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Thursday July 17th - Vesicles and Cytoskeleton

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December 2018 		Zhang, X., Avellaneda, J., Spletter, M. L., Lemke, S. B., Mangeol, P., Habermann, B. H., Schnorrer, F. (2024). Mechanoresponsive regulation of myogenesis by the force-sensing transcriptional regulator Tono. Curr Biol, PubMed ID: 39163855
Summary:
Muscle morphogenesis is a multi-step program, starting with myoblast fusion, followed by myotube-tendon attachment and sarcomere assembly, with subsequent sarcomere maturation, mitochondrial amplification, and specialization. The correct chronological order of these steps requires precise control of the transcriptional regulators and their effectors. How this regulation is achieved during muscle development is not well understood. In a genome-wide RNAi screen in Drosophila, this study identified the BTB-zinc-finger protein Tono (CG32121) as a muscle-specific transcriptional regulator. tono mutant flight muscles display severe deficits in mitochondria and sarcomere maturation, resulting in uncontrolled contractile forces causing muscle rupture and degeneration during development. Tono protein is expressed during sarcomere maturation and localizes in distinct condensates in flight muscle nuclei. Interestingly, internal pressure exerted by the maturing sarcomeres deforms the muscle nuclei into elongated shapes and changes the Tono condensates, suggesting that Tono senses the mechanical status of the muscle cells. Indeed, external mechanical pressure on the muscles triggers rapid liquid-liquid phase separation of Tono utilizing its BTB domain. Thus, it is proposed that Tono senses high mechanical pressure to adapt muscle transcription, specifically at the sarcomere maturation stages. Consistently, tono mutant muscles display specific defects in a transcriptional switch that represses early muscle differentiation genes and boosts late ones. It is hypothesized that a similar mechano-responsive regulation mechanism may control the activity of related BTB-zinc-finger proteins that, if mutated, can result in uncontrolled force production in human muscle. 		Kos, P., Baumann, O. (2024). Spatial arrangement, polarity, and posttranslational modifications of the microtubule system in the Drosophila eye. Cell and tissue research, PubMed ID: 39152365 PMID
Summary:
This study examined the organization of the microtubule system in photoreceptor cells and pigment cells within the adult Drosophila compound eye. Immunofluorescence localization of tubulin and of Short stop, a spectraplakin that has been reported to be involved in the anchorage of microtubule minus ends at the membrane, suggests the presence of non-centrosomal microtubule-organizing centers at the distal tip of the visual cells. Ultrastructural analyses confirm that microtubules emanate from membrane-associated plaques at the site of contact with cone cells and that all microtubules are aligned in distal-proximal direction within the photoreceptor cells. Determination of microtubule polarities demonstrated that about 95% of the microtubules in photoreceptor cells are oriented with their plus end in the direction of the synapse. Pigment cells in the eye contain only microtubules aligned in distal-proximal direction, with their plus end pointing towards the retinal floor. There, two populations of microtubules can be distinguished, single microtubules and bundled microtubules, the latter associated with actin filaments. Whereas microtubules in both photoreceptor cells and pigment cells are acetylated and mono/bi-glutamylated on α-tubulin, bundled microtubules in pigment cells are apparently also mono/bi-glutamylated on &β;-tubulin, providing the possibility of binding different microtubule-associated proteins.
Niwa, S., Watanabe, T., Chiba, K. (2024). The FHA domain is essential for the autoinhibition of KIF1A/UNC-104. J Cell Sci, PubMed ID: 39239883
Summary:
KIF1A/UNC-104, a member of the kinesin superfamily motor proteins, plays a pivotal role in the axonal transport of synaptic vesicles and their precursors. Drosophila melanogaster UNC-104 (DmUNC-104) is a relatively recently discovered Drosophila kinesin. Although some point mutations that disrupt synapse formation have been identified, the biochemical properties of DmUNC-104 protein have not been investigated. Recombinant full-length DmUNC-104 protein was prepared and its biochemical features were determined. The effect was anayzed of a previously identified missense mutation in the forkhead-associated (FHA) domain, called bristly (bris). The bris mutation strongly promoted the dimerization of DmUNC-104 protein, whereas wild-type DmUNC-104 was a mixture of monomers and dimers. The G618R mutation near the FHA domain which was previously shown to disrupt the autoinhibition of C. elegans UNC-104. The biochemical properties of the G618R mutant recapitulated those of the bris mutant. Finally, this study found that disease-associated mutations also promote the dimerization of DmUNC-104. Collectively, this results suggest that the FHA domain is essential for the autoinhibition of KIF1A/UNC-104, and that abnormal dimerization of KIF1A is linked to human diseases. 		Zein-Sabatto, H., Brockett, J. S., Jin, L., Husbands, C. A., Lee, J., Fang, J., Buehler, J., Bullock, S. L., Lerit, D. A. (2024). Centrocortin potentiates co-translational localization of its mRNA to the centrosome via dynein. bioRxiv, PubMed ID: 39149256
Summary:
Centrosomes rely upon proteins within the pericentriolar material to nucleate and organize microtubules. Several mRNAs also reside at centrosomes, although less is known about how and why they accumulate there. Previous studies showed that local Centrocortin (Cen) mRNA supports centrosome separation, microtubule organization, and viability in Drosophila embryos. Using Cen mRNA as a model, this study examined mechanisms of centrosomal mRNA localization. While the Cen N'-terminus is sufficient for protein enrichment at centrosomes, multiple domains cooperate to concentrate Cen mRNA at this location. This study further identified an N'-terminal motif within Cen that is conserved among dynein cargo adaptor proteins, and its contribution to RNA localization was assessed. These results support a model whereby Cen protein enables the accumulation of its own mRNA to centrosomes through a mechanism requiring active translation, microtubules, and the dynein motor complex. Taken together, these data uncover the basis of translation-dependent localization of a centrosomal RNA required for mitotic integrity.
Voelzmann, A., Nuhu-Soso, L., Roof, A., Patel, S., Bennett, H., Adamson, A., Bentley, M., Evans, G. J. O., Hahn, I. (2025). GSK-3β coordinates axonal microtubule organisation through Shot and Tau. bioRxiv, PubMed ID: 40654684
Summary:
Glycogen Synthase Kinase 3β (GSK-3β) is a key coordinator of neuronal development and maintenance; hyperactive GSK-3β is linked to neurodevelopmental and -degenerative diseases and therefore a promising therapeutic target. In neurons, GSK-3β coordinates the cytoskeleton by phosphorylating microtubule-binding proteins. This study found that tight regulation of GSK-3β kinase activity is required for the maintenance of parallel microtubule bundles in Drosophila and rat axons. Up- or down-regulation of GSK-3β led to axons forming pathological swellings in which microtubule bundles disintegrated into disorganised, curled microtubules. The microtubule bundling proteins Shot and Tau were identified as key GSK-3β targets, and GSK-3β was found to exert its regulatory effect on microtubule bundling through them. GSK-3β regulates the ability of Shot and Tau to attach to microtubules and/or the plus-end protein Eb1. Mis-regulation of GSK-3β leads to the loss of Eb1-Shot-mediated guidance of polymerising microtubules into parallel bundles, thus causing disorganisation. Microtubule disorganisation is proposed as a new explanation for how GSK-3β hyperactivity leads to neurodegeneration and why global inhibition of GSK-3β has not been successful in clinical trials for neuronal disorders. 		Delaney, M., Zhao, Y., van de Leemput, J., Lee, H., Han, Z. (2024). Actin Cytoskeleton and Integrin Components Are Interdependent for Slit Diaphragm Maintenance in Drosophila Nephrocytes. Cells, 13(16) PubMed ID: 39195240
Summary:
In nephrotic syndrome, the podocyte filtration structures are damaged in a process called foot process effacement. This is mediated by the actin cytoskeleton; however, which actins are involved and how they interact with other filtration components, like the basement membrane, remains poorly understood. This study used the well-established Drosophila pericardial nephrocyte-the equivalent of podocytes in flies-knockdown models (RNAi) to study the interplay of the actin cytoskeleton (Act5C, Act57B, Act42A, and Act87E), alpha- and β-integrin (basement membrane), and the slit diaphragm (Sns and Pyd). Knockdown of an actin gene led to variations of formation of actin stress fibers, the internalization of Sns, and a disrupted slit diaphragm cortical pattern. Notably, deficiency of Act5C, which resulted in complete absence of nephrocytes, could be partially mitigated by overexpressing Act42A or Act87E, suggesting at least partial functional redundancy. Integrin localized near the actin cytoskeleton as well as slit diaphragm components, but when the nephrocyte cytoskeleton or slit diaphragm was disrupted, this switched to colocalization, both at the surface and internalized in aggregates. Altogether, the data show that the interdependence of the slit diaphragm, actin cytoskeleton, and integrins is key to the structure and function of the Drosophila nephrocyte.
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Tuesday July 15th - Stress

Herranz, H., Ye, Z., Olsen, J. V., Li, Y., Nygaard, M., Christensen, K., Tong, X., Bohr, V. A., Rasmussen, L. J., Dai, F. (2024). FOXO-regulated OSER1 reduces oxidative stress and extends lifespan in multiple species.. Nat Commun, 15(1):7144 PubMed ID: 39164296
Summary:
FOXO transcription factors modulate aging-related pathways and influence longevity in multiple species, but the transcriptional targets that mediate these effects remain largely unknown. This study identified an evolutionarily conserved FOXO target gene, Oxidative stress-responsive serine-rich protein 1 (OSER1), whose overexpression extends lifespan in silkworms, nematodes, and flies, while its depletion correspondingly shortens lifespan. In flies, overexpression of OSER1 increases resistance to oxidative stress, starvation, and heat shock, while OSER1-depleted flies are more vulnerable to these stressors. In silkworms, hydrogen peroxide both induces and is scavenged by OSER1 in vitro and in vivo. Knockdown of OSER1 in Caenorhabditis elegans leads to increased ROS production and shorter lifespan, mitochondrial fragmentation, decreased ATP production, and altered transcription of mitochondrial genes. Human proteomic analysis suggests that OSER1 plays roles in oxidative stress response, cellular senescence, and reproduction, which is consistent with the data and suggests that OSER1 could play a role in fertility in silkworms and nematodes. Human studies demonstrate that polymorphic variants in OSER1 are associated with human longevity. In summary, OSER1 is an evolutionarily conserved FOXO-regulated protein that improves resistance to oxidative stress, maintains mitochondrial functional integrity, and increases lifespan in multiple species. Additional studies will clarify the role of OSER1 as a critical effector of healthy aging. 		Fulton, T. L., Johnstone, J. N., Tan, J. J., Balagopal, K., Dedman, A., Chan, A. Y., Johnson, T. K., Mirth, C. K., Piper, M. D. W. (2024). Transiently restricting individual amino acids protects Drosophila melanogaster against multiple stressors. Open biology, 14(8):240093 PubMed ID: 39106944
Summary:
Nutrition and resilience are linked, though it is not yet clear how diet confers stress resistance or the breadth of stressors that it can protect against. Previous WORK shoD that transiently restricting an essential amino acid can protect Drosophila melanogaster against nicotine poisoning. This study sought to characterize the nature of this dietary-mediated protection and determine whether it was sex, amino acid and/or nicotine specific. When between sexes were compared, isoleucine deprivation was found to increases female, but not male, nicotine resistance. Surprirsingly, it was found that this protection afforded to females was not replicated by dietary protein restriction and was instead specific to individual amino acid restriction. To understand whether these beneficial effects of diet were specific to nicotine or were generalizable across stressors, flies were pre-treated with amino acid restriction diets and then exposed to other types of stress. Some of ome of the diets protected against nicotine also protected against oxidative and starvation stress, and improved survival following cold shock. Interestingly, it was found that a diet lacking isoleucine was the only diet to protect against all these stressors. These data point to isoleucine as a critical determinant of robustness in the face of environmental challenges.
Glineburg, M. R., Yildirim, E., Gomez, N., Rodriguez, G., Pak, J., Li, X., Altheim, C., Waksmacki, J., McInerney, G. M., Barmada, S. J., Todd, P. K. (2024). Stress granule formation helps to mitigate neurodegeneration. Nucleic Acids Res, PubMed ID: 39106168
Summary:
Cellular stress pathways that inhibit translation initiation lead to transient formation of cytoplasmic RNA/protein complexes known as stress granules. Many of the proteins found within stress granules and the dynamics of stress granule formation and dissolution are implicated in neurodegenerative disease. Whether stress granule formation is protective or harmful in neurodegenerative conditions is not known. To address this, advantage was taken of the alphavirus protein nsP3, which selectively binds dimers of the central stress granule nucleator protein 6G3BP and markedly reduces stress granule formation without directly impacting the protein translational inhibitory pathways that trigger stress granule formation. In Drosophila and rodent neurons, reducing stress granule formation with nsP3 had modest impacts on lifespan even in the setting of serial stress pathway induction. In contrast, reducing stress granule formation in models of ataxia, amyotrophic lateral sclerosis and frontotemporal dementia largely exacerbated disease phenotypes. These data support a model whereby stress granules mitigate, rather than promote, neurodegenerative cascades. 		Esteban-Collado, J., Fernandez-Manas, M., Fernandez-Moreno, M., Maeso, I., Corominas, M., Serras, F. (2024). Reactive oxygen species activate the Drosophila TNF receptor Wengen for damage-induced regeneration. The EMBO journal, 43(17):3604-3626 PubMed ID: 39020149
Summary:
Tumor necrosis factor receptors (TNFRs) control pleiotropic pro-inflammatory functions that range from apoptosis to cell survival. The ability to trigger a particular function will depend on the upstream cues, association with regulatory complexes, and downstream pathways. In Drosophila melanogaster, two TNFRs have been identified, Wengen (Wgn) and Grindelwald (Grnd). Although several reports associate these receptors with JNK-dependent apoptosis, it has recently been found that Wgn activates a variety of other functions. This study demonstrates that Wgn is required for survival by protecting cells from apoptosis. This is mediated by dTRAF1 and results in the activation of p38 MAP kinase. Remarkably, Wgn is required for apoptosis-induced regeneration and is activated by the reactive oxygen species (ROS) produced following apoptosis. This ROS activation is exclusive for Wgn, but not for Grnd, and can occur after knocking down Eiger/TNFα. The extracellular cysteine-rich domain of Grnd is much more divergent than that of Wgn, which is more similar to TNFRs from other animals, including humans. These results show a novel TNFR function that responds to stressors by ensuring p38-dependent regeneration.
Sun, X., Zhou, D., Sun, Y., Zhao, Y., Deng, Y., Pang, X., Liu, Q., Zhou, Z. (2024). Oxidative stress reprograms the transcriptional coactivator Yki to suppress cell proliferation. Cell Rep, 43(8):114584 PubMed ID: 39106181
Summary:
The transcriptional coactivator Yorkie (Yki) regulates organ size by promoting cell proliferation. It is unclear how cells control Yki activity when exposed to harmful stimuli such as oxidative stress. This study shows that oxidative stress inhibits the binding of Yki to Scalloped (Sd) but promotes the interaction of Yki with another transcription factor, forkhead box O (Foxo), ultimately leading to a halt in cell proliferation. Mechanistically, Foxo normally exhibits a low binding affinity for Yki, allowing Yki to form a complex with Sd and activate proliferative genes. Under oxidative stress, Usp7 deubiquitinates Foxo to promote its interaction with Yki, thereby activating the expression of proliferation suppressors. Finally, Yki was shown to be essential for Drosophila survival under oxidative stress. In summary, these findings suggest that oxidative stress reprograms Yki from a proliferation-promoting factor to a proliferation suppressor, forming a self-protective mechanism. 		Camilleri-Robles, C., Climent-Canto, P., Llorens-Giralt, P., Klein, C. C., Serras, F., Corominas, M. (2024). A shift in chromatin binding of phosphorylated p38 precedes transcriptional changes upon oxidative stress. FEBS letters, PubMed ID: 39218622
Summary:
P38 mitogen-activated protein kinases are key in the regulation of the cellular response to stressors. P38 is known to regulate transcription, mRNA processing, stability, and translation. The transcriptional changes mediated by phosphorylated p38 (P-p38) in response to extracellular stimuli have been thoroughly analyzed in many tissues and organisms. However, the genomic localization of chromatin-associated P-p38 remains poorly understood. This study analyzed the chromatin binding of activated P-p38 and its role in the response to reactive oxygen species (ROS) in Drosophila S2 cells. P-p38 was found to be already bound to chromatin in basal conditions. After ROS exposure, chromatin-associated P-p38 relocates towards genes involved in the recovery process. These findings highlight the role of P-p38 dynamic chromatin binding in orchestrating gene expression responses to oxidative stress.
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Friday July 11th - Gonads

Fang, Y., Zhang, F., Zhao, F., Wang, J., Cheng, X., Ye, F., He, J., Zhao, L., Su, Y. (2024). RpL38 modulates germ cell differentiation by controlling Bam expression in Drosophila testis. Science China Life sciences, PubMed ID: 39187660
Summary:
Switching from mitotic spermatogonia to meiotic spermatocytes is critical to producing haploid sperms during male germ cell differentiation. However, the underlying mechanisms of this switch remain largely unexplored. In Drosophila melanogaster, the gene RpL38 encodes the ribosomal protein L38, one component of the 60S subunit of ribosomes. Its depletion in spermatogonia was found to severely diminished the production of mature sperms and thus led to the infertility of male flies. By examining the germ cell differentiation in testes, RpL38-knockdown was found to block the transition from spermatogonia to spermatocytes and accumulated spermatogonia in the testis. To understand the intrinsic reason for this blockage, proteomic analysis was conducted for these spermatogonia populations. Differing from the control spermatogonia, the accumulated spermatogonia in RpL38-knockdown testes already expressed many spermatocyte markers but lacked many meiosis-related proteins, suggesting that spermatogonia need to prepare some important proteins for meiosis to complete their switch into spermatocytes. Mechanistically, the expression of bag of marbles (bam), a crucial determinant in the transition from spermatogonia to spermatocytes, was found to be inhibited at both the mRNA and protein levels upon RpL38 depletion. The bam loss phenocopied RpL38 RNAi in the testis phenotype and transcriptomic profiling. Strikingly, overexpressing bam was able to fully rescue the testis abnormality and infertility of RpL38-knockdown flies, indicating that bam is the key effector downstream of RpL38 to regulate spermatogonia differentiation. Overall, these data suggested that germ cells start to prepare meiosis-related proteins as early as the spermatogonial stage, and RpL38 in spermatogonia is required to regulate their transition toward spermatocytes in a bam-dependent manner, providing new knowledge for understanding of the transition process from spermatogonia to spermatocytes in Drosophila spermatogenesis. 		Mahmoudzadeh, N. H., Heidarian, Y., Tourigny, J. P., Fitt, A. J., Beebe, K., Li, H., Luhur, A., Buddika, K., Mungcal, L., Kundu, A., Policastro, R. A., Brinkley, G. J., Zentner, G. E., Nemkov, T., Pepin, R., Chawla, G., Sudarshan, S., Rodan, A. R., D'Alessandro, A., Tennessen, J. M. (2024). Renal L-2-hydroxyglutarate dehydrogenase activity promotes hypoxia tolerance and mitochondrial metabolism in Drosophila melanogaster. Molecular metabolism, 89:102013 PubMed ID: 39182840
Summary:
The mitochondrial enzyme L-2-hydroxyglutarate dehydrogenase (L2HGDH) regulates the abundance of L-2-hydroxyglutarate (L-2HG), a potent signaling metabolite capable of influencing chromatin architecture, mitochondrial metabolism, and cell fate decisions. Loss of L2hgdh activity in humans induces ectopic L-2HG accumulation, resulting in neurodevelopmental defects, altered immune cell function, and enhanced growth of clear cell renal cell carcinomas. To better understand the molecular mechanisms that underlie these disease pathologies, the fruit fly Drosophila melanogaster was used to investigate the endogenous functions of L2hgdh. L2hgdh mutant adult male flies were analyzed under normoxic and hypoxic conditions using a combination of semi-targeted metabolomics and RNA-seq. These multi-omic analyses were complemented by tissue-specific genetic studies that examined the effects of L2hgdh mutations on the Drosophila renal system (Malpighian tubules; MTs). These studies revealed that while L2hgdh is not essential for growth or viability under standard culture conditions, LL2hgdhmutants are hypersensitive to hypoxia and expire during the reoxygenation phase with severe disruptions of mitochondrial metabolism. Moreover, this study found that the fly renal system is a key site of L2hgdh activity, as L2hgdh mutants that express a rescuing transgene within the MTs survive hypoxia treatment and exhibit normal levels of mitochondrial metabolites. Even under normoxic conditions, L2hgdh mutant MTs experience significant metabolic stress and are sensitized to aberrant growth upon Egfr activation. CONCLUSIONS: These findings present a model in which renal L2hgdh activity limits systemic L-2HG accumulation, thus indirectly regulating the balance between glycolytic and mitochondrial metabolism, enabling successful recovery from hypoxia exposure, and ensuring renal tissue integrity.
Buglak, D. B., Holmes, K. H. M., Galletta, B. J., Rusan, N. M. (2024). The proximal centriole-like structure maintains nucleus-centriole architecture in sperm. J Cell Sci, 137(17) PubMed ID: 39166297
Summary:
Proper connection between the sperm head and tail is critical for sperm motility and fertilization. Head-tail linkage is mediated by the head-tail coupling apparatus (HTCA), which secures the axoneme (tail) to the nucleus (head). However, the molecular architecture of the HTCA is poorly understood. This study used Drosophila to investigate formation and remodeling of the HTCA throughout spermiogenesis by visualizing key components of this complex. Using structured illumination microscopy, Key HTCA proteins Spag4 and Yuri were shown to form a 'centriole cap' that surrounds the centriole (or basal body) as it invaginates into the surface of the nucleus. As development progresses, the centriole is laterally displaced to the side of the nucleus while the HTCA expands under the nucleus, forming what we term the 'nuclear shelf'. The proximal centriole-like (PCL) structure is positioned under the nuclear shelf, functioning as a crucial stabilizer of centriole-nucleus attachment. Together, our data indicate that the HTCA is a complex, multi-point attachment site that simultaneously engages the PCL, the centriole and the nucleus to ensure proper head-tail connection during late-stage spermiogenesis. 		Zhang, L., Ten Hagen, K. G. (2024). O-Glycosylation of a male seminal fluid protein influences sperm binding and female postmating behavior. PNAS nexus, 3(8):pgae322 PubMed ID: 39189023
Summary:
Glycoproteins are abundant within the human reproductive system and alterations in glycosylation lead to reproductive disorders, suggesting that glycans play an important role in reproductive function. In this study, the Drosophila reproductive system was used as a model to investigate the biological functions of O-glycosylation. O-glycosylation in the male accessory glands, an organ responsible for secreting seminal fluid proteins, plays important roles in female postmating behavior. The loss of one O-glycosyltransferase, PGANT9, in the male reproductive system resulted in decreased egg production in mated females. One substrate of PGANT9 was detected, lectin-46Ca (CG1656), which is known to affect female postmating responses. Loss of lectin-46Ca O-glycosylation affects its ability to associate with sperm tails, resulting in reduced transfer within the female reproductive system. These results provide the first example that O-glycosylation of a seminal fluid protein affects its ability to associate with sperm in vivo. These studies may shed light on the biological function of O-glycans in mammalian reproduction.
Guay, S. Y., Patel, P. H., Thomalla, J. M., McDermott, K. L., O'Toole, J. M., Arnold, S. E., Obrycki, S. J., Wolfner, M. F., Findlay, G. D. (2024). A newly evolved gene is essential for efficient sperm entry into eggs in Drosophila melanogaster.bioRxiv, PubMed ID: 39149251
Summary:
New genes arise through a variety of evolutionary processes and provide raw material for adaptation in the face of both natural and sexual selection. De novo evolved genes emerge from previously non-protein-coding DNA sequences, and many such genes are expressed in male reproductive structures. In Drosophila melanogaster, several putative de novo genes have evolved essential roles in spermatogenesis, but whether such genes can also impact sperm function beyond the male has not been investigated. This study identified a putative de novo gene, katherine johnson (kj; CG43167), that is required for high levels of male fertility. Males that do not express kj produce and transfer sperm that are stored normally in females, but sperm from these males enter eggs with severely reduced efficiency. Using a tagged transgenic rescue construct, KJ protein was observed to localize to the nuclear periphery in various stages of spermatogenesis, but is not detectable in mature sperm. These data suggest that kj exerts an effect on sperm development, the loss of which results in reduced fertilization ability. While previous bioinformatic analyses suggested the kj gene was restricted to the melanogaster group of Drosophila,putative orthologs with conserved synteny, male-biased expression, and predicted protein features were observed across the genus, as well as instances of gene loss in some lineages. Thus, kj potentially arose in the Drosophila common ancestor and subsequently evolved an essential role in D. melanogaster. These results demonstrate a new aspect of male reproduction that has been shaped by new gene evolution and provide a molecular foothold for further investigating the mechanism of sperm entry into eggs in Drosophila. 		Kagemann, C. H., Colocho, G. M., Aquadro, C. F. (2024). Non-ovarian Wolbachia pipientis titer correlates with fertility rescue of a Drosophila melanogaster bag of marbles hypomorph. microPublication biology, 2024 PubMed ID: 39157806
Summary:
Bag of marbles (bam<) is an essential gene that regulates germline stem cell maintenance and germline stem cell daughter cell differentiation in Drosophila melanogaster . When bam is partially functional (hypomorphic), the introduction of Wolbachia pipientis rescues the mutant fertility phenotype that would otherwise result in partial sterility. Infection by different W. pipientis variants results in differential rescue of the bam hypomorph fertility phenotype. The varying degrees of rescue exhibited in the bam hypomorph when exposed to different W. pipientis variants was intreguing, prompting an investigation into whether this phenomenon is attributable to variations in the titers of W. pipientis variants. No significant difference in ovarian titer was found between two W. pipientis variant groups, w Mel-like (low bam hypomorph fertility rescue) and w MelCS-like variants (higher bam hypomorph fertility rescue), at bam hypomorph peak fertility. However, carcass (whole flies without the ovaries) titer between w Mel-like and w MelCS-like infected bam hypomorph differed during peak fertility rescue. A positive correlation emerged between the combined titers of ovarian and carcass infections and fertility, implying a more extensive influence that extends beyond ovarian infection alone.
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Thursday July 10th - Enhancers and Transcriptional Regulation

