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Current papers in developmental biology and gene function


Monday, October 30th 2019 - Cytoskeleton and Junctions

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Izumi, Y., Furuse, K. and Furuse, M. (2019). Septate junctions regulate gut homeostasis through regulation of stem cell proliferation and enterocyte behavior in Drosophila. J Cell Sci. PubMed ID: 31444286
Smooth septate junctions (sSJs) contribute to the epithelial barrier, which restricts leakage of solutes through the paracellular route of epithelial cells in the Drosophila midgut. Previous work identified three sSJ-associated membrane proteins, Ssk, Mesh, and Tsp2A and showed that these proteins were required for sSJ formation and intestinal barrier function in the larval midgut. This study investigated the roles of sSJs in the Drosophila adult midgut. Depletion of any of the sSJ-proteins from enterocytes resulted in remarkably shortened lifespan and intestinal barrier dysfunction in flies. Interestingly, the sSJ-protein-deficient flies showed intestinal hypertrophy accompanied by accumulation of morphologically abnormal enterocytes. The phenotype was associated with increased stem cell proliferation and activation of the MAP kinase and Jak-Stat pathways in stem cells. Loss of cytokines Unpaired2 and Unpaired3, which are involved in Jak-Stat pathway activation, reduced the intestinal hypertrophy, but not the increased stem cell proliferation, in flies lacking Mesh. The present findings suggest that SJs play a crucial role in maintaining tissue homeostasis through regulation of stem cell proliferation and enterocyte behavior in the Drosophila adult midgut.
Karabasheva, D. and Smyth, J. T. (2019). A novel, dynein-independent mechanism focuses the endoplasmic reticulum around spindle poles in dividing Drosophila spermatocytes. Sci Rep 9(1): 12456. PubMed ID: 31462700
In dividing animal cells the endoplasmic reticulum (ER) concentrates around the poles of the spindle apparatus by associating with astral microtubules (MTs), and this association is essential for proper ER partitioning to progeny cells. The mechanisms that associate the ER with astral MTs are unknown. Because astral MT minus-ends are anchored by centrosomes at spindle poles, it is hypothesized that the MT minus-end motor dynein mediates ER concentration around spindle poles. Live in vivo imaging of Drosophila spermatocytes revealed that dynein is required for ER concentration around centrosomes during late interphase. In marked contrast, however, dynein suppression had no effect on ER association with astral MTs and concentration around spindle poles in early M-phase. In fact, there was a sudden onset of ER association with astral MTs in dynein RNAi cells, revealing activation of an M-phase specific mechanism of ER-MT association. ER redistribution to spindle poles also did not require non-claret disjunctional (ncd), the other known Drosophila MT minus-end motor, nor Klp61F, a MT plus-end motor that generates spindle poleward forces. Collectively, these results suggest that a novel, M-phase specific mechanism of ER-MT association that is independent of MT minus-end motors is required for proper ER partitioning in dividing cells.
Ahmed, Z., Doodhi, H., Bhaumik, A., Mazumdar, S. and Ray, K. (2019). The structural dynamics of the kinesin-2 stalk heterodimer and its biological relevance. Biochem Biophys Res Commun 518(1): 171-177. PubMed ID: 31420166
Association between two motor subunits through the rod/stalk domain enables molecular motors to walk processively on protein filaments. Previous studies suggested that structural flexibility in the coiled-coil stalk of kinesins is essential for processive runs. The stalk of heterotrimeric kinesin-2, a comparatively less processive motor, is unstable at ambient temperature. How this structural instability impacts the motor function is unclear. This study used the Forster Resonance Energy Transfer based assays to show that the Drosophila kinesin-2alpha/beta stalk heterodimer is dynamic at physiological conditions. Insertion of a missense mutation (Glu551-Lys) at the C-terminal half of kinesin-2alpha stalk reduces the dynamics of the heterodimeric stalk in vitro. The mutation, isolated as a recessive lethal allele in a forward genetic screen, is reported to disrupt the motor function in axonal transport and cilia development. Together these two results suggest that the dynamic instability of the kinesin-2 stalk could play a crucial role in maintaining its biological function.
Balaji, R., Weichselberger, V. and Classen, A. K. (2019). Response of Drosophila epithelial cell and tissue shape to external forces in vivo. Development 146(17). PubMed ID: 31399470
How actomyosin generates forces at epithelial adherens junctions has been extensively studied. However, less is known about how a balance between internal and external forces establishes epithelial cell, tissue and organ shape. This study used the Drosophila egg chamber to investigate how contractility at adherens junctions in the follicle epithelium is modulated to accommodate and resist forces arising from the growing germ line. Between stages 6 and 9, adherens junction tension in the post-mitotic epithelium decreases, suggesting that the junctional network relaxes to accommodate germline growth. At that time, a prominent medial Myosin II network coupled to corrugating adherens junctions develops. Local enrichment of medial Myosin II in main body follicle cells resists germline-derived forces, thus constraining apical areas and, consequently, cuboidal cell shapes at stage 9. At the tissue and organ level, local reinforcement of medial junction architecture ensures the timely contact of main body cells with the expanding oocyte and imposes circumferential constraints on the germ line guiding egg elongation. This study provides insight into how adherens junction tension promotes cell and tissue shape transitions while integrating the growth and shape of an internally enclosed structure in vivo.
Fan, A., Joy, M. S. H. and Saif, T. (2019). A connected cytoskeleton network generates axonal tension in embryonic Drosophila. Lab Chip 19(18): 3133-3139. PubMed ID: 31435630
Axons of neurons are contractile, i.e., they actively maintain a rest tension. However, the spatial origin of this contractility along the axon and the role of the cytoskeleton in generating tension and sustaining rigidity are unknown. Using a microfluidic platform, this study exposed a small segment of the axons of embryonic Drosophila motor neurons to specific cytoskeletal disruption drugs. A local actomyosin disruption led to a total loss in axonal tension, with the stiffness of the axon remaining unchanged. A local disruption of microtubules led to a local reduction in bending stiffness, while tension remained unchanged. These observations demonstrated that contractile forces are generated and transferred along the entire length of the axon in a serial fashion. Thus, a local force disruption results in a collapse of tension of the entire axon. This mechanism potentially provides a pathway for rapid tension regulation to facilitate physiological processes that are influenced by axonal tension.
Goldman, C. H., Neiswender, H., Veeranan-Karmegam, R. and Gonsalvez, G. B. (2019). The Egalitarian binding partners Dynein light chain and Bicaudal-D act sequentially to link mRNA to the Dynein motor. Development 146(15). PubMed ID: 31391195
A conserved mechanism of polarity establishment is the localization of mRNA to specific cellular regions. Although it is clear that many mRNAs are transported along microtubules, much less is known about the mechanism by which these mRNAs are linked to microtubule motors. The RNA binding protein Egalitarian (Egl) is necessary for localization of several mRNAs in Drosophila oocytes and embryos. Egl also interacts with Dynein light chain (Dlc) and Bicaudal-D (BicD). The role of Dlc and BicD in mRNA localization has remained elusive. Both proteins are required for oocyte specification, as is Egl. Null alleles in these genes result in an oogenesis block. This report used an shRNA-depletion strategy to overcome the oogenesis block. The findings reveal that the primary function of Dlc is to promote Egl dimerization. Loss of dimerization compromises the ability of Egl to bind RNA. Consequently, Egl is not bound to cargo, and is not able to efficiently associate with BicD and the Dynein motor. The results therefore identify the key molecular steps required for assembling a localization-competent mRNP.

Friday, September 27th - Adult Neural Development and Function

Hill, A. S., Jain, P., Folan, N. E. and Ben-Shahar, Y. (2019). The Drosophila ERG channel seizure plays a role in the neuronal homeostatic stress response. PLoS Genet 15(8): e1008288. PubMed ID: 31393878
Neuronal physiology is particularly sensitive to acute stressors that affect excitability, many of which can trigger seizures and epilepsies. Although intrinsic neuronal homeostasis plays an important role in maintaining overall nervous system robustness and its resistance to stressors, the specific genetic and molecular mechanisms that underlie these processes are not well understood. This study used a reverse genetic approach in Drosophila to test the hypothesis that specific voltage-gated ion channels contribute to neuronal homeostasis, robustness, and stress resistance. The activity of the voltage-gated potassium channel seizure (sei), an ortholog of the mammalian ERG channel family, is essential for protecting flies from acute heat-induced seizures. Although sei is broadly expressed in the nervous system, the data indicate that its impact on the organismal robustness to acute environmental stress is primarily mediated via its action in excitatory neurons, the octopaminergic system, as well as neuropile ensheathing and perineurial glia. Furthermore, these studies suggest that human mutations in the human ERG channel (hERG), which have been primarily implicated in the cardiac Long QT Syndrome (LQTS), may also contribute to the high incidence of seizures in LQTS patients via a cardiovascular-independent neurogenic pathway.
Honegger, K. S., Smith, M. A., Churgin, M. A., Turner, G. C. and de Bivort, B. L. (2019). Idiosyncratic neural coding and neuromodulation of olfactory individuality in Drosophila. Proc Natl Acad Sci U S A. PubMed ID: 31455738
Innate behavioral biases and preferences can vary significantly among individuals of the same genotype. Though individuality is a fundamental property of behavior, it is not currently understood how individual differences in brain structure and physiology produce idiosyncratic behaviors. This study presents evidence for idiosyncrasy in olfactory behavior and neural responses in Drosophila. Individual female Drosophila from a highly inbred laboratory strain exhibit idiosyncratic odor preferences that persist for days. In vivo calcium imaging of neural responses were used to compare projection neuron (second-order neurons that convey odor information from the sensory periphery to the central brain) responses to the same odors across animals. While odor responses appear grossly stereotyped, upon closer inspection, many individual differences are apparent across antennal lobe (AL) glomeruli (compact microcircuits corresponding to different odor channels). Moreover, this study shows that neuromodulation, environmental stress in the form of altered nutrition, and activity of certain AL local interneurons affect the magnitude of interfly behavioral variability. Taken together, this work demonstrates that individual Drosophila exhibit idiosyncratic olfactory preferences and idiosyncratic neural responses to odors, and that behavioral idiosyncrasies are subject to neuromodulation and regulation by neurons in the AL.
Harish, R. K., Tendulkar, S., Deivasigamani, S., Ratnaparkhi, A. and Ratnaparkhi, G. S. (2019). Monensin Sensitive 1 regulates dendritic arborization in Drosophila by modulating endocytic flux. Front Cell Dev Biol 7: 145. PubMed ID: 31428611
Monensin Sensitive 1 (Mon1) is a component of the Mon1:Ccz1 complex that mediates Rab5 to Rab7 conversion in eukaryotic cells by serving as a guanine nucleotide exchange factor for Rab7 during vesicular trafficking. Mon1 activity modulates the complexity of Class IV dendritic arborization (da) neurons during larval development. Loss of Mon1 function leads to an increase in arborization and complexity, while increased expression, leads to reduced arborization. The ability of Mon1 to influence dendritic development is possibly a function of its interactions with Rab family GTPases that are central players in vesicular trafficking. Earlier, these GTPases, specifically Rab1, Rab5, Rab10, and Rab11 have been shown to regulate dendritic arborization. This study has conducted genetic epistasis experiments, by modulating the activity of Rab5, Rab7, and Rab11 in da neurons, in Mon1 mutants, and has demonstrated that the ability of Mon1 to regulate arborization is possibly due to its effect on the recycling pathway. Dendritic branching is critical for proper connectivity and physiological function of the neuron. An understanding of regulatory elements, such as Mon1, as demonstrated in this study, is essential to understand neuronal function.
Kacsoh, B. Z., Bozler, J., Hodge, S. and Bosco, G. (2019). Neural circuitry of social learning in Drosophila requires multiple inputs to facilitate inter-species communication. Commun Biol 2: 309. PubMed ID: 31428697
Drosophila species communicate the threat of parasitoid wasps to naive individuals. Communication of the threat between closely related species is efficient, while more distantly related species exhibit a dampened, partial communication. Partial communication between D. melanogaster and D. ananassae about wasp presence is enhanced following a period of cohabitation, suggesting that species-specific natural variations in communication 'dialects' can be learned through socialization. This study identified six regions of the Drosophila brain essential for dialect training. Subgroups of neurons in these regions were identified, including motion detecting neurons in the optic lobe, layer 5 of the fan-shaped body, the D glomerulus in the antennal lobe, and the odorant receptor Or69a, where activation of each component is necessary for dialect learning. These results reveal functional neural circuits that underlie complex Drosophila social behaviors, and these circuits are required for integration several cue inputs involving multiple regions of the Drosophila brain.
Kaladchibachi, S., Negelspach, D. C., Zeitzer, J. M. and Fernandez, F. (2019). Optimization of circadian responses with shorter and shorter millisecond flashes. Biol Lett 15(8): 20190371. PubMed ID: 31387472
Recent work suggests that the circadian pacemaker responds optimally to millisecond flashes of light, not continuous light exposure as has been historically believed. It is unclear whether these responses are influenced by the physical characteristics of the pulsing. In the present study, Drosophila (n = 2199) were stimulated with 8, 16 or 120 ms flashes. For each duration, the energy content of the exposure was systematically varied by changing the pulse irradiance and the number of stimuli delivered over a fixed 15 min administration window (64 protocols surveyed in all). Results showed that per microjoule invested, 8 ms flashes were more effective at resetting the circadian activity rhythm than 16- and 120 ms flashes (i.e. left shift of the dose-response curve, as well as a higher estimated maximal response). These data suggest that the circadian pacemaker's photosensitivity declines within milliseconds of light contact. Further introduction of light beyond a floor of (at least) 8 ms leads to diminishing returns on phase-shifting.
Guo, P., Xu, X., Wang, F., Yuan, X., Tu, Y., Zhang, B., Zheng, H., Yu, D., Ge, W., Gong, Z., Yang, X. and Xi, Y. (2019). A novel neuroprotective role of phosphatase of Regenerating Liver-1 against CO2 stimulation in Drosophila. iScience 19: 291-302. PubMed ID: 31404830
Neuroprotection is essential for the maintenance of normal physiological functions in the nervous system. This is especially true under stress conditions. This study demonstrates a novel protective function of PRL-1 against CO2 stimulation in Drosophila. In the absence of PRL-1, flies exhibit a permanent held-up wing phenotype upon CO2 exposure. Knockdown of the CO2 olfactory receptor, Gr21a, suppresses the phenotype. Genetic data indicate that the wing phenotype is due to a neural dysfunction. PRL-1 physically interacts with Uex and controls Uex expression levels. Knockdown of Uex alone leads to a similar wing held-up phenotype to that of PRL-1 mutants. Uex acts downstream of PRL-1. Elevated Uex levels in PRL-1 mutants prevent the CO2-induced phenotype. PRL-1 and Uex are required for a wide range of neurons to maintain neuroprotective functions. Expression of human homologs of PRL-1 could rescue the phenotype in Drosophila, suggesting a similar function in humans.

