerous theoretical population genetic models predict that mating assortatively by genetic "quality" will enhance the efficiency of purging of deleterious mutations and/or the spread of beneficial alleles in the gene pool, empirical examples of assortative mating by quality are surprisingly rare and often inconclusive. This study set out to examine whether fruit flies (Drosophila melanogaster) engage in assortative mating by body-size phenotype, a composite trait strongly associated with both reproductive success and survival and is considered a reliable indicator of natural genetic quality. Male and female flies of different body-size classes (large and small) were obtained under typical culture conditions, which allows use of standing variation of body size without involving artificial nutritional manipulation, so that their interactions and mating patterns could be measured. While flies did not exhibit assortative courtship behavior, when patterns of offspring production were analyzed, it was found that individuals produced more offspring with partners of similar quality/body size, resulting produced from disassortative mating. Together, these results validate theoretical predictions that sexual selection can enhance the effects of natural selection and consequently the rate of adaptive evolution in a positive correlation in fitness between mates. Subsequent assays of offspring fitness indicated that assortative mating produced sons and daughters that had greater or equal reproductive success than those.

Bladen, J., Cooper, J. C., Ridges, J. T., Guo, P., Phadnis, N. (2024). A new hybrid incompatibility locus between D. melanogaster and D. sechellia. Genetics, PubMed ID: 38184848
Summary:
Despite the fundamental importance of hybrid incompatibilities to the process of speciation, there are few cases where the evolution and genetic architecture of hybrid incompatibilities are understood. One of the longest studied hybrid incompatibilities causes F1 hybrid male inviability in crosses between Drosophila melanogaster females and males from the D. simulans clade of species - D. simulans, D. mauritiana, and D. sechellia. This study discovered dramatic differences in the manifestation of this lethal hybrid incompatibility among the D. simulans clade of species. In particular, F1 hybrid males between D. melanogaster and D. sechellia are resistant to hybrid rescue through RNAi knockdown of an essential hybrid incompatibility gene. To understand the genetic basis of this inter-species difference in hybrid rescue, a triple hybrid mapping method was developed. The results show that two discrete large effect loci and many dispersed small effect changes across the genome underlie D. sechellia aversion to hybrid rescue. The large effect loci encompass a known incompatibility gene Lethal hybrid rescue (Lhr) and previously unknown factor, Sechellia aversion to hybrid rescue (Satyr). These results show that the genetic architecture of F1 hybrid male inviability is overlapping but not identical in the three inter-species crosses. These results raise questions about whether new hybrid incompatibility genes can integrate into an existing hybrid incompatibility thus increasing in complexity over time, or if the continued evolution of genes can gradually strengthen an existing hybrid incompatibility.

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