Ke, W., Fujioka, M., Schedl, P., Jaynes, J. B. (2024). Stem-loop and circle-loop TADs generated by directional pairing of boundary elements have distinct physical and regulatory properties. Elife, 13 PubMed ID: 39110491
Summary:
The chromosomes in multicellular eukaryotes are organized into a series of topologically independent loops called TADs. In flies, TADs are formed by physical interactions between neighboring boundaries. Fly boundaries exhibit distinct partner preferences, and pairing interactions between boundaries are typically orientation-dependent. Pairing can be head-to-tail or head-to-head. The former generates a stem-loop TAD, while the latter gives a circle-loop TAD. The TAD that encompasses the Drosophila even skipped (eve) gene is formed by the head-to-tail pairing of the nhomie and homie boundaries. To explore the relationship between loop topology and the physical and regulatory landscape, the nhomie boundary region was flanked with two attP sites. The attP sites were then used to generate four boundary replacements: λ DNA, nhomie forward (WT orientation), nhomie reverse (opposite of WT orientation), and homie forward (same orientation as WT homie). The nhomie forward replacement restores the WT physical and regulatory landscape: in MicroC experiments, the eve TAD is a 'volcano' triangle topped by a plume, and the eve gene and its regulatory elements are sequestered from interactions with neighbors. The λ DNA replacement lacks boundary function: the endpoint of the 'new' eve TAD on the nhomie side is ill-defined, and eve stripe enhancers activate a nearby gene, eIF3j. While nhomie reverse and homie forward restore the eve TAD, the topology is a circle-loop, and this changes the local physical and regulatory landscape. In MicroC experiments, the eve TAD interacts with its neighbors, and the plume at the top of the eve triangle peak is converted to a pair of 'clouds' of contacts with the next-door TADs. Consistent with the loss of isolation afforded by the stem-loop topology, the eve enhancers weakly activate genes in the neighboring TADs. Conversely, eve function is partially disrupted. 		Glaser-Schmitt, A., Lemoine, M., Kaltenpoth, M., Parsch, J. (2024). Pervasive tissue-, genetic background-, and allele-specific gene expression effects in Drosophila melanogaster. PLoS Genet, 20(8):e1011257 PubMed ID: 39178312
Summary:
The pervasiveness of gene expression variation and its contribution to phenotypic variation and evolution is well known. This gene expression variation is context dependent, with differences in regulatory architecture often associated with intrinsic and environmental factors, and is modulated by regulatory elements that can act in cis (linked) or in trans (unlinked) relative to the genes they affect. So far, little is known about how this genetic variation affects the evolution of regulatory architecture among closely related tissues during population divergence. To address this question, gene expression was analyzed in the midgut, hindgut, and Malpighian tubule as well as microbiome composition in the two gut tissues in four Drosophila melanogaster strains and their F1 hybrids from two divergent populations: one from the derived, European range and one from the ancestral, African range. In both the transcriptome and microbiome data, extensive tissue- and genetic background-specific effects were detected, including effects of genetic background on overall tissue specificity. Tissue-specific effects were typically stronger than genetic background-specific effects, although the two gut tissues were not more similar to each other than to the Malpighian tubules. An examination of allele specific expression revealed that, while both cis and trans effects were more tissue-specific in genes expressed differentially between populations than genes with conserved expression, trans effects were more tissue-specific than cis effects. Despite there being highly variable regulatory architecture, this observation was robust across tissues and genetic backgrounds, suggesting that the expression of trans variation can be spatially fine-tuned as well as or better than cis variation during population divergence and yielding new insights into cis and trans regulatory evolution.
Monastirioti, M., Koltsaki, I., Pitsidianaki, I., Skafida, E., Batsiotos, N., Delidakis, C. (2024). Notch-Dependent Expression of the Drosophila Hey Gene Is Supported by a Pair of Enhancers with Overlapping Activities. Genes, 15(8) PubMed ID: 39202431
Summary:
Drosophila Hey is a basic helix-loop-helix-orange (bHLH-O) protein with an important role in the establishment of distinct identities of postmitotic cells. Previously Hey was identified as a transcriptional target and effector of Notch signalling during the asymmetric division of neuronal progenitors, generating neurons of two types, and Notch-dependent expression of Hey also marks a subpopulation of the newborn enteroendocrine (EE) cells in the midgut primordium of the embryo. This study investigated the transcriptional regulation of Hey in neuronal and intestinal tissues. Two genomic regions upstream of the promoter (HeyUP) and in the second intron (HeyIN2) of the Hey gene were isolated based on the presence of binding motifs for Su(H), the transcription factor that mediates Notch activity. Both regions can direct the overlapping expression patterns of reporter transgenes recapitulating endogenous Hey expression. Moreover, while HeyIN2 represents a Notch-dependent enhancer, HeyUP confers both Notch-dependent and independent transcriptional regulation. Mutations were induced that removed the Su(H) binding motifs in either region, and then the the enhancer functionality was studied in the respective Hey mutant lines. The results provide direct evidence that although both enhancers support Notch-dependent regulation of the Hey gene, their role is redundant, as a Hey loss-of-function lethal phenotype is observed only after deletion of all their Su(H) binding motifs by CRISPR/Cas9. 		Villarreal-Puente, A., Altamirano-Torres, C., Jimenez-Mejia, G., Hernandez-Bautista, C., Montalvo-Mendez, R., Vazquez, M., Zurita, M., Resendez-Perez, D. (2024). Novel Antennapedia and Ultrabithorax trimeric complexes with TBP and Exd regulate transcription. Hereditas, 161(1):25 PubMed ID: 39080786
Summary:
Hox proteins interact with DNA and many other proteins, co-factors, transcriptional factors, chromatin remodeling components, non-coding RNAs and even the extracellular matrix that assembles the Hox complexes. The number of interacting partners continues to grow with diverse components and more transcriptional factors than initially thought. Hox complexes present many activities, but their molecular mechanisms to modulate their target genes remain unsolved. This study showed the protein-protein interaction of Antp with Ubx through the homeodomain using BiFC in Drosophila. Analysis of Antp-deletional mutants showed that AntpHD helixes 1 and 2 are required for the interaction with Ubx. Also, a novel interaction of Ubx with TBP was found, in which the PolyQ domain of TBP is required for the interaction. Moreover, the formation of two new trimeric complexes of Antp with Ubx, TBP and Exd was found using BiFC-FRET; these proteins, however, do not form a trimeric interaction with BIP2 or TFIIEβ. The novel trimeric complexes reduced Antp transcriptional activity, indicating that they could confer specificity for repression. These results increase the number of transcriptional factors in the Antp and Ubx interactomes that form two novel trimeric complexes with TBP and Exd. A new Ubx interaction with TBP was also found. These novel interactions provide important clues of the dynamics of Hox-interacting complexes involved in transcriptional regulation, contributing to better understand Hox function.
Zhang, L., Hodgins, L., Sakib, S., Verbeem, A., Mahmood, A., Perez-Romero, C., Marmion, R. A., Dostatni, N., Fradin, C. (2024)s. Both the transcriptional activator, Bcd, and repressor, Cic, form small mobile oligomeric clusters. Biophys J, PubMed ID: 39164967
Summary:
Transcription factors play an essential role in pattern formation during early embryo development, generating a strikingly fast and precise transcriptional response that results in sharp gene expression boundaries. To characterize the steps leading up to transcription, a side-by-side comparison was performed of the nuclear dynamics of two morphogens, a transcriptional activator, Bicoid (Bcd), and a transcriptional repressor, Capicua (Cic), both involved in body patterning along the anterior-posterior axis of the early Drosophila embryo. A combination of fluorescence recovery after photobleaching, fluorescence correlation spectroscopy, and single particle tracking was used to access a wide range of dynamical timescales. Despite their opposite effects on gene transcription, Bcd and Cic have very similar nuclear dynamics, characterized by the co-existence of a freely diffusing monomer population with a number of oligomeric clusters, which range from low stoichiometry and high mobility clusters to larger, DNA-bound hubs. These observations are consistent with the inclusion of both Bcd and Cic into transcriptional hubs or condensates, while putting constraints on the mechanism by which these form. These results fit in with the recent proposal that many transcription factors might share a common search strategy for target genes regulatory regions that makes use of their large unstructured regions, and may eventually help explain how the transcriptional response they elicit can be at the same time so fast and so precise. 		Jalal, M. S., Duttaroy, A. (2024). Maternal Spargel/dPGC-1 is critical for embryonic development and influences chorion gene amplification via Cyclin E activity. Dev Biol, 516:158-166 PubMed ID: 39173813
Summary:
The function of spargel/dPGC-1 in Drosophila oogenesis has been unequivocally established. This study sought to assess whether Spargel protein or RNA is essential for developmentally competent eggs. The trans-heterozygotic combination of two spargel mutant alleles allowed a decrease Spargel expression to very low levels. Using this model, this study demonstrated the requirement for Spargel in eggshell patterning and embryonic development, which led to the finding that spargel is a maternal effect gene. Further examination of Spargel's potential mechanism of action in eggshell biogenesis revealed that low levels of Spargel in the adult ovary cause diminished Cyclin E activity, resulting in reduced chorion gene amplification levels, leading to eggshell biogenesis defects. Thus, another novel role for Cyclin E/dPGC-1 is established whereby, through Cyclin E activity, this conserved transcriptional coactivator regulates the chorion gene amplification process.
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Wednesday July 9th - Adult Physiology and Metabolism

Chattopadhyay, D., Philip, S. M., Prabhakar, G., Machamada Bheemaiah, M. (2024). Influence of α-lipoic acid on longevity and stress resistance in Drosophila melanogaster fed with a high-fat diet. Biogerontology, PubMed ID: 39046586
Summary:
Consumption of a high-fat diet is accompanied by the risks of obesity and early onset of age-associated complications for which dietary interventions are imperative to combat. α-lipoic acid has been shown to hinder diet-induced obesity and induce lifespan-extending efficacy in model organisms. In this study, α-lipoic acid was investigated for its efficacy in improving lifespan and stress resistance in the Canton-S strain of Drosophila melanogaster fed with a high-fat diet. Furthermore, as mating status significantly impacts survival in fruit flies, flies were reared in two experimental groups-group one, in which males and females were bred together, and group two, in which males and females were bred separately. In group one, α-lipoic acid improved the mean lifespan, reduced the fecundity of females, and reduced the mean body weight of flies at a dose range of 2-2.5 mM, respectively. In group two, α-lipoic acid improved the mean lifespan, reduced the fecundity of females, and reduced the mean body weight of flies at a dose range of 1-2.5 mM, respectively. Improved climbing efficiency was observed with α-lipoic acid at the dose range of 1.5-2.5 mM in flies of group one and 1-2.5 mM in flies of group two, respectively. Administration of α-lipoic acid improved resistance to oxidative stress in only female flies of group one at 2.5 mM, whereas in group two, both male and female flies exhibited enhanced resistance to oxidative stress with α-lipoic acid at a dose range of 2-2.5 mM, respectively. Male and female flies of only group one showed improved resistance to heat shock stress with α-lipoic acid at a dose range of 2-2.5 mM. Only female flies of group two exhibited a slight improvement in recovery time following cold shock with α-lipoic acid only at 2.5 mM. No significant change in resistance to starvation stress was observed with any dose of α-lipoic acid in either group of flies. To summarize, data from this study suggested a probable dose and gender-dependent efficacy of α-lipoic acid in flies fed with a high-fat diet, which was significantly influenced by the mating status of flies due to varied rearing conditions. 		Nicol, A., Ahmed, M., Fischer, C., Garces, J. G., Magnus, S., Maung, N., Molisani, N., Petrov, S., Palu, R. A. S. (2024). Larval density can be used to predict genetic modifiers of glucagon signaling in Drosophila melanogaster. PLoS One, 19(8):e0302565 PubMed ID: 39196987
Summary:
Obesity is a growing concern. 42.3% of people in the U.S were considered obese between 2017-2018. Much is still unknown about the genetic components that contribute to weight gain. In humans, the hormone glucagon is a major contributor to the body's energy regulation as it signals for the breakdown of lipids. Treatments targeting the glucagon pathway have helped patients with both weight loss and appetite suppression. Understanding the genetic modifiers of glucagon signaling and its downstream pathways could enable the development of a wider variety of effective therapeutics. This study blocked the glucagon pathway in Drosophila melanogaster by reducing the expression of the fly ortholog of the glucagon receptor (AKHR). This model was then crossed to the Drosophila Genetic Reference Panel (DGRP) and looked for natural variation in fat content. We used variation in larval density to identify candidate modifier genes through a genome-wide association study. We then tested these modifier genes by increasing or decreasing their expression in the AKHR model. These candidates were initially screened with the same density assay used in the original study to narrow down to four candidate genes that substantially impacted the density of the larvae: THADA, AmyD, GluRIIC, and CG9826. These candidates were further characterized using biochemical assays to analyze stored metabolites such as triglycerides, glucose, glycogen, and protein under control, high sugar, and high fat conditions to see if the larvae are resistant to environmental changes. Our results indicate consistency between the results of the density assay and direct measurement of metabolite levels. In particular, THADA and AmyD are highlighted as interesting genes for additional study. This study should to improve understanding of the glucagon signaling pathway, obesity, and lipid metabolism. We also aim to provide candidate genes that can be regarded as future therapeutic targets.
Kauffman, M. R., DiAngelo, J. R. (2024). Glut1 Functions in Insulin-Producing Neurons to Regulate Lipid and Carbohydrate Storage in Drosophila. Biomolecules, 14(8) PubMed ID: 39199423
Summary:
Obesity remains one of the largest health problems in the world, arising from the excess storage of triglycerides (TAGs). However, the full complement of genes that are important for regulating TAG storage is not known. The Glut1 gene encodes a Drosophila glucose transporter that has been identified as a potential obesity gene through genetic screening. Yet, the tissue-specific metabolic functions of Glut1 are not fully understood. This study characterized the role of Glut1 in the fly brain by decreasing neuronal Glut1 levels with RNAi and measuring glycogen and TAGs. Glut1RNAi flies had decreased TAG and glycogen levels, suggesting a nonautonomous role of Glut1 in the fly brain to regulate nutrient storage. A group of hormones that regulate metabolism and are expressed in the fly brain are Drosophila insulin-like peptides (Ilps) 2, 3, and 5. Interestingly, this study observed blunted Ilp3 and Ilp5 expression in neuronal Glut1RNAi flies, suggesting Glut1 functions in insulin-producing neurons (IPCs) to regulate whole-organism TAG and glycogen storage. Consistent with this hypothesis, we also saw fewer TAGs and glycogens and decreased expression of Ilp3 and Ilp5 in flies with IPC-specific Glut1RNAi. Together, these data suggest Glut1 functions as a nutrient sensor in IPCs, controlling TAG and glycogen storage and regulating systemic energy homeostasis. 		Vonolfen, M. C., Meyer Zu Altenschildesche, F. L., Nam, H. J., Brodesser, S., Gyenis, A., Buellesbach, J., Lam, G., Thummel, C. S., Storelli, G. (2024). Drosophila HNF4 acts in distinct tissues to direct a switch between lipid storage and export in the gut. Cell Rep, 43(9):114693 PubMed ID: 39235946
Summary:
Nutrient digestion, absorption, and export must be coordinated in the gut to meet the nutritional needs of the organism. This study used the Drosophila intestine to characterize the mechanisms that coordinate the fate of dietary lipids. Enterocytes specialized in absorbing and exporting lipids to peripheral organs. Distinct hepatocyte-like cells, called oenocytes, communicate with these enterocytes to adjust intestinal lipid storage and export. A single transcription factor, Drosophila hepatocyte nuclear factor 4 (dHNF4), supports this gut-liver axis. In enterocytes, dHNF4 maximizes dietary lipid export by preventing their sequestration in cytoplasmic lipid droplets. In oenocytes, dHNF4 promotes the expression of the insulin antagonist ImpL2 to activate Foxo and suppress lipid retention in enterocytes. Disruption of this switch between lipid storage and export is associated with intestinal inflammation, suggesting a lipidic origin for inflammatory bowel diseases. These studies establish dHNF4 as a central regulator of intestinal metabolism and inter-organ lipid trafficking.
Wu, G., Ma, T., Hancock, C. E., Gonzalez, S., Aryal, B., Vaz, S., Chan, G., Palarca-Wong, M., Allen, N., Chung, C. I., Shu, X., Liu, Q. (2024). Opposing GPCR signaling programs protein intake setpoint in Drosophila. Cell, PubMed ID: 39197448
Summary:
Animals defend a target level for their fundamental needs, including food, water, and sleep. Deviation from the target range, or "setpoint," triggers motivated behaviors to eliminate that difference. Whether and how the setpoint itself is encoded remains enigmatic for all motivated behaviors. Employing a high-throughput feeding assay in Drosophila, this study demonstrate dthat the protein intake setpoint is set to different values in male, virgin female, and mated female flies to meet their varying protein demands. Leveraging this setpoint variability, this study found, remarkably, that the information on the intake setpoint is stored within the protein hunger neurons as the resting membrane potential. Two RFamide G protein-coupled receptor (GPCR) pathways, by tuning the resting membrane potential in opposite directions, coordinately program and adjust the protein intake setpoint. The FMRFa-FMRFaR-PKC pathway was found to signal to lower the protein intake setpoint, and the Myosuppressin-MSR2-PKA pathway signals were found to raise the protein intake setpoint. Together, these studies map the protein intake setpoint to a single trackable physiological parameter and elucidate the cellular and molecular mechanisms underlying setpoint determination and modulation. 		Kosakamoto, H., Sakuma, C., Okada, R., Miura, M., Obata, F. (2024). Context-dependent impact of the dietary non-essential amino acid tyrosine on Drosophila physiology and longevity.. Sci Adv, 10(35):eadn7167 PubMed ID: 39213345
Summary:
Dietary protein intake modulates growth, reproduction, and longevity by stimulating amino acid (AA)-sensing pathways. Essential AAs are often considered as limiting nutrients during protein scarcity, and the role of dietary non-essential AAs (NEAAs) is less explored. Although tyrosine has been reported to be crucial for sensing protein restriction in Drosophila larvae, its effect on adult physiology and longevity remains unclear. Using a synthetic diet, a systematic investigation was performed of the effect of single NEAA deprivation on nutrient-sensing pathways, reproductive ability, starvation resistance, feeding behavior, and life span in adult female flies. Specifically, dietary tyrosine deprivation decreases internal tyrosine levels and fecundity, influences AA-sensing machineries, and extends life span. These nutritional responses are not observed under higher total AA intake or in infertile female flies, suggesting a context-dependent influence of dietary tyrosine. These findings highlight the unique role of tyrosine as a potentially limiting nutrient, underscoring its value for dietary interventions aimed at enhancing health span.
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Monday July 7th - Disease Models

Landaverde, S., Sleep, M., Lacoste, A., Tan, S., Schuback, R., Reiter, L. T., Iyengar, A. (2024). Glial expression of Drosophila UBE3A causes spontaneous seizures that can be modulated by 5-HT signaling. Neurobiol Dis, 200:106651 PubMed ID: 39197537
Summary:
Misexpression of the E3 ubiquitin ligase gene UBE3A is thought to contribute to a range of neurological disorders. In the context of Dup15q syndrome, additional genomic copies of UBE3A give rise to the autism, muscle hypotonia and spontaneous seizures characteristics of the disorder. In a Drosophila model of Dup 15q syndrome, it was recently shown that 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 compare the consequences of glial- and neuronal-driven dube3a expression on motor coordination and seizure susceptibility in Drosophila. To quantify seizure-related behavioral events, a hidden Markov model was developed and trained: driven dube3a expression led to clear motor phenotypes. However, only glial-driven dube3a expression displayed spontaneous seizure-associated immobilization events, that were clearly observed at high-temperature (38 °C). Using a tethered fly preparation amenable to electrophysiological monitoring of seizure activity, this study found glial-driven dube3a flies display aberrant spontaneous spike discharges which are bilaterally synchronized. Neither neuronal-dube3a overexpressing flies, nor control flies displayed these firing patterns. A previous drug screen was performed for FDA approved compounds that can suppress bang-sensitivity in glial-driven dube3a expressing flies; certain 5-HT modulators were identified as strong seizure suppressors. Glial-driven dube3a flies fed the serotonin reuptake inhibitor vortioxetine and the 5-HT(2A) antagonist ketanserin displayed reduced immobilization and spike bursting, consistent with the previous study. Together these findings highlight the potential for glial pathophysiology to drive Dup15q syndrome-related seizure activity. 		Bitar, S., Baumann, T., Weber, C., Abusaada, M., Rojas-Charry, L., Ziegler, P., Schettgen, T., Randerath, I. E., Venkataramani, V., Michalke, B., Hanschmann, E. M., Arena, G., Krueger, R., Zhang, L., Methner, A. (2024). Iron-sulfur cluster loss in mitochondrial CISD1 mediates PINK1 loss-of-function phenotypes. Elife, 13 PubMed ID: 39159312
Summary:
Parkinson's disease (PD) is characterized by the progressive loss of dopaminergic neurons in the substantia nigra of the midbrain. Familial cases of PD are often caused by mutations of PTEN-induced kinase 1 (PINK1) and the ubiquitin ligase Parkin, both pivotal in maintaining mitochondrial quality control. CISD1, a homodimeric mitochondrial iron-sulfur-binding protein, is a major target of Parkin-mediated ubiquitination. This study discovered a heightened propensity of CISD1 to form dimers in Pink1 mutant flies and in dopaminergic neurons from PINK1 mutation patients. The dimer consists of two monomers that are covalently linked by a disulfide bridge. In this conformation CISD1 cannot coordinate the iron-sulfur cofactor. Overexpressing Cisd, the Drosophila orthologue of CISD1, and a mutant Cisd incapable of binding the iron-sulfur cluster in Drosophila reduced climbing ability and lifespan. This was more pronounced with mutant Cisd and aggravated in Pink1 mutant flies. Complete loss of Cisd, in contrast, rescued all detrimental effects of Pink1 mutation on climbing ability, wing posture, dopamine levels, lifespan, and mitochondrial ultrastructure. These results suggest that Cisd, probably iron-depleted Cisd, operates downstream of Pink1 shedding light on PD pathophysiology and implicating CISD1 as a potential therapeutic target.
Nitta, Y., Osaka, J., Maki, R., Hakeda-Suzuki, S., Suzuki, E., Ueki, S., Suzuki, T., Sugie, A. (2024). Drosophila model to clarify the pathological significance of OPA1 in autosomal dominant optic atrophy. Elife, 12 PubMed ID: 39177028
Summary:
Autosomal dominant optic atrophy (DOA) is a progressive form of blindness caused by degeneration of retinal ganglion cells and their axons, mainly caused by mutations in the OPA1 mitochondrial dynamin like GTPase (OPA1) gene. OPA1 encodes a dynamin-like GTPase present in the mitochondrial inner membrane. When associated with OPA1 mutations, DOA can present not only ocular symptoms but also multi-organ symptoms (DOA plus). DOA plus often results from point mutations in the GTPase domain, which are assumed to have dominant-negative effects. However, the presence of mutations in the GTPase domain does not always result in DOA plus. Therefore, an experimental system to distinguish between DOA and DOA plus is needed. This study found that loss-of-function mutations of the dOPA1 gene in Drosophila can imitate the pathology of optic nerve degeneration observed in DOA. This degeneration was successfully rescued by expressing the human OPA1 (hOPA1) gene, indicating that hOPA1 is functionally interchangeable with dOPA1 in the fly system. However, mutations previously identified did not ameliorate the dOPA1 deficiency phenotype. By expressing both WT and DOA plus mutant hOPA1 forms in the optic nerve of dOPA1 mutants, DOA plus mutations suppressed the rescue, facilitating the distinction between loss-of-function and dominant-negative mutations in hOPA1. This fly model aids in distinguishing DOA from DOA plus and guides initial hOPA1 mutation treatment strategies. 		Layalle, S., Aimond, F., Brugioti, V., Guissart, C., Raoul, C., Soustelle, L. (2024). The ALS-associated KIF5A P986L variant is not pathogenic for Drosophila motoneurons. Sci Rep, 14(1):19540 PubMed ID: 39174694
Summary:
Amyotrophic lateral sclerosis (ALS) is a devastating paralytic disorder caused by the death of motoneurons. Several mutations in the KIF5A gene have been identified in patients with ALS. Some mutations affect the splicing sites of exon 27 leading to its deletion (Δ27 mutation). KIF5A Δ27 is aggregation-prone and pathogenic for motoneurons due to a toxic gain of function. Another mutation found to be enriched in ALS patients is a proline/leucine substitution at position 986 (P986L mutation). Bioinformatic analyses strongly suggest that this variant is benign. This study aimed to conduct functional studies in Drosophila to classify the KIF5A P986L variant. When expressed in motoneurons, KIF5A P986L does not modify the morphology of larval NMJ or the synaptic transmission. In addition, KIF5A P986L is uniformly distributed in axons and does not disturb mitochondria distribution. Locomotion at larval and adult stages is not affected by KIF5A P986L. Finally, both KIF5A WT and P986L expression in adult motoneurons extend median lifespan compared to control flies. Altogether, these data show that the KIF5A P986L variant is not pathogenic for motoneurons and may represent a hypomorphic allele, although it is not causative for ALS.
Oka, M., Nakajima, S., Suzuki, E., Yamamoto, S., Ando, K. (2024). Glucose uptake in pigment glia suppresses tau-induced inflammation and photoreceptor degeneration in Drosophila. bioRxiv, PubMed ID: 39229232
Summary:
Brain inflammation contributes to the pathogenesis of neurodegenerative diseases such as Alzheimer's disease (AD). Glucose hypometabolism and glial activation are pathological features seen in AD brains; however, the connection between the two is not fully understood. Using a Drosophila model of AD, this study identified that glucose metabolism in glia plays a critical role in neuroinflammation under disease conditions. Expression of human tau in the retinal cells, including photoreceptor neurons and pigment glia, causes photoreceptor degeneration accompanied by inclusion formation and swelling of lamina glial cells. We found that inclusions are formed by glial phagocytosis, and swelling of the laminal cortex correlates with the expression of antimicrobial peptides (AMPs). Co-expression of human glucose transporter 3 (GLUT3) with tau in the retina does not affect tau levels but suppresses these inflammatory responses and photoreceptor degeneration. This study also found that expression of GLUT3, specifically in the pigment glia, is sufficient to suppress inflammatory phenotypes and mitigate photoreceptor degeneration in the /tau-expressing retina. These results suggest that glial glucose metabolism contributes to inflammatory responses and neurodegeneration in tauopathy. 		Chechenova, M., McLendon, L., Dallas, B., Stratton, H., Kiani, K., Gerberich, E., Alekseyenko, A., Tamba, N., An, S., Castillo, L., Czajkowski, E., Talley, C., Brown, A., Bryantsev, A. L. (2024). Muscle degeneration in aging Drosophila flies: the role of mechanical stress. Skeletal muscle, 14(1):20 PubMed ID: 39164781
Summary:
Muscle wasting is a universal hallmark of aging which is displayed by a wide range of organisms, although the causes and mechanisms of this phenomenon are not fully understood. This study used Drosophila to characterize the phenomenon of spontaneous muscle fiber degeneration (SMFD) during aging. SMFD occurs across diverse types of somatic muscles, progresses with chronological age, and positively correlates with functional muscle decline. Data from vital dyes and morphological markers imply that degenerative fibers most likely die by necrosis. Mechanistically, SMFD is driven by the damage resulting from muscle contractions, and the nervous system may play a significant role in this process. This quantitative model of SMFD assessment can be useful in identifying and validating novel genetic factors that influence aging-related muscle wasting.
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Wednesday July 2nd - Larval and Adult Neural Development, Structure and Function