Thursday, September 26th - Signaling

Cheng, Y. Y., Huang, Y. F., Lin, H. H., Chang, W. W. and Lyu, P. C. (2019). The ligand-mediated affinity of brain-type fatty acid-binding protein for membranes determines the directionality of lipophilic cargo transport. Biochim Biophys Acta Mol Cell Biol Lipids 1864(12): 158506. PubMed ID: 31404652
The intracellular transport of lipophilic cargoes is a highly dynamic process. In eukaryotic cells, the uptake and release of long-chain fatty acids (LCFAs) are executed by fatty-acid binding proteins. However, how these carriers control the directionality of cargo trafficking remains unclear. This study has revealed that the unliganded archetypal Drosophila brain-type fatty acid-binding protein (dFABP) possesses a stronger binding affinity than its liganded counterpart for empty nanodiscs (ND). Titrating unliganded dFABP and nanodiscs with LCFAs rescued the broadening of FABP cross-peak intensities in HSQC spectra from a weakened protein-membrane interaction. Two out of the 3 strongest LCFA contacting residues in dFABP identified by NMR HSQC chemical shift perturbation (CSP) are also part of the 30 ND-contacting residues (out of the total 130 residues in dFABP), revealed by attenuated TROSY signal in the presence of lipid ND to apo-like dFABP. Crystallographic temperature factor data suggest enhanced alphaII helix dynamics upon LCFA binding, compensating for the entropic loss in the betaC-D/betaE-F loops. The aliphatic tail of bound LCFA impedes the charge-charge interaction between dFABP and the head groups of the membrane, and dFABP is prone to dissociate from the membrane upon ligand binding. It is therefore concluded that lipophilic ligands participate directly in the control of the functionally required membrane association and dissociation of FABPs.
Byun, P. K., Zhang, C., Yao, B., Wardwell-Ozgo, J., Terry, D., Jin, P. and Moberg, K. (2019). The Taiman transcriptional coactivator engages Toll signals to promote apoptosis and intertissue invasion in Drosophila. Curr Biol 29(17): 2790-2800.e2794. PubMed ID: 31402304
The Drosophila Taiman (Tai) protein is homologous to the human steroid-receptor coactivators SRC1-3 and activates transcription in complex with the 20-hydroxyecdysone (20E) receptor (EcR). Tai has roles in intestinal homeostasis, germline maintenance, cell motility, and proliferation through interactions with EcR and the coactivator Yorkie (Yki). Tai also promotes invasion of tumor cells in adjacent organs, but this pro-invasive mechanism is undefined. This study shows that Tai expression transforms sessile pupal wing cells into an invasive mass that penetrates the adjacent thorax during a period of high 20E. Candidate analysis confirms a reliance on elements of the 20E and Hippo pathways, such as Yki and the Yki-Tai target dilp8. Screening the Tai-induced wing transcriptome detects enrichment for innate immune factors, including the Spatzle (Spz) family of secreted Toll ligands that induce apoptosis during cell competition. Tai-expressing wing cells induce immune signaling and apoptosis among adjacent thoracic cells, and genetic reduction of spz, Toll, or the rpr/hid/grim pro-apoptotic factors each suppresses invasion, suggesting an intercellular Spz-Toll circuit supports killing-mediated invasion. Modeling these interactions in larval epithelia confirms that Tai kills neighboring cells via a mechanism involving Toll, Spz factors, and the Spz inhibitor Necrotic. Tai-expressing cells evade death signals by repressing the immune deficiency (IMD) pathway, which operates in parallel to Toll to control nuclear factor kappaB (NF-kappaB) activity and independently regulates JNK activity. In sum, these findings suggest that Tai promotes competitive cell killing via Spz-Toll and that this killing mechanism supports pathologic intertissue invasion in Drosophila.
Chaudhary, V., Hingole, S., Frei, J., Port, F., Strutt, D. and Boutros, M. (2019). Robust Wnt signaling is maintained by a Wg protein gradient and Fz2 receptor activity in the developing Drosophila wing. Development 146(15). PubMed ID: 31399474
Wnts are secreted proteins that regulate cell fate during development of all metazoans. Wnt proteins were proposed to spread over several cells to activate signaling directly at a distance. In the Drosophila wing epithelium, an extracellular gradient of the Wnt1 homolog Wingless (Wg) was observed extending over several cells away from producing cells. Surprisingly, however, it was also shown that a membrane-tethered Neurotactin-Wg fusion protein (NRT-Wg) can largely replace endogenous Wg, leading to proper patterning of the wing. Therefore, the functional range of Wg and whether Wg spreading is required for correct tissue patterning remains controversial. In this study, by capturing secreted Wg on cells away from the source, it was shown that Wg acts over a distance of up to 11 cell diameters to induce signaling. Furthermore, cells located outside the reach of extracellular Wg depend on the Frizzled2 receptor to maintain signaling. Frizzled2 expression is increased in the absence of Wg secretion and is required to maintain signaling and cell survival in NRT-wg wing discs. Together, these results provide insight into the mechanisms by which robust Wnt signaling is achieved in proliferating tissues.
Hao, Y., Waller, T. J., Nye, D. M., Li, J., Zhang, Y., Hume, R. I., Rolls, M. M. and Collins, C. A. (2019). Degeneration of injured axons and dendrites requires restraint of a protective JNK signaling pathway by the transmembrane protein Raw. J Neurosci. PubMed ID: 31492772
The degeneration of injured axons involves a self-destruction pathway whose components and mechanism are not fully understood. This study reports a new regulator of axonal resilience. The transmembrane protein Raw is cell autonomously required for the degeneration of injured axons, dendrites and synapses in Drosophila melanogaster. In both male and female raw hypomorphic mutant or knockdown larvae, the degeneration of injured axons, dendrites and synapses from motoneurons and sensory neurons is strongly inhibited. This protection is insensitive to reduction in the levels of the NAD+ synthesis enzyme Nmnat (nicotinamide mononucleotide adenylyl transferase), but requires the JNK MAP Kinase and -transcription factors Fos and Jun (AP-1). While these factors are previously known to function in axonal injury signaling and regeneration, Raw's function can be genetically separated from other axonal injury responses: Raw does not modulate JNK-dependent axonal injury signaling and regenerative responses, but instead restrains a protective pathway that inhibits the degeneration of axons, dendrites and synapses. While protection in raw mutants requires JNK, Fos and Jun, JNK also promotes axonal degeneration. These findings suggest the existence of multiple independent pathways that share modulation by JNK, Fos and Jun that influence how axons respond to stress and injury.
Bertinelli, M., Paesen, G. C., Grimes, J. M. and Renner, M. (2019). High-resolution crystal structure of arthropod Eiger TNF suggests a mode of receptor engagement and altered surface charge within endosomes. Commun Biol 2: 293. PubMed ID: 31396573
The tumour necrosis factor alpha (TNFalpha) superfamily of proteins are critical in numerous biological processes, such as in development and immunity. Eiger is the sole TNFalpha member described in arthropods such as in the important model organism Drosophila. To date there are no structural data on any Eiger protein. This study presents the structure of the TNF domain of Eiger from the fall armyworm Spodoptera frugiperda (SfEiger) to 1.7 A from a serendipitously obtained crystal without prior knowledge of the protein sequence. The structure confirms that canonical trimerization is conserved from ancestral TNFs and points towards a mode of receptor engagement. Furthermore, numerous surface histidines were observed on SfEiger, potentially acting as pH switches following internalization into endosomes. These data contributes to the genome annotation of S. frugiperda, a voracious agricultural pest, and can serve as a basis for future structure-function investigations of the TNF system in related arthropods such as Drosophila.
Hu, D. J. and Jasper, H. (2019). Control of intestinal cell fate by dynamic mitotic spindle repositioning influences epithelial homeostasis and longevity. Cell Rep 28(11): 2807-2823. PubMed ID: 31509744
Tissue homeostasis depends on precise yet plastic regulation of stem cell daughter fates. During growth, Drosophila intestinal stem cells (ISCs) adjust fates by switching from asymmetric to symmetric lineages to scale the size of the ISC population. Using a combination of long-term live imaging, lineage tracing, and genetic perturbations, this study demonstrates that this switch is executed through the control of mitotic spindle orientation by Jun-N-terminal kinase (JNK) signaling. JNK interacts with the WD40-repeat protein Wdr62 at the spindle and transcriptionally represses the kinesin Kif1a to promote planar spindle orientation. In stress conditions, this function becomes deleterious, resulting in overabundance of symmetric fates and contributing to the loss of tissue homeostasis in the aging animal. Restoring normal ISC spindle orientation by perturbing the JNK/Wdr62/Kif1a axis is sufficient to improve intestinal physiology and extend lifespan. These findings reveal a critical role for the dynamic control of SC spindle orientation in epithelial maintenance.

Wednesday, September 25th - Stem cells

Chang, Y. C., Tu, H., Chen, J. Y., Chang, C. C., Yang, S. Y. and Pi, H. (2019). Reproduction disrupts stem cell homeostasis in testes of aged male Drosophila via an induced microenvironment. PLoS Genet 15(7): e1008062. PubMed ID: 31295251
Stem cells rely on instructive cues from their environment. Alterations in microenvironments might contribute to tissue dysfunction and disease pathogenesis. Germline stem cells (GSCs) and cyst stem cells (CySC) in Drosophila testes are normally maintained in the apical area by the testicular hub. This study found that reproduction leads to accumulation of early differentiating daughters of CySCs and GSCs in the testes of aged male flies, due to hyperactivation of Jun-N-terminal kinase (JNK) signaling to maintain self-renewal gene expression in the differentiating cyst cells. JNK activity is normally required to maintain CySCs in the apical niche. A muscle sheath surrounds the Drosophila testis to maintain its long coiled structure. Importantly, reproduction triggers accumulation of the tumor necrosis factor (TNF) Eiger in the testis muscle to activate JNK signaling via the TNF receptor Grindelwald in the cyst cells. Reducing Eiger activity in the testis muscle sheath suppressed reproduction-induced differentiation defects, but had little effect on testis homeostasis of unmated males. These results reveal that reproduction in males provokes a dramatic shift in the testicular microenvironment, which impairs tissue homeostasis and spermatogenesis in the testes.
Angulo, B., Srinivasan, S., Bolival, B. J., Olivares, G. H., Spence, A. C. and Fuller, M. T. (2019). DREF genetically counteracts Mi-2 and Caf1 to regulate adult stem cell maintenance. PLoS Genet 15(6): e1008187. PubMed ID: 31226128
Active adult stem cells maintain a bipotential state with progeny able to either self-renew or initiate differentiation depending on extrinsic signals from the surrounding microenvironment. However, the intrinsic gene regulatory networks and chromatin states that allow adult stem cells to make these cell fate choices are not entirely understood. This study shows that the transcription factor DNA Replication-related Element Factor (DREF) regulates adult stem cell maintenance in the Drosophila male germline. A temperature-sensitive allele of DREF described in this study genetically separated a role for DREF in germline stem cell self-renewal from the general roles of DREF in cell proliferation. The DREF temperature-sensitive allele caused defects in germline stem cell self-renewal but allowed viability and division of germline stem cells as well as cell viability, growth and division of somatic cyst stem cells in the testes and cells in the Drosophila eye. Germline stem cells mutant for the temperature sensitive DREF allele exhibited lower activation of a TGF-beta reporter, and their progeny turned on expression of the differentiation factor Bam prematurely. Results of genetic interaction analyses revealed that Mi-2 and Caf1/p55, components of the Nucleosome Remodeling and Deacetylase (NuRD) complex, genetically antagonize the role of DREF in germline stem cell maintenance. Taken together, these data suggest that DREF contributes to intrinsic components of the germline stem cell regulatory network that maintains competence to self-renew.
Gambarotto, D., Pennetier, C., Ryniawec, J. M., Buster, D. W., Gogendeau, D., Goupil, A., Nano, M., Simon, A., Blanc, D., Racine, V., Kimata, Y., Rogers, G. C. and Basto, R. (2019). Plk4 regulates centriole asymmetry and spindle orientation in neural stem cells. Dev Cell. PubMed ID: 31130353
Defects in mitotic spindle orientation (MSO) disrupt the organization of stem cell niches impacting tissue morphogenesis and homeostasis. Mutations in centrosome genes reduce MSO fidelity, leading to tissue dysplasia and causing several diseases such as microcephaly, dwarfism, and cancer. Whether these mutations perturb spindle orientation solely by affecting astral microtubule nucleation or whether centrosome proteins have more direct functions in regulating MSO is unknown. To investigate this question, the consequences were analyzed of deregulating Plk4 (the master centriole duplication kinase) activity in Drosophila asymmetrically dividing neural stem cells. Plk4 functions upstream of MSO control, orchestrating centriole symmetry breaking and consequently centrosome positioning. Mechanistically, Plk4 was shown to act through Spd2 phosphorylation, which induces centriole release from the apical cortex. Overall, this work not only reveals a role for Plk4 in regulating centrosome function but also links the centrosome biogenesis machinery with the MSO apparatus.
Gil-Ranedo, J., Gonzaga, E., Jaworek, K. J., Berger, C., Bossing, T. and Barros, C. S. (2019). STRIPAK Members Orchestrate Hippo and Insulin Receptor Signaling to Promote Neural Stem Cell Reactivation. Cell Rep 27(10): 2921-2933.e2925. PubMed ID: 31167138
Adult stem cells reactivate from quiescence to maintain tissue homeostasis and in response to injury. How the underlying regulatory signals are integrated is largely unknown. Drosophila neural stem cells (NSCs) also leave quiescence to generate adult neurons and glia, a process that is dependent on Hippo signaling inhibition and activation of the insulin-like receptor (InR)/PI3K/Akt cascade. A transcriptome analysis was performed of individual quiescent and reactivating NSCs harvested directly from Drosophila brains; and the conserved STRIPAK complex members mob4, cka, and PP2A (microtubule star, mts) were identified. PP2A/Mts phosphatase, with its regulatory subunit Widerborst, maintains NSC quiescence, preventing premature activation of InR/PI3K/Akt signaling. Conversely, an increase in Mob4 and Cka levels promotes NSC reactivation. Mob4 and Cka are essential to recruit PP2A/Mts into a complex with Hippo kinase, resulting in Hippo pathway inhibition. It is proposed that Mob4/Cka/Mts functions as an intrinsic molecular switch coordinating Hippo and InR/PI3K/Akt pathways and enabling NSC reactivation.
Tracy Cai, X., Li, H., Safyan, A., Gawlik, J., Pyrowolakis, G. and Jasper, H. (2019). AWD regulates timed activation of BMP signaling in intestinal stem cells to maintain tissue homeostasis. Nat Commun 10(1): 2988. PubMed ID: 31278345
Precise control of stem cell (SC) proliferation ensures tissue homeostasis. In the Drosophila intestine, injury-induced regeneration involves initial activation of intestinal SC (ISC) proliferation and subsequent return to quiescence. These two phases of the regenerative response are controlled by differential availability of the BMP type I receptor Thickveins (Tkv), yet how its expression is dynamically regulated remains unclear. This study shows that during homeostasis, the E3 ubiquitin ligase Highwire and the ubiquitin-proteasome system maintain low Tkv protein expression. After ISC activation, Tkv is stabilized by proteasome inhibition and undergoes endocytosis due to the induction of the nucleoside diphosphate kinase Abnormal Wing Disc (AWD). Tkv internalization is required for the activation of the Smad protein Mad, and for the return to quiescence after a regenerative episode. These data provide insight into the mechanisms ensuring tissue homeostasis by dynamic control of somatic stem cell activity.
Hu, D. J. and Jasper, H. (2019). Control of intestinal cell fate by dynamic mitotic spindle repositioning influences epithelial homeostasis and longevity. Cell Rep 28(11): 2807-2823. PubMed ID: 31509744
Tissue homeostasis depends on precise yet plastic regulation of stem cell daughter fates. During growth, Drosophila intestinal stem cells (ISCs) adjust fates by switching from asymmetric to symmetric lineages to scale the size of the ISC population. Using a combination of long-term live imaging, lineage tracing, and genetic perturbations, this study demonstrates that this switch is executed through the control of mitotic spindle orientation by Jun-N-terminal kinase (JNK) signaling. JNK interacts with the WD40-repeat protein Wdr62 at the spindle and transcriptionally represses the kinesin Kif1a to promote planar spindle orientation. In stress conditions, this function becomes deleterious, resulting in overabundance of symmetric fates and contributing to the loss of tissue homeostasis in the aging animal. Restoring normal ISC spindle orientation by perturbing the JNK/Wdr62/Kif1a axis is sufficient to improve intestinal physiology and extend lifespan. These findings reveal a critical role for the dynamic control of SC spindle orientation in epithelial maintenance.