Prelic, S., Keesey, I. W., Lavista-Llanos, S., Hansson, B. S., Wicher, D. (2024). Innexin expression and localization in the Drosophila antenna indicate gap junction or hemichannel involvement in antennal chemosensory sensilla. Cell and tissue research, PubMed ID: 39174822
Summary:
Odor detection in insects is largely mediated by structures on antennae called sensilla, which feature a strongly conserved architecture and repertoire of olfactory sensory neurons (OSNs) and various support cell types. In Drosophila, OSNs are tightly apposed to supporting cells, whose connection with neurons and functional roles in odor detection remain unclear. Coupling mechanisms between these neuronal and non-neuronal cell types have been suggested based on morphological observations, concomitant physiological activity during odor stimulation, and known interactions that occur in other chemosensory systems. For instance, it is not known whether cell-cell coupling via gap junctions between OSNs and neighboring cells exists, or whether hemichannels interconnect cellular and extracellular sensillum compartments. This study showed that innexins, which form hemichannels and gap junctions in invertebrates, are abundantly expressed in adult drosophilid antennae. By surveying antennal transcriptomes and performing various immunohistochemical stainings in antennal tissues, innexin-specific patterns of expression and localization were discovered, with a majority of innexins strongly localizing to glial and non-neuronal cells, likely support and epithelial cells. Finally, by injecting gap junction-permeable dye into a pre-identified sensillum, no dye coupling was discovered between neuronal and non-neuronal cells. Together with evidence of non-neuronal innexin localization, it is concluded that innexins likely do not conjoin neurons to support cells, but that junctions and hemichannels may instead couple support cells among each other or to their shared sensillum lymph to achieve synchronous activity. How coupling of sensillum microenvironments or compartments may potentially contribute to facilitate chemosensory functions of odor sensing and sensillum homeostasis is discussed. 		Rohrbach, E. W., Asuncion, J. D., Meera, P., Kralovec, M., Deshpande, S. A., Schweizer, F. E., Krantz, D. E. (2024). Heterogeneity in the projections and excitability of tyraminergic/octopaminergic neurons that innervate the Drosophila reproductive tract. Frontiers in molecular neuroscience, 17:1374896 PubMed ID: 39156129
Summary:
Aminergic nuclei in mammals are generally composed of relatively small numbers of cells with broad projection patterns. Despite the gross similarity of many individual neurons, recent transcriptomic, anatomic and behavioral studies suggest previously unsuspected diversity. Smaller clusters of aminergic neurons in the model organism Drosophila melanogaster provide an opportunity to explore the ramifications of neuronal diversity at the level of individual cells. A group of approximately 10 tyraminergic/octopaminergic neurons innervates the female reproductive tract in flies and has been proposed to regulate multiple activities required for fertility. The projection patterns of individual neurons within the cluster are not known and it remains unclear whether they are functionally heterogenous. Using a single cell labeling technique, this study showed that each region of the reproductive tract is innervated by a distinct subset of tyraminergic/octopaminergic cells. Optogenetic activation of one subset stimulates oviduct contractions, indicating that the cluster as a whole is not required for this activity, and underscoring the potential for functional diversity across individual cells. Using whole cell patch clamp, two adjacent and morphologically similar cells were shown to be tonically inhibited, but each responds differently to injection of current or activation of the inhibitory GluCl receptor. GluCl appears to be expressed at relatively low levels in tyraminergic/octopaminergic neurons within the cluster, suggesting that it may regulate their excitability via indirect pathways. Together, these data indicate that specific tyraminergic/octopaminergic cells within a relatively homogenous cluster have heterogenous properties and provide a platform for further studies to determine the function of each cell.
Peng, L., Wang, T. (2024). Histamine synthesis and transport are coupled in axon terminals via a dual quality control system. The EMBO journal, PubMed ID: 39242788
Summary:
Monoamine neurotransmitters generated by de novo synthesis are rapidly transported and stored into synaptic vesicles at axon terminals. This transport is essential both for sustaining synaptic transmission and for limiting the toxic effects of monoamines. In this study, synthesis of the monoamine histamine by histidine decarboxylase (HDC) and subsequent loading of histamine into synaptic vesicles are shown to be physically and functionally coupled within Drosophila photoreceptor terminals. This process requires HDC anchoring to synaptic vesicles via interactions with N-ethylmaleimide-sensitive fusion protein 1 (NSF1). Disassociating HDC from synaptic vesicles disrupts visual synaptic transmission and causes somatic accumulation of histamine, which leads to retinal degeneration. We further identified a proteasome degradation system mediated by the E3 ubiquitin ligase, purity of essence (POE), which clears mislocalized HDC from the soma, thus eliminating the cytotoxic effects of histamine. Taken together, these results reveal a dual mechanism for translocation and degradation of HDC that ensures restriction of histamine synthesis to axonal terminals and at the same time rapid loading into synaptic vesicles. This is crucial for sustaining neurotransmission and protecting against cytotoxic monoamines. 		Sung, H. H., Li, H., Huang, Y. C., Ai, C. L., Hsieh, M. Y., Jan, H. M., Peng, Y. J., Lin, H. Y., Yeh, C. H., Lin, S. Y., Yeh, C. Y., Cheng, Y. J., Khoo, K. H., Lin, C. H., Chien, C. T. (2024). Galectins induced from hemocytes bridge phosphatidylserine and N-glycosylated Drpr/CED-1 receptor during dendrite pruning. Nat Commun, 15(1):7402 PubMed ID: 39191750
Summary:
During neuronal pruning, phagocytes engulf shed cellular debris to avoid inflammation and maintain tissue homeostasis. How phagocytic receptors recognize degenerating neurites had been unclear. This study identified two glucosyltransferases Alg8 and Alg10 of the N-glycosylation pathway required for dendrite fragmentation and clearance through genetic screen. The scavenger receptor Draper (Drpr) is N-glycosylated with complex- or hybrid-type N-glycans that interact specifically with galectins. We also identify the galectins Crouching tiger (Ctg) and Hidden dragon (Hdg) that interact with N-glycosylated Drpr and function in dendrite pruning via the Drpr pathway. Ctg and Hdg are required in hemocytes for expression and function, and are induced during dendrite injury to localize to injured dendrites through specific interaction with exposed phosphatidylserine (PS) on the surface membrane of injured dendrites. Thus, the galectins Ctg and Hdg bridge the interaction between PS and N-glycosylated Drpr, leading to the activation of phagocytosis.
Warren, B., Gopfert, M. C. (2024). Mechanically-evoked spike responses of pentascolopidial chordotonal organs of Drosophila larvae. The Journal of experimental biology, PubMed ID: 39206682
Summary:
Mechano-sensitive ensembles of neurons in insects, known as Chordotonal organs (COs), function in proprioception, the detection of sound and substrate vibrations. This study characterized the mechanical sensitivity of the lateral pentascolopidial CO (lch5) of Drosophila larvae to establish its postulated role in proprioception. A physiologically realistic method was developed to replicate proprioceptive input to lch5 by pulling the apodeme (tendon) to which the tips of the neurons attach. Lch5 sensory neurons were found to respond transiently with a short latency to the velocity-component of stretch displacements and the release of stretch (relaxation). In the mechanosensory mutant inactive, lch5 has a decreased response to mechanical stimuli and a lower overall spontaneous spike rate. Finally, the input that lch5 receives was stimulated during crawling; spike rate was found to change of peristaltic body contraction. This study provides a characterization of proprioceptive feedback in Drosophila larvae and firmly establish the proprioceptive function of lch5 in larval locomotion. 		Delescluse, J., Simonnet, M. M., Ziegler, A. B., Piffaretti, K., Alves, G., Grosjean, Y., Maniere, G. (2024). A LAT1-Like Amino Acid Transporter Regulates Neuronal Activity in the Drosophila Mushroom Bodies. Cells, 13(16) PubMed ID: 39195231
Summary:
The proper functioning of neural circuits that integrate sensory signals is essential for individual adaptation to an ever-changing environment. Many molecules can modulate neuronal activity, including neurotransmitters, receptors, and even amino acids. This study asked whether amino acid transporters expressed by neurons can influence neuronal activity. minidiscs (mnd), which encodes a light chain of a heterodimeric amino acid transporter, was found to be expressed in different cell types of the adult Drosophila brain: in mushroom body neurons (MBs) and in glial cells. Using live calcium imaging, MND was found to be expressed in α/β MB neurons and is essential for sensitivity to the L-amino acids: Leu, Ile, Asp, Glu, Lys, Thr, and Arg. We found that the Target Of Rapamycin (TOR) pathway but not the Glutamate Dehydrogenase (GDH) pathway is involved in the Leucine-dependent response of α/β MB neurons. This study strongly supports the key role of MND in regulating MB activity in response to amino acids.
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Friday June 27th - RNAs and Transposons

Adashev, V. E., Kotov, A. A., Bazylev, S. S., Kombarov, I. A., Olenkina, O. M., Shatskikh, A. S., Olenina, L. V. (2024). Essential functions of RNA helicase Vasa in maintaining germline stem cells and piRNA-guided Stellate silencing in Drosophila spermatogenesis. Frontiers in cell and developmental biology, 12:1450227 PubMed ID: 39184915
Summary:
DEAD-box RNA helicase Vasa is required for gonad development and fertility in multiple animals. Data about Vasa functions in Drosophila spermatogenesis remain controversial. This study showed that loss-of-function vasa mutations led to failures of GSC maintenance in the testes, a severe loss of total germ cell content, and a cessation of male fertility over time. Defects in GSC maintenance in vasa mutant testes were not associated with an increasing frequency of programmed cell death, indicating that a premature loss of GSCs occurred via entering differentiation. Vasa is implicated in the positive regulation of rhino expression both in the testes and ovaries. The introduction of a transgene copy of rhino, encoding a nuclear component of piRNA pathway machinery, in vasa mutant background allowed us to restore premeiotic stages of spermatogenesis, including the maintenance of GSCs and the development of spermatogonia and spermatocytes. However, piRNA-guided repression of Stellate genes in spermatocytes of vasa mutant testes with additional rhino copy was not restored, and male fertility was disrupted. This study uncovered a novel mechanistic link involving Vasa and Rhino in a regulatory network that mediates GSC maintenance but is dispensable for the perfect biogenesis of Su(Ste) piRNAs in testes. Thus, this study has shown that Vasa functions in spermatogenesis are essential at two distinct developmental stages: in GSCs for their maintenance and in spermatocytes for piRNA-mediated silencing of Stellate genes. 		Tang, T., Pei, M., Xiao, Y., Deng, Y., Lu, Y., Yu, X. Q., Wen, L., Hu, Q. (2024). Functional Analysis of Forkhead Transcription Factor Fd59a in the Spermatogenesis of Drosophila melanogaster. Insects, 15(7) PubMed ID: 39057213
Summary:
Spermatogenesis is critical for insect reproduction and is regulated by many different genes. This study found that Forkhead transcription factor Fd59a functions as a key factor in the spermatogenesis of Drosophila melanogaster. Fd59a contains a conversed Forkhead domain, and it is clustered to the FoxD subfamily with other FoxD members from some insect and vertebrate species. Mutations in Fd59a caused swelling in the apical region of the testis. More importantly, fewer mature sperm were present in the seminal vesicle of Fd59a mutant flies compared to the control flies, and the fertility of Fd59a2/2 mutant males was significantly lower than that of the control flies. Immunofluorescence staining showed that the homeostasis of the testis stem cell niche in Fd59a2/2) mutant and Fd59a RNAi flies was disrupted and the apoptosis of sperm bundles was increased. Furthermore, results from RNA sequencing and qRT-PCR suggested that Fd59a can regulate the expression of genes related to reproductive process and cell death. Taken together, these results indicated that Fd59a plays a key role in the spermatogenesis of Drosophila.
Jansen, G., Gebert, D., Kumar, T. R., Simmons, E., Murphy, S., Teixeira, F. K. (2024). Tolerance thresholds underlie responses to DNA damage during germline development. Genes Dev, 38(13-14):631-654 PubMed ID: 39054057
Summary:
Selfish DNA modules like transposable elements (TEs) are particularly active in the germline, the lineage that passes genetic information across generations. New TE insertions can disrupt genes and impair the functionality and viability of germ cells. However, this study found that in P-M hybrid dysgenesis in Drosophila, a sterility syndrome triggered by the P-element DNA transposon, germ cells harbor unexpectedly few new TE insertions despite accumulating DNA double-strand breaks (DSBs) and inducing cell cycle arrest. Using an engineered CRISPR-Cas9 system, this study showed that generating DSBs at silenced P-elements or other noncoding sequences is sufficient to induce germ cell loss independently of gene disruption. Indeed, it was demonstrated that both developing and adult mitotic germ cells are sensitive to DSBs in a dosage-dependent manner. Following the mitotic-to-meiotic transition, however, germ cells become more tolerant to DSBs, completing oogenesis regardless of the accumulated genome damage. These findings establish DNA damage tolerance thresholds as crucial safeguards of genome integrity during germline development. 		Shaikh, U., Sherlock, K., Wilson, J., Gilliland, W., Lewellyn, L. (2024). Lineage-based scaling of germline intercellular bridges during oogenesis. Development, 151(16) PubMed ID: 39190553
Summary:
The size of subcellular structures must be tightly controlled to maintain normal cell function. Despite its importance, few studies have determined how the size of organelles or other structures is maintained during development, when cells are growing, dividing and rearranging. The developing Drosophila egg chamber is a powerful model in which to study the relative growth rates of subcellular structures. The egg chamber contains a cluster of 16 germline cells, which are connected through intercellular bridges called ring canals. As the egg chamber grows, the germline cells and the ring canals that connect them increase in size. This study demonstrates that ring canal size scaling is related to lineage; the largest, 'first-born' ring canals increase in size at a relatively slower rate than ring canals derived from subsequent mitotic divisions. This lineage-based scaling relationship is maintained even if directed transport is reduced, ring canal size is altered, or in egg chambers with twice as many germline cells. Analysis of lines that produce larger or smaller mature eggs reveals that different strategies could be used to alter final egg size.
Netherton, J. K., Ogle, R. A., Robinson, B. R., Molloy, M., Krisp, C., Velkov, T., Casagranda, F., Dominado, N., Silva Balbin Villaverde, A. I., Zhang, X. D., Hime, G. R., Baker, M. A. (2024). The role of HnrnpF/H as a driver of oligoteratozoospermia. iScience, 27(7):110198 PubMed ID: 39092172
Summary:
Male subfertility or infertility is a common condition often characterized by men producing a low number of sperm with poor quality. To gain insight into this condition, a quantitative proteomic analysis was performed of semen samples obtained from infertile and fertile men. At least 6 proteins showed significant differences in regulation of alternatively spliced isoforms. To investigate this link between aberrant alternative splicing and production of poor-quality spermatozoa, TShe hnrnpH/F-orthologue Glorund (Glo) was overexpressed in Drosophila; overexpression of the human orthollog was also found to be abundant in poor quality human sperm. Transgenic animals produced low numbers of morphologically defective spermatozoa and aberrant formation of the "dense body," an organelle akin to the mammalian manchette. Furthermore, fertility trials demonstrated that transgenic flies were either completely infertile or highly subfertile. These findings suggest that dysregulation of hnrnpH/F is likely to result in the production of low-quality semen, leading to subfertility or infertility in men. 		Matzkin, L. M., Bono, J. M., Pigage, H. K., Allan, C. W., Diaz, F., McCoy, J. R., Green, C. C., Callan, J. B., Delahunt, S. P., (2024). Females translate male mRNA transferred during mating. iScience, 27(8):110442 PubMed ID: 39108707
Summary:
Although RNA is found in the seminal fluid of diverse organisms, it is unknown whether it is functional within females. This study developed a proteomic method (VESPA, Variant Enabled SILAC Proteomic Analysis) to test the hypothesis that Drosophila male seminal fluid RNA is translated by females. 67 male-derived, female-translated proteins (mdFTPs) were found in female lower reproductive tracts, many with predicted functions relevant to reproduction. Knockout experiments indicate that mdFTPs play diverse roles in postmating interactions, affecting fertilization success, and the formation/persistence of the insemination reaction mass, a trait hypothesized to be involved in sexual conflict. These findings advance understanding of reproduction by revealing a mechanism of postmating molecular interactions between the sexes that strengthens and extends male influences on reproduction in previously unrecognized ways. Given the diverse species that carry RNA in seminal fluid, this discovery has broad significance for understanding molecular mechanisms of cooperation and conflict during reproduction.
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Friday June 27th - RNAs and Transposons

Kaul, N., Pradhan, S. J., Boin, N. G., Mason, M. M., Rosales, J., Starke, E. L., Wilkinson, E. C., Chapman, E. G., Barbee, S. A. (2024). FMRP cooperates with miRISC components to repress translation and regulate neurite morphogenesis in Drosophila. RNA Biol, 21(1):11-22 PubMed ID: 39190491
Summary:
Fragile X Syndrome (FXS) is the most common inherited form of intellectual disability and is caused by mutations in the gene encoding the Fragile X messenger ribonucleoprotein (FMRP). FMRP is an evolutionarily conserved and neuronally enriched RNA-binding protein (RBP) with functions in RNA editing, RNA transport, and protein translation. Specific target RNAs play critical roles in neurodevelopment, including the regulation of neurite morphogenesis, synaptic plasticity, and cognitive function. The different biological functions of FMRP are modulated by its cooperative interaction with distinct sets of neuronal RNA and protein-binding partners. This study focused on interactions between FMRP and components of the microRNA (miRNA) pathway. Using the Drosophila S2 cell model system, this study showed that the Drosophila ortholog of FMRP (dFMRP) can repress translation when directly tethered to a reporter mRNA. This repression requires the activity of AGO1, GW182, and MOV10/Armitage, conserved proteins associated with the miRNA-containing RNA-induced silencing complex (miRISC). Additionally, untagged dFMRP can interact with a short stem-loop sequence in the translational reporter, a prerequisite for repression by exogenous miR-958. Finally, dFmr1 interacts genetically with GW182 to control neurite morphogenesis. These data suggest that dFMRP may recruit the miRISC to nearby miRNA binding sites and repress translation via its cooperative interactions with evolutionarily conserved components of the miRNA pathway. 		He, Q., Chen, S., Hou, T., Chen, J. (2024). Juvenile hormone-induced microRNA miR-iab-8 regulates lipid homeostasis and metamorphosis in Drosophila melanogaster. Insect Mol Biol, PubMed ID: 39005109
Summary:
Metamorphosis plays an important role in the evolutionary success of insects. Accumulating evidence indicated that microRNAs (miRNAs) are involved in the regulation of processes associated with insect metamorphosis. However, the miRNAs coordinated with juvenile hormone (JH)-regulated metamorphosis remain poorly reported. In the present study, using high-throughput miRNA sequencing combined with Drosophila genetic approaches, it was demonstrated that miR-iab-8, which primarily targets homeotic genes to modulate haltere-wing transformation and sterility was up-regulated by JH and involved in JH-mediated metamorphosis. Overexpression of miR-iab-8 in the fat body resulted in delayed development and failure of larval-pupal transition. Furthermore, metabolomic analysis results revealed that overexpression of miR-iab-8 caused severe energy metabolism defects especially the lipid metabolism, resulting in significantly reduced triacylglycerol (TG) content and glycerophospholipids but enhanced accumulation of phosphatidylcholine (PC) and phosphatidylethanolamine (PE). In line with this, Nile red staining demonstrated that during the third larval development, the TG content in the miR-iab-8 overexpression larvae was continuously decreased, which is opposite to the control. Additionally, the transcription levels of genes committed to TG synthesis and breakdown were found to be significantly increased and the expression of genes responsible for glycerophospholipids metabolism were also altered. Overall, it is proposed that JH induced miR-iab-8 expression to perturb the lipid metabolism homeostasis especially the TG storage in the fat body, which in turn affected larval growth and metamorphosis.
Zhang, Q., Li, L., Zhang, Q., Zhang, Y., Yan, L., Wang, Y., Wang, Y., Zhao, S. (2024). Genetic circuitry controlling Drosophila female germline overgrowth. Dev Biol, 515:160-168 PubMed ID: 39067502
Summary:
Germ cells mutant for bam or bgcn are locked in a germline stem cell (GSC)-like state, leading to tumor-like overgrowth in Drosophila ovaries. Previous studies have demonstrated that germline overgrowth in bam mutants can be suppressed by defects in the miRNA pathway but enhanced by a null mutation in hippo. However, the genetic epistasis between the miRNA and Hippo pathways still remains unknown. Here, we determined that the miRNA pathway acts downstream of the Hippo pathway in regulating this process. Germ cells mutant for bam or bgcn and defective in both pathways divide very slowly, phenocopying those defective only in the miRNA pathway. In addition, we found that Yki, a key oncoprotein in the Hippo pathway, promotes the growth of both wild-type germ cells and bam mutant GSC-like cells. Like wild-type GSCs, bam mutant GSC-like cells predominantly stay in the G2 phase. Remarkably, many of those defective in the miRNA pathway are arrested before entering this phase. Furthermore, these studies identified bantam as a critical miRNA promoting germline overgrowth in bam or bgcn mutants. Taken together, these findings establish a genetic circuitry controlling Drosophila female germline overgrowth. 		Li, Z., Iida, J., Shiimori, M., Okamura, K. (2024). Exportin-5 binding precedes 5'- and 3'-end processing of tRNA precursors in Drosophila. J Biol Chem, 300(9):107632 PubMed ID: 39098529
Summary:
Exportin5 (Exp5) is the major miRNA nuclear export factor and recognizes structural features of pre-miRNA hairpins, while it also exports other minihelix-containing RNAs. In Drosophila, Exp5 is suggested to play a major role in tRNA export because the gene encoding the canonical tRNA export factor Exportin-t is missing in its genome. To understand molecular functions of fly Exp5, the Exp5/RNA interactome was studied in the cell line S2R + using the crosslinking and immunoprecipitation (CLIP) technology. The CLIP experiment captured known substrates such as tRNAs and miRNAs and detected candidates of novel Exp5 substrates including various mRNAs and long non-coding RNAs (lncRNAs). Some mRNAs and lncRNAs enriched PAR-CLIP tags compared to their expression levels, suggesting selective binding of Exp5 to them. Intronless mRNAs tended to enrich PAR-CLIP tags; therefore, it was proposed that Exp5 might play a role in the export of specific classes of mRNAs/lncRNAs. This result suggested that Drosophila Exp5 might have a wider variety of substrates than initially thought. Surprisingly, Exp5 CLIP reads often contained sequences corresponding to the flanking 5'-leaders and 3'-trailers of tRNAs, which were thought to be removed prior to nuclear export. In fact, pre-tRNAs before end-processing were present in the cytoplasm, supporting the idea that tRNA end-processing is a cytoplasmic event. In summary, theseresults provide a genome-wide list of Exp5 substrate candidates and suggest that flies may lack a mechanism to distinguish pre-tRNAs with or without the flanking sequences.
Mostoufi, S. L., Singh, N. D. (2024)s. Pathogen infection alters the gene expression landscape of transposable elements in Drosophila melanogaster. G3 (Bethesda), 14(9) PubMed ID: 39129654
Summary:
Transposable elements make up substantial proportions of eukaryotic genomes, and many are thought to be remnants of ancient viral infections. Current research has begun to highlight the role transposable elements can play in the immune system response to infections. However, most of our knowledge about transposable element expression during infection is limited by the specific host and pathogen factors from each study, making it difficult to compare studies and develop broader patterns regarding the role of transposable elements during infection. This study used the tools and resources available in the model, Drosophila melanogaster, to analyze multiple gene expression datasets of flies subject to bacterial, fungal, and viral infections. Differences in pathogen species, host genotype, host tissue, and sex were analyzed to understand how these factors impact transposable element expression during infection. The results highlight both shared and unique transposable element expression patterns between pathogens and suggest a larger effect of pathogen factors over host factors for influencing transposable element expression. 		Chen, R., Stainier, W., Dufourt, J., Lagha, M., Lehmann, R. (2024). Direct observation of translational activation by a ribonucleoprotein granule. Nat Cell Biol, 26(8):1322-1335 PubMed ID: 38965420
Summary:
Biomolecular condensates organize biochemical processes at the subcellular level and can provide spatiotemporal regulation within a cell. Among these, ribonucleoprotein (RNP) granules are storage hubs for translationally repressed mRNA. Whether RNP granules can also activate translation and how this could be achieved remains unclear. Using single-molecule imaging, this study demonstrated that the germ cell-determining RNP granules in Drosophila embryos are sites for active translation of nanos mRNA. Nanos translation occurs preferentially at the germ granule surface with the 3' UTR buried within the granule. Smaug, a cytosolic RNA-binding protein, represses nanos translation, which is relieved when Smaug is sequestered to the germ granule by the scaffold protein Oskar. Together, these findings uncover a molecular process by which RNP granules achieve localized protein synthesis through the compartmentalized loss of translational repression.
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Thursday June 26th - Larval and Adult Neural Development, Structure and Function

Zhu, J., Boivin, J. C., Garner, A., Ning, J., Zhao, Y. Q., Ohyama, T. (2024). Feedback inhibition by a descending GABAergic neuron regulates timing of escape behavior in Drosophila larvae. Elife, 13 PubMed ID: 39196635
Summary:
Escape behaviors help animals avoid harm from predators and other threats in the environment. Successful escape relies on integrating information from multiple stimulus modalities (of external or internal origin) to compute trajectories toward safe locations, choose between actions that satisfy competing motivations, and execute other strategies that ensure survival. To this end, escape behaviors must be adaptive. When a Drosophila melanogaster larva encounters a noxious stimulus, such as the focal pressure a parasitic wasp applies to the larval cuticle via its ovipositor, it initiates a characteristic escape response. The escape sequence consists of an initial abrupt bending, lateral rolling, and finally rapid crawling. Previous work has shown that the detection of noxious stimuli primarily relies on class IV multi-dendritic arborization neurons (Class IV neurons) located beneath the body wall, and more recent studies have identified several important components in the nociceptive neural circuitry involved in rolling. However, the neural mechanisms that underlie the rolling-escape sequence remain unclear. This study presents both functional and anatomical evidence suggesting that bilateral descending neurons within the subesophageal zone of D. melanogaster larva play a crucial role in regulating the termination of rolling and subsequent transition to escape crawling. These descending neurons (designated SeIN128) are inhibitory and receive inputs from a second-order interneuron upstream (Basin-2) and an ascending neuron downstream of Basin-2. Together with optogenetic experiments showing that co-activation of SeIN128 neurons and Basin-2 influence the temporal dynamics of rolling, these findings collectively suggest that the ensemble of SeIN128, Basin-2, and A00c neurons forms a GABAergic feedback loop onto Basin-2, which inhibits rolling and thereby facilitates the shift to escape crawling. 		Chu, L. A., Tai, C. Y., Chiang, A. S. (2024). Thirst-driven hygrosensory suppression promotes water seeking in Drosophila. Proc Natl Acad Sci U S A, 121(34):e2404454121 PubMed ID: 39145936
Summary:
Survival in animals relies on navigating environments aligned with physiological needs. In Drosophila melanogaster, antennal ionotropic receptors (IRs) sensing humidity changes govern hygrotaxis behavior. This study sheds light on the crucial role of R8a neurons in the transition from high humidity avoidance to water-seeking behavior when the flies become thirsty. These neurons demonstrate a heightened calcium response toward high humidity stimuli in satiated flies and a reduced response in thirsty flies, modulated by fluctuating levels of the neuropeptide leucokinin, which monitors the internal water balance. Optogenetic activation of IR8a neurons in thirsty flies triggers an avoidance response similar to the moisture aversion in adequately hydrated flies. Furthermore, this study identifies IR40a neurons as associated with dry avoidance, while IR68a neurons are linked to moist attraction. The dynamic interplay among these neurons, each with opposing valences, establishes a preference for approximately 30% relative humidity in well-hydrated flies and facilitates water-seeking behavior in thirsty individuals. This research unveils the intricate interplay between sensory perception, neuronal plasticity, and internal states, providing valuable insights into the adaptive mechanisms governing hygrotaxis in Drosophila.
Zhao, Q., Li, X., Wen, J., He, Y., Zheng, N., Li, W., Cardona, A., Gong, Z. (2024). A two-layer neural circuit controls fast forward locomotion in Drosophila. Curr Biol, 34(15):3439-3453 PubMed ID: 39053465
Summary:
Fast forward locomotion is critical for animal hunting and escaping behaviors. However, how the underlying neural circuit is wired at synaptic resolution to decide locomotion direction and speed remains poorly understood. This study identified in the ventral nerve cord (VNC) a set of ascending cholinergic neurons (AcNs) to be command neurons capable of initiating fast forward peristaltic locomotion in Drosophila larvae. Targeted manipulations revealed that AcNs are necessary and sufficient for fast forward locomotion. AcNs can activate their postsynaptic partners, A01j and A02j; both are interneurons with locomotory rhythmicity. Activated A01j neurons form a posterior-anteriorly descendent gradient in output activity along the VNC to launch forward locomotion from the tail. Activated A02j neurons exhibit quicker intersegmental transmission in activity that enables fast propagation of motor waves. This work revealed a global neural mechanism that coordinately controls the launch direction and propagation speed of Drosophila locomotion, furthering the understanding of the strategy for locomotion control. 		Lehman, M., Barre, C., Hasan, M. A., Flament, B., Autran, S., Dhiman, N., Soba, P., Masson, J. B., Jovanic, T. (2025). Neural circuits underlying context-dependent competition between defensive actions in Drosophila larvae. Nat Commun, 16(1):1120 PubMed ID: 39875414
Summary:
To ensure their survival, animals must be able to respond adaptively to threats within their environment. However, the precise neural circuit mechanisms that underlie flexible defensive behaviors remain poorly understood. Using neuronal manipulations, machine learning-based behavioral detection, electron microscopy (EM) connectomics and calcium imaging in Drosophila larvae, this study mapped second-order interneurons that are differentially involved in the competition between defensive actions in response to competing aversive cues. Mechanosensory stimulation inhibits escape behaviors in favor of startle behaviors by influencing the activity of escape-promoting second-order interneurons. Stronger activation of those neurons inhibits startle-like behaviors. This suggests that competition between startle and escape behaviors occurs at the level of second-order interneurons. Finally, a pair of descending neurons was identified that promote startle behaviors and could modulate the escape sequence. Taken together, these results characterize the pathways involved in startle and escape competition, which is modulated by the sensory context.
Shiozaki, H. M., Wang, K., Lillvis, J. L., Xu, M., Dickson, B. J., Stern, D. L. (2024). Activity of nested neural circuits drives different courtship songs in Drosophila. Nat Neurosci, PubMed ID: 39198658
Summary:
Motor systems implement diverse motor programs to pattern behavioral sequences, yet how different motor actions are controlled on a moment-by-moment basis remains unclear. This study investigated the neural circuitcourtship songs in Drosophila. Courting males rapidly alternate between two types of song: pulse and sine. By recording calcium signals in the ventral nerve cord in singing flies, one neural population was found to be active during both songs, whereas an expanded neural population, which includes neurons from the first population, is active during pulse song. Brain recordings showed that this nested activation pattern is present in two descending pathways required for singing. Connectomic analysis reveals that these two descending pathways provide structured input to ventral nerve cord neurons in a manner consistent with their activation patterns. These results suggest that nested premotor circuit activity, directed by distinct descending signals, enables rapid switching between motor actions. 		Prieto, D., Egger, B., Cantera, R. (2024). Atypical soluble guanylyl cyclases control brain size in Drosophila. . microPublication biology, 2024. PubMed ID: 39185012
Summary:
Hypoxia-induced proliferation of neural stem cells has a crucial role in brain development. In the brain of Drosophila melanogaster, the optic lobe exhibits progressive hypoxia during larval development. This study investigated an alternative oxygen-sensing mechanism within this brain compartment, distinct from the canonical hypoxia signaling pathway mediated by HIF. Using genetic tools, immunostaining, and confocal microscopy, the loss of the atypical soluble guanylyl cyclase (asGC) subunit Gyc88E, or the ectopic expression of Gyc89Db in neural stem cells was shown to lead to increased optic lobe volume. The existence of a link between cGMP signaling and neurogenesis in the developing brain is proposed.
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Wednesday June 25th - Adult Development

Zhang, X., Wang, Y., Zhao, W., Yang, S., Moussian, B., Zhao, Z., Zhang, J., Dong, W. (2024). Excess Dally-like Induces Malformation of Drosophila Legs. Cells, 13(14) PubMed ID: 39056781
Summary:
Glypicans are closely associated with organ development and tumorigenesis in animals. Dally-like (Dlp), a membrane-bound glypican, plays pivotal roles in various biological processes in Drosophila. This study observed that an excess of Dlp led to the malformation of legs, particularly affecting the distal part. Accordingly, the leg disc was shrunken and frequently exhibited aberrant morphology. In addition, elevated Dlp levels induced ectopic cell death with no apparent cell proliferation changes. Furthermore, Dlp overexpression in the posterior compartment significantly altered Wingless (Wg) distribution. A marked expansion of Wg distribution was observed within the posterior compartment, accompanied by a corresponding decrease in the anterior compartment. It appears that excess Dlp guides Wg to diffuse to cells with higher Dlp levels. In addition, the distal-less (dll) gene, which is crucial for leg patterning, was up-regulated significantly. Notably, dachshund (dac) and homothorax (hth) expression, also essential for leg patterning and development, only appeared to be negligibly affected. Based on these findings, it is speculated that excess Dlp may contribute to malformations of the distal leg region of Drosophila, possibly through its influence on Wg distribution, dll expression and induced cell death. Our research advances the understanding of Dlp function in Drosophila leg development. 		Walther, R. F., Lancaster, C., Burden, J. J., Pichaud, F. (2024). A dystroglycan-laminin-integrin axis coordinates cell shape remodeling in the developing Drosophila retina. PLoS Biol, 22(9):e3002783 PubMed ID: 39226305
Summary:
Cell shape remodeling is a principal driver of epithelial tissue morphogenesis. While progress continues to be made in our understanding of the pathways that control the apical (top) geometry of epithelial cells, comparatively little is known about those that control cell basal (bottom) geometry. To examine this, the Drosophila ommatidium, which is the basic visual unit of the compound eye, was used. The ommatidium is shaped as a hexagonal prism, and generating this 3D structure requires ommatidial cells to adopt specific apical and basal polygonal geometries. Using this model system generating cell type-specific basal geometries was found to start with patterning of the basal extracellular matrix, whereby Laminin<>/a> accumulates at discrete locations across the basal surface of the retina. The Dystroglycan receptor complex (DGC) is required for this patterning by promoting localized Laminin accumulation at the basal surface of cells. Moreover, the results reveal that localized accumulation of Laminin and the DGC are required for directing Integrin adhesion. This induces cell basal geometry remodeling by anchoring the basal surface of cells to the extracellular matrix at specific, Laminin-rich locations. It is proposed that patterning of a basal extracellular matrix by generating discrete Laminin domains can direct Integrin adhesion to induce cell shape remodeling in epithelial morphogenesis.
Teeters, G., Weasner, B. M., Ordway, A. J., Weasner, B. P., Kumar, J. P. (2024). Control of fate specification within the dorsal head of Drosophila melanogaster. Development, 151(16) PubMed ID: 39190554
Summary:
During development, unique combinations of transcription factors and signaling pathways carve the nascent eye-antennal disc of the fruit fly Drosophila melanogaster into several territories that will eventually develop into the compound eye, ocelli, head epidermis, bristles, antenna and maxillary palpus of the adult head. Juxtaposed patterns of Hedgehog (Hh) and Decapentaplegic (Dpp) initiate compound eye development, while reciprocal domains of Dpp and Wingless (Wg) induce formation of the antennal and maxillary palp fields. Hh and Wg signaling, but not Dpp, contribute to the patterning of the dorsal head vertex. This study shows that combinatorial reductions of the Pax6 transcription factor Twin of Eyeless and either the Wg pathway or the Mirror (Mirr)z transcription factor trigger a transformation of the ocelli into a compound eye and the neighboring head epidermis into an antenna. These changes in fate are accompanied by the ectopic expression of Dpp, which might be expected to trigger these changes in fate. However, the transformation of the field cannot be replicated by increasing Dpp levels alone despite the recreation of adjacent Hh-Dpp and Wg-Dpp domains. As such, the emergence of these ectopic organs occurs through a unique regulatory path. 		Ehrhardt, B., Angstmann, H., Hoschler, B., Kovacevic, D., Hammer, B., Roeder, T., Rabe, K. F., Wagner, C., Uliczka, K., Krauss-Etschmann, S. (2024). Airway specific deregulation of asthma-related serpins impairs tracheal architecture and oxygenation in D. melanogaster. Sci Rep, 14(1):16567 PubMed ID: 39019933
Summary:
Serine proteases are important regulators of airway epithelial homeostasis. Altered serum or cellular levels of two serpins, Scca1 and Spink5, have been described for airway diseases but their function beyond antiproteolytic activity is insufficiently understood. To close this gap, fly lines were generated with overexpression or knockdown for each gene in the airways. Overexpression of both fly homologues of Scca1 (Spn42DdCG32354) induced the growth of additional airway branches, with more variable results for the respective knockdowns. Dysregulation of Scca1 resulted in a general delay in fruit fly development, with increases in larval and pupal mortality following overexpression of this gene. In addition, the morphological changes in the airways were concomitant with lower tolerance to hypoxia. In conclusion, the observed structural changes of the airways evidently had a strong impact on the airway function in our model as they manifested in a lower physical fitness of the animals. We assume that this is due to insufficient tissue oxygenation. Future work will be directed at the identification of key molecular regulators following the airway-specific dysregulation of Scca1 and Spink5 expression.
Hiraiwa, S., Takeshita, S., Terano, T., Hayashi, R., Suzuki, K., Tajiri, R., Kojima, T. (2024). Unveiling the cell dynamics during the final shape formation of the tarsus in Drosophila adult leg by live imaging. Dev Genes Evol, PubMed ID: 38977431
Summary:
Organisms display a remarkable diversity in their shapes. Although substantial progress has been made in unraveling the mechanisms that govern cell fate determination during development, the mechanisms by which fate-determined cells give rise to the final shapes of organisms remain largely unknown. This study describes in detail the process of the final shape formation of the tarsus, which is near the distal tip of the adult leg, during the pupal stage in Drosophila melanogaster. Days-long live imaging revealed unexpectedly complicated cellular dynamics. The epithelial cells transiently form the intriguing structure, which was named the Parthenon-like structure. The basal surface of the epithelial cells and localization of the basement membrane protein initially show a mesh-like structure and rapidly shrink into the membranous structure during the formation and disappearance of the Parthenon-like structure. Furthermore, macrophage-like cells are observed moving around actively in the Parthenon-like structure and engulfing epithelial cells. The findings in this research are expected to significantly contribute to our understanding of the mechanisms involved in shaping the final structure of the adult tarsus. 		Fuhrmann, J. F., Krishna, A., Paijmans, J., Duclut, C., Cwikla, G., Eaton, S., Popovic, M., Julicher, F., Modes, C. D., Dye, N. A. (2024). Active shape programming drives Drosophila wing disc eversion. Sci Adv, 10(32):eadp0860 PubMed ID: 39121221
Summary:
How complex 3D tissue shape emerges during animal development remains an important open question in biology and biophysics. This study discovered a mechanism for 3D epithelial shape change based on active, in-plane cellular events that is analogous to inanimate "shape programmable" materials, which undergo blueprinted 3D shape transformations from in-plane gradients of spontaneous strains. Eversion of the Drosophila wing disc pouch, when the epithelium transforms from a dome into a curved fold, quantifying 3D tissue shape changes and mapping spatial patterns of cellular behaviors on the evolving geometry using cellular topology. Using a physical model inspired by shape programming, active cell rearrangements were found that are the major contributor to pouch eversion and validate this conclusion using a knockdown of MyoVI, which reduces rearrangements and disrupts morphogenesis. This work shows that shape programming is a mechanism for animal tissue morphogenesis and suggests that patterns in nature could present design strategies for shape-programmable materials.
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Tuesday June 24th - Signaling

Nair, S., Baker, N. E. (2024). Extramacrochaetae regulates Notch signaling in the Drosophila eye through non-apoptotic caspase activity. bioRxiv, PubMed ID: 39131389
Summary:
Many cell fate decisions are determined transcriptionally. Accordingly, some fate specification is prevented by Inhibitor of DNA binding (Id) proteins that interfere with DNA binding by master regulatory transcription factors. The Drosophila Id protein Extra macrochaetae (Emc) also affect developmental decisions by regulating caspase activity. Emc, which prevents proneural bHLH transcription factors from specifying neural cell fate, also prevents homodimerization of another bHLH protein, Daughterless (Da), and thereby maintains expression of the Death-Associated Inhibitor of Apoptosis (diap1) gene. Multiple effects of emc mutations on cell growth and on eye development were all caused by reduced Diap1 levels and corresponding activation of caspases. These effects included acceleration of the morphogenetic furrow, failure of R7 photoreceptor cell specification, and delayed differentiation of non-neuronal cone cells. Within emc mutant clones, Notch signaling was elevated in the morphogenetic furrow, increasing morphogenetic furrow speed. This was associated with caspase-dependent increase in levels of Delta protein, the transmembrane ligand for Notch. Posterior to the morphogenetic furrow, elevated Delta cis-inhibited Notch signaling that was required for R7 specification and cone cell differentiation. Thus, emc mutations reveal the importance of restraining caspase activity even in non-apoptotic cells to prevent abnormal development, in the Drosophila eye through effects on Notch signaling. 		Ewen-Campen, B., Perrimon, N. (2024). Wnt signaling modulates the response to DNA damage in the Drosophila wing imaginal disc by regulating the EGFR pathway. PLoS Biol, 22(7):e3002547 PubMed ID: 39047051
Summary:
Despite the deep conservation of the DNA damage response (DDR) pathway, cells in different contexts vary widely in their susceptibility to DNA damage and their propensity to undergo apoptosis as a result of genomic lesions. One of the cell signaling pathways implicated in modulating the DDR is the highly conserved Wnt pathway, which is known to promote resistance to DNA damage caused by ionizing radiation in a variety of human cancers. However, the mechanisms linking Wnt signal transduction to the DDR remain unclear. Heuse a genetically encoded system in Drosophila to reliably induce consistent levels of DNA damage in vivo, and demonstrate that canonical Wnt signaling in the wing imaginal disc buffers cells against apoptosis in the face of DNA double-strand breaks. Wg, the primary Wnt ligand in Drosophila, activates epidermal growth factor receptor (EGFR) signaling via the ligand-processing protease Rhomboid, which, in turn, modulates the DDR in a Chk2<.a>-, p53-, and E2F1-dependent manner. These studies provide mechanistic insight into the modulation of the DDR by the Wnt and EGFR pathways in vivo in a highly proliferative tissue. Furthermore, they reveal how the growth and patterning functions of Wnt signaling are coupled with prosurvival, antiapoptotic activities, thereby facilitating developmental robustness in the face of genomic damage.
Turley, J., Robertson, F., Chenchiah, I. V., Liverpool, T. B., Weavers, H., Martin, P. (2024). AI reveals a damage signalling hierarchy that coordinates different cell behaviours driving wound re-epithelialisation. Development, PubMed ID: 39177163
Summary:
One of the key tissue movements driving closure of a wound is re-epithelialisation. Earlier wound healing studies have described the dynamic cell behaviours that contribute to wound re-epithelialisation, including cell division, cell shape changes and cell migration, as well as the signals that might regulate these cell behaviours. This study used a series of deep learning tools to quantify the contributions of each of these cell behaviours from movies of repairing wounds in the Drosophila pupal wing epithelium. Tests were performed to show how each is altered following knockdown of the conserved wound repair signals, Ca2+ and JNK, as well as ablation of macrophages which supply growth factor signals believed to orchestrate aspects of the repair process. These genetic perturbation experiments provide quantifiable insights regarding how these wound signals impact cell behaviours. This study found that Ca2+ signalling is a master regulator required for all contributing cell behaviours; JNK signalling primarily drives cell shape changes and divisions, whereas signals from macrophages regulate largely cell migration and proliferation. These studies show AI to be a valuable tool for unravelling complex signalling hierarchies underlying tissue repair. 		Al Asafen, H., Beseli, A., Chen, H. Y., Hiremath, S., Williams, C. M., Reeves, G. T. (2024). Dynamics of BMP signaling and stable gene expression in the early Drosophila embryo. Biol Open, 13(9) PubMed ID: 39207258
Summary:
In developing tissues, morphogen gradients are thought to initialize gene expression patterns. However, the relationship between the dynamics of morphogen-encoded signals and gene expression decisions is largely unknown. This study examined the dynamics of the Bone Morphogenetic Protein (BMP) pathway in Drosophila blastoderm-stage embryos. In this tissue, the BMP pathway is highly dynamic: it begins as a broad and weak signal on the dorsal half of the embryo, then 20-30 min later refines into a narrow, intense peak centered on the dorsal midline. This dynamical progression of the BMP signal raises questions of how it stably activates target genes. Therefore, live imaging of the BMP signal was performed; dorsal-lateral cells were found to experience only a short transient in BMP signaling, after which the signal is lost completely. Moreover, the transcriptional response of the BMP target gene pannier was measured in live embryos, and it was found to remain activated in dorsal-lateral cells, even after the BMP signal is lost. These findings may suggest that the BMP pathway activates a memory, or 'ratchet' mechanism that may sustain gene expression.
Nourisanami, F., Sobol, M., Li, Z., Horvath, M., Kowalska, K., Kumar, A., Vlasak, J., Koupilova, N., Luginbuhl, D. J., Luo, L., Rozbesky, D. (2024). Molecular mechanisms of proteoglycan-mediated semaphorin signaling in axon guidance. Proc Natl Acad Sci U S A, 121(31):e2402755121 PubMed ID: 39042673
Summary:
The precise assembly of a functional nervous system relies on axon guidance cues. Beyond engaging their cognate receptors and initiating signaling cascades that modulate cytoskeletal dynamics, guidance cues also bind components of the extracellular matrix, notably proteoglycans, yet the role and mechanisms of these interactions remain poorly understood. This study found that Drosophila secreted semaphorins bind specifically to glycosaminoglycan (GAG) chains of proteoglycans, showing a preference based on the degree of sulfation. Structural analysis of Sema2b unveiled multiple GAG-binding sites positioned outside canonical plexin-binding site, with the highest affinity binding site located at the C-terminal tail, characterized by a lysine-rich helical arrangement that appears to be conserved across secreted semaphorins. In vivo studies revealed a crucial role of the Sema2b C-terminal tail in specifying the trajectory of olfactory receptor neurons. It is proposed that secreted semaphorins tether to the cell surface through interactions with GAG chains of proteoglycans, facilitating their presentation to cognate receptors on passing axons. 		Guo, P., Li, B., Dong, W., Zhou, H., Wang, L., Su, T., Carl, C., Zheng, Y., Hong, Y., Deng, H., Pan, D. (2024). PI4P-mediated solid-like Merlin condensates orchestrate Hippo pathway regulation. Science, 385(6709):eadf4478 PubMed ID: 39116228
Summary:
Despite recent studies implicating liquid-like biomolecular condensates in diverse cellular processes, many biomolecular condensates exist in a solid-like state, and their function and regulation are less understood. This study shows that the tumor suppressor Merlin, an upstream regulator of the Hippo pathway, localizes to both cell junctions and medial apical cortex in Drosophila epithelia, with the latter forming solid-like condensates that activate Hippo signaling. Merlin condensation required phosphatidylinositol-4-phosphate (PI4P)-mediated plasma membrane targeting and was antagonistically controlled by Pez and cytoskeletal tension through plasma membrane PI4P regulation. The solid-like material properties of Merlin condensates are essential for physiological function and protect the condensates against external perturbations. Collectively, these findings uncover an essential role for solid-like condensates in normal physiology and reveal regulatory mechanisms for their formation and disassembly.
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Monday June 23rd - Cell Cycle