Tuesday, September 24th - Evolution

Chen, A. L., Chen, C. C., Katoh, T., Katoh, T. K., Watada, M., Toda, M. J., Ritchie, M. G. and Wen, S. Y. (2019). Evolution and diversity of the courtship repertoire in the Drosophila montium species group (Diptera: Drosophilidae). J Evol Biol. PubMed ID: 31386239
Changes in elements of courtship behaviour can influence sexual isolation between species. Large-scale analyses of changes, including loss and gain of courtship elements, across a relatively complete phylogenetic group are rare but needed to understand the significance of such changes, for example whether the gain and loss of courtship elements are essentially arbitrary or equally reversible. In most species of Drosophila, courtship, including singing, mainly occurs before mounting as premounting courtship. The Drosophila montium species group is unusual because loss of premounting courtship and gain of post-mounting one has been detected in this group. This study provides an extensive analysis on the courtship repertoire and songs of 42 species in this group. Synchronously captured video and audio recordings were analysed to describe courtship patterns and male courtship songs, and changes were analysed in a phylogenetic context. Ancestral state reconstruction suggests that a gain of post-mounting courtship singing at the ancestor of this species group has been accompanied by a concurrent decrease in the incidence of premounting courtship singing and has led to subsequent further decrease and eventually complete loss of premounting courtship song in several lineages. Alongside this evolutionary trend towards post-mounting courtship, sine song and a special type of "high pulse repetition song" have become more widely used for courtship during species diversification in the montium group. It is likely that the elaboration of post-mounting courtship behaviours is associated with changes in the relative importance of pre- and post-mounting components of mating systems, such as sperm competition or cryptic female choice.
Hagen, J. F. D., Mendes, C. C., Blogg, A., Payne, A., Tanaka, K. M., Gaspar, P., Figueras Jimenez, J., Kittelmann, M., McGregor, A. P. and Nunes, M. D. S. (2019). tartan underlies the evolution of Drosophila male genital morphology. Proc Natl Acad Sci U S A. PubMed ID: 31484761
Male genital structures are among the most rapidly evolving morphological traits and are often the only features that can distinguish closely related species. This process is thought to be driven by sexual selection and may reinforce species separation. However, while the genetic bases of many phenotypic differences have been identified, knowledge about the genes underlying evolutionary differences in male genital organs and organ size more generally is still lacking. The claspers (surstyli) are periphallic structures that play an important role in copulation in insects. This study shows that divergence in clasper size and bristle number between Drosophila mauritiana and Drosophila simulans is caused by evolutionary changes in tartan (trn), which encodes a transmembrane leucine-rich repeat domain protein that mediates cell-cell interactions and affinity. There are no fixed amino acid differences in trn between D. mauritiana and D. simulans, but differences in the expression of this gene in developing genitalia suggest that cis-regulatory changes in trn underlie the evolution of clasper morphology in these species. Finally, analyses of reciprocal hemizygotes that are genetically identical, except for the species from which the functional allele of trn originates, determined that the trn allele of D. mauritiana specifies larger claspers with more bristles than the allele of D. simulans. Therefore, this study has identified a gene underlying evolutionary change in the size of a male genital organ, which will help to better understand not only the rapid diversification of these structures, but also the regulation and evolution of organ size more broadly.
Chapman, J. R., Hill, T. and Unckless, R. L. (2019). Balancing selection drives the maintenance of genetic variation in Drosophila antimicrobial peptides. Genome Biol Evol. PubMed ID: 31504505
Genes involved in immune defense against pathogens provide some of the most well-known examples of both directional and balancing selection. Antimicrobial peptides (AMPs) are innate immune effector genes, playing a key role in pathogen clearance in many species, including Drosophila. Conflicting lines of evidence have suggested AMPs may be under directional, balancing or purifying selection. This study used both a linear model and control gene-based approach to show that balancing selection is an important force shaping AMP diversity in Drosophila. In D. melanogaster, this is most clearly observed in ancestral African populations. Furthermore, the signature of balancing selection is even more striking once background selection has been accounted for. Balancing selection also acts on AMPs in D. mauritiana, an isolated island endemic separated from D. melanogaster by about 4 million years of evolution. This suggests that balancing selection may be broadly acting to maintain adaptive diversity in Drosophila AMPs, as has been found in other taxa.
Dominguez-Garcia, S., Garcia, C., Quesada, H. and Caballero, A. (2019). Accelerated inbreeding depression suggests synergistic epistasis for deleterious mutations in Drosophila melanogaster. Heredity (Edinb). PubMed ID: 31477803
Synergistic epistasis for deleterious alleles is relevant to the mutation load paradox and the evolution of sex and recombination. Some studies have shown evidence of synergistic epistasis for spontaneous or induced deleterious mutations appearing in mutation-accumulation experiments. However, many newly arising mutations may not actually be segregating in natural populations because of the erasing action of natural selection. A demonstration of synergistic epistasis for naturally segregating alleles can be achieved by means of inbreeding depression studies, as deleterious recessive allelic effects are exposed in inbred lines. This paper reports the results of two independent inbreeding experiments carried out with two different populations of Drosophila melanogaster. The results show a consistent accelerated inbreeding depression for fitness, suggesting synergistic epistasis among deleterious alleles. Computer simulations were performed assuming different possible models of epistasis and mutational parameters for fitness, finding some of them to be compatible with the results observed. These results suggest that synergistic epistasis for deleterious mutations not only occurs among newly arisen spontaneous or induced mutations, but also among segregating alleles in natural populations.

Monday, September 23rd - Adult Physiology

He, Z., Zheng, Y., Yu, W. J., Fang, Y., Mao, B. and Wang, Y. F. (2019). How do Wolbachia modify the Drosophila ovary? New evidences support the "titration-restitution" model for the mechanisms of Wolbachia-induced CI.. BMC Genomics 20(1): 608. PubMed ID: 31340757
Cytoplasmic incompatibility (CI) is the most common phenotype induced by endosymbiont Wolbachia and results in embryonic lethality when Wolbachia-modified sperm fertilize eggs without Wolbachia. However, eggs carrying the same strain of Wolbachia can rescue this embryonic death, thus producing viable Wolbachia-infected offspring. Hence Wolbachia can be transmitted mainly by hosts' eggs. By RNA-seq analyses, the transcription profiles of Drosophila melanogaster adult ovaries were first compared with and without the wMel Wolbachia, and 149 differentially expressed genes (DEGs) were identified, of which 116 genes were upregulated and 33 were downregulated by Wolbachia infection. Seven microRNAs (miRNAs) were identified that were all upregulated in fly ovaries by Wolbachia infection. Matching of miRNA and mRNA data showed that these seven miRNAs regulated 15 DEGs. Most of the DEGs showed variation in opposite directions in ovaries versus testes in the presence of Wolbachia, which generally supports the "titration-restitution" model for CI. Furthermore, genes related to metabolism were upregulated, which may benefit maximum proliferation and transmission of Wolbachia. This provides new insights into the molecular mechanisms of Wolbachia-induced CI and Wolbachia dependence on host ovaries.
Keebaugh, E. S., Yamada, R. and Ja, W. W. (2019). The nutritional environment influences the impact of microbes on Drosophila melanogaster life span. MBio 10(4). PubMed ID: 31289176
Microbes can extend Drosophila melanogaster life span by contributing to the nutritional value of malnourishing fly culture medium. The beneficial effect of microbes during malnutrition is dependent on their individual ability to proliferate in the fly environment and is mimicked by lifelong supplementation of equivalent levels of heat-killed microbes or dietary protein, suggesting that microbes can serve directly as a protein-rich food source. This study used nutritionally rich fly culture medium to demonstrate how changes in dietary composition influence monocolonized fly life span; microbes that extend fly life span on malnourishing diets can shorten life on rich diets. The mechanisms employed by microbes to affect host health likely differ on low- or high-nutrient diets. The results demonstrate how Drosophila-associated microbes can positively or negatively influence fly life span depending on the nutritional environment. Although controlled laboratory environments allow focused investigations on the interaction between fly microbiota and nutrition, the relevance of these studies is not straightforward, because it is difficult to mimic the nutritional ecology of natural Drosophila-microbe interactions. As such, caution is needed in designing and interpreting fly-microbe experiments and before categorizing microbes into specific symbiotic roles based on results obtained from experiments testing limited conditions.
Austin, C. J. and Moehring, A. J. (2019). Local thermal adaptation detected during multiple life stages across populations of Drosophila melanogaster. J Evol Biol. PubMed ID: 31454449
Thermal adaptation is typically detected by examining the tolerance of a few populations to extreme temperatures within a single life stage. However, the extent to which adaptation occurs among many different populations might depend on the tolerance of multiple life stages and the average temperature range that the population experiences. This study examined local adaptation to native temperature conditions in eleven populations of the well-known cosmopolitan fruit fly, Drosophila melanogaster. These populations were sampled from across the global range of D. melanogaster. Traits related to fitness were measured during each life stage to determine if certain stages are more sensitive to changes in temperature than others. D. melanogaster appeared to show local adaptation to native temperatures during the egg, larval, and adult life stages, but not the pupal stage. This suggests that across the entire distribution of D. melanogaster, certain life stages might be locally adapted to native temperatures, while other stages might use phenotypic plasticity or tolerance to a wide range of temperatures experienced in the native environment of this species.
Baumers, M., Klose, S., Bruser, C., Haag, C., Hansch, S., Pannen, H., Weidtkamp-Peters, S., Feldbrugge, M. and Klein, T. (2019). The auxiliary ESCRT complexes provide robustness to cold in poikilothermic organisms. Biol Open 8(9). PubMed ID: 31412999
The ESCRT pathway, comprising the in sequence acting ESCRT-0, -I, -II, -III and Vps4 complexes, conducts the abscission of membranes away from the cytosol. Whereas the components of the central ESCRT-III core complex have been thoroughly investigated, the function of the components of the associated two auxiliary ESCRT sub-complexes are not well-understood in metazoans, especially at the organismal level. This paper presents the developmental analysis of the Drosophila orthologs of the auxiliary ESCRTs Chmp5 and Ist1, DChmp5 and DIst1, which belong to the two auxiliary sub-complexes. While each single null mutant displayed mild defects in development, the Dist1 Dchmp5 double mutant displayed a severe defect, indicating that the two genes act synergistically, but in separate pathways. Moreover, the presented results indicate that the auxiliary ESCRTs provide robustness against cold during development of diverse poikilothermic organisms, probably by preventing the accumulation of the ESCRT-III core component Shrub on the endosomal membrane.
Camus, M. F., Piper, M. D. and Reuter, M. (2019). Sex-specific transcriptomic responses to changes in the nutritional environment. Elife 8. PubMed ID: 31436529
Males and females typically pursue divergent reproductive strategies and accordingly require different dietary compositions to maximise their fitness. This study moves from identifying sex-specific optimal diets to understanding the molecular mechanisms that underlie male and female responses to dietary variation in Drosophila melanogaster. Male and female gene expression was examined on male-optimal (carbohydrate-rich) and female-optimal (protein-rich) diets. The sexes share a large core of metabolic genes that are concordantly regulated in response to dietary composition. However, smaller sets of genes were observed with divergent and opposing regulation, most notably in reproductive genes which are over-expressed on each sex's optimal diet. These results suggest that nutrient sensing output emanating from a shared metabolic machinery are reversed in males and females, leading to opposing diet-dependent regulation of reproduction in males and females. Further analysis and experiments suggest that this reverse regulation occurs within the IIS/TOR network.
Gururaja Rao, S., Bednarczyk, P., Towheed, A., Shah, K., Karekar, P., Ponnalagu, D., Jensen, H. N., Addya, S., Reyes, B. A. S., Van Bockstaele, E. J., Szewczyk, A., Wallace, D. C. and Singh, H. (2019). BKCa (Slo) Channel Regulates Mitochondrial Function and Lifespan in Drosophila melanogaster. Cells 8(9). PubMed ID: 31438578
BKCa channels, originally discovered in Drosophila melanogaster as slowpoke (slo), are recognized for their roles in cellular and organ physiology. Pharmacological approaches implicated BKCa channels in cellular and organ protection possibly for their ability to modulate mitochondrial function. However, the direct role of BKCa channels in regulating mitochondrial structure and function is not deciphered. This study demonstrates that BKCa channels are present in fly mitochondria, and slo mutants show structural and functional defects in mitochondria. slo mutants display an increase in reactive oxygen species and the modulation of ROS affected their survival. The absence of BKCa channels reduced the lifespan of Drosophila, and overexpression of human BKCa channels in flies extends life span in males. This study establishes the presence of BKCa channels in mitochondria of Drosophila and ascertains its novel physiological role in regulating mitochondrial structural and functional integrity, and lifespan.