Leca, N., Barbosa, F., Rodriguez-Calado, S., Moura, M., Pedroso, P. D., Pinto, I., Verza, A. E., Bange, T., Sunkel, C. E., Barisic, M., Maresca, T. J., Conde, C. (2024). Proximity-based activation of AURORA A by MPS1 potentiates error correction. bioRxiv, PubMed ID: 38948877
Summary:
Faithfull cell division relies on mitotic chromosomes becoming bioriented with each pair of sister kinetochores bound to microtubules oriented toward opposing spindle poles. Erroneous kinetochore-microtubule attachments often form during early mitosis, but are destabilized through the phosphorylation of outer kinetochore proteins by centromeric AURORA B kinase (ABK) and centrosomal AURORA A kinase (AAK), thus allowing for re-establishment of attachments until biorientation is achieved. MPS1-mediated phosphorylation of NDC80 has also been shown to directly weaken the kinetochore-microtubule interface in yeast. In human cells, MPS1 has been proposed to transiently accumulate at end-on attached kinetochores and phosphorylate SKA3 to promote microtubule release. Whether MPS1 directly targets NDC80 and/or promotes the activity of AURORA kinases in metazoans remains unclear. This study reports a novel mechanism involving communication between kinetochores and centrosomes, wherein MPS1 acts upstream of AAK to promote error correction. MPS1 on pole-proximal kinetochores phosphorylates the C-lobe of AAK thereby increasing its activation at centrosomes. This proximity-based activation ensures the establishment of a robust AAK activity gradient that locally destabilizes mal-oriented kinetochores near spindle poles. Accordingly, MPS1 depletion from Drosophila cells causes severe chromosome misalignment and erroneous kinetochore-microtubule attachments, which can be rescued by tethering either MPS1 or constitutively active AAK mutants to centrosomes. Proximity-based activation of AAK by MPS1 also occurs in human cells to promote AAK-mediated phosphorylation of the NDC80 N-terminal tail. These findings uncover an MPS1-AAK cross-talk that is required for efficient error correction, showcasing the ability of kinetochores to modulate centrosome outputs to ensure proper chromosome segregation. 		Haseeb, M. A., Weng, K. A., Bickel, S. E. (2024). Chromatin-associated cohesin turns over extensively and forms new cohesive linkages in Drosophila oocytes during meiotic prophase. Curr Biol, 34(13):2868-2879. PubMed ID: 38870933
Summary:
In dividing cells, accurate chromosome segregation depends on sister chromatid cohesion, protein linkages that are established during DNA replication. Faithful chromosome segregation in oocytes requires that cohesion, first established in S phase, remain intact for days to decades, depending on the organism. Premature loss of meiotic cohesion in oocytes leads to the production of aneuploid gametes and contributes to the increased incidence of meiotic segregation errors as women age (maternal age effect). The prevailing model is that cohesive linkages do not turn over in mammalian oocytes. However, it has been previously reported that cohesion-related defects arise in Drosophila oocytes when individual cohesin subunits or cohesin regulators are knocked down after meiotic S phase. This study used two strategies to express a tagged cohesin subunit exclusively during mid-prophase in Drosophila oocytes; newly expressed cohesin was found to be used used to form de novo linkages after meiotic S phase. Cohesin along the arms of oocyte chromosomes appears to completely turn over within a 2-day window during prophase, whereas replacement is less extensive at centromeres. Unlike S-phase cohesion establishment, the formation of new cohesive linkages during meiotic prophase does not require acetylation of conserved lysines within the Smc head. These findings indicate that maintenance of cohesion between S phase and chromosome segregation in Drosophila oocytes requires an active cohesion rejuvenation program that generates new cohesive linkages during meiotic prophase.
Shapiro, J. G., Changela, N., Jang, J. K., Joshi, J. N., McKim, K. S. (2024). Distinct checkpoint and homolog biorientation pathways regulate meiosis I in Drosophila oocytes. bioRxiv, PubMed ID: 39229242
Summary:
Mitosis and meiosis have two mechanisms for regulating the accuracy of chromosome segregation: error correction and the spindle assembly checkpoint (SAC). This study investigated the function of several checkpoint proteins in meiosis I of Drosophila oocytes. Evidence of a SAC response by several of these proteins is found upon depolymerization of microtubules by colchicine. However, unattached kinetochores or errors in biorientation of homologous chromosomes does not induce a SAC response. Furthermore, the metaphase I arrest does not depend on SAC genes, suggesting the APC is inhibited even if the SAC is silenced. Two SAC proteins, ROD of the ROD-ZW10-Zwilch (RZZ) complex and MPS1, are also required for the biorientation of homologous chromosomes during meiosis I, suggesting an error correction function. Both proteins aid in preventing or correcting erroneous attachments and depend on SPC105R for localization to the kinetochore. This study has defined a region of SPC105R, amino acids 123-473, that is required for ROD localization and biorientation of homologous chromosomes at meiosis I. Surprisingly, ROD removal, or "streaming", is independent of the dynein adaptor Spindly and is not linked to the stabilization of end-on attachments. Instead, meiotic RZZ streaming appears to depend on cell cycle stage and may be regulated independently of kinetochore attachment or biorientation status. Dynein adaptor Spindly is also required for biorientation at meiosis I, and surprisingly, the direction of RZZ streaming. 		Joshi, J. N., Changela, N., Mahal, L., Jang, J., Defosse, T., Wang, L. I., Das, A., Shapiro, J. G., McKim, K. (2024). Meiosis-specific functions of kinetochore protein SPC105R required for chromosome segregation in Drosophila oocytes. Mol Biol Cell, 35(8):ar105 PubMed ID: 38865189
Summary:
The reductional division of meiosis I requires the separation of chromosome pairs towards opposite poles. The outer kinetochore protein SPC105R/KNL1 has been implemented in driving meiosis I chromosome segregation through lateral attachments to microtubules and coorientation of sister centromeres. To identify the domains of SPC105R that are critical for meiotic chromosome segregation, an RNAi-resistant gene expression system was developed. The SPC105R C-terminal domain (aa 1284-1960) was shown to be is necessary and sufficient for recruiting NDC80 to the kinetochore and building the outer kinetochore. Furthermore, the C-terminal domain recruits BUBR1, which in turn recruits the cohesion protection proteins MEI-S332 and PP2A. Of the remaining 1283 amino acids, the first 473 are most important for meiosis. The first 123 amino acids of the N-terminal half of SPC105R contain the conserved SLRK and RISF motifs that are targets of PP1 and Aurora B kinase and are most important for regulating the stability of microtubule attachments and maintaining metaphase I arrest. The region between amino acids 124 and 473 are required for lateral microtubule attachments and biorientation of homologues, which are critical for accurate chromosome segregation in meiosis I.
Lake, C. M., Gardner, J., Briggs, S., Yu, Z., McKown, G., Hawley, R. S. (2024). The deubiquitinase Usp7 in Drosophila melanogaster is required for synaptonemal complex maintenance. Proc Natl Acad Sci U S A, 121(36):e2409346121 PubMed ID: 39190345
Summary:
Meiosis is a form of cell division that is essential to sexually reproducing organisms and is therefore highly regulated. Each event of meiosis must occur at the correct developmental stage to ensure that chromosomes are segregated properly during both meiotic divisions. One unique meiosis-specific structure that is tightly regulated in terms of timing of assembly and disassembly is the synaptonemal complex (SC). While the mechanism(s) for assembly and disassembly of the SC are poorly understood in Drosophila melanogaster, posttranslational modifications, including ubiquitination and phosphorylation, are known to play a role. This study identified a role for the deubiquitinase Usp7 in the maintenance of the SC in early prophase and showed that its function in SC maintenance is independent of the meiotic recombination process. Using two usp7 shRNA constructs that result in different knockdown levels, the presence of SC through early/mid-pachytene was shown to be critical for normal levels and placement of crossovers. 		Zhang, Q., Wang, Y., Bu, Z., Zhang, Y., Zhang, Q., Li, L., Yan, L., Wang, Y., Zhao, S. (2024). Ras promotes germline stem cell division in Drosophila ovaries. Stem cell reports, 19(8):1205-1216 PubMed ID: 39029459
Summary:
The Ras family genes are proto-oncogenes that are highly conserved from Drosophila to humans. In Drosophila, Ras(V12) is a constitutively activated form of the Ras oncoprotein, and its function in cell-cycle progression is context dependent. However, how it influences the cell cycle of female germline stem cells (GSCs) still remains unknown. Using both wild-type GSCs and bam mutant GSC-like cells as model systems, this study determined that Ras(V12) overexpression promotes GSC division, not growth, opposite to that in somatic wing disc cells. Ras performs this function through activating the mitogen-activated protein kinase (MAPK) signaling. This signaling is activated specifically in the M phase of mitotic germ cells, including both wild-type GSCs and bam mutant GSC-like cells. Furthermore, Ras(V12) overexpression triggers polyploid nurse cells to die through inducing mitotic stress. Given the similarities between Drosophila and mammalian GSCs, it is proposed that the Ras/MAPK signaling also promotes mammalian GSC division.
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Thursday, June 19th - Behavior

Kubrak, O., Jorgensen, A. F., Koyama, T., Lassen, M., Nagy, S., Hald, J., Mazzoni, G., Madsen, D., Hansen, J. B., Larsen, M. R., Texada, M. J., Hansen, J. L., Halberg, K. V., Rewitz, K. (2024). LGR signaling mediates muscle-adipose tissue crosstalk and protects against diet-induced insulin resistance. Nat Commun, 15(1):6126 PubMed ID: 39033139
Summary:
Obesity impairs tissue insulin sensitivity and signaling, promoting type-2 diabetes. Although improving insulin signaling is key to reversing diabetes, the multi-organ mechanisms regulating this process are poorly defined. This study screen the secretome and receptome in Drosophila to identify the hormonal crosstalk affecting diet-induced insulin resistance and obesity. This study discovered a complex interplay between muscle, neuronal, and adipose tissues, mediated by Bone Morphogenetic Protein (BMP) signaling and the hormone Bursicon, that enhances insulin signaling and sugar tolerance. Muscle-derived BMP signaling, induced by sugar, governs neuronal Bursicon signaling. Bursicon, through its receptor Rickets, a Leucine-rich-repeat-containing G-protein coupled receptor (LGR), improves insulin secretion and insulin sensitivity in adipose tissue, mitigating hyperglycemia. In mouse adipocytes, loss of the Rickets ortholog LGR4 blunts insulin responses, showing an essential role of LGR4 in adipocyte insulin sensitivity.These findings reveal a muscle-neuronal-fat-tissue axis driving metabolic adaptation to high-sugar conditions, identifying LGR4 as a critical mediator in this regulatory network. 		Stupski, S. D., van Breugel, F. (2024). Wind gates olfaction-driven search states in free flight. Curr Biol, PubMed ID: 39067453
Summary:
For organisms tracking a chemical cue to its source, the motion of their surrounding fluid provides crucial information for success. Swimming and flying animals engaged in olfaction-driven search often start by turning into the direction of an oncoming wind or water current. However, it is unclear how organisms adjust their strategies when directional cues are absent or unreliable, as is often the case in nature. This study used the genetic toolkit of Drosophila melanogaster to develop an optogenetic paradigm to deliver temporally precise "virtual" olfactory experiences for free-flying animals in either laminar wind or still air. It was first confirmed that in laminar wind flies turn upwind. Furthermore, it was shown that they achieve this using a rapid (~100 ms) turn, implying that flies estimate the ambient wind direction prior to "surging" upwind. In still air, flies adopt a remarkably stereotyped "sink and circle" search state characterized by ~60° turns at 3-4 Hz, biased in a consistent direction. Together, these results show that Drosophila melanogaster assesses the presence and direction of ambient wind prior to deploying a distinct search strategy. In both laminar wind and still air, immediately after odor onset, flies decelerate and often perform a rapid turn. Both maneuvers are consistent with predictions from recent control theoretic analyses for how insects may estimate properties of wind while in flight. It is suggested that flies may use their deceleration and "anemometric" turn as active sensing maneuvers to rapidly gauge properties of their wind environment before initiating a proximal or upwind search routine.
Booth, J. H., Meek, A. T., Kronenberg, N. M., Pulver, S. R., Gather, M. C. (2024). Optical mapping of ground reaction force dynamics in freely behaving Drosophila melanogaste larvaer. Elife, 12 PubMed ID: 39042447
Summary:
During locomotion, soft-bodied terrestrial animals solve complex control problems at substrate interfaces, but understanding of how they achieve this without rigid components remains incomplete. This study developed new all-optical methods based on optical interference in a deformable substrate to measure ground reaction forces (GRFs) with micrometre and nanonewton precision in behaving Drosophila larvae. Combining this with a kinematic analysis of substrate-interfacing features, new light is shed onto the biomechanical control of larval locomotion. Crawling in larvae measuring ~1 mm in length involves an intricate pattern of cuticle sequestration and planting, producing GRFs of 1-7 μN. Larvae insert and expand denticulated, feet-like structures into substrates as they move, a process not previously observed in soft-bodied animals. These 'protopodia' form dynamic anchors to compensate counteracting forces. This work provides a framework for future biomechanics research in soft-bodied animals and promises to inspire improved soft-robot design. 		Poissonnier, L. A., Danchin, E., Isabel, G. (2024). Male attractiveness is subjective to exposure to males of different attractiveness in fruit flies. Sci Rep, 14(1):16463 PubMed ID: 39014083
Summary:
Mate choice is a crucial decision in any animal. In terms of fitness, the best mate is the one that leads to the most abundant and productive offspring. Pairing with a low-quality mate would reduce fitness, generating selection for accurate and subtle mate choice in all animal species. Hence, mate choice is expected to be highly context dependent, and should depend on other potential options. For instance, a medium-quality male can constitute the best option when all other males are in poorer condition, but not when there are better-quality males available. Therefore, animals are predicted to gather information about their social context and adapt their mate choice to it. This study reports on experiments in which the social environment of females of Drosophila melanogaster were manipulated; after encountering a high or a low-quality male, females take more or less time to accept copulation with another male, suggesting that females adapt their mating strategy to their social context. This study also reports on a similar effect in D. biarmiceps. Thus, male attractiveness appears to depend on the quality of recently met males, suggesting that male attractiveness is subjective, indicating plastic and context dependent mate choice.
Lynn, K., Ichinose, T., Tanimoto, H. (2024). Peer-induced quiescence of male Drosophila melanogaster following copulation.. Front Behav Neurosci, 18:1414029 PubMed ID: 39081685
Summary:
Mating experience impacts the physiology and behavior of animals. Although mating effects of female Drosophila melanogaster have been studied extensively, the behavioral changes of males following copulation have not been fully understood. This study characterized the mating-dependent behavioral changes of male flies, especially focusing on fly-to-fly interaction, and their dependence on rearing conditions. The data demonstrate that male flies quiesce their courtship toward both females and males, as well as their locomotor activity. This post-copulatory quiescence appears to be contingent upon the presence of a peer, as minimal variation is noted in locomotion when the male is measured in isolation. Interestingly, copulated males influence a paired male without successful copulation to reduce his locomotion. These findings point to a conditional behavioral quiescence following copulation, influenced by the presence of other flies. 		Otarola-Jimenez, J., Nataraj, N., Bisch-Knaden, S., Hansson, B. S., Knaden, M. (2024). Oviposition experience affects oviposition preference in Drosophila melanogaster. iScience, 27(8):110472 PubMed ID: 39129830
Summary:
Learning, memorizing, and recalling of potential ovipositing sites can influence oviposition preference. Classical conditioning experiments have shown that vinegar flies can learn the association of olfactory, gustatory, or visual stimuli with either positive or negative unconditioned stimuli. However, less is known about whether similar associations are formed in an ecologically more relevant context like during oviposition. The current experiments reveal that Drosophila melanogaster females increase their preference for substrates they have already experienced. However, this change of preference requires that the flies not only smelled or touched the substrates but also oviposited on them. It was furthermore shown that such an experience results in long-term memory lasting for at least 4 days, i.e., a duration that so far was shown only for aversive conditioning. This study thus reveals a different form of associative learning in D. melanogaster that might be highly relevant for settling novel ecological niches.
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Wednescay, June 18th - Disease Models

Yadav, S., Graham, A., Al Hammood, F., Garbark, C., Vasudevan, D., Pandey, U., Asara, J. M., Rajasundaram, D., Parkhitko, A. A. (2024). Unique tau- and synuclein-dependent metabolic reprogramming in neurons distinct from normal aging. Aging Cell:e14277 PubMed ID: 39137949
Summary:
Neuronal cells are highly specialized cells and have a specific metabolic profile to support their function. It has been demonstrated that the metabolic profiles of different cells/tissues undergo significant reprogramming with advancing age, which has often been considered a contributing factor towards aging-related diseases including Alzheimer's (AD) and Parkinson's (PD) diseases. However, it is unclear if the metabolic changes associated with normal aging predispose neurons to disease conditions or a distinct set of metabolic alterations happen in neurons in AD or PD which might contribute to disease pathologies. To decipher the changes in neuronal metabolism with age, in AD, or in PD, high-throughput steady-state metabolite profiling was performed on heads in wildtype Drosophila and in Drosophila models relevant to AD and PD. Intriguingly, it was found that the spectrum of affected metabolic pathways is dramatically different between normal aging, Tau, or Synuclein overexpressing neurons. Genetic targeting of the purine and glutamate metabolism pathways, which were dysregulated in both old age and disease conditions partially rescued the neurodegenerative phenotype associated with the overexpression of wildtype and mutant tau. These findings support a "two-hit model" to explain the pathological manifestations associated with AD where both aging- and Tau/Synuclein- driven metabolic reprogramming events cooperate with each other, and targeting both could be a potent therapeutic strategy. 		Ruchitha, B. G., Kumar, N., Sura, C., Tung, S. (2024). Selection for greater dispersal in early life leads to faster age-dependent decline in locomotor activity and shorter lifespan. J Evol Biol, PubMed ID: 39105302
Summary:
Locomotor activity is one of the major traits that is affected by age. Greater locomotor activity is also known to evolve in the course of dispersal evolution. However, the impact of dispersal evolution on the functional senescence of locomotor activity is largely unknown. This knowledge gap using large outbred populations of Drosophila melanogaster selected for increased dispersal. Locomotor activity of these flies was assessed at regular intervals until a late age. Longevity of these flies was also recorded. Locomotor activity was shown to decline with age in general. However interestingly, activity level of dispersal selected populations never drops below the ancestry-matched-controls, despite the rate of age-dependent decline in activity of the dispersal selected populations being greater than their respective controls. Dispersal selected population was also found to have shorter lifespan as compared to its control, a potential cost of elevated level of activity throughout their life. These results are crucial in the context of invasion biology as contemporary climate change, habitat degradation, and destruction provide congenial conditions for dispersal evolution. Such controlled and tractable studies investigating the ageing pattern of important functional traits are important in the field of biogerontology as well.
Lim, J. J., Noh, S., Kang, W., Hyun, B., Lee, B. H., Hyun, S. (2024). Pharmacological inhibition of USP14 delays proteostasis-associated aging in a proteasome-dependent but foxo-independent manner. Autophagy:1-17 PubMed ID: 39113571
Summary:
Aging is often accompanied by a decline in proteostasis, manifested as an increased propensity for misfolded protein aggregates, which are prevented by protein quality control systems, such as the ubiquitin-proteasome system (UPS) and macroautophagy/autophagy. Although the role of the UPS and autophagy in slowing age-induced proteostasis decline has been elucidated, limited information is available on how these pathways can be activated in a collaborative manner to delay proteostasis-associated aging. This study showed that activation of the UPS via the pharmacological inhibition of USP14 (ubiquitin specific peptidase 14) using IU1 improves proteostasis and autophagy decline caused by aging or proteostatic stress in Drosophila and human cells. Treatment with IU1 not only alleviated the aggregation of polyubiquitinated proteins in Drosophila flight muscles but also extended the fly lifespan with enhanced locomotive activity via simultaneous activation of the UPS and autophagy. Interestingly, the effect of this drug disappeared when proteasomal activity was inhibited, but was evident upon proteostasis disruption by foxo mutation. Overall, these findings shed light on potential strategies to efficiently ameliorate age-associated pathologies associated with perturbed proteostasis. 		Dondi, C., Vogler, G., Gupta, A., Walls, S. M., Kervadec, A., Marchant, J., Romero, M. R., Diop, S., Goode, J., Thomas, J. B., Colas, A. R., Bodmer, R., Montminy, M., Ocorr, K. (2024). The nutrient sensor CRTC and Sarcalumenin/thinman represent an alternate pathway in cardiac hypertrophy. Cell Rep, 43(8):114549 PubMed ID: 39093699
Summary:
CREB-regulated transcription co-activator (CRTC) is activated by Calcineurin (CaN) to regulate gluconeogenic genes. CaN also has roles in cardiac hypertrophy. This study explored a cardiac-autonomous role for CRTC in cardiac hypertrophy. In Drosophila, CRTC mutants exhibit severe cardiac restriction, myofibrillar disorganization, fibrosis, and tachycardia. Cardiac-specific CRTC knockdown (KD) phenocopies mutants, and cardiac overexpression causes hypertrophy. CaN-induced hypertrophy in Drosophila is reduced in CRTC mutants, suggesting that CRTC mediates the effects. RNA sequencing (RNA-seq) of CRTC-KD and overexpressing hearts reveals contraregulation of metabolic genes. Genes with conserved CREB sites include the fly ortholog of Sarcalumenin, a Ca(2+)-binding protein. Cardiac manipulation of this gene recapitulates the CRTC-KD and -overexpression phenotypes. CRTC KD in zebrafish also causes cardiac restriction, and CRTC KD in human induced cardiomyocytes causes a reduction in Srl expression and increased action potential duration. These data from three model systems suggest that CaN-CRTC-Sarcalumenin signaling represents an alternate, conserved pathway underlying cardiac function and hypertrophy.
Vannelli, A., Mariano, V., Bagni, C., Kanellopoulos, A. K. (2024). Activation of the 5-HT1A Receptor by Eltoprazine Restores Mitochondrial and Motor Deficits in a Drosophila Model of Fragile X Syndrome. Int J Mol Sci, 25(16) PubMed ID: 39201473
Summary:
Neurons rely on mitochondrial energy metabolism for essential functions like neurogenesis, neurotransmission, and synaptic plasticity. Mitochondrial dysfunctions are associated with neurodevelopmental disorders including Fragile X syndrome (FXS), the most common cause of inherited intellectual disability, which also presents with motor skill deficits. However, the precise role of mitochondria in the pathophysiology of FXS remains largely unknown. Notably, previous studies have linked the serotonergic system and mitochondrial activity to FXS. This study investigates the potential therapeutic role of serotonin receptor 1A (5-HT1A) in FXS. Using the Drosophila model of FXS, it was demonstrated that treatment with eltoprazine, a 5-HT1A agonist, can ameliorate synaptic transmission, correct mitochondrial deficits, and ultimately improve motor behavior. While these findings suggest that the 5-HT1A-mitochondrial axis may be a promising therapeutic target, further investigation is needed in the context of FXS. 		Wei, F., Liu, S., Liu, J., Sun, Y., Allen, A. E., Reid, M. A., Locasale, J. W. (2024). Separation of reproductive decline from lifespan extension during methionine restriction. Nature aging, 4(8):1089-1101 PubMed ID: 39060538
Summary:
Lifespan-extending interventions are generally thought to result in reduced fecundity. The generality of this principle and how it may extend to nutrition and metabolism is not understood. This study considered dietary methionine restriction (MR), a lifespan-extending intervention linked to Mediterranean and plant-based diets. Using a chemically defined diet that was developed for Drosophila melanogaster, this study surveyed the nutritional landscape in the background of MR and found that folic acid, a vitamin linked to one-carbon metabolism, notably was the lone nutrient that restored reproductive capacity while maintaining lifespan extension. In vivo isotope tracing, metabolomics and flux analysis identified the tricarboxylic cycle and redox coupling as major determinants of the MR-folic acid benefits, in part, as they related to sperm function. Together these findings suggest that dietary interventions optimized for longevity may be separable from adverse effects such as reproductive decline.
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Tuesday, June 17th - Adult Neural Development, Structure and Function