Friday, September 20th - Adult Nervous System Development and Function

Gao, Y., Shuai, Y., Zhang, X., Peng, Y., Wang, L., He, J., Zhong, Y. and Li, Q. (2019). Genetic dissection of active forgetting in labile and consolidated memories in Drosophila. Proc Natl Acad Sci U S A. PubMed ID: 31488722
Different memory components are forgotten through distinct molecular mechanisms. In Drosophila, the activation of 2 Rho GTPases (Rac1 and Cdc42), respectively, underlies the forgetting of an early labile memory (anesthesia-sensitive memory, ASM) and a form of consolidated memory (anesthesia-resistant memory, ARM). This study dissected the molecular mechanisms that tie Rac1 and Cdc42 to the different types of memory forgetting. Two WASP family proteins, SCAR/WAVE and WASp, act downstream of Rac1 and Cdc42 separately to regulate ASM and ARM forgetting in mushroom body neurons. Arp2/3 complex, which organizes branched actin polymerization, is a canonical downstream effector of WASP family proteins. However, this study found that Arp2/3 complex is required in Cdc42/WASp-mediated ARM forgetting but not in Rac1/SCAR-mediated ASM forgetting. Instead, Rac1/SCAR may function with formin Diaphanous (Dia), a nucleator that facilitates linear actin polymerization, in ASM forgetting. The present study, complementing the previously identified Rac1/cofilin pathway that regulates actin depolymerization, suggests that Rho GTPases regulate forgetting by recruiting both actin polymerization and depolymerization pathways. Moreover, Rac1 and Cdc42 may regulate different types of memory forgetting by tapping into different actin polymerization mechanisms.
Goyal, G., Zierau, A., Lattemann, M., Bergkirchner, B., Javorski, D., Kaur, R. and Hummel, T. (2019). Inter-axonal recognition organizes Drosophila olfactory map formation. Sci Rep 9(1): 11554. PubMed ID: 31399611
Olfactory systems across the animal kingdom show astonishing similarities in their morphological and functional organization. In mouse and Drosophila, olfactory sensory neurons are characterized by the selective expression of a single odorant receptor (OR) type and by the OR class-specific connection in the olfactory brain center. Monospecific OR expression in mouse provides each sensory neuron with a unique recognition identity underlying class-specific axon sorting into synaptic glomeruli. This study shows that in Drosophila, although OR genes are not involved in sensory neuron connectivity, afferent sorting via OR class-specific recognition defines a central mechanism of odortopic map formation. Sensory neurons mutant for the Ig-domain receptor Dscam converge into ectopic glomeruli with single OR class identity independent of their target cells. Mosaic analysis showed that Dscam prevents premature recognition among sensory axons of the same OR class. Single Dscam isoform expression in projecting axons revealed the importance of Dscam diversity for spatially restricted glomerular convergence. These data support a model in which the precise temporal-spatial regulation of Dscam activity controls class-specific axon sorting thereby indicating convergent evolution of olfactory map formation via self-patterning of sensory neurons.
Guo, H., Kunwar, K. and Smith, D. (2019). Multiple channels of DEET repellency in Drosophila. Pest Manag Sci. PubMed ID: 31429190
DEET (N,N-diethyl-m-toluamide) is the most widely used prophylactic insect repellent to inhibit insect bites. Despite its use since 1944, the mechanism for DEET repellency remains controversial. This study revisited the role of smell and taste in DEET repellence using Drosophila as a model. Analysis of the responses of individual olfactory receptor neuron (ORN) classes to DEET reveals that 11 ORNs are activated and 2 ORNs are inhibited by this compound. Blocking individual ORN classes in the antenna does not block DEET repellence. This argues against the existence of a single ORN mediating DEET repellence in Drosophila. Activation of all ORCO-expressing neurons using channelrhodopsin favors attraction, not repellence, in behavioral valence. It was also demonstrated that gustatory neurons are highly sensitive to DEET. RNAi was used to screen candidate receptors encoded by gene families involved in the detection of bitter compounds, including 34 gustatory receptors (Grs), 14 ionotropic receptors (Irs), 5 pick-pocket subunits (PPKs), 3 transient receptor potential ion channels (TrpA, TrpL, Painless) and 1 metabotropic glutamate receptors gene (DmXR). Striking defects in DEET-mediated oviposition behavior were seen when expression of either Gr32a or Gr33a were inhibited. These findings support a multimodal mechanism for DEET detection in fruit flies and indicate a prominent role for taste detection mediating DEET repellence.
Ayoub, A. E., Dominguez, M. H., Benoit, J., Ortega, J. A., Radonjic, N., Zecevic, N. and Rakic, P. (2019). Coordination of neuron production in mouse and human cerebral cortex by the homolog of Drosophila Mastermind Protein. Brain Behav Evol 93(2-3): 152-165. PubMed ID: 31416089
The coordination of progenitor self-renewal, neuronal production, and migration is essential to the normal development and evolution of the cerebral cortex. Numerous studies have shown that the Notch, Wnt/beta-catenin, and Neurogenin pathways contribute separately to progenitor expansion, neurogenesis, and neuronal migration, but it is unknown how these signals are coordinated. In vitro studies suggested that the mastermind-like 1 (MAML1) gene, homologue of the Drosophila mastermind, plays a role in coordinating the aforementioned signaling pathways, yet its role during cortical development remains largely unknown. This study shows that ectopic expression of dominant-negative MAML (dnMAML) causes exuberant neuronal production in the mouse cortex without disrupting neuronal migration. Comparing the transcriptional consequences of dnMAML and Neurog2 ectopic expression revealed a complex genetic network controlling the balance of progenitor expansion versus neuronal production. Manipulation of MAML and Neurog2 in cultured human cerebral stem cells exposed interactions with the same set of signaling pathways. Thus, these data suggest that evolutionary changes that affect the timing, tempo, and density of successive neuronal layers of the small lissencephalic rodent and large convoluted primate cerebral cortex depend on similar molecular mechanisms that act from the earliest developmental stages.
Papanikolopoulou, K., Roussou, I. G., Gouzi, J. Y., Samiotaki, M., Panayotou, G., Turin, L. and Skoulakis, E. M. (2019). Drosophila Tau negatively regulates translation and olfactory Long-Term Memory, but facilitates footshock habituation and cytoskeletal homeostasis. J Neurosci. PubMed ID: 31488613
Although the involvement of pathological Tau in neurodegenerative dementias is indisputable, its physiological roles have remained elusive in part because its abrogation has been reported without overt phenotypes in mice and Drosophila. This was addressed using the recently described Drosophila tau(KO) and Mi mutants; Focus was placed on molecular and behavioural analyses. Initially it was shown that dTau loss precipitates dynamic cytoskeletal changes in the adult Drosophila CNS and translation upregulation. Significantly, distinct roles were demonstrated for dTau in adult Mushroom Body (MB)-dependent neuroplasticity as its down-regulation within alpha beta neurons impairs habituation. In accord with its negative regulation of translation, dTau loss specifically enhances Protein Synthesis Dependent Long-Term Memory (PSD-LTM), but not Anaesthesia-Resistant Memory. In contrast, elevation of the protein in the MBs yielded premature habituation and depressed PSD-LTM. Therefore, Tau loss in Drosophila dynamically alters brain cytoskeletal dynamics and profoundly affects neuronal proteostasis and plasticity.
Chen, Y. D., Ahmad, S., Amin, K. and Dahanukar, A. (2019). A subset of brain neurons controls regurgitation in adult Drosophila melanogaster. J Exp Biol. PubMed ID: 31511344
Taste is essential for animals to evaluate food quality and make important decisions about food choice and intake. How complex brains process sensory information to produce behavior is an essential question in the field of sensory neurobiology. Currently, little is known about higher order taste circuits in the brain as compared to those of other sensory systems. This study used the common vinegar fly, Drosophila melanogaster, to screen for candidate neurons labeled by different transgenic GAL4 lines in controlling feeding behaviors. Activation of one line (VT041723-GAL4) produces "proboscis holding" behavior (extrusion of the mouthpart without withdrawal). Further analysis shows that the proboscis holding phenotype indicates an aversive response, since flies pre-fed with either sucrose or water prior to neuronal activation exhibit regurgitation. Anatomical characterization of VT041723-GAL4 labeled neurons suggests that they receive sensory input from peripheral taste neurons. Overall, this study identifies a subset of brain neurons labeled by VT041723-GAL4 that may be involved in a taste circuit that controls regurgitation.

Thursday, September 19th - Behavior

Pauls, D., Hamarat, Y., Trufasu, L., Schendzielorz, T. M., Gramlich, G., Kahnt, J., Vanselow, J. T., Schlosser, A. and Wegener, C. (2019). Drosophila carboxypeptidase D (SILVER) is a key enzyme in neuropeptide processing required to maintain locomotor activity levels and survival rate. Eur J Neurosci. PubMed ID: 31309630
Neuropeptides are processed from larger preproproteins by a dedicated set of enzymes. In contrast to mammals, Drosophila melanogaster lacks a gene for carboxypeptidase E (CPE), a key enzyme for mammalian peptide processing. By combining peptidomics and neurogenetics, this study addressed the role of carboxypeptidase D (dCPD) in global neuropeptide processing and selected peptide-regulated behaviours in Drosophila. A deficiency in dCPD results in C-terminally extended peptides across the peptidome, suggesting that dCPD took over CPE function in the fruit fly. dCPD is widely expressed throughout the nervous system, including peptidergic neurons in the mushroom body and neuroendocrine cells expressing adipokinetic hormone. Conditional hypomorphic mutation in the dCPD-encoding gene silver in the larva causes lethality, and leads to deficits in starvation-induced hyperactivity and appetitive gustatory preference, as well as to reduced viability and activity levels in adults. A phylogenomic analysis suggests that loss of CPE is not common to insects, but only occurred in Hymenoptera and Diptera. These results show that dCPD is a key enzyme for neuropeptide processing and peptide-regulated behaviour in Drosophila. dCPD thus appears as a suitable target to genetically shut down total neuropeptide production in peptidergic neurons. The persistent occurrence of CPD in insect genomes may point to important further CPD functions beyond neuropeptide processing which cannot be fulfilled by CPE.
Krittika, S., Indhumathi, P., Vedha Hari, B. N., Ramya Devi, D. and Yadav, P. (2019). Evidence of nanoemulsion as an effective control measure for fruit flies Drosophila melanogaster. Sci Rep 9(1): 10578. PubMed ID: 31332229
Pesticide resistance is a common concern. It exerts close association with economic and health associated problems in various plants and other organisms. Several approaches have been trialled for attracting and trapping the insects and flies that are acting as vectors for transmission of communicable diseases. Although Drosophila melanogaster (fruit flies) is not an agricultural pest, its presence in consumer dwelling areas is an objection to human, as it indicates signs of an unhealthy environment or products. The current study focuses on the development of nanoemulsion with synthetic attractants and entrapping in sticky glue formulation that could provide prolonged effect for attracting and trapping the fruit flies. The results of this study showed the efficient attractive ability of exposed nanoemulsion (A3E1T) containing amyl acetate, ammonia, ethanol and Tween 80 compared to that of control. While the sex-based effect was not very prominent, the nanoemulsion showed a higher relative response index to the flies and increased activity even during their siesta time. Therefore, the nanoemulsion-based approach could be identified as one of the promising lines of attack and a suitable alternative for the existing fruit fly control measures. The present study is the first of its kind in reporting the ability of nanoemulsion formulation to attract and influence the activity of fruit flies D. melanogaster.
Sato, K., Ahsan, M. T., Ote, M., Koganezawa, M. and Yamamoto, D. (2019). Calmodulin-binding transcription factor shapes the male courtship song in Drosophila. PLoS Genet 15(7): e1008309. PubMed ID: 31344027
Males of the Drosophila melanogaster mutant croaker (cro) generate a polycyclic pulse song dissimilar to the monocyclic songs typical of wild-type males during courtship. However, cro has not been molecularly mapped to any gene in the genome. This study demonstrates that cro is a mutation in the gene encoding the Calmodulin-binding transcription factor (Camta) by genetic complementation tests with chromosomal deficiencies, molecular cloning of genomic fragments that flank the cro-mutagenic P-insertion, and phenotypic rescue of the cro mutant phenotype by Camta+-encoding cDNA as well as a BAC clone containing the gene for Camta. It was shown that knockdown of the Camta-encoding gene phenocopies cro mutant songs when targeted to a subset of fruitless-positive neurons that include the mcALa and AL1 clusters in the brain. cro-GAL4 and an anti-Camta antibody labeled a large number of brain neurons including mcALa. It is concluded that the Camta-encoding gene represents the cro locus, which has been implicated in a species-specific difference in courtship songs between D. sechellia and simulans.
Musso, P. Y., Junca, P., Jelen, M., Feldman-Kiss, D., Zhang, H., Chan, R. C. and Gordon, M. D. (2019). Closed-loop optogenetic activation of peripheral or central neurons modulates feeding in freely moving Drosophila. Elife 8. PubMed ID: 31322499
Manipulating feeding circuits in freely moving animals is challenging, in part because the timing of sensory inputs is affected by the animal's behavior. To address this challenge in Drosophila, the Sip-Triggered Optogenetic Behavior Enclosure ('STROBE') was developed. The STROBE is a closed-looped system for real-time optogenetic activation of feeding flies, designed to evoke neural excitation coincident with food contact. Previous work has demonstrated the STROBE's utility in probing the valence of fly sensory neurons. This study provides a thorough characterization of the STROBE system, demonstrates that STROBE-driven behavior is modified by hunger and the presence of taste ligands, and found that mushroom body dopaminergic input neurons and their respective post-synaptic partners drive opposing feeding behaviors following activation. Together, these results establish the STROBE as a new tool for dissecting fly feeding circuits and suggest a role for mushroom body circuits in processing naive taste responses.
Akhund-Zade, J., Ho, S., O'Leary, C. and de Bivort, B. (2019). The effect of environmental enrichment on behavioral variability depends on genotype, behavior, and type of enrichment. J Exp Biol. PubMed ID: 31413102
Non-genetic individuality in behavior, also termed intragenotypic variability, has been observed across many different organisms. A potential cause of intragenotypic variability is sensitivity to minute environmental differences during development, even as major environmental parameters are kept constant. Animal enrichment paradigms often include the addition of environmental diversity, whether in the form of social interaction, novel objects, or exploratory opportunities. Enrichment could plausibly affect intragenotypic variability in opposing ways: it could cause an increase in variability due to the increase in microenvironmental variation, or a decrease in variability due to elimination of aberrant behavior as animals are taken out of impoverished laboratory conditions. In order to test this hypothesis, five isogenic Drosophila melanogaster lines raised in control and mild enrichment conditions were assayed, and one isogenic line was assayed under both mild and intense enrichment conditions. The mean and variability were compared of six behavioral metrics between the enriched fly populations and the laboratory housing control. Enrichment often caused a small increase in variability across most of the behaviors, but the ultimate effect of enrichment on both behavioral means and variabilities was highly dependent on genotype and its interaction with the particular enrichment treatment. These results support previous work on enrichment that presents a highly variable picture of its effects on both behavior and physiology.
Gunther, C. S., Knight, S. J., Jones, R. and Goddard, M. R. (2019). Are Drosophila preferences for yeasts stable or contextual?. Ecol Evol 9(14): 8075-8086. PubMed ID: 31380072
Whether there are general mechanisms, driving interspecific chemical communication is uncertain. Saccharomycetaceae yeast and Drosophila fruit flies, both extensively studied research models, share the same fruit habitat, and it has been suggested their interaction comprises a facultative mutualism that is instigated and maintained by yeast volatiles. Using choice tests, experimental evolution, and volatile analyses, this study investigated the maintenance of this relationship and reveal little consistency between behavioral responses of two isolates of sympatric Drosophila species. While D. melanogaster was attracted to a range of different Saccharomycetaceae yeasts and this was independent of fruit type, D. simulans preference appeared specific to a particular S. cerevisiae genotype isolated from a vineyard fly population. This response, however, was not consistent across fruit types and is therefore context-dependent. In addition, D. simulans attraction to an individual S. cerevisiae isolate was pliable over ecological timescales. Volatile candidates were analyzed to identify a common signal for yeast attraction, and while D. melanogaster generally responded to fermentation profiles, D. simulans preference was more discerning and likely threshold-dependent. Overall, there is no strong evidence to support the idea of bespoke interactions with specific yeasts for either of these Drosophila genotypes. Rather the data support the idea Drosophila are generally adapted to sense and locate fruits infested by a range of fungal microbes and/or that yeast-Drosophila interactions may evolve rapidly.

Wednesday, September 18th - Synapse and Vesicles

Blanco-Redondo, B., Nuwal, N., Kneitz, S., Nuwal, T., Halder, P., Liu, Y., Ehmann, N., Scholz, N., Mayer, A., Kleber, J., Kahne, T., Schmitt, D., Sadanandappa, M. K., Funk, N., Albertova, V., Helfrich-Forster, C., Ramaswami, M., Hasan, G., Kittel, R. J., Langenhan, T., Gerber, B. and Buchner, E. (2019). Implications of the Sap47 null mutation for synapsin phosphorylation, longevity, climbing, and behavioural plasticity in adult Drosophila. J Exp Biol. PubMed ID: 31488622
The Sap47 gene of Drosophila melanogaster encodes a highly abundant 47 kDa synaptic vesicle-associated protein. Sap47 null mutants show defects in synaptic plasticity and larval olfactory associative learning but the molecular function of Sap47 at the synapse is unknown. This study demonstrates that Sap47 modulates the phosphorylation of another highly abundant conserved presynaptic protein, Synapsin. Site-specific phosphorylation of Drosophila Synapsin has repeatedly been shown to be important for behavioural plasticity but information of where in the brain phospho-synapsin isoforms are localized has been lacking. This study reports the distribution of serine-6-phosphorylated Synapsin in the adult brain and shows that it is highly enriched in rings of synapses in the ellipsoid body, and in large synapses near the lateral triangle. Effects of knock-out of Sap47 or Synapsin on olfactory associative learning/memory are compatible with the hypothesis that both proteins operate in the same molecular pathway. It was therefore asked if this might also hold for other aspects of their function. Knock-out of Sap47 but not synapsin reduces life span, whereas knock-out of Sap47, synapsin or both affects climbing as well as plasticity in circadian rhythms and sleep. Furthermore, electrophysiological assessment of synaptic properties at the larval neuromuscular junction (NMJ) reveals increased spontaneous synaptic vesicle fusion and reduced paired pulse facilitation in Sap47 and synapsin single and double mutants. These results imply that Sap47 and Synapsin cooperate non-uniformly in the control of synaptic properties in different behaviourally relevant neuronal networks of the fruitfly.
Troup, M., Zalucki, O. H., Kottler, B. D., Karunanithi, S., Anggono, V. and van Swinderen, B. (2019). Syntaxin1A neomorphic mutations promote rapid recovery from isoflurane anesthesia in Drosophila melanogaster. Anesthesiology. PubMed ID: 31356232
Syntaxin1A is a presynaptic molecule that plays a key role in vesicular neurotransmitter release. Mutations of syntaxin1A result in resistance to both volatile and intravenous anesthetics. Truncated syntaxin1A isoforms confer drug resistance in cell culture and nematode models of anesthesia Resistance to isoflurane anesthesia can be produced by transiently expressing truncated syntaxin1A proteins in adult Drosophila flies. Electrophysiologic and behavioral studies in Drosophila show that mutations in syntaxin1A facilitate recovery from isoflurane anesthesia. These observations suggest that presynaptic mechanisms, via syntaxin1A-mediated regulation of neurotransmitter release, are involved in general anesthesia maintenance and recovery Mutations in the presynaptic protein syntaxin1A modulate general anesthetic effects in vitro and in vivo. Coexpression of a truncated syntaxin1A protein confers resistance to volatile and intravenous anesthetics, suggesting a target mechanism distinct from postsynaptic inhibitory receptor processes. Hypothesizing that recovery from anesthesia may involve a presynaptic component, whether synatxin1A mutations facilitated recovery from isoflurane anesthesia in Drosophila melanogaster was tested. The same neomorphic syntaxin1A mutation that confers isoflurane resistance in cell culture and nematodes also produces isoflurane resistance in Drosophila. Resistance in Drosophila is, however, most evident at the level of recovery from anesthesia, suggesting that the syntaxin1A target affects anesthesia maintenance and recovery processes rather than induction. The absence of truncated syntaxin1A from the presynaptic complex suggests that the resistance-promoting effect of this molecule occurs before core complex formation.
Xiong, L., Zhang, L., Yang, Y., Li, N., Lai, W., Wang, F., Zhu, X. and Wang, T. (2019). ER complex proteins are required for rhodopsin biosynthesis and photoreceptor survival in Drosophila and mice. Cell Death Differ. PubMed ID: 31263175
Defective rhodopsin homeostasis is one of the major causes of retinal degeneration, including the disease Retinitis pigmentosa. To identify cellular factors required for the biosynthesis of rhodopsin, a genome-wide genetic screen was performed in Drosophila for mutants with reduced levels of rhodopsin. Loss-of-function alleles were investigated in endoplasmic reticulum membrane protein complex 3 (emc3), emc5, and emc6, each of which exhibited defective phototransduction and photoreceptor cell degeneration. EMC3, EMC5, and EMC6 were essential for rhodopsin synthesis independent of the ER associated degradation (ERAD) pathway, which eliminates misfolded proteins. Null mutations were generated for all EMC subunits; it was further demonstrated that different EMC subunits play roles in different cellular functions. Conditional knockout of the Emc3 gene in mice led to mislocalization of rhodopsin protein and death of cone and rod photoreceptor cells. These data indicate conserved roles for EMC subunits in maintaining rhodopsin homeostasis and photoreceptor function, and suggest that retinal degeneration may also be caused by defects in early biosynthesis of rhodopsin.
Bulgari, D., Deitcher, D. L., Schmidt, B. F., Carpenter, M. A., Szent-Gyorgyi, C., Bruchez, M. P. and Levitan, E. S. (2019). Activity-evoked and spontaneous opening of synaptic fusion pores. Proc Natl Acad Sci U S A 116(34): 17039-17044. PubMed ID: 31383765
Synaptic release of neuropeptides packaged in dense-core vesicles (DCVs) regulates synapses, circuits, and behaviors including feeding, sleeping, and pain perception. This study imaged synaptic DCV fusion pore openings without interference from cotransmitting small synaptic vesicles (SSVs) with the use of a fluorogen-activating protein (FAP). Activity-evoked kiss and run exocytosis opens synaptic DCV fusion pores away from active zones that readily conduct molecules larger than most native neuropeptides (i.e., molecular weight [MW] up to, at least, 4.5 kDa). Remarkably, these synaptic fusion pores also open spontaneously in the absence of stimulation and extracellular Ca(2+) SNARE perturbations demonstrate different mechanisms for activity-evoked and spontaneous fusion pore openings with the latter sharing features of spontaneous small molecule transmitter release by active zone-associated SSVs. Fusion pore opening at resting synapses provides a mechanism for activity-independent peptidergic transmission.
Wang, S., Zhao, Z. and Rodal, A. A. (2019). Higher-order assembly of Sorting Nexin 16 controls tubulation and distribution of neuronal endosomes. J Cell Biol. PubMed ID: 31253649
The activities of neuronal signaling receptors depend heavily on the maturation state of the endosomal compartments in which they reside. However, it remains unclear how the distribution of these compartments within the uniquely complex morphology of neurons is regulated and how this distribution itself affects signaling. This study identified mechanisms by which Sorting Nexin 16 (SNX16) controls neuronal endosomal maturation and distribution. Higher-order assembly of SNX16 via its coiled-coil (CC) domain was found to drive membrane tubulation in vitro and endosome association in cells. In Drosophila melanogaster motor neurons, activation of Rab5 and CC-dependent self-association of SNX16 lead to its endosomal enrichment, accumulation in Rab5- and Rab7-positive tubulated compartments in the cell body, and concomitant depletion of SNX16-positive endosomes from the synapse. This results in accumulation of synaptic growth-promoting bone morphogenetic protein receptors in the cell body and correlates with increased synaptic growth. These results indicate that Rab regulation of SNX16 assembly controls the endosomal distribution and signaling activities of receptors in neurons.
Chen, P. Y., Tsai, Y. W., Cheng, Y. J., Giangrande, A. and Chien, C. T. (2019). Glial response to hypoxia in mutants of NPAS1/3 homolog Trachealess through Wg signaling to modulate synaptic bouton organization. PLoS Genet 15(8): e1007980. PubMed ID: 31381576
Synaptic structure and activity are sensitive to environmental alterations. Modulation of synaptic morphology and function is often induced by signals from glia. However, the process by which glia mediate synaptic responses to environmental perturbations such as hypoxia remains unknown. In the mutant for Trachealess (Trh), the Drosophila homolog for NPAS1 and NPAS3, smaller synaptic boutons form clusters named satellite boutons appear at larval neuromuscular junctions (NMJs), which is induced by the reduction of internal oxygen levels due to defective tracheal branches. Thus, the satellite bouton phenotype in the trh mutant is suppressed by hyperoxia, and recapitulated in wild-type larvae raised under hypoxia. It was further shown that hypoxia-inducible factor (HIF)-1alpha/Similar (Sima) is critical in mediating hypoxia-induced satellite bouton formation. Sima upregulates the level of the Wnt/Wingless (Wg) signal in glia, leading to reorganized microtubule structures within presynaptic sites. Finally, hypoxia-induced satellite boutons maintain normal synaptic transmission at the NMJs, which is crucial for coordinated larval locomotion.