Koch, S., Kandimalla, P., Padilla, E., Kaur, S., Kaur, R., Nguyen, M., Nelson, A., Khalsa, S., Younossi-Hartenstein, A., Hartenstein, V. (2024). Structural changes shaping the Drosophila ellipsoid body ER-neurons during development and aging. Dev Biol, 516:96-113 PubMed ID: 39089472
Summary:
The ellipsoid body (EB) of the insect brain performs pivotal functions in controlling navigation. Input and output of the EB is provided by multiple classes of R-neurons (now referred to as ER-neurons) and columnar neurons which interact with each other in a stereotypical and spatially highly ordered manner. The developmental mechanisms that control the connectivity and topography of EB neurons are largely unknown. One indispensable prerequisite to unravel these mechanisms is to document in detail the sequence of events that shape EB neurons during their development. This study analyzed the development of the Drosophila EB. In addition to globally following the ER-neuron and columnar neuron (sub)classes in the spatial context of their changing environment a single cell analysis was performed using the multi-color flip out (MCFO) system to analyze the developmental trajectory of ER-neurons at different pupal stages, young adults (4d) and aged adults (∼60d). The EB was shown to develop as a merger of two distinct elements, a posterior and anterior EB primordium (prEBp and prEBa, respectively. ER-neurons belonging to different subclasses form growth cones and filopodia that associate with the prEBp and prEBa in a pattern that, from early pupal stages onward, foreshadows their mature structure. Filopodia of all ER-subclasses are initially much longer than the dendritic and terminal axonal branches they give rise to, and are pruned back during late pupal stages. Interestingly, extraneous branches, particularly significant in the dendritic domain, are a hallmark of ER-neuron structure in aged brains. Aging is also associated with a decline in synaptic connectivity from columnar neurons, as well as upregulation of presynaptic protein (Brp) in ER-neurons. These findings advance the EB (and ER-neurons) as a favorable system to visualize and quantify the development and age-related decline of a complex neuronal circuitry. 		Someya, W., Akutsu, T., Nacher, J. C. (2024). Target control of linear directed networks based on the path cover problem. Sci Rep, 14(1):16881 PubMed ID: 39043768
Summary:
Securing complete control of complex systems comprised of tens of thousands of interconnected nodes holds immense significance across various fields, spanning from cell biology and brain science to human-engineered systems. However, depending on specific functional requirements, it can be more practical and efficient to focus on a pre-defined subset of nodes for control, a concept known as target control. While some methods have been proposed to find the smallest driver node set for target control, they either rely on heuristic approaches based on k-walk theory, lacking a guarantee of optimal solutions, or they are overly complex and challenging to implement in real-world networks. To address this challenge, this study introduced a simple and elegant algorithm, inspired by the path cover problem, which efficiently identifies the nodes required to control a target node set within polynomial time. To practically apply the algorithm in real-world systems, several networks were selected, in which a specific set of nodes with functional significance can be designated as a target control set. The analysed systems include the complete connectome of the nematode worm C. elegans, the recently disclosed connectome of the Drosophila larval brain, as well as dozens of genome-wide metabolic networks spanning major plant lineages. The target control analysis shed light on distinctions between neural systems in nematode worms and larval brain insects, particularly concerning the number of nodes necessary to regulate specific functional systems. Furthermore, this analysis uncovers evolutionary trends within plant lineages, notably when examining the proportion of nodes required to control functional pathways.
Liu, J., Wang, Y., Liu, X., Han, J., Tian, Y. (2024). Spatiotemporal changes in Netrin/Dscam1 signaling dictate axonal projection direction in Drosophila small ventral lateral clock neurons. Elife, 13 PubMed ID: 39052321
Summary:
Axon projection is a spatial- and temporal-specific process in which the growth cone receives environmental signals guiding axons to their final destination. However, the mechanisms underlying changes in axonal projection direction without well-defined landmarks remain elusive. This study presents evidence showcasing the dynamic nature of axonal projections in Drosophila's small ventral lateral clock neurons (s-LNvs). The findings reveal that these axons undergo an initial vertical projection in the early larval stage, followed by a subsequent transition to a horizontal projection in the early-to-mid third instar larvae. The vertical projection of s-LNv axons correlates with mushroom body calyx expansion, while the s-LNv-expressed Down syndrome cell adhesion molecule (Dscam1) interacts with Netrins to regulate the horizontal projection. During a specific temporal window, locally newborn dorsal clock neurons secrete Netrins, facilitating the transition of axonal projection direction in s-LNvs. This study establishes a compelling in vivo model to probe the mechanisms of axonal projection direction switching in the absence of clear landmarks. These findings underscore the significance of dynamic local microenvironments in the complementary regulation of axonal projection direction transitions. 		Sharma, P. N., Sheeba, V. (2024). Reorganization of circadian activity and the pacemaker circuit under novel light regimes. Proceedings Biological sciences, 291(2027):20241190 PubMed ID: 39043245
Summary:
Many environmental features are cyclic, with predictable changes across the day, seasons and latitudes. Additionally, anthropogenic, artificial-light-induced changes in photoperiod or shiftwork-driven novel light/dark cycles also occur. Endogenous timekeepers or circadian clocks help organisms cope with such changes. The remarkable plasticity of clocks is evident in the waveforms of behavioural and molecular rhythms they govern. Despite detailed mechanistic insights into the functioning of the circadian clock, practical means to manipulate activity waveform are lacking. Previous studies using a nocturnal rodent model showed that novel light regimes caused locomotor activity to bifurcate such that mice showed two bouts of activity restricted to the dimly lit phases. This study explored the generalizability of these findings and leveraged the genetic toolkit of Drosophila melanogaster to obtain mechanistic insights into this unique phenomenon. Dim scotopic illumination of specific durations induces circadian photoreceptor CRYPTOCHROME-dependent activity bifurcation in male flies. Circadian reorganization was shown of the pacemaker circuit, wherein the 'evening' neurons regulate the timing of both bouts of activity under novel light regimes. These findings indicate that such environmental regimes can be exploited to design light cycles, which can ease the circadian waveform into synchronizing with challenging conditions.
Neitz, A. F., Carter, B. M., Ceriani, M. F., Ellisman, M. H., de la Iglesia, H. O. (2024). Suprachiasmatic nucleus VIPergic fibers show a circadian rhythm of expansion and retraction. Curr Biol, PubMed ID: 39127047
Summary:
In animals, overt circadian rhythms of physiology and behavior are centrally regulated by a circadian clock located in specific brain regions. In the fruit fly Drosophila and in mammals, these clocks rely on single-cell oscillators, but critical for their function as central circadian pacemakers are network properties that change dynamically throughout the circadian cycle as well as in response to environmental stimuli. In the fly, this plasticity involves circadian rhythms of expansion and retraction of clock neuron fibers. Whether these drastic structural changes are a universal property of central neuronal pacemakers is unknown. To address this question, neurons of the mouse suprachiasmatic nucleus (SCN) were studied that express vasoactive intestinal polypeptide (VIP), which are critical for the SCN to function as a central circadian pacemaker. By targeting the expression of the fluorescent protein tdTomato to these neurons and using tissue clearing techniques to visualize all SCN VIPergic neurons and their fibers, this study showed that, similar to clock neurons in the fly, VIPergic fibers undergo a daily rhythm of expansion and retraction, with maximal branching during the day. This rhythm is circadian, as it persists under constant environmental conditions and is present in both males and females. It is proposed that circadian structural remodeling of clock neurons represents a key feature of central circadian pacemakers that is likely critical to regulate network properties, the response to environmental stimuli, and the regulation of circadian outputs. 		Guillemin, J., Li, J., Li, V., McDowell, S. A. T., Audette, K., Davis, G., Jelen, M., Slamani, S., Kelliher, L., Gordon, M. D., Stanley, M. (2024). Taste cells expressing Ionotropic Receptor 94e reciprocally impact feeding and egg laying in Drosophila.. Cell Rep, 43(8):114625 PubMed ID: 39141516
Summary:
Chemosensory cells across the body of Drosophila melanogaster evaluate the environment to prioritize certain behaviors. Previous mapping of gustatory receptor neurons (GRNs) on the fly labellum identified a set of neurons in L-type sensilla that express Ionotropic Receptor 94e (IR94e), but the impact of IR94e GRNs on behavior remains unclear. This study used optogenetics and chemogenetics to activate IR94e neurons and found that they drive mild feeding suppression but enhance egg laying. In vivo calcium imaging revealed that IR94e GRNs respond strongly to certain amino acids, including glutamate, and that IR94e plus co-receptors IR25a and IR76b are required for amino acid detection. Furthermore, IR94e mutants show behavioral changes to solutions containing amino acids, including increased consumption and decreased egg laying. Overall, these results suggest that IR94e GRNs on the fly labellum discourage feeding and encourage egg laying as part of an important behavioral switch in response to certain chemical cues.
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Thursday, June 12th - Genes, Enzymes, and Proteins

Khodaverdian, V., Sano, T., Maggs, L. R., Tomarchio, G., Dias, A., Tran, M., Clairmont, C., McVey, M. (2024). REV1 coordinates a multi-faceted tolerance response to DNA alkylation damage and prevents chromosome shattering in Drosophila melanogaster. PLoS Genet, 20(7):e1011181 PubMed ID: 39074150
Summary:
When replication forks encounter damaged DNA, cells utilize damage tolerance mechanisms to allow replication to proceed. These include translesion synthesis at the fork, postreplication gap filling, and template switching via fork reversal or homologous recombination. The extent to which these different damage tolerance mechanisms are utilized depends on cell, tissue, and developmental context-specific cues, the last two of which are poorly understood. To address this gap, damage tolerance responses were investigated in Drosophila melanogaster. Tolerance of DNA alkylation damage in rapidly dividing larval tissues depends heavily on translesion synthesis. Furthermore, it was shown that the REV1 protein plays a multi-faceted role in damage tolerance in Drosophila. Larvae lacking REV1 are hypersensitive to methyl methanesulfonate (MMS) and have highly elevated levels of γ-H2Av (Drosophila γ-H2AX) foci and chromosome aberrations in MMS-treated tissues. Loss of the REV1 C-terminal domain (CTD), which recruits multiple translesion polymerases to damage sites, sensitizes flies to MMS. In the absence of the REV1 CTD, DNA polymerase eta and DNA polymerase zeta become critical for MMS tolerance. In addition, flies lacking REV3, the catalytic subunit of polymerase zeta, require the deoxycytidyl transferase activity of REV1 to tolerate MMS. Together, the results demonstrate that Drosophila prioritize the use of multiple translesion polymerases to tolerate alkylation damage and highlight the critical role of REV1 in the coordination of this response to prevent genome instability. 		Klemm, J., Van Hazel, C., Harris, R. (2024). Regeneration following tissue necrosis is mediated by non-apoptotic caspase activity. bioRxiv, PubMed ID: 39091851
Summary:
Tissue necrosis is a devastating complication for many human diseases and injuries. Unfortunately, understanding of necrosis and how it impacts surrounding healthy tissue - an essential consideration when developing methods to treat such injuries - has been limited by a lack of robust genetically tractable models. Previous work established a method to study necrosis-induced regeneration in the Drosophila wing imaginal disc, which revealed a unique phenomenon whereby cells at a distance from the injury upregulate caspase activity in a process called Necrosis-induced Apoptosis (NiA) that is vital for regeneration. This study further investigated this phenomenon, showing that NiA is predominantly associated with the highly regenerative pouch region of the disc, shaped by genetic factors present in the presumptive hinge. Furthermore, this study found that a proportion of NiA fail to undergo apoptosis, instead surviving effector caspase activation to persist within the tissue and stimulate reparative proliferation late in regeneration. This proliferation relies on the initiator caspase Dronc, and occurs independent of JNK, ROS or mitogens associated with the previously characterized Apoptosis-induced Proliferation (AiP) mechanism. These data reveal a new means by which non-apoptotic Dronc signaling promotes regenerative proliferation in response to necrotic damage.
Vicidomini, R., Choudhury, S. D., Han, T. H., Nguyen, T. H., Nguyen, P., Opazo, F., Serpe, M. (2024). Versatile nanobody-based approach to image, track and reconstitute functional Neurexin-1 in vivo. Nat Commun, 15(1):6068 PubMed ID: 39025931
Summary:
Neurexins are key adhesion proteins that coordinate extracellular and intracellular synaptic components. Nonetheless, the low abundance of these multidomain proteins has complicated any localization and structure-function studies. This study combines an ALFA tag (AT)/nanobody (NbALFA) tool with classic genetics, cell biology and electrophysiology to examine the distribution and function of the Drosophila Nrx-1 in vivo. Full-length and ΔPDZ ALFA-tagged Nrx-1 variants were generated, and it was found that the PDZ binding motif is key to Nrx-1 surface expression. A PDZ binding motif provided in trans, via genetically encoded cytosolic NbALFA-PDZ chimera, fully restores the synaptic localization and function of Nrx(ΔPDZ-AT). Using cytosolic NbALFA-mScarlet intrabody, compartment-specific detection was achieved of endogenous Nrx-1, live Nrx-1 transport was tracked along the motor neuron axons, and Nrx-1 was demonstrated to co-migrate with Rab2-positive vesicles. These findings illustrate the versatility of the ALFA system and pave the way towards dissecting functional domains of complex proteins in vivo. 		Schwartz, M., Petiot, N., Chaloyard, J., Senty-Segault, V., Lirussi, F., Senet, P., Nicolai, A., Heydel, J. M., Canon, F., Sonkaria, S., Khare, V., Didierjean, C., Neiers, F. (2024). Structural and Thermodynamic Insights into Dimerization Interfaces of Drosophila Glutathione Transferases. Biomolecules, 14(7) PubMed ID: 39062472
Summary:
This study presents a comprehensive analysis of the dimerization interfaces of fly Glutathione S-Transferases (GSTs) through sequence alignment. GSTE1 is revealed to be a particularly intriguing target, providing valuable insights into the variations within Delta and Epsilon GST interfaces. The X-ray structure of GSTE1 was determined, unveiling remarkable thermal stability and a distinctive dimerization interface. Utilizing circular dichroism, the thermal stability of GSTE1 and other Drosophila GSTs with resolved X-ray structures was assessed. The subsequent examination of GST dimer stability correlated with the dimerization interface supported by findings from X-ray structural analysis and thermal stability measurements. A discussion extends to the broader context of GST dimer interfaces, offering a generalized perspective on their stability. This research enhances understanding of the structural and thermodynamic aspects of GST dimerization, contributing valuable insights to the field.
Ostale, C. M., Pulido, D., Vega-Cuesta, P., Lopez-Varea, A., de Celis, J. F. (2024). Developmental analysis of Spalt function in the Drosophila prothoracic gland. Development, 151(16) PubMed ID: 39087588
Summary:
Spalt transcriptional regulators participate in a variety of cell fate specification processes during development, regulating transcription through interactions with DNA AT-rich regions. Spalt proteins also bind to heterochromatic regions, and some of their effects require interactions with the NuRD chromatin remodeling and deacetylase complex. Most of the biological roles of Spalt proteins have been characterized in diploid cells engaged in cell proliferation. This study addresses the function of Drosophila Spalt genes in the development of a larval tissue formed by polyploid cells, the prothoracic gland, the cells of which undergo several rounds of DNA replication without mitosis during larval development. Prothoracic glands depleted of Spalt expression display severe changes in the size of the nucleolus, the morphology of the nuclear envelope and the disposition of the chromatin within the nucleus, leading to a failure in the synthesis of ecdysone. It is proposed that loss of ecdysone production in the prothoracic gland of Spalt mutants is primarily caused by defects in nuclear pore complex function that occur as a consequence of faulty interactions between heterochromatic regions and the nuclear envelope. 		Bellah, S. F., Yang, F., Xiong, F., Dou, Z., Yao, X., Liu, X. (2024). ZW10: an emerging orchestrator of organelle dynamics during the cell division cycle. J Mol Cell Biol, PubMed ID: 38830800
Summary:
Zeste white 10 (ZW10) was first identified as a centromere/kinetochore protein encoded by the ZW10 gene in Drosophila. ZW10 guides the spindle assembly checkpoint signaling during mitotic chromosome segregation in metazoans. Recent studies have shown that ZW10 is also involved in membranous organelle interactions during interphase and plays a vital role in membrane transport between the endoplasmic reticulum and Golgi apparatus. Despite these findings, the precise molecular mechanisms by which ZW10 regulates interactions between membranous organelles in interphase and the assembly of membraneless organelle kinetochore in mitosis remain elusive. This study highlights how ZW10 forms context-dependent protein complexes during the cell cycle. These complexes are essential for mediating membrane trafficking in interphase and ensuring the accurate segregation of chromosomes in mitosis.
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Wednesday, June 11th - Adult Neural Structure, Development and Function

Chang, Y. C., Peng, Y. J., Lee, J. Y., Chag, K. T. (2024). Peripheral glia and neurons jointly regulate activity-induced synaptic remodeling at the Drosophila neuromuscular junction. bioRxiv, PubMed ID: 39005352
Summary:
In the nervous system, reliable communication depends on the ability of neurons to adaptively remodel their synaptic structure and function in response to changes in neuronal activity. While neurons are the main drivers of synaptic plasticity, glial cells are increasingly recognized for their roles as active modulators. However, the underlying molecular mechanisms remain unclear. Using Drosophila neuromuscular junction as a model system for a tripartite synapse, this study showed that peripheral glial cells collaborate with neurons at the NMJ to regulate activity-induced synaptic remodeling, in part through a protein called shriveled (Shv). Shv is an activator of integrin signaling previously shown to be released by neurons during intense stimulation at the fly NMJ to regulate activity-induced synaptic remodeling. This study demonstrated that Shv is also present in peripheral glia, and glial Shv is both necessary and sufficient for synaptic remodeling. However, unlike neuronal Shv, glial Shv does not activate integrin signaling at the NMJ. Instead, it regulates synaptic plasticity in two ways: 1) maintaining the extracellular balance of neuronal Shv proteins to regulate integrin signaling, and 2) controlling ambient extracellular glutamate concentration to regulate postsynaptic glutamate receptor abundance. Loss of glial cells showed the same phenotype as loss of Shv in glia. Together, these results reveal that neurons and glial cells homeostatically regulate extracellular Shv protein levels to control activity-induced synaptic remodeling. Additionally, peripheral glia maintains postsynaptic glutamate receptor abundance and contribute to activity-induced synaptic remodeling by regulating ambient glutamate concentration at the fly NMJ. 		Kaneko, T., Li, R., He, Q., Yang, L., Ye, B. (2024). Transsynaptic BMP Signaling Regulates Fine-Scale Topography between Adjacent Sensory Neurons. eNeuro, 11(8) PubMed ID: 39137988
Summary:
Sensory axons projecting to the central nervous system are organized into topographic maps that represent the locations of sensory stimuli. In some sensory systems, even adjacent sensory axons are arranged topographically, forming "fine-scale" topographic maps. Although several broad molecular gradients are known to instruct coarse topography, little is known about the molecular signaling that regulates fine-scale topography at the level of two adjacent axons. This study provides evidence that transsynaptic bone morphogenetic protein (BMP) signaling mediates local interneuronal communication to regulate fine-scale topography in the nociceptive system of Drosophila larvae. The topographic separation of the axon terminals of adjacent nociceptors requires their common postsynaptic target, the A08n neurons. This phenotype is recapitulated by knockdown of the BMP ligand, Decapentaplegic (Dpp), in these neurons. In addition, removing the Type 2 BMP receptors or their effector (Mad transcription factor) in single nociceptors impairs the fine-scale topography, suggesting the contribution of BMP signaling originated from A08n. This signaling is likely mediated by phospho-Mad in the presynaptic terminals of nociceptors to ensure local interneuronal communication. Finally, reducing Dpp levels in A08n reduces the nociceptor-A08n synaptic contacts. These data support that transsynaptic BMP signaling establishes the fine-scale topography by facilitating the formation of topographically correct synapses. Local BMP signaling for synapse formation may be a developmental strategy that independently regulates neighboring axon terminals for fine-scale topography.
Boerner, J., Robbins, K., Murphey, R. (2024). Laser Cell Ablation in Intact Drosophila Larvae Reveals Synaptic Competition. J Vis Exp, (209) PubMed ID: 39141541
Summary:
The protocol describes single-neuron ablation with a 2-photon laser system in the central nervous system (CNS) of intact Drosophila melanogaster larvae. Using this non-invasive method, the developing nervous system can be manipulated in a cell-specific manner. Disrupting the development of individual neurons in a network can be used to study how the nervous system can compensate for the loss of synaptic input. Individual neurons were specifically ablated in the giant fiber system of Drosophila, with a focus on two neurons: the presynaptic giant fiber (GF) and the postsynaptic tergotrochanteral motor neuron (TTMn). The GF synapses with the ipsilateral TTMn, which is crucial to the escape response. Ablating one of the GFs in the 3(rd) instar brain, just after the GF starts axonal growth, permanently removes the cell during the development of the CNS. The remaining GF reacts to the absent neighbor and forms an ectopic synaptic terminal to the contralateral TTMn. This atypical, bilaterally symmetric terminal innervates both TTMns, as demonstrated by dye coupling, and drives both motor neurons, as demonstrated by electrophysiological assays. In summary, the ablation of a single interneuron demonstrates synaptic competition between a bilateral pair of neurons that can compensate for the loss of one neuron and restore normal responses to the escape circuit. 		Welch, L. G., Muschalik, N., Munro, S. (2024). The FAM114A proteins are adaptors for the recycling of Golgi enzymes. J Cell Sci, 137(17) PubMed ID: 39129673
Summary:
Golgi-resident enzymes remain in place while their substrates flow through from the endoplasmic reticulum to elsewhere in the cell. COPI-coated vesicles bud from the Golgi to recycle Golgi residents to earlier cisternae. Different enzymes are present in different parts of the stack, and one COPI adaptor protein, GOLPH3 (see Drosophila sauron), acts to recruit enzymes into vesicles in part of the stack. This study used proximity biotinylation to identify further components of intra-Golgi vesicles and found FAM114A2, a cytosolic protein. Affinity chromatography with FAM114A2, and its paralogue FAM114A1, showed that they bind to Golgi-resident membrane proteins, with membrane-proximal basic residues in the cytoplasmic tail being sufficient for the interaction. Deletion of both proteins from U2OS cells did not cause substantial defects in Golgi function. However, a Drosophila orthologue of these proteins (CG9590/FAM114A) is also localised to the Golgi and binds directly to COPI. Drosophila mutants lacking FAM114A have defects in glycosylation of glue proteins in the salivary gland. Thus, the FAM114A proteins bind Golgi enzymes and are candidate adaptors to contribute specificity to COPI vesicle recycling in the Golgi stack.
Xu, C., Li, Z., Lyu, C., Hu, Y., McLaughlin, C. N., Wong, K. K. L., Xie, Q., Luginbuhl, D. J., Li, H., Udeshi, N. D., Svinkina, T., Mani, D. R., Han, S., Li, T., Li, Y., Guajardo, R., Ting, A. Y., Carr, S. A., Li, J., Luo, L. (2024). Molecular and cellular mechanisms of teneurin signaling in synaptic partner matching. Cell, 187(18):5081-5101 PubMed ID: 38996528
Summary:
In developing brains, axons exhibit remarkable precision in selecting synaptic partners among many non-partner cells. Evolutionarily conserved teneurins are transmembrane proteins that instruct synaptic partner matching. However, how intracellular signaling pathways execute teneurins' functions is unclear. HThis study used in situ proximity labeling to obtain the intracellular interactome of a teneurin (Ten-m) in the Drosophila brain. Genetic interaction studies using quantitative partner matching assays in both olfactory receptor neurons (ORNs) and projection neurons (PNs) reveal a common pathway: Ten-m binds to and negatively regulates a RhoGAP, thus activating the Rac1 small GTPases to promote synaptic partner matching. Developmental analyses with single-axon resolution identify the cellular mechanism of synaptic partner matching: Ten-m signaling promotes local F-actin levels and stabilizes ORN axon branches that contact partner PN dendrites. Combining spatial proteomics and high-resolution phenotypic analyses, this study advanced understanding of both cellular and molecular mechanisms of synaptic partner matching. 		Mou, W., Cui, Y. (2024). Enhancing neuronal reticulophagy: a strategy for combating aging and APP toxicity. Autophagy, 20(12):2819-2820 PubMed ID: 38963012
Summary:
Reticulophagy, which directs the endoplasmic reticulum (ER) to the phagophore for sequestration within an autophagosome and subsequent lysosomal degradation via specific receptors, is essential for ER quality control and is implicated in various diseases. This study utilizes Drosophila to establish an in vivo model for reticulophagy. Starvation-induced reticulophagy is detected across multiple tissues in Drosophila. Whole-body upregulation or downregulation of the expression of reticulophagy receptors, atl and Rtnl1, negatively affects fly health. Notably, moderate upregulation of reticulophagy in neuronal tissues by overexpressing these receptors reduces age-related degeneration. In a Drosophila Alzheimer model expressing human APP (amyloid beta precursor protein), reticulophagy is compromised. Correcting reticulophagy by enhancing atl and Rtnl1 expression in the neurons promotes APP degradation, significantly reducing neurodegenerative symptoms. However, overexpression of mutated atl and Rtnl1 , which disrupts the interaction of the corresponding proteins with Atg8, does not alleviate these symptoms, emphasizing the importance of receptor functionality. These findings support modulating reticulophagy as a therapeutic strategy for aging and neurodegenerative diseases associated with ER protein accumulation.