Tuesday, September 17th - Disease Models

Penserga, T., Kudumala, S. R., Poulos, R. and Godenschwege, T. A. (2019). A role for Drosophila Amyloid precursor protein in retrograde trafficking of L1-Type cell adhesion molecule Neuroglian. Front Cell Neurosci 13: 322. PubMed ID: 31354437
The normal function of APP in the nervous system is poorly understood. This study characterized the role of the Drosophila homolog (APPL) in the adult giant fiber (GF) neurons. Endogenous APPL is transported from the synapse to the soma in the adult. Live-imaging revealed that retrograde moving APPL vesicles co-traffic with L1-type cell adhesion molecule Neuroglian (Nrg). In APPL null mutants, stationary Nrg vesicles were increased along the axon, and the number of Nrg vesicles moving in retrograde but not anterograde direction was reduced. In contrast, trafficking of endo-lysosomal vesicles, which did not co-localize with APPL in GF axons, was not affected. This suggests that APPL loss of function does not generally disrupt axonal transport but that APPL has a selective role in the effectiveness of retrograde transport of proteins it co-traffics with. While the GF terminals of Loss of function animals exhibited pruning defects. Cell-autonomous developmental expression of a secretion-deficient form of APPL (APPL-SD), lacking the alpha-, beta-, and, gamma-secretase cleavage sites, resulted in progressive retraction of the GF terminals. Conditional expression of APPL-SD in mature GFs caused accumulation of Nrg in normal sized synaptic terminals, which was associated with severely reduced retrograde flux of Nrg labeled vesicles in the axons. Albeit beta-secretase null mutants developed GF terminals they also exhibited Nrg accumulations. This suggests that cleavage defective APPL has a toxic effect on retrograde trafficking and that beta-secretase cleavage has a function in Nrg sorting in endosomal compartments at the synapse.
Chatterjee, S., Ambegaokar, S. S., Jackson, G. R. and Mudher, A. (2019). Insulin-mediated changes in Tau hyperphosphorylation and autophagy in a Drosophila model of tauopathy and neuroblastoma cells. Front Neurosci 13: 801. PubMed ID: 31427921
Almost 50 million people in the world are affected by dementia; the most prevalent form of which is Alzheimer's disease (AD). Although aging is considered to be the main risk factor for AD, growing evidence from epidemiological studies suggests that type 2 diabetes mellitus (T2DM) increases the risk of dementia including AD. Defective brain insulin signaling has been suggested as an early event in AD and other tauopathies but the mechanisms that link these diseases are largely unknown. Tau hyperphosphorylation is a hallmark of neurofibrillary pathology and insulin resistance increases the number of neuritic plaques particularly in AD. Utilizing a combination of Drosophila models of tauopathy (expressing the 2N4R-Tau) and neuroblastoma cells, this study has attempted to decipher the pathways downstream of the insulin signaling cascade that lead to tau hyperphosphorylation, aggregation and autophagic defects. Using cell-based, genetic, and biochemical approaches this study demonstrated that tau phosphorylation at AT8 and PHF1 residues is enhanced in an insulin-resistant environment. It was also shown that insulin-induced changes in total and phospho-tau are mediated by the crosstalk of AKT, glycogen synthase kinase-3beta, and extracellular regulating kinase located downstream of the insulin receptor pathway. Finally, this study demonstrated a significant change in the levels of the key proteins in the mammalian target of rapamycin/autophagy pathway, implying an increased impairment of aggregated protein clearance in the transgenic Drosophila models and cultured neuroblastoma cells.
Olsen, A. L. and Feany, M. B. (2019). Glial alpha-synuclein promotes neurodegeneration characterized by a distinct transcriptional program in vivo. Glia. PubMed ID: 31267577
alpha-Synucleinopathies are neurodegenerative diseases that are characterized pathologically by alpha-synuclein inclusions in neurons and glia. The pathologic contribution of glial alpha-synuclein in these diseases is not well understood. Glial alpha-synuclein may be of particular importance in multiple system atrophy (MSA), which is defined pathologically by glial cytoplasmic alpha-synuclein inclusions. Previous work has described Drosophila models of neuronal alpha-synucleinopathy, which recapitulate key features of the human disorders. This study now expands this model to express human alpha-synuclein in glia. Expression of alpha-synuclein in glia alone results in alpha-synuclein aggregation, death of dopaminergic neurons, impaired locomotor function, and autonomic dysfunction. Furthermore, co-expression of alpha-synuclein in both neurons and glia worsens these phenotypes as compared to expression of alpha-synuclein in neurons alone. Unique transcriptomic signatures induced by glial as opposed to neuronal alpha-synuclein were identified. These results suggest that glial alpha-synuclein may contribute to the burden of pathology in the alpha-synucleinopathies through a cell type-specific transcriptional program. This new Drosophila model system enables further mechanistic studies dissecting the contribution of glial and neuronal alpha-synuclein in vivo, potentially shedding light on mechanisms of disease that are especially relevant in MSA but also the alpha-synucleinopathies more broadly.
Wu, W., Du, S., Shi, W., Liu, Y., Hu, Y., Xie, Z., Yao, X., Liu, Z., Ma, W., Xu, L., Ma, C. and Zhong, Y. (2019). Inhibition of Rac1-dependent forgetting alleviates memory deficits in animal models of Alzheimer's disease. Protein Cell. PubMed ID: 31321704
Accelerated forgetting has been identified as a feature of Alzheimer's disease (AD), but the therapeutic efficacy of the manipulation of biological mechanisms of forgetting has not been assessed in AD animal models. Ras-related C3 botulinum toxin substrate 1 (Rac1), a small GTPase, has been shown to regulate active forgetting in Drosophila and mice. This study has shown that Rac1 activity is aberrantly elevated in the hippocampal tissues of AD patients and AD animal models. Moreover, amyloid-beta 42 could induce Rac1 activation in cultured cells. The elevation of Rac1 activity not only accelerated 6-hour spatial memory decay in 3-month-old APP/PS1 mice, but also significantly contributed to severe memory loss in aged APP/PS1 mice. A similar age-dependent Rac1 activity-based memory loss was also observed in an AD fly model. Moreover, inhibition of Rac1 activity could ameliorate cognitive defects and synaptic plasticity in AD animal models. Finally, two novel compounds, identified through behavioral screening of a randomly selected pool of brain permeable small molecules for their positive effect in rescuing memory loss in both fly and mouse models, were found to be capable of inhibiting Rac1 activity. Thus, multiple lines of evidence corroborate in supporting the idea that inhibition of Rac1 activity is effective for treating AD-related memory loss.
Cabasso, O., Paul, S., Dorot, O., Maor, G., Krivoruk, O., Pasmanik-Chor, M., Mirzaian, M., Ferraz, M., Aerts, J. and Horowitz, M. (2019). Drosophila melanogaster mutated in its GBA1b ortholog recapitulates Neuronopathic Gaucher disease. J Clin Med 8(9). PubMed ID: 31505865
Gaucher disease (GD) results from mutations in the GBA1 gene, which encodes lysosomal glucocerebrosidase (GCase). The two fly GBA1 orthologs, GBA1a and GBA1b each contains a Minos element insertion, which truncates its coding sequence. In the GBA1a(m/m) flies, which express a mutant protein, missing 33 C-terminal amino acids, there was no decrease in GCase activity or substrate accumulation. However, GBA1b(m/m) mutant flies presented a significant decrease in GCase activity with concomitant substrate accumulation, which included C14:1 glucosylceramide and C14:0 glucosylsphingosine. GBA1b(m/m) mutant flies showed activation of the Unfolded Protein Response (UPR) and presented inflammation and neuroinflammation that culminated in development of a neuronopathic disease. Treatment with ambroxol did not rescue GCase activity or reduce substrate accumulation; however, it ameliorated UPR, inflammation and neuroinflammation, and increased life span. These results highlight the resemblance between the phenotype of the GBA1b(m/m) mutant fly and neuronopathic GD and underlie its relevance in further GD studies as well as a model to test possible therapeutic modalities.
Chang, Y. H. and Dubnau, J. (2019). The gypsy endogenous retrovirus drives non-cell-autonomous propagation in a Drosophila TDP-43 model of neurodegeneration. Curr Biol. PubMed ID: 31495585
A hallmark of neurodegenerative disease is focal onset of pathological protein aggregation, followed by progressive spread of pathology to connected brain regions. In amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD), pathology is often associated with aggregation of TAR DNA-binding protein 43 (TDP-43). Although aggregated TDP-43 protein moves between cells, it is not clear whether and how this movement propagates the degeneration. A Drosophila model of human TDP-43 was established in which toxic expression of human TDP-43 was initiated focally within small groups of glial cells. This focal onset kills adjacent neurons. Surprisingly, this spreading death is caused by an endogenous retrovirus within the glia, which leads to DNA damage and death in adjacent neurons. These findings suggest a possible mechanism by which human retroviruses such as HERV-K might contribute to TDP-43-mediated propagation of neurodegeneration.

Monday, September 16th - Apoptosis

Aparicio, R., Rana, A. and Walker, D. W. (2019). Upregulation of the autophagy adaptor p62/SQSTM1 prolongs health and lifespan in middle-aged Drosophila. Cell Rep 28(4): 1029-1040. PubMed ID: 31340141
Autophagy, a lysosomal degradation pathway, plays crucial roles in health and disease. p62/SQSTM1 (hereafter p62) is an autophagy adaptor protein that can shuttle ubiquitinated cargo for autophagic degradation. This study shows that upregulating the Drosophila p62 homolog ref(2)P/dp62, starting in midlife, delays the onset of pathology and prolongs healthy lifespan. Midlife induction of dp62 improves proteostasis, in aged flies, in an autophagy-dependent manner. Previous studies have reported that p62 plays a role in mediating the clearance of dysfunctional mitochondria via mitophagy. However, the causal relationships between p62 expression, mitochondrial homeostasis, and aging remain largely unexplored. This study shows that upregulating dp62, in midlife, promotes mitochondrial fission, facilitates mitophagy, and improves mitochondrial function in aged flies. Finally, this study shows that mitochondrial fission is required for the anti-aging effects of midlife dp62 induction. These findings indicate that p62 represents a potential therapeutic target to counteract aging and prolong health in aged mammals.
Ghosh, N., Bakshi, A., Khandelwal, R., Rajan, S. G. and Joshi, R. (2019). Hox gene Abdominal-B uses Doublesex(F) as a cofactor to promote neuroblast apoptosis in Drosophila central nervous system. Development. PubMed ID: 31371379
Highly conserved DM domain containing transcription factors (Doublesex/MAB-3/DMRT1) are responsible for generating sexually dimorphic features. In Drosophila CNS a set of Doublesex (Dsx) expressing neuroblasts undergo apoptosis in females while their male counterparts proliferate and give rise to serotonergic neurons crucial for adult mating behaviour. This study study demonstrates that female specific isoform of Doublesex collaborates with Hox gene Abdominal-B (AbdB) to bring about this apoptosis. Biochemical results suggest AbdB and Dsx interact through their highly conserved Homeodomain and DM domains respectively. This interaction is translated into a cooperative binding of the two proteins (AbdB and Dsx) on the apoptotic enhancer in case of females but not in case of males, resulting in female specific activation of apoptotic genes. The capacity of AbdB to utilize sex specific isoform of Dsx as a cofactor underlines the possibility that two classes of proteins are capable of cooperating in selection and regulation of target genes in tissue and sex specific manner. It is proposed that this interaction could be a common theme in generating sexual dimorphism in different tissues across different species.
Zhang, P., Holowatyj, A. N., Ulrich, C. M. and Edgar, B. A. (2019). Tumor suppressive autophagy in intestinal stem cells controls gut homeostasis. Autophagy: 1-3. PubMed ID: 31213134
Re-routing intracellular vesicle traffic by suppressing macroautophagy/autophagy or endocytosis genes drastically deregulates Drosophila intestinal stem cell (ISC) proliferation, leading to massive gut hyperplasia that has a negative impact upon lifespan. Beginning with the poorly characterized Snx (sorting nexin) genes, this study surveyed a broad set of genes in the endocytosis-autophagy network and found that most of them have this effect. Deregulated Egfr-Ras85D/Ras1-mitogen-activated protein kinase signaling is the primary trigger for ISC proliferation upon disruption of this network; in the mutants, ligand-activated receptors were stabilized and recycled to the cell surface via Rab11-dependent endosomes, rather than being degraded via autophagosomes. This study profiled the mutational landscape for orthologous network genes in human cancers using The Cancer Genome Atlas (TCGA), and revealed strong, novel associations with distinct genomic and epigenomic subtypes of colorectal cancer.
Delos Santos, K., Kim, M., Yergeau, C., Jean, S. and Moon, N. S. (2019). Pleiotropic role of Drosophila phosphoribosyl pyrophosphate synthetase in autophagy and lysosome homeostasis. PLoS Genet 15(9): e1008376. PubMed ID: 31487280
Phosphoribosyl pyrophosphate synthetase (PRPS) is a rate-limiting enzyme whose function is important for the biosynthesis of purines, pyrimidines, and pyridines. Importantly, while missense mutations of PRPS1 have been identified in neurological disorders such as Arts syndrome, how they contribute to neuropathogenesis is still unclear. This study identified the Drosophila ortholog of PRPS (dPRPS) as a direct target of RB/E2F in Drosophila, a vital cell cycle regulator, and engineered dPRPS alleles carrying patient-derived mutations. Interestingly, while they are able to develop normally, dPRPS mutant flies have a shortened lifespan and locomotive defects, common phenotypes associated with neurodegeneration. Careful analysis of the fat body revealed that patient-derived PRPS mutations result in profound defects in lipolysis, macroautophagy, and lysosome function. Significantly, evidence is shown that the nervous system of dPRPS mutant flies is affected by these defects. Overall, this study has uncovered an unexpected link between nucleotide metabolism and autophagy/lysosome function, providing a possible mechanism by which PRPS-dysfunction contributes to neurological disorders.
Harsh, S., Heryanto, C. and Eleftherianos, I. (2019). Intestinal lipid droplets as novel mediators of host-pathogen interaction in Drosophila. Biol Open. PubMed ID: 31278163
Lipid droplets (LDs) are lipid carrying multifunctional organelles, which might also interact with pathogens and influence the host immune response. However, the exact nature of these interactions remains currently unexplored. This study shows that systemic infection of Drosophila adult flies with non-pathogenic E. coli, the extracellular bacterial pathogen P. luminescens or the facultative intracellular pathogen P. asymbiotica results in intestinal steatosis marked by lipid accumulation in the midgut. Accumulation of LDs in the midgut also correlates with increased whole-body lipid levels characterized by increased expression of genes regulating lipogenesis. The lipid enriched midgut further displays reduced expression of enteroendocrine secreted hormone, Tachykinin. The observed lipid accumulation requires the Gram-negative cell wall pattern recognition molecule PGRP-LC, but not PGRP-LE, for the humoral immune response. Altogether, these findings indicate that Drosophila LDs are inducible organelles, which can serve as marker for inflammation and depending on the nature of the challenge they can dictate the outcome of the infection.
Yalonetskaya, A., Mondragon, A. A., Hintze, Z. J., Holmes, S. and McCall, K. (2019). Nuclear degradation dynamics in a nonapoptotic programmed cell death. Cell Death Differ. PubMed ID: 31285547
Nuclear degradation is a major event during programmed cell death (PCD). The breakdown of nuclear components has been well characterized during apoptosis, one form of PCD. Many nonapoptotic forms of PCD have been identified, but understanding of nuclear degradation during those events is limited. This study took advantage of Drosophila oogenesis to investigate nuclear degeneration during stress-induced apoptotic and developmental nonapoptotic cell death in the same cell type in vivo. Nuclear Lamin, a caspase substrate, dissociates from the nucleus as an early event during apoptosis, but remains associated with nuclei during nonapoptotic cell death. Lamin reveals a series of changes in nuclear architecture during nonapoptotic death, including nuclear crenellations and involutions. Stretch follicle cells contribute to these architecture changes, and phagocytic and lysosome-associated machinery in stretch follicle cells promote Lamin degradation. More specifically, this study found that the lysosomal cathepsin CP1 facilitates Lamin degradation.