Tuesday June 10th - Immune Response

Brantley, S. E., Stouthamer, C. M., Kr, P., Fischer, M. L., Hill, J., Schlenke, T. A., Mortimer, N. T. (2024). Host JAK-STAT activity is a target of parasitoid wasp virulence strategies. PLoS pathogens, 20(7):e1012349 PubMed ID: 38950076
Summary:
Innate immune responses that allow hosts to survive infection depend on the action of multiple conserved signaling pathways. Pathogens and parasites in turn have evolved virulence factors to target these immune signaling pathways in an attempt to overcome host immunity. Consequently, the interactions between host immune molecules and pathogen virulence factors play an important role in determining the outcome of an infection. The immune responses of Drosophila melanogaster provide a valuable model to understand immune signaling and host-pathogen interactions. Flies are commonly infected by parasitoid wasps and mount a coordinated cellular immune response following infection. This response is characterized by the production of specialized blood cells called lamellocytes that form a tight capsule around wasp eggs in the host hemocoel. The conserved JAK-STAT signaling pathway has been implicated in lamellocyte proliferation and is required for successful encapsulation of wasp eggs. Activity of Stat92E, the D. melanogaster STAT ortholog, is induced in immune tissues following parasitoid infection. Virulent wasp species are able to suppress Stat92E activity during infection, suggesting they target JAK-STAT pathway activation as a virulence strategy. Furthermore, two wasp species (Leptopilina guineaensis and Ganaspis xanthopoda) suppress phenotypes associated with a gain-of-function mutation in hopscotch, the D. melanogaster JAK ortholog, indicating that they inhibit the activity of the core signaling components of the JAK-STAT pathway. These data suggest that parasitoid wasp virulence factors block JAK-STAT signaling to overcome fly immune defenses. Chauhan, M., Martinak, P. E., Hollenberg, B. M., Goodman, A. G. (2024). Drosophila melanogaster Toll-9 elicits antiviral immunity against Drosophila C virus. bioRxiv, PubMed ID: 38948804
Summary:
The Toll pathway plays a pivotal role in innate immune responses against pathogens. The evolutionary conserved pathogen recognition receptors (PRRs), including Toll like receptors (TLRs), play a crucial role in recognition of pathogen associated molecular patterns (PAMPs). The Drosophila genome encodes nine Toll receptors that are orthologous to mammalian TLRs. While mammalian TLRs directly recognize PAMPs, most Drosophila Tolls recognize the proteolytically cleaved ligand Spatzle to activate downstream signaling cascades. This study demonstrated that Toll-9 is crucial for antiviral immunity against Drosophila C virus (DCV), a natural pathogen of Drosophila A transposable element insertion in the Toll-9 gene renders the flies more susceptible to DCV. The stable expression of Toll-9 in S2 cells confers resistance against DCV infection by upregulation of the RNAi pathway. Toll-9 promotes the dephosphorylation of AKT, resulting in the induction of antiviral RNAi genes to inhibit DCV replication. Toll-9 localizes to the endosome where it binds dsRNA, suggesting its role to detect viral dsRNA. Toll-9 also induces apoptosis during DCV infection, contributing to its antiviral role. Together, this work identifies the role of Toll-9 in antiviral immunity against DCV infection through its ability to bind dsRNA and induce AKT-mediated RNAi antiviral immunity.
Chasse, A. Y., Bandyadka, S., Wertheimer, M. C., Serizier, S. B., McCall, K. (2024). Professional phagocytes are recruited for the clearance of obsolete nonprofessional phagocytes in the Drosophila ovary. Frontiers in immunology 15:1389674 PubMed ID: 38994369
Summary:
Cell death is an important process in the body, as it occurs throughout every tissue during development, disease, and tissue regeneration. Phagocytes are responsible for clearing away dying cells and are typically characterized as either professional or nonprofessional phagocytes. Professional phagocytes, such as macrophages, are found in nearly every part of the body while nonprofessional phagocytes, such as epithelial cells, are found in every tissue type. However, there are organs that are considered "immune-privileged" as they have little to no immune surveillance and rely on nonprofessional phagocytes to engulf dying cells. These organs are surrounded by barriers to protect the tissue from viruses, bacteria, and perhaps even immune cells. The Drosophila ovary is considered immune-privileged, however the presence of hemocytes, the macrophages of Drosophila, around the ovary suggests they may have a potential function. This study analyze hemocyte localization and potential functions in response to starvation-induced cell death in the ovary. Hemocytes were found to accumulate in the oviduct in the vicinity of mature eggs and follicle cell debris. Genetic ablation of hemocytes revealed that the presence of hemocytes affects oogenesis and that they phagocytose ovarian cell debris and in their absence fecundity decreases. Unpaired3, an IL-6 like cytokine, was found to be required for the recruitment of hemocytes to the oviduct to clear away obsolete follicle cells. These findings demonstrate a role for hemocytes in the ovary, providing a more thorough understanding of phagocyte communication and cell clearance in a previously thought immune-privileged organ. 		Yao, X., He, Y., Zhu, C., Yang, S., Wu, J., Ma, F., Jin, P. (2024). miR-190 restores the innate immune homeostasis of Drosophila by directly inhibiting Tab2 in Imd pathway. Microbes and infection:105399 PubMed ID: 39084397
Summary:
The Drosophila Imd pathways are well-known mechanisms involved in innate immunity responsible for Gram-negative (G-) bacterial infection. The intensity and durability of immunity need to be finely regulated to keep sufficient immune activation meanwhile avoid excessive immune response. This study firstly demonstrated that miR-190 can downregulate the expression levels of antimicrobial peptides (AMPs) in the Imd immune pathway after Escherichia coli infection using the miR-190 overexpression flies and the miR-190KO/+ flies. Secondly, miR-190 overexpression significantly reduces while miR-190 KO increases Drosophila survival rates upon lethal Enterobacter cloacae infection. Thirdly, this study further demonstrated that miR-190 negatively regulates innate immune responses by directly targeting both RA/RB and RC isoforms of Tab2. In addition, the dynamic expression pattern of AMPs (Dpt, AttA, CecA1), miR-190 and Tab2 in the wild-type flies reveals that miR-190 play an important role in Drosophila immune homeostasis restoration at the late stage of E. coli infection. Collectively, this study reveals that miR-190 can downregulate the expression of AMPs by targeting Tab2 and promote immune homeostasis restoration in Drosophila Imd pathway. This study provides new insights into the regulatory mechanism of animal innate immune homeostasis.
Mallick, S., Kenney, E., Eleftherianos, I. (2024). The Activin Branch Ligand Daw Regulates the Drosophila melanogaster Immune Response and Lipid Metabolism against the Heterorhabditis bacteriophora Serine Carboxypeptidase. Int J Mol Sci, 25(14) PubMed ID: 39063211
Summary:
Despite impressive advances in the broad field of innate immunity, understanding of the molecules and signaling pathways that control the host immune response to nematode infection remains incomplete. Recent work has shown that Transforming Growth Factor-β (TGF-β) signaling in the fruit fly Drosophila melanogaster is activated by nematode infection and certain TGF-β superfamily members regulate the D. melanogaster anti-nematode immune response. This study investigated the effect of an entomopathogenic nematode infection factor on host TGF-β pathway regulation and immune function. Heterorhabditis bacteriophora serine carboxypeptidase activates the Activin branch in D. melanogaster adults and the immune deficiency pathway in Activin-deficient flies. The carboxypeptidase affects hemocyte numbers and survival in flies deficient for Activin signaling, and causes increased intestinal steatosis in Activin-deficient flies. Thus, insights into the D. melanogaster signaling pathways and metabolic processes interacting with H. bacteriophora pathogenicity factors will be applicable to entomopathogenic nematode infection of important agricultural insect pests and vectors of disease. 		Wang, X., Wei, D., Pan, Y., Liu, J., Xiao, X., Xia, Q., Wang, F. (2024). A cryptic homotypic interaction motif of insect STING is required for its antiviral signaling. Dev Comp Immunol, 159:105224 PubMed ID: 38969190
Summary:
Stimulator of interferon genes (STING) mediates innate immune response upon binding to cyclic GMP-AMP (cGAMP). It recruits tank-binding kinase 1 (TBK1) and transcription factor interferon regulatory factor 3 (IRF3) through its C-terminal tail and facilitates TBK1-dependent phosphorylation of IRF3 via forming STING polymers in mammalian cells. However, the mechanism behind STING-mediated activation of NF-kappaB transcription factor, Relish, in insect cells is unknown. This study revealed that insect STING formed oligomers and the cryptic RIP homotypic interaction motif (cRHIM) was required for its oligomerization and its anti-viral functions. Cells expressing cRHIM-deficient mutants exhibited lower levels of anti-viral molecules, higher viral load after viral infection and weak activation of Relish. Moreover, under cGAMP stimulation, insect STING interacted with IMD, and deletion of the cRHIM motif on either protein prevented this interaction. Finally, this study demonstrated that cGAMP enhanced the amyloid-like property of insect STING aggregates by ThT staining. In summary, this research showed that insect STING employed a homotypic motif to form intermolecular interactions that are essential for its antiviral signaling.
Brantley, S. E., Stouthamer, C. M., Kr, P., Fischer, M. L., Hill, J., Schlenke, T. A., Mortimer, N. T. (2024). Host JAK-STAT activity is a target of parasitoid wasp virulence strategies. PLoS pathogens, 20(7):e1012349 PubMed ID: 38950076
Summary:
Innate immune responses that allow hosts to survive infection depend on the action of multiple conserved signaling pathways. Pathogens and parasites in turn have evolved virulence factors to target these immune signaling pathways in an attempt to overcome host immunity. Consequently, the interactions between host immune molecules and pathogen virulence factors play an important role in determining the outcome of an infection. The immune responses of Drosophila melanogaster provide a valuable model to understand immune signaling and host-pathogen interactions. Flies are commonly infected by parasitoid wasps and mount a coordinated cellular immune response following infection. This response is characterized by the production of specialized blood cells called lamellocytes that form a tight capsule around wasp eggs in the host hemocoel. The conserved JAK-STAT signaling pathway has been implicated in lamellocyte proliferation and is required for successful encapsulation of wasp eggs. Activity of Stat92E, the D. melanogaster STAT ortholog, is induced in immune tissues following parasitoid infection. Virulent wasp species are able to suppress Stat92E activity during infection, suggesting they target JAK-STAT pathway activation as a virulence strategy. Furthermore, two wasp species (Leptopilina guineaensis and Ganaspis xanthopoda) suppress phenotypes associated with a gain-of-function mutation in hopscotch, the D. melanogaster JAK ortholog, indicating that they inhibit the activity of the core signaling components of the JAK-STAT pathway. These data suggest that parasitoid wasp virulence factors block JAK-STAT signaling to overcome fly immune defenses. 		Chasse, A. Y., Bandyadka, S., Wertheimer, M. C., Serizier, S. B., McCall, K. (2024). Professional phagocytes are recruited for the clearance of obsolete nonprofessional phagocytes in the Drosophila ovary. Frontiers in immunology 15:1389674 PubMed ID: 38994369
Summary:
Cell death is an important process in the body, as it occurs throughout every tissue during development, disease, and tissue regeneration. Phagocytes are responsible for clearing away dying cells and are typically characterized as either professional or nonprofessional phagocytes. Professional phagocytes, such as macrophages, are found in nearly every part of the body while nonprofessional phagocytes, such as epithelial cells, are found in every tissue type. However, there are organs that are considered "immune-privileged" as they have little to no immune surveillance and rely on nonprofessional phagocytes to engulf dying cells. These organs are surrounded by barriers to protect the tissue from viruses, bacteria, and perhaps even immune cells. The Drosophila ovary is considered immune-privileged, however the presence of hemocytes, the macrophages of Drosophila, around the ovary suggests they may have a potential function. This study analyze hemocyte localization and potential functions in response to starvation-induced cell death in the ovary. Hemocytes were found to accumulate in the oviduct in the vicinity of mature eggs and follicle cell debris. Genetic ablation of hemocytes revealed that the presence of hemocytes affects oogenesis and that they phagocytose ovarian cell debris and in their absence fecundity decreases. Unpaired3, an IL-6 like cytokine, was found to be required for the recruitment of hemocytes to the oviduct to clear away obsolete follicle cells. These findings demonstrate a role for hemocytes in the ovary, providing a more thorough understanding of phagocyte communication and cell clearance in a previously thought immune-privileged organ.
Yao, X., He, Y., Zhu, C., Yang, S., Wu, J., Ma, F., Jin, P. (2024). miR-190 restores the innate immune homeostasis of Drosophila by directly inhibiting Tab2 in Imd pathway. Microbes and infection:105399 PubMed ID: 39084397
Summary:
The Drosophila Imd pathways are well-known mechanisms involved in innate immunity responsible for Gram-negative (G-) bacterial infection. The intensity and durability of immunity need to be finely regulated to keep sufficient immune activation meanwhile avoid excessive immune response. This study firstly demonstrated that miR-190 can downregulate the expression levels of antimicrobial peptides (AMPs) in the Imd immune pathway after Escherichia coli infection using the miR-190 overexpression flies and the miR-190KO/+ flies. Secondly, miR-190 overexpression significantly reduces while miR-190 KO increases Drosophila survival rates upon lethal Enterobacter cloacae infection. Thirdly, this study further demonstrated that miR-190 negatively regulates innate immune responses by directly targeting both RA/RB and RC isoforms of Tab2. In addition, the dynamic expression pattern of AMPs (Dpt, AttA, CecA1), miR-190 and Tab2 in the wild-type flies reveals that miR-190 play an important role in Drosophila immune homeostasis restoration at the late stage of E. coli infection. Collectively, this study reveals that miR-190 can downregulate the expression of AMPs by targeting Tab2 and promote immune homeostasis restoration in Drosophila Imd pathway. This study provides new insights into the regulatory mechanism of animal innate immune homeostasis. 		Chauhan, M., Martinak, P. E., Hollenberg, B. M., Goodman, A. G. (2024). Drosophila melanogaster Toll-9 elicits antiviral immunity against Drosophila C virus. bioRxiv, PubMed ID: 38948804
Summary:
The Toll pathway plays a pivotal role in innate immune responses against pathogens. The evolutionary conserved pathogen recognition receptors (PRRs), including Toll like receptors (TLRs), play a crucial role in recognition of pathogen associated molecular patterns (PAMPs). The Drosophila genome encodes nine Toll receptors that are orthologous to mammalian TLRs. While mammalian TLRs directly recognize PAMPs, most Drosophila Tolls recognize the proteolytically cleaved ligand Spatzle to activate downstream signaling cascades. This study demonstrated that Toll-9 is crucial for antiviral immunity against Drosophila C virus (DCV), a natural pathogen of Drosophila A transposable element insertion in the Toll-9 gene renders the flies more susceptible to DCV. The stable expression of Toll-9 in S2 cells confers resistance against DCV infection by upregulation of the RNAi pathway. Toll-9 promotes the dephosphorylation of AKT, resulting in the induction of antiviral RNAi genes to inhibit DCV replication. Toll-9 localizes to the endosome where it binds dsRNA, suggesting its role to detect viral dsRNA. Toll-9 also induces apoptosis during DCV infection, contributing to its antiviral role. Together, this work identifies the role of Toll-9 in antiviral immunity against DCV infection through its ability to bind dsRNA and induce AKT-mediated RNAi antiviral immunity.
Mallick, S., Kenney, E., Eleftherianos, I. (2024). The Activin Branch Ligand Daw Regulates the Drosophila melanogaster Immune Response and Lipid Metabolism against the Heterorhabditis bacteriophora Serine Carboxypeptidase. Int J Mol Sci, 25(14) PubMed ID: 39063211
Summary:
w Despite impressive advances in the broad field of innate immunity, understanding of the molecules and signaling pathways that control the host immune response to nematode infection remains incomplete. Recent work has shown that Transforming Growth Factor-β (TGF-β) signaling in the fruit fly Drosophila melanogaster is activated by nematode infection and certain TGF-β superfamily members regulate the D. melanogaster anti-nematode immune response. This study investigated the effect of an entomopathogenic nematode infection factor on host TGF-β pathway regulation and immune function. Heterorhabditis bacteriophora serine carboxypeptidase activates the Activin branch in D. melanogaster adults and the immune deficiency pathway in Activin-deficient flies. The carboxypeptidase affects hemocyte numbers and survival in flies deficient for Activin (Dawdle) signaling, and causes increased intestinal steatosis in Activin-deficient flies. Thus, insights into the D. melanogaster signaling pathways and metabolic processes interacting with H. bacteriophora pathogenicity factors will be applicable to entomopathogenic nematode infection of important agricultural insect pests and vectors of disease. 		Wang, X., Wei, D., Pan, Y., Liu, J., Xiao, X., Xia, Q., Wang, F. (2024). A cryptic homotypic interaction motif of insect STING is required for its antiviral signaling. Dev Comp Immunol, 159:105224 PubMed ID: 38969190
Summary:
Stimulator of interferon genes (STING) mediates innate immune response upon binding to cyclic GMP-AMP (cGAMP). It recruits tank-binding kinase 1 (TBK1) and transcription factor interferon regulatory factor 3 (IRF3) through its C-terminal tail and facilitates TBK1-dependent phosphorylation of IRF3 via forming STING polymers in mammalian cells. However, the mechanism behind STING-mediated activation of NF-kappaB transcription factor, Relish, in insect cells is unknown. This study revealed that insect STING formed oligomers and the cryptic RIP homotypic interaction motif (cRHIM) was required for its oligomerization and its anti-viral functions. Cells expressing cRHIM-deficient mutants exhibited lower levels of anti-viral molecules, higher viral load after viral infection and weak activation of Relish. Moreover, under cGAMP stimulation, insect STING interacted with IMD, and deletion of the cRHIM motif on either protein prevented this interaction. Finally, this study demonstrated that cGAMP enhanced the amyloid-like property of insect STING aggregates by ThT staining. In summary, this research showed that insect STING employed a homotypic motif to form intermolecular interactions that are essential for its antiviral signaling.

Monday, June 9th - Enhancers and Transcriptional regulation

Hodkinson, L. J., Rieder, L. E. (2024). Cis element length variability does not confer differential transcription factor occupancy at the D. melanogaster histone locus. bioRxiv, PubMed ID: 38979213
Summary:
Histone genes require precise regulation to maintain histone homeostasis and ensure nucleosome stoichiometry. Animal histone genes often have unique clustered genomic organization. However, there is variability of histone gene number and organization as well as differential regulation of the histone genes across species. The Drosophila melanogaster histone locus has unique organizational characteristics as it exists as a series of ~100 highly regular, tandemly repeated arrays of the 5 replication-dependent histone genes at a single locus. Yet D. melanogaster are viable with only 12 transgenic histone gene arrays. It was hypothesized that the histone genes across the locus are differentially regulated. The GA-repeat within the H3/H4 promoter was discovered to be the only variable sequence across the histone gene arrays. The H3/H4 promoter GA-repeat is targeted by CLAMP to promote histone gene expression. This study also showed two additional GA-binding transcription factors, GAGA Factor and Pipsqueak, target the GA-repeat. When CLAMP and GAF targeting was examined, neither CLAMP nor GAF show bias for any GA-repeat lengths. Furthermore, it was found that the distribution of GA-repeats targeted by both CLAMP and GAF do not change throughout early development. Together the results suggest that the transcription factors targeting the H3/H4 GA-repeat do not impact differential regulation of the histone genes, but indicate that future studies should interrogate additional cis elements or factors that impact histone gene regulation. 		Fogarty, E. A., Buchert, E. M., Ma, Y., Nicely, A. B., Buttitta, L. A. (2024). Transcriptional repression and enhancer decommissioning silence cell cycle genes in postmitotic tissues. bioRxiv, PubMed ID: 38766255
Summary:
The mechanisms that maintain a non-cycling status in postmitotic tissues are not well understood. Many cell cycle genes have promoters and enhancers that remain accessible even when cells are terminally differentiated and in a non-cycling state, suggesting their repression must be maintained long term. In contrast, enhancer decommissioning has been observed for rate-limiting cell cycle genes in the Drosophila wing, a tissue where the cells die soon after eclosion, but it has been unclear if this also occurs in other contexts of terminal differentiation. This study shows that enhancer decommissioning also occurs at specific, rate-limiting cell cycle genes in the long-lived tissues of the Drosophila eye and brain, and it is proposed that this loss of chromatin accessibility may help maintain a robust postmitotic state.The decommissioned enhancers were examined at specific rate-limiting cell cycle genes; they were shown to encode dynamic temporal and spatial expression patterns that include shared, as well as tissue-specific elements, resulting in broad gene expression with developmentally controlled temporal regulation. This analysis was extended to cell cycle gene expression and chromatin accessibility in the mammalian retina using a published dataset, and the principles of cell cycle gene regulation identified in terminally differentiating Drosophila tissues were found to be conserved in the differentiating mammalian retina. A robust, non-cycling status is proposed to be maintained in long-lived postmitotic tissues through a combination of stable repression at most cell cycle gens, alongside enhancer decommissioning at specific rate-limiting cell cycle genes.
Versluis, P., Graham, T. G. W., Eng, V., Ebenezer, J., Darzacq, X., Zipfel, W. R., Lis, J. T. (2024). Live-cell imaging of RNA Pol II and elongation factors distinguishes competing mechanisms of transcription regulation. Mol Cell, 84(15):2856-2869 PubMed ID: 39121843
Summary:
RNA polymerase II (RNA Pol II)-mediated transcription is a critical, highly regulated process aided by protein complexes at distinct steps. To investigate RNA Pol II and transcription-factor-binding and dissociation dynamics, endogenous photoactivatable-GFP (PA-GFP) and HaloTag knockins were constructed using CRISPR-Cas9, allowing tracking a population of molecules at the induced .htm">Hsp70 loci in Drosophila melanogaster polytene chromosomes. Early in the heat-shock response, little RNA Pol II and DRB sensitivity-inducing factor (DSIF) are reused for iterative rounds of transcription. Surprisingly, although PAF1 and Spt6 are found throughout the gene body by chromatin immunoprecipitation (ChIP) assays, they show markedly different binding behaviors. Additionally, PAF1 and Spt6 are only recruited after positive transcription elongation factor (P-TEFb)-mediated phosphorylation and RNA Pol II promoter-proximal pause escape. Finally, PAF1 may be expendable for transcription of highly expressed genes where nucleosome density is low. Thus, live-cell imaging data provide key constraints to mechanistic models of transcription regulation. 		Sloutskin, A., Itzhak, D., Vogler, G., Pozeilov, H., Ideses, D., Alter, H., Adato, O., Shachar, H., Doniger, T., Shohat-Ophir, G., Frasch, M., Bodmer, R., Duttke, S. H., Juven-Gershon, T. (2024). From promoter motif to cardiac function: a single DPE motif affects transcription regulation and organ function in vivo. Development, 151(14) PubMed ID: 38958007
Summary:
Transcription initiates at the core promoter, which contains distinct core promoter elements. This study highlights the complexity of transcriptional regulation by outlining the effect of core promoter-dependent regulation on embryonic development and the proper function of an organism. The importance is demonstrate in vivo of the downstream core promoter element (DPE) in complex heart formation in Drosophila. Pioneering a novel approach using both CRISPR and nascent transcriptomics, the effects are shown of mutating a single core promoter element within the natural context. Specifically, the downstream core promoter element (DPE) was targeted of the endogenous tin gene, encoding the Tinman transcription factor, a homologue of human NKX2-5 associated with congenital heart diseases. The 7 bp substitution mutation results in massive perturbation of the Tinman regulatory network that orchestrates dorsal musculature, which is manifested as physiological and anatomical changes in the cardiac system, impaired specific activity features, and significantly compromised viability of adult flies. Thus, a single motif can have a critical impact on embryogenesis and, in the case of DPE, functional heart formation.
Hodkinson, L. J., Gross, J., Schmidt, C. A., Diaz-Saldana, P. P., Aoki, T., Rieder, L. E. (2024). Sequence reliance of the Drosophila context-dependent transcription factor CLAMP. Genetics, 227(3) PubMed ID: 38775472
Summary:
Despite binding similar cis elements in multiple locations, a single transcription factor (TF) often performs context-dependent functions at different loci. How factors integrate cis sequence and genomic context is still poorly understood and has implications for off-target effects in genetic engineering. The Drosophila context-dependent TF chromatin-linked adaptor for male-specific lethal proteins (CLAMP) targets similar GA-rich cis elements on the X-chromosome and at the histone gene locus but recruits very different, locus-specific factors. This study discovered that CLAMP leverages information from both cis element and local sequence to perform context-specific functions. These observations imply the importance of other cues, including protein-protein interactions and the presence of additional cofactors. 		DeHaro-Arbona, F. J., Roussos, C., Baloul, S., Townson, J., Gomez Lamarca, M. J., Bray, S. (2024). Dynamic modes of Notch transcription hubs conferring memory and stochastic activation revealed by live imaging the co-activator Mastermind. Elife, 12 PubMed ID: 38727722 Developmental programming involves the accurate conversion of signalling levels and dynamics to transcriptional outputs. The transcriptional relay in the Notch pathway relies on nuclear complexes containing the co-activator Mastermind (Mam). By tracking these complexes in real time, this study revealed that they promote the formation of a dynamic transcription hub in Notch ON nuclei which concentrates key factors including the Mediator CDK module. The composition of the hub is labile and persists after Notch withdrawal conferring a memory that enables rapid reformation. Surprisingly, only a third of Notch ON hubs progress to a state with nascent transcription, which correlates with polymerase II and core Mediator recruitment. This probability is increased by a second signal. The discovery that target-gene transcription is probabilistic has far-reaching implications because it implies that stochastic differences in Notch pathway output can arise downstream of receptor activation.

Friday, June 6th - Synapse and Vesicles

Chang, Y. C., Peng, Y. J., Lee, J. Y., Chag, K. T. (2024). Peripheral glia and neurons jointly regulate activity-induced synaptic remodeling at the Drosophila neuromuscular junction. bioRxiv, PubMed ID: 39005352
Summary:
In the nervous system, reliable communication depends on the ability of neurons to adaptively remodel their synaptic structure and function in response to changes in neuronal activity. While neurons are the main drivers of synaptic plasticity, glial cells are increasingly recognized for their roles as active modulators. However, the underlying molecular mechanisms remain unclear. In this study, using Drosophila neuromuscular junction as a model system for a tripartite synapse, this study showed that peripheral glial cells collaborate with neurons at the NMJ to regulate activity-induced synaptic remodeling, in part through a protein called shriveled (Shv). Shv is an activator of integrin signaling previously shown to be released by neurons during intense stimulation at the fly NMJ to regulate activity-induced synaptic remodeling. This study demonstrate that Shv is also present in peripheral glia, and glial Shv is both necessary and sufficient for synaptic remodeling. However, unlike neuronal Shv, glial Shv does not activate integrin signaling at the NMJ. Instead, it regulates synaptic plasticity in two ways: 1) maintaining the extracellular balance of neuronal Shv proteins to regulate integrin signaling, and 2) controlling ambient extracellular glutamate concentration to regulate postsynaptic glutamate receptor abundance. Loss of glial cells showed the same phenotype as loss of Shv in glia. Together, these results reveal that neurons and glial cells homeostatically regulate extracellular Shv protein levels to control activity-induced synaptic remodeling. Additionally, peripheral glia maintains postsynaptic glutamate receptor abundance and contribute to activity-induced synaptic remodeling by regulating ambient glutamate concentration at the fly NMJ. 		Kaneko, T., Li, R., He, Q., Yang, L., Ye, B. (2024). Transsynaptic BMP Signaling Regulates Fine-Scale Topography between Adjacent Sensory Neurons. eNeuro, 11(8) PubMed ID: 39137988
Summary:
Sensory axons projecting to the central nervous system are organized into topographic maps that represent the locations of sensory stimuli. In some sensory systems, even adjacent sensory axons are arranged topographically, forming "fine-scale" topographic maps. Although several broad molecular gradients are known to instruct coarse topography, little is known about the molecular signaling that regulates fine-scale topography at the level of two adjacent axons. This study provides evidence that transsynaptic bone morphogenetic protein (BMP) signaling mediates local interneuronal communication to regulate fine-scale topography in the nociceptive system of Drosophila larvae. The topographic separation of the axon terminals of adjacent nociceptors requires their common postsynaptic target, the A08n neurons. This phenotype is recapitulated by knockdown of the BMP ligand, Decapentaplegic (Dpp), in these neurons. In addition, removing the Type 2 BMP receptors or their effector (Mad transcription factor) in single nociceptors impairs the fine-scale topography, suggesting the contribution of BMP signaling originated from A08n. This signaling is likely mediated by phospho-Mad in the presynaptic terminals of nociceptors to ensure local interneuronal communication. Finally, reducing Dpp levels in A08n reduces the nociceptor-A08n synaptic contacts. These data support that transsynaptic BMP signaling establishes the fine-scale topography by facilitating the formation of topographically correct synapses. Local BMP signaling for synapse formation may be a developmental strategy that independently regulates neighboring axon terminals for fine-scale topography.
Boerner, J., Robbins, K., Murphey, R. (2024). Laser Cell Ablation in Intact Drosophila Larvae Reveals Synaptic Competition. J Vis Exp, (209) PubMed ID: 39141541
Summary:
The protocol describes single-neuron ablation with a 2-photon laser system in the central nervous system (CNS) of intact Drosophila melanogaster larvae. Using this non-invasive method, the developing nervous system can be manipulated in a cell-specific manner. Disrupting the development of individual neurons in a network can be used to study how the nervous system can compensate for the loss of synaptic input. Individual neurons were specifically ablated in the giant fiber system of Drosophila, with a focus on two neurons: the presynaptic giant fiber (GF) and the postsynaptic tergotrochanteral motor neuron (TTMn). The GF synapses with the ipsilateral TTMn, which is crucial to the escape response. Ablating one of the GFs in the 3(rd) instar brain, just after the GF starts axonal growth, permanently removes the cell during the development of the CNS. The remaining GF reacts to the absent neighbor and forms an ectopic synaptic terminal to the contralateral TTMn. This atypical, bilaterally symmetric terminal innervates both TTMns, as demonstrated by dye coupling, and drives both motor neurons, as demonstrated by electrophysiological assays. In summary, the ablation of a single interneuron demonstrates synaptic competition between a bilateral pair of neurons that can compensate for the loss of one neuron and restore normal responses to the escape circuit. 		Welch, L. G., Muschalik, N., Munro, S. (2024). The FAM114A proteins are adaptors for the recycling of Golgi enzymes. J Cell Sci, 137(17) PubMed ID: 39129673
Summary:
Golgi-resident enzymes remain in place while their substrates flow through from the endoplasmic reticulum to elsewhere in the cell. COPI-coated vesicles bud from the Golgi to recycle Golgi residents to earlier cisternae. Different enzymes are present in different parts of the stack, and one COPI adaptor protein, GOLPH3 (see Drosophila sauron), acts to recruit enzymes into vesicles in part of the stack. This study used proximity biotinylation to identify further components of intra-Golgi vesicles and found FAM114A2, a cytosolic protein. Affinity chromatography with FAM114A2, and its paralogue FAM114A1, showed that they bind to Golgi-resident membrane proteins, with membrane-proximal basic residues in the cytoplasmic tail being sufficient for the interaction. Deletion of both proteins from U2OS cells did not cause substantial defects in Golgi function. However, a Drosophila orthologue of these proteins (CG9590/FAM114A) is also localised to the Golgi and binds directly to COPI. Drosophila mutants lacking FAM114A have defects in glycosylation of glue proteins in the salivary gland. Thus, the FAM114A proteins bind Golgi enzymes and are candidate adaptors to contribute specificity to COPI vesicle recycling in the Golgi stack.
Xu, C., Li, Z., Lyu, C., Hu, Y., McLaughlin, C. N., Wong, K. K. L., Xie, Q., Luginbuhl, D. J., Li, H., Udeshi, N. D., Svinkina, T., Mani, D. R., Han, S., Li, T., Li, Y., Guajardo, R., Ting, A. Y., Carr, S. A., Li, J., Luo, L. (2024). Molecular and cellular mechanisms of teneurin signaling in synaptic partner matching. Cell, 187(18):5081-5101 PubMed ID: 38996528
Summary:
In developing brains, axons exhibit remarkable precision in selecting synaptic partners among many non-partner cells. Evolutionarily conserved teneurins are transmembrane proteins that instruct synaptic partner matching. However, how intracellular signaling pathways execute teneurins' functions is unclear. HThis study used in situ proximity labeling to obtain the intracellular interactome of a teneurin (Ten-m) in the Drosophila brain. Genetic interaction studies using quantitative partner matching assays in both olfactory receptor neurons (ORNs) and projection neurons (PNs) reveal a common pathway: Ten-m binds to and negatively regulates a RhoGAP, thus activating the Rac1 small GTPases to promote synaptic partner matching. Developmental analyses with single-axon resolution identify the cellular mechanism of synaptic partner matching: Ten-m signaling promotes local F-actin levels and stabilizes ORN axon branches that contact partner PN dendrites. Combining spatial proteomics and high-resolution phenotypic analyses, this study advanced understanding of both cellular and molecular mechanisms of synaptic partner matching. 		Mou, W., Cui, Y. (2024). Enhancing neuronal reticulophagy: a strategy for combating aging and APP toxicity. Autophagy, 20(12):2819-2820 PubMed ID: 38963012
Summary:
Reticulophagy, which directs the endoplasmic reticulum (ER) to the phagophore for sequestration within an autophagosome and subsequent lysosomal degradation via specific receptors, is essential for ER quality control and is implicated in various diseases. This study utilizes Drosophila to establish an in vivo model for reticulophagy. Starvation-induced reticulophagy is detected across multiple tissues in Drosophila. Whole-body upregulation or downregulation of the expression of reticulophagy receptors, atl and Rtnl1, negatively affects fly health. Notably, moderate upregulation of reticulophagy in neuronal tissues by overexpressing these receptors reduces age-related degeneration. In a Drosophila Alzheimer model expressing human APP (amyloid beta precursor protein), reticulophagy is compromised. Correcting reticulophagy by enhancing atl and Rtnl1 expression in the neurons promotes APP degradation, significantly reducing neurodegenerative symptoms. However, overexpression of mutated atl and Rtnl1 , which disrupts the interaction of the corresponding proteins with Atg8, does not alleviate these symptoms, emphasizing the importance of receptor functionality. These findings support modulating reticulophagy as a therapeutic strategy for aging and neurodegenerative diseases associated with ER protein accumulation.

Wednesday June 4th - Disease Models

Yuan, Y., Yu, L., Zhuang, X., Wen, D., He, J., Hong, J., Xie, J., Ling, S., Du, X., Chen, W., Wang, X. (2025). Drosophila models used to simulate human ATP1A1 gene mutations that cause Charcot-Marie-Tooth type 2 disease and refractory seizures. Neural Regen Res, 20(1):265-276 PubMed ID: 38767491
Summary:
Certain amino acids changes in the human Na+/K+-ATPase pump, ATPase Na+/K+ transporting subunit alpha 1 (ATP1A1), cause Charcot-Marie-Tooth disease type 2 (CMT2) disease and refractory seizures. To develop in vivo models to study the role of Na+/K+-ATPase in these diseases, the Drosophila gene homolog, Atpα, was modified to mimic the human ATP1A1 gene mutations that cause CMT2. Mutations located within the helical linker region of human ATP1A1 (I592T, A597T, P600T, and D601F) were simultaneously introduced into endogenous Drosophila Atpα by CRISPR/Cas9-mediated genome editing, generating the AtpαTTTF model. In addition, the same strategy was used to generate the corresponding single point mutations in flies (AtpαI571T, AtpαA576T, AtpαP579T, and AtpαD580F). Moreover, a deletion mutation (Atpαmut) that causes premature termination of translation was generated as a positive control. Of these alleles, two were found that could be maintained as homozygotes (AtpαI571T and AtpαP579T). Three alleles (AtpαA576T, AtpαP579 and AtpαD580F) can form heterozygotes with the Atpαmut allele. The Atpα allele carrying these CMT2-associated mutations showed differential phenotypes in Drosophila. Flies heterozygous for AtpαTTTF mutations have motor performance defects, a reduced lifespan, seizures, and an abnormal neuronal morphology. These Drosophila models will provide a new platform for studying the function and regulation of the sodium-potassium pump. 		Catterson, J. H., Mouofo, E. N., Lopez De Toledo Soler, I., Lean, G., Dlamini, S., Liddell, P., Voong, G., Katsinelos, T., Wang, Y. C., Schoovaerts, N., Verstreken, P., Spires-Jones, T. L., Durrant, C. S. (2024). Drosophila appear resistant to trans-synaptic tau propagation. Brain communications, 6(4):fcae256 PubMed ID: 39130515
Summary:
Alzheimer's disease is the most common cause of dementia in the elderly, prompting extensive efforts to pinpoint novel therapeutic targets for effective intervention. Among the hallmark features of Alzheimer's disease is the development of neurofibrillary tangles comprised of hyperphosphorylated tau protein, whose progressive spread throughout the brain is associated with neuronal death. Trans-synaptic propagation of tau has been observed in mouse models, and indirect evidence for tau spread via synapses has been observed in human Alzheimer's disease. Halting tau propagation is a promising therapeutic target for Alzheimer's disease; thus, a scalable model system to screen for modifiers of tau spread would be very useful for the field. To this end, this study sought to emulate the trans-synaptic spread of human tau in Drosophila melanogaster. Employing the trans-Tango circuit mapping technique, this study investigated whether tau spreads between synaptically connected neurons. Immunohistochemistry and confocal imaging were used to look for tau propagation. Examination of hundreds of flies expressing four different human tau constructs in two distinct neuronal populations reveals a robust resistance in Drosophila to the trans-synaptic spread of human tau. This resistance persisted in lines with concurrent expression of amyloid-β, in lines with global human tau knock-in to provide a template for human tau in downstream neurons, and with manipulations of temperature. These negative data are important for the field as this study established that Drosophila expressing human tau in subsets of neurons are unlikely to be useful to perform screens to find mechanisms to reduce the trans-synaptic spread of tau. The inherent resistance observed in Drosophila may serve as a valuable clue, offering insights into strategies for impeding tau spread in future studies.
Lewis, S. A., Forstrom, J., Tavani, J., Schafer, R., Tiede, Z., Padilla-Lopez, S. R., Kruer, M. C. (2024). eIF2alpha phosphorylation evokes dystonia-like movements with D2-receptor and cholinergic origin and abnormal neuronal connectivity. bioRxiv, PubMed ID: 38798458
Summary:
Dystonia is the 3(rd) most common movement disorder. Dystonia is acquired through either injury or genetic mutations, with poorly understood molecular and cellular mechanisms. Eukaryotic initiation factor alpha (eIF2α) controls cell state including neuronal plasticity via protein translation control and expression of ATF4. Dysregulated eIF2α phosphorylation (eIF2α-P) occurs in dystonia patients and models including DYT1 (Drosophila Torsin), but the consequences are unknown. This study increased/decreased eIF2α-P and tested motor control and neuronal properties in a Drosophila model. Bidirectionally altering eIF2α-P produced dystonia-like abnormal posturing and dyskinetic movements in flies. These movements were also observed with expression of the DYT1 risk allele. Cholinergic and D2-receptor neuroanatomical origins were identified of these dyskinetic movements caused by genetic manipulations to dystonia molecular candidates eIF2α-P, ATF4, or DYT1, with evidence for decreased cholinergic release. In vivo, increased and decreased eIF2α-P increase synaptic connectivity at the NMJ with increased terminal size and bouton synaptic release sites. Long-term treatment of elevated eIF2α-P with ISRIB restored adult longevity, but not performance in a motor assay. Disrupted eIF2α-P signaling may alter neuronal connectivity, change synaptic release, and drive motor circuit changes in dystonia. 		Hendricks, E. L., Linskey, N., Smith, I. R., Liebl, F. L. W. (2024). Kismet/CHD7/CHD8 and Amyloid Precursor Protein-like Regulate Synaptic Levels of Rab11 at the Drosophila Neuromuscular Junction. Int J Mol Sci, 25(15) PubMed ID: 39125997
Summary:
The transmembrane protein β-amyloid precursor protein (APP) is central to the pathophysiology of Alzheimer's disease (AD). The β-amyloid hypothesis posits that aberrant processing of APP forms neurotoxic β-amyloid aggregates, which lead to the cognitive impairments observed in AD. Although numerous additional factors contribute to AD, there is a need to better understand the synaptic function of APP. This study found that Drosophila APP-like (APPL) has both shared and non-shared roles at the synapse with Kismet (Kis), a chromatin helicase binding domain (CHD) protein. Kis is the homolog of CHD7 and CHD8, both of which are implicated in neurodevelopmental disorders including CHARGE Syndrome and autism spectrum disorders, respectively. Loss of function mutations in kis> and animals expressing human APP and BACE in their central nervous system show reductions in the glutamate receptor subunit, GluRIIC, the GTPase Rab11, and the bone morphogenetic protein (BMP), pMad, at the Drosophila larval neuromuscular junction (NMJ). Similarly, processes like endocytosis, larval locomotion, and neurotransmission are deficient in these animals. These pharmacological and epistasis experiments indicate that there is a functional relationship between Kis and APPL, but Kis does not regulate appl expression at the larval NMJ. Instead, Kis likely influences the synaptic localization of APPL, possibly by promoting rab11 transcription. These data identify a potential mechanistic connection between chromatin remodeling proteins and aberrant synaptic function in AD.
Nair, S., Jiang, Y., Marchal, I. S., Chernobelsky, E., Huang, H. W., Suh, S., Pan, R., Kong, X. P., Ryoo, H. D., Sigurdsson, E. M. (2024). Anti-tau single domain antibodies clear pathological tau and attenuate its toxicity and related functional defects. Cell Death Dis, 15(7):543 PubMed ID: 39079958
Summary:
Tauopathies are a group of neurodegenerative diseases characterized by the presence of tau inclusions. This study has developed over fifty anti-tau single-domain antibodies (sdAbs) derived from phage display libraries of a llama immunized with recombinant and pathological tau immunogens. This study examined the therapeutic potential of four of these sdAbs in a Drosophila tauopathy model following their transgenic expression either in all neurons or neuronal subtypes. Three of these sdAbs showed therapeutic potential in various assays, effectively clearing pathological tau and attenuating or preventing tau-induced phenotypes that typically manifest as defects in neuronal axonal transport, neurodegeneration, functional impairments, and shortened lifespan. Of these three, one sdAb was superior in every assay, which may at least in part be attributed to its tau-binding epitope. These findings support its development as a gene therapy for tauopathies. 		Singh, M. K., Ryu, T. H., Nguyen, M. N., Yu, K. (2024). Inhibition of high-fat diet-induced miRNA ameliorates tau toxicity in Drosophila. Biochem Biophys Res Commun, 733:150446 PubMed ID: 39067249
Summary:
Alzheimer's disease (AD), caused by amyloid β (Aβ) plaques and Tau tangles, is a neurodegenerative disease characterized by progressive memory impairment and cognitive dysfunction. High-fat diet (HFD), which induces type 2 diabetes, exacerbates Aβ plaque deposition in the brain. To investigate the function of HFD in Tau-mediated AD, an HFD was fed to the Drosophila Tau model and found that HFD aggravates Tau-induced neurological phenotypes. Since microRNAs (miRNAs) are biomarkers for diabetes and AD, the expression levels of common miRNAs of HFD and AD was evaluated in HFD-fed Tau model fly brains. Among the common miRNAs, the expression levels of Let-7 and miR-34 were increased. The inhibition of these miRNAs alleviates Tau-mediated AD phenotypes. This research provides valuable insights into how HFD accelerates tau toxicity. Additionally, this work highlights the therapeutic potential of targeting Let-7 and miR-34 to develop innovative treatment approaches for AD.

Tuesday June 3rd - Adult physiology, metabolism, and homeostasis

Xie, N., Bickley, B. A., Gross, A. D. (2024). GABA-gated chloride channel mutation (Rdl) induces cholinergic physiological compensation resulting in cross resistance in Drosophila melanogaster. Pesticide biochemistry and physiology, 203:105972 PubMed ID: 39084765
Summary:
The Drosophila melanogaster MD-RR strain contains an Rdl mutation (A301S) resulting in resistance to several insecticide classes viz. phenyl pyrazoles (e.g., fipronil), cyclodienes (e.g., dieldrin), and chlorinated aliphatic hydrocarbons (e.g., lindane). Fitness costs are commonly observed with resistant insect populations as side effects of the genetic change conferring the resistant phenotype. Because of fitness costs, reversion from the resistant to susceptible genotype and phenotype is common. However, the Rdl genotype in D. melanogaster appears to allow the flies to maintain the resistant genotype/phenotype without selective pressure and with minimal fitness costs. Evidence is provided that compensation for the Rdl mutation influences the cholinergic system, where an increase in acetylcholineesterase gene expression and enzyme activity results in neurophysiological changes and cross resistance to a carbamate insecticide (propoxur oral resistance ratio (RR) of 63) and an organophosphate insecticide (dichlorvos oral RR of 7). Such cross resistance was not previously reported with the initial collection and testing of this strain. In addition to acetylcholinesterase, the Rdl mutation influences the expression of the muscarinic acetylcholine receptor subtype-B, resulting in resistance to non-selective muscarinic compounds (pilocarpine and atropine). Collectively, these results indicate that the Rdl mutation (A301S) at GABA-gated ionophore complex influences the physiology of the cholinergic system, leading to resistance to established insecticide classes. Additionally, this mutation may impact the effectiveness of insecticides targeting novel sites, like muscarinic receptors. 		Zhao, B., Luo, J., Wang, H., Li, Y., Li, D., Bi, X. (2024). In vivo RNAi screening identifies multiple deubiquitinases required for the maintenance of intestinal homeostasis in Drosophila. Insect biochemistry and molecular biology, 172:104162 PubMed ID: 39067716
Summary:
Deubiquitinases (DUBs) are essential for the maintenance of protein homeostasis and assembly of proteins into functional complexes. Despite growing interest in DUBs biological functions, the roles of DUBs in regulating intestinal stem cells (ISCs) and gut homeostasis remain largely unknown. This study performed an in vivo RNAi screen through induced knock-down of DUBs expression in adult midgut ISCs and enteroblasts (EBs) to identify DUB regulators of intestinal homeostasis in Drosophila. 43 DUBs were identified and 8 DUBs were identified that are required for ISCs homeostasis. Knocking-down of usp1, CG7857, usp5, rpn8, usp10 and csn5 decreases the number of ISCs/EBs, while knocking-down of CG4968 and usp8 increases the number of ISCs/EBs. Moreover, knock-down of usp1, CG4968, CG7857, or rpn8 in ISCs/EBs disrupts the intestinal barrier integrity and shortens the lifespan, indicating the requirement of these DUBs for the maintenance of gut homeostasis. Furthermore, evidences are provided that USP1 mediates ISC lineage differentiation via modulating the Notch signaling activity. This study identifies, for the first time, the deubiquitinases required for the maintenance of intestinal homeostasis in Drosophila, and provide new insights into the functional links between the DUBs and intestinal homeostasis.
Lenhart, A., Ahsan, A., McHaty, M., Bergland, A. O. (2024). Improvement of starvation resistance via periodic fasting is genetically variable in Drosophila melanogaster. Physiological entomology, 49(3):270-278 PubMed ID: 39130127
Summary:
Organisms subjected to periodic nutrient limitation early in life exhibit improvements in aspects of survival, including resistance to some environmental stressors. Recent findings indicate that forms of periodic fasting such as intermittent fasting and time restricted feeding can improve starvation resistance. However, it remains unclear to what extent this survival improvement persists across different genetic backgrounds. This study examined fasting-induced starvation resistance across a broad survey of wild-derived lineages and documented genetic variation within this trait. A standard dietary intervention was adopted and improvement was shown to starvation resistance within a common laboratory lineage, replicating previous results. Next, fasting-induced starvation resistance was adopted across isofemale lines collected across latitudes and in different seasons, and among inbred lines derived from flies collected on different continents. Genetic variation was discovered of fasting-induced starvation resistance, and it was shown that fasting improved starvation resistance as often as it worsened starvation resistance. Fasted flies generally showed reduced fat concentration, and their starvation survival varied with sex, season of collection, and geographic origin. While specific lineages common to the laboratory can show a specific fasting-induced phenotype, this study showed that this result is not consistent across genetic backgrounds, reinforcing the idea that phenotypes observed in historic laboratory strains may not be conserved across a species. 		Darby, A. M., Okoro, D. O., Aredas, S., Frank, A. M., Pearson, W. H., Dionne, M. S., Lazzaro, B. P. (2024). High sugar diets can increase susceptibility to bacterial infection in Drosophila melanogaster. PLoS pathogens, 20(8):e1012447 PubMed ID: 39133760
Summary:
Overnutrition with dietary sugar can worsen infection outcomes in diverse organisms including insects and humans, through generally unknown mechanisms. In the present study, adult Drosophila melanogaster fed high-sugar diets became more susceptible to infection by the Gram-negative bacteria Providencia rettgeri and Serratia marcescens. P. rettgeri and S. marcescens were found to proliferate more rapidly in D. melanogaster fed a high-sugar diet, resulting in increased probability of host death. D. melanogaster become hyperglycemic on the high-sugar diet, and evidence was shown that the extra carbon availability may promote S. marcescens growth within the host. However, no evidence was found that increased carbon availability directly supports greater P. rettgeri growth. D. melanogaster on both diets fully induce transcription of antimicrobial peptide (AMP) genes in response to infection, but D. melanogaster provided with high-sugar diets show reduced production of AMP protein. Thus, overnutrition with dietary sugar may impair host immunity at the level of AMP translation. These results demonstrate that dietary sugar can shape infection dynamics by impacting both host and pathogen, depending on the nutritional requirements of the pathogen and by altering the physiological capacity of the host to sustain an immune response.
Bretscher, H., O'Connor, M. B. (2024). Glycogen homeostasis and mtDNA expression require motor neuron to muscle TGFβ Activin Signaling in Drosophila. bioRxiv, PubMed ID: 39131342
Summary:
>Maintaining metabolic homeostasis requires coordinated nutrient utilization between intracellular organelles and across multiple organ systems. Many organs rely heavily on mitochondria to generate (ATP) from glucose, or stored glycogen. Proteins required for ATP generation are encoded in both nuclear and mitochondrial DNA (mtDNA). Motoneuron to muscle signaling by the TGFβ/Activin family member Actβ positively regulates glycogen levels during Drosophila development. Remarkably, this study found that levels of stored glycogen are unaffected by altering cytoplasmic glucose catabolism. Instead, Actβ loss reduces levels of mtDNA and nuclearly encoded genes required for mtDNA replication, transcription and translation. Direct RNAi mediated knockdown of these same nuclearly encoded mtDNA expression factors also results in decreased glycogen stores. Lastly, expressing an activated form of the type I receptor Baboon in muscle was shown to restore both glycogen and mtDNA levels in actβ mutants, thereby confirming a direct link between Actβ signaling, glycogen homeostasis and mtDNA expression. 		Tanaka, T., Yano, T., Usuki, S., Seo, Y., Mizuta, K., Okaguchi, M., Yamaguchi, M., Hanyu-Nakamura, K., Toyama-Sorimachi, N., Bruckner, K., Nakamura, A. (2024). Endocytosed dsRNAs induce lysosomal membrane permeabilization that allows cytosolic dsRNA translocation for Drosophila RNAi responses. Nat Commun, 15(1):6993 PubMed ID: 39143098
Summary:
RNA interference (RNAi) is a gene-silencing mechanism triggered by the cytosolic entry of double-stranded RNAs (dsRNAs). Many animal cells internalize extracellular dsRNAs via endocytosis for RNAi induction. However, it is not clear how the endocytosed dsRNAs are translocated into the cytosol across the endo/lysosomal membrane. This study showed that in Drosophila S2 cells, endocytosed dsRNAs induce lysosomal membrane permeabilization (LMP) that allows cytosolic dsRNA translocation. LMP mediated by dsRNAs requires the lysosomal Cl(-)/H(+) antiporter DmOstm1. In clc-b or dmostm1 knockout S2 cells, extracellular dsRNAs are endocytosed and reach the lysosomes normally but fail to enter the cytosol. Pharmacological induction of LMP restores extracellular dsRNA-directed RNAi in clc-b or dmostm1-knockout cells. Furthermore, clc-b or dmostm1 mutant flies are defective in extracellular dsRNA-directed RNAi and its associated antiviral immunity. Therefore, endocytosed dsRNAs have an intrinsic ability to induce ClC-b/DmOstm1-dependent LMP that allows cytosolic dsRNA translocation for RNAi responses in Drosophila cells.
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