Friday, September 13th - Adult neural development and function

Stern, U., Srivastava, H., Chen, H. L., Mohammad, F., Claridge-Chang, A. and Yang, C. H. (2019). Learning a spatial task by trial and error in Drosophila. Curr Biol. PubMed ID: 31327716
The ability to use memory to return to specific locations for foraging is advantageous for survival. Although recent reports have demonstrated that the fruit flies Drosophila melanogaster are capable of visual cue-driven place learning and idiothetic path integration, the depth and flexibility of Drosophila's ability to solve spatial tasks and the underlying neural substrate and genetic basis have not been extensively explored. This study shows that Drosophila can remember a reward-baited location through reinforcement learning and do so quickly and without requiring vision. This study found that both sighted and blind flies can learn-by trial and error-to repeatedly return to an unmarked location where a brief stimulation of the 0273-GAL4 neurons was available for each visit. Optogenetic stimulation of these neurons enabled learning by employing both a cholinergic pathway that promoted self-stimulation and a dopaminergic pathway that likely promoted association of relevant cues with reward. Lastly, inhibiting activities of specific neurons time-locked with stimulation of 0273-GAL4 neurons showed that mushroom bodies (MB) and central complex (CX) both play a role in promoting learning of the task. This work uncovered new depth in flies' ability to learn a spatial task and established an assay with a level of throughput that permits a systematic genetic interrogation of flies' ability to learn spatial tasks.
Xu, C., Theisen, E., Maloney, R., Peng, J., Santiago, I., Yapp, C., Werkhoven, Z., Rumbaut, E., Shum, B., Tarnogorska, D., Borycz, J., Tan, L., Courgeon, M., Meinertzhagen, I. A., de Bivort, B., Drugowitsch, J. and Pecot, M. Y. (2019). Control of synaptic specificity by establishing a relative preference for synaptic partners. Neuron. PubMed ID: 31300277
The ability of neurons to identify correct synaptic partners is fundamental to the proper assembly and function of neural circuits. Relative to other steps in circuit formation such as axon guidance, knowledge of how synaptic partner selection is regulated is severely limited. Drosophila Dpr and DIP immunoglobulin superfamily (IgSF) cell-surface proteins bind heterophilically and are expressed in a complementary manner between synaptic partners in the visual system. This study shows that in the lamina, DIP mis-expression is sufficient to promote synapse formation with Dpr-expressing neurons and that disrupting DIP function results in ectopic synapse formation. These findings indicate that DIP proteins promote synapses to form between specific cell types and that in their absence, neurons synapse with alternative partners. It is proposed that neurons have the capacity to synapse with a broad range of cell types and that synaptic specificity is achieved by establishing a preference for specific partners.
Qin, B., Humberg, T. H., Kim, A., Kim, H. S., Short, J., Diao, F., White, B. H., Sprecher, S. G. and Yuan, Q. (2019). Muscarinic acetylcholine receptor signaling generates OFF selectivity in a simple visual circuit. Nat Commun 10(1): 4093. PubMed ID: 31501438
ON and OFF selectivity in visual processing is encoded by parallel pathways that respond to either light increments or decrements. Despite lacking the anatomical features to support split channels, Drosophila larvae effectively perform visually-guided behaviors. To understand principles guiding visual computation in the larval visual system, focus was placed on investigating the physiological properties and behavioral relevance of larval visual interneurons. The ON vs. OFF discrimination in the larval visual circuit emerges through light-elicited cholinergic signaling that depolarizes a cholinergic interneuron (cha-lOLP) and hyperpolarizes a glutamatergic interneuron (glu-lOLP). Genetic studies further indicate that muscarinic acetylcholine receptor (mAchR)/Galphao signaling produces the sign-inversion required for OFF detection in glu-lOLP, the disruption of which strongly impacts both physiological responses of downstream projection neurons and dark-induced pausing behavior. Together, these studies identify the molecular and circuit mechanisms underlying ON vs. OFF discrimination in the Drosophila larval visual system.
Raad, H. and Robichon, A. (2019). The pleiotropic effects of Innexin genes expressed in Drosophila glia encompass wing chemosensory sensilla. J Neurosci Res. PubMed ID: 31257643
The neuroanatomy of Drosophila wing chemosensilla and the analysis of their sensory organ precursor cell lineage have demonstrated that they are surprisingly related to taste perception. The microarchitecture of wing bristles limits the use of electrophysiology methods to investigate wing chemosensory mechanisms. However, by monitoring the fluorescence of the complex calcium/GCaMP, calcium flux triggered upon tastant stimulation was observed within sensilla aligned along the wing anterior nerve. This string of fluorescent puncta was impaired in wings of Innexin 2 (Inx2) mutant flies; although it is unclear whether the Innexin proteins act at the level of the wing imaginal disc, adult wing and/or at both levels. Glial cells known to shelter Innexin(s) expression have no documented role in adult chemosensory sensilla. These data suggest that Innexin(s) are likely required for the maturation of functional wing chemosensilla in adulthood. The unexpected presence of most Innexin transcripts in adult wing RNAseq data set argues for the expression of Innexin proteins in the larval imaginal wing disc that are continued in wing chemosensilla at adulthood.
Roessingh, S., Rosing, M., Marunova, M., Ogueta, M., George, R., Lamaze, A. and Stanewsky, R. (2019). Temperature synchronization of the Drosophila circadian clock protein PERIOD is controlled by the TRPA channel PYREXIA. Commun Biol 2: 246. PubMed ID: 31286063
Circadian clocks are endogenous molecular oscillators that temporally organize behavioral activity thereby contributing to the fitness of organisms. To synchronize the fly circadian clock with the daily fluctuations of light and temperature, these environmental cues are sensed both via brain clock neurons, and by light and temperature sensors located in the peripheral nervous system. This study demonstrates that the TRPA channel PYREXIA (PYX) is required for temperature synchronization of the key circadian clock protein PERIOD. A molecular synchronization defect was observed explaining the previously reported defects of pyx mutants in behavioral temperature synchronization. Surprisingly, surgical ablation of pyx-mutant antennae partially rescues behavioral synchronization, indicating that antennal temperature signals are modulated by PYX function to synchronize clock neurons in the brain. These results suggest that PYX protects antennal neurons from faulty signaling that would otherwise interfere with temperature synchronization of the circadian clock neurons in the brain.
Zhang, X., Coates, K., Dacks, A., Gunay, C., Lauritzen, J. S., Li, F., Calle-Schuler, S. A., Bock, D. and Gaudry, Q. (2019). Local synaptic inputs support opposing, network-specific odor representations in a widely projecting modulatory neuron. Elife 8. PubMed ID: 31264962
Serotonin plays different roles across networks within the same sensory modality. Previous whole-cell electrophysiology in Drosophila has shown that serotonergic neurons innervating the first olfactory relay are inhibited by odorants. This study shows that network-spanning serotonergic neurons segregate information about stimulus features, odor intensity and identity, by using opposing coding schemes in different olfactory neuropil. A pair of serotonergic neurons (the CSDns) innervate the antennal lobe and lateral horn, which are first and second order neuropils. CSDn processes in the antennal lobe are inhibited by odors in an identity independent manner. In the lateral horn, CSDn processes are excited in an odor identity dependent manner. Using functional imaging, modeling, and EM reconstruction, it was demonstrated that antennal lobe derived inhibition arises from local GABAergic inputs and acts as a means of gain control on branch-specific inputs that the CSDns receive within the lateral horn.

Thursday, September 12th - Immune Response

Yadav, S. and Eleftherianos, I. (2019). Participation of the serine protease Jonah66Ci in the Drosophila anti-nematode immune response. Infect Immun. PubMed ID: 31182620
Serine proteases and serine protease homologs form the second largest gene family in the Drosophila melanogaster genome. Certain genes in the Jonah multi-gene family encoding serine proteases have been implicated in the fly antiviral immune response. This study reports the involvement of Jonah66Ci in the Drosophila immune defense against Steinernema carpocapsae nematode infection. Jonah66Ci is upregulated in response to symbiotic (carrying the mutualistic bacteria Xenorhabdus nematophila) or axenic (lacking Xenorhabdus) Steinernema nematodes and is expressed exclusively in the gut of Drosophila larvae. Inactivation of Jonah66Ci provides a survival advantage to larvae against axenic nematodes and results in differential expression of Toll and Imd pathway effector genes, specifically in the gut. Also, inactivation of Jonah66Ci increases the numbers of enteroendocrine and mitotic cells in the gut of uninfected larvae and infection with Steinernema nematodes reduces their numbers, whereas the numbers of intestinal stem cells are unaffected by nematode infection. Jonah66Ci knockdown further reduces nitric oxide levels in response to infection with Steinernema symbiotic nematodes. Finally, Jonah66Ci knockdown does not alter the feeding rates of uninfected Drosophila larvae, however infection with Steinernema axenic nematodes lowers larval feeding. In conclusion, this study reports that Jonah66Ci participates in maintaining homeostasis of certain physiological processes in Drosophila larvae in the context of Steinernema nematode infection. Similar findings will ultimately lead to an understanding the molecular and physiological mechanisms that take place during parasitic nematode infection in insects.
Patrnogic, J., Heryanto, C., Ozakman, Y. and Eleftherianos, I. (2019). Transcript analysis reveals the involvement of NF-kappaB transcription factors for the activation of TGF-beta signaling in nematode-infected Drosophila. Immunogenetics. PubMed ID: 31147740
The common fruit fly Drosophila melanogaster is a powerful model for studying signaling pathway regulation. Conserved signaling pathways underlying physiological processes signify evolutionary relationship between organisms and the nature of the mechanisms they control. This study explores the cross-talk between the well-characterized nuclear factor kappa B (NF-kappaB) innate immune signaling pathways and transforming growth factor beta (TGF-beta) signaling pathway in response to parasitic nematode infection in Drosophila. To understand the link between signaling pathways, a transcript-level analysis of different TGF-beta signaling components was performed following infection of immune-compromised Drosophila adult flies with the nematode parasites Heterorhabditis gerrardi and H. bacteriophora. The findings demonstrate the requirement of NF-kappaB transcription factors for activation of TGF-beta signaling pathway in Drosophila in the context of parasitic nematode infection. Significant decrease were observed in transcript level of glass bottom boat (gbb) and screw (scw), components of the bone morphogenic protein (BMP) branch, as well as Activinbeta (actbeta) which is a component of the Activin branch of the TGF-beta signaling pathway. These results are observed only in H. gerrardi nematode-infected flies compared to uninfected control. Also, this significant decrease in transcript level is found only for extracellular ligands. Future research examining the mechanisms regulating the interaction of these signaling pathways could provide further insight into Drosophila anti-nematode immune function against infection with potent parasitic nematodes.
West, C., Rus, F., Chen, Y., Kleino, A., Gangloff, M., Gammon, D. B. and Silverman, N. (2019). IIV-6 inhibits NF-kappaB responses in Drosophila. Viruses 11(5). PubMed ID: 31052481
The host immune response and virus-encoded immune evasion proteins pose constant, mutual selective pressure on each other. Virally encoded immune evasion proteins also indicate which host pathways must be inhibited to allow for viral replication. This study shows that Invertebrate iridescent virus 6 (IIV-6) is capable of inhibiting the two Drosophila NF-kappaB signaling pathways, Imd and Toll. Antimicrobial peptide (AMP) gene induction downstream of either pathway is suppressed when cells infected with IIV-6 are also stimulated with Toll or Imd ligands. Cleavage of both Imd and Relish, as well as Relish nuclear translocation, three key points in Imd signal transduction, occur in IIV-6 infected cells, indicating that the mechanism of viral inhibition is farther downstream, at the level of Relish promoter binding or transcriptional activation. Additionally, flies co-infected with both IIV-6 and the Gram-negative bacterium, Erwinia carotovora carotovora, succumb to infection more rapidly than flies singly infected with either the virus or the bacterium. These findings demonstrate how pre-existing infections can have a dramatic and negative effect on secondary infections, and establish a Drosophila model to study confection susceptibility.
Houtz, P., Bonfini, A., Bing, X. and Buchon, N. (2019). Recruitment of adult precursor cells underlies limited repair of the infected larval midgut in Drosophila. Cell Host Microbe. PubMed ID: 31492656
Surviving infection requires immune and repair mechanisms. Developing organisms face the additional challenge of integrating these mechanisms with tightly controlled developmental processes. The larval Drosophila midgut lacks dedicated intestinal stem cells. This study shows that, upon infection, larvae perform limited repair using adult midgut precursors (AMPs). AMPs differentiate in response to damage to generate new enterocytes, transiently depleting their pool. Developmental delay allows for AMP reconstitution, ensuring the completion of metamorphosis. Notch signaling is required for the differentiation of AMPs into the encasing, niche-like peripheral cells (PCs), but not to differentiate PCs into enterocytes. Dpp (TGF-beta) signaling is sufficient, but not necessary, to induce PC differentiation into enterocytes. Infection-induced JAK-STAT pathway is both required and sufficient for differentiation of AMPs and PCs into new enterocytes. Altogether, this work highlights the constraints imposed by development on an organism's response to infection and demonstrates the transient use of adult precursors for tissue repair.

Wednesday, September 11th - RNA

Ustaoglu, P., Haussmann, I. U., Liao, H., Torres-Mendez, A., Arnold, R., Irimia, M. and Soller, M. (2019). Srrm234, but not canonical SR and hnRNP proteins drive inclusion of Dscam exon 9 variable exons. RNA. PubMed ID: 31292260
In the Drosophila Dscam, which is important for neuronal wiring up to 38,016 isoforms, can be generated by mutually exclusive alternative splicing in four clusters of variable exons. However, it is not understood how a specific exon is chosen from the many variables and how variable exons are prevented from being spliced together. A main role in the regulation of Dscam alternative splicing has been attributed to RNA binding proteins, but how they impact on exon selection is not well understood. Serine-arginine-rich (SR) proteins and hnRNP proteins are the two main types of RNA binding proteins with major roles in exon definition and splice site selection. This study analyzed the role of SR and hnRNP proteins in Dscam exon 9 alternative splicing in mutant Drosophila embryos because of their essential function for development. Strikingly, loss or overexpression of canonical SR and hnRNP proteins even when multiple proteins are depleted together, does not affect Dscam alternative exon selection very dramatically. Conversely, non-canonical SR protein Serine-arginine repetitive matrix 2/3/4 (Srrm234) is a main determinant of exon inclusion in Dscam exon 9 cluster. Since long-range base-pairings are absent in the exon 9 cluster, these data argue for a small complement of regulatory factors as main determinants of exon inclusion in the Dscam exon 9 cluster.
Reichholf, B., Herzog, V. A., Fasching, N., Manzenreither, R. A., Sowemimo, I. and Ameres, S. L. (2019). Time-resolved small RNA sequencing unravels the molecular principles of microRNA homeostasis. Mol Cell. PubMed ID: 31350118
Argonaute-bound microRNAs silence mRNA expression in a dynamic and regulated manner to control organismal development, physiology, and disease. This study employed metabolic small RNA sequencing for a comprehensive view on intracellular microRNA kinetics in Drosophila. Based on absolute rate of biogenesis and decay, microRNAs rank among the fastest produced and longest-lived cellular transcripts, disposing up to 10(5) copies per cell at steady-state. Mature microRNAs are produced within minutes, revealing tight intracellular coupling of biogenesis that is selectively disrupted by pre-miRNA-uridylation. Control over Argonaute protein homeostasis generates a kinetic bottleneck that cooperates with non-coding RNA surveillance to ensure faithful microRNA loading. Finally, regulated small RNA decay enables the selective rapid turnover of Ago1-bound microRNAs, but not of Ago2-bound small interfering RNAs (siRNAs), reflecting key differences in the robustness of small RNA silencing pathways. Time-resolved small RNA sequencing opens new experimental avenues to deconvolute the timescales, molecular features, and regulation of small RNA silencing pathways in living cells.
Zhang, Z., So, K., Peterson, R., Bauer, M., Ng, H., Zhang, Y., Kim, J. H., Kidd, T. and Miura, P. (2019). Elav-mediated exon skipping and alternative polyadenylation of the Dscam1 gene are required for axon outgrowth. Cell Rep 27(13): 3808-3817.e3807. PubMed ID: 31242415
Many metazoan genes express alternative long 3' UTR isoforms in the nervous system, but their functions remain largely unclear. In Drosophila melanogaster, the Dscam1 gene generates short and long (Dscam1-L) 3' UTR isoforms because of alternative polyadenylation (APA). This study found that the RNA-binding protein Embryonic Lethal Abnormal Visual System (Elav) impacts Dscam1 biogenesis at two levels, including regulation of long 3' UTR biogenesis and skipping of an upstream exon (exon 19). MinION long-read sequencing confirmed the connectivity of this alternative splicing event to the long 3' UTR. Knockdown or CRISPR deletion of Dscam1-L impaired axon outgrowth in Drosophila. The Dscam1 long 3' UTR was found to be required for correct Elav-mediated skipping of exon 19. Elav thus co-regulates APA and alternative splicing to generate specific Dscam1 transcripts that are essential for neural development. This coupling of APA to alternative splicing might represent a new class of regulated RNA processing.
Russo, A., Gatti, G., Alfieri, R., Pesce, E., Soanes, K., Ricciardi, S., Mancino, M., Cheroni, C., Vaccari, T., Biffo, S. and Calamita, P. (2019). Modulating eIF6 levels unveils the role of translation in ecdysone biosynthesis during Drosophila development. Dev Biol. PubMed ID: 31283922
During development, ribosome biogenesis and translation reach peak activities, due to impetuous cell proliferation. Current models predict that protein synthesis elevation is controlled by transcription factors and signalling pathways. Developmental models addressing translation factors overexpression effects are lacking. Eukaryotic Initiation Factor 6 (eIF6) is necessary for ribosome biogenesis and efficient translation. eIF6 is a single gene, conserved from yeasts to mammals, suggesting a tight regulation need. This study generated a Drosophila melanogaster model of eIF6 upregulation, leading to a boost in general translation and the shut-down of the ecdysone biosynthetic pathway. Indeed, translation modulation in S2 cells showed that translational rate and ecdysone biosynthesis are inversely correlated. In vivo, eIF6-driven alterations delayed Programmed Cell Death (PCD), resulting in aberrant phenotypes, partially rescued by ecdysone administration. These data show that eIF6 triggers a translation program with far-reaching effects on metabolism and development, stressing the driving and central role of translation.
Zirin, J., Ni, X., Sack, L. M., Yang-Zhou, D., Hu, Y., Brathwaite, R., Bulyk, M. L., Elledge, S. J. and Perrimon, N. (2019). Interspecies analysis of MYC targets identifies tRNA synthetases as mediators of growth and survival in MYC-overexpressing cells. Proc Natl Acad Sci U S A 116(29): 14614-14619. PubMed ID: 31262815
Aberrant MYC oncogene activation is one of the most prevalent characteristics of cancer. By overlapping datasets of Drosophila genes that are insulin-responsive and also regulate nucleolus size, this study enriched for Myc target genes required for cellular biosynthesis. Among these, the aminoacyl tRNA synthetases (aaRSs) were identified as essential mediators of Myc growth control in Drosophila; their pharmacologic inhibition is sufficient to kill MYC-overexpressing human cells, indicating that aaRS inhibitors might be used to selectively target MYC-driven cancers. A general principle is suggested in which oncogenic increases in cellular biosynthesis sensitize cells to disruption of protein homeostasis.
Redmond, W., Allen, D., Elledge, M. C., Arellanes, R., Redmond, L., Yeahquo, J., Zhang, S., Youngblood, M., Reiner, A. and Seo, J. (2019). Screening of microRNAs controlling body fat in Drosophila melanogaster and identification of miR-969 and its target, Gr47b. PLoS One 14(7): e0219707. PubMed ID: 31318925
MicroRNAs (miRNAs) are small non-protein coding RNAs and post-transcriptionally regulate cellular gene expression. In animal development, miRNAs play essential roles such as stem cell maintenance, organogenesis, and apoptosis. Using gain-of-function (GOF) screening with 160 miRNA lines in Drosophila melanogaster, this study identified a set of miRNAs which regulates body fat contents and named them microCATs (microRNAs Controlling Adipose Tissue). Further examination of egg-to-adult developmental kinetics of selected miRNA lines showed a negative correlation between fat content and developmental time. Comparison of microCATs with loss-of-function miRNA screening data uncovered miR-969 as an essential regulator of adiposity. Subsequently, adipose tissue-specific knock-down of gustatory receptor 47b (Gr47b), a miR-969 target, was found to greatly reduced the amount of body fat, recapitulating the miR-969 GOF phenotype.

Tuesday, September 10th - Cytoskeleton and Junctions

Uechi, H. and Kuranaga, E. (2019). The tricellular junction protein Sidekick regulates vertex dynamics to promote bicellular junction extension. Dev Cell. PubMed ID: 31353316
Remodeling of cell-cell junctions drives cell intercalation that causes tissue movement during morphogenesis through the shortening and growth of bicellular junctions. The growth of new junctions is essential for continuing and then completing cellular dynamics and tissue shape sculpting; however, the mechanism underlying junction growth remains obscure. This study investigated Drosophila genitalia rotation where continuous cell intercalation occurs to show that myosin II accumulating at the vertices of a new junction is required for the junction growth. This myosin II accumulation requires the adhesive transmembrane protein Sidekick (Sdk), which localizes to the adherens junctions (AJs) of tricellular contacts (tAJs). Sdk also localizes to and blocks the accumulation of E-Cadherin at newly formed growing junctions, which maintains the growth rate. It is proposed that Sdk facilitates tAJ movement by mediating myosin II-driven contraction and altering the adhesive properties at the tAJs, leading to cell-cell junction extension during persistent junction remodeling.
Pavlova, G. A., et al. (2019). The role of Patronin in Drosophila mitosis. BMC Mol Cell Biol 20(Suppl 1): 7. PubMed ID: 31284878
The calmodulin-regulated spectrin-associated proteins (CAMSAPs) belong to a conserved protein family, which includes members that bind the polymerizing microtubule (MT) minus ends and remain associated with the MT lattice formed by minus end polymerization. In Drosophila, there is only one CAMSAP, named Patronin. Previous work has shown that Patronin stabilizes the minus ends of non-mitotic MTs and is required for proper spindle elongation. However, the precise role of Patronin in mitotic spindle assembly is poorly understood. This study has explored the role of Patronin in Drosophila mitosis using S2 tissue culture cells. Patronin was shown to associate with different types of MT bundles within the Drosophila mitotic spindle and that it is required for their stability. In prometaphase cells, Patronin accumulates on short segments of MT bundles located near the chromosomes. These Patronin "seeds" extend towards the cell poles and stop growing just before reaching the poles. It is proposed that Patronin binds the minus ends within MT bundles, including those generated from the walls of preexisting MTs via the augmin-mediated pathway. This would help maintaining MT association within the mitotic bundles, thereby stabilizing the spindle structure. The data also raise the intriguing possibility that the minus ends of bundled MTs can undergo a limited polymerization.
Wang, Y., Xu, R., Cheng, Y., Cao, H., Wang, Z., Zhu, T., Jiang, J., Zhang, H., Wang, C., Qi, L., Liu, M., Guo, X., Huang, J. and Sha, J. (2019). RSBP15 interacts with and stabilizes dRSPH3 during sperm axoneme assembly in Drosophila. J Genet Genomics 46(6): 281-290. PubMed ID: 31281031
Flagellum in sperm is composed of over 200 different proteins and is essential for sperm motility. In particular, defects in the assembly of the radial spoke in the flagellum result in male infertility due to loss of sperm motility. However, mechanisms regulating radial spoke assembly remain unclear in metazoans. This study has identified a novel Drosophila protein radial spoke binding protein 15 (RSBP15) which plays an important role in regulating radial spoke assembly. Loss of RSBP15 results in complete lack of mature sperms in seminal vesicles (SVs), asynchronous individualization complex (IC) and defective "9 + 2" structure in flagella. RSBP15 is colocalized with dRSPH3 in sperm flagella, and interacts with dRSPH3 through its DD_R_PKA superfamily domain which is important for the stabilization of dRSPH3. Moreover, loss of dRSPH3, as well as dRSPH1, dRSPH4a and dRSPH9, showed similar phenotypes to rsbp15(KO) mutant. Together, these results suggest that RSBP15 acts in stabilizing the radial spoke protein complex to anchor and strengthen the radial spoke structures in sperm flagella.
Verma, V. and Maresca, T. J. (2019). Direct observation of branching MT nucleation in living animal cells. J Cell Biol. PubMed ID: 31340987
Centrosome-mediated microtubule (MT) nucleation has been well characterized; however, numerous noncentrosomal MT nucleation mechanisms exist. The branching MT nucleation pathway envisages that the gamma-tubulin ring complex (gamma-TuRC) is recruited to MTs by the augmin complex to initiate nucleation of new MTs. While the pathway is well conserved at a molecular and functional level, branching MT nucleation by core constituents has never been directly observed in animal cells. In this study, multicolor TIRF microscopy was applied to visualize and quantitatively define the entire process of branching MT nucleation in dividing Drosophila cells during anaphase. The steps of a stereotypical branching nucleation event entailed augmin binding to a mother MT and recruitment of gamma-TuRC after 15 s, followed by nucleation 16 s later of a daughter MT at a 36 degrees branch angle. Daughters typically remained attached throughout their approximately 40-s lifetime unless the mother depolymerized past the branch point. Assembly of branched MT arrays, which did not require Drosophila TPX2, enhanced localized RhoA activation during cytokinesis.
Yevick, H. G., Miller, P. W., Dunkel, J. and Martin, A. C. (2019). Structural redundancy in supracellular actomyosin networks enables robust tissue folding. Dev Cell. PubMed ID: 31353314
Tissue morphogenesis is strikingly robust. Yet, how tissues are sculpted under challenging conditions is unknown. This study combined network analysis, experimental perturbations, and computational modeling to determine how network connectivity between hundreds of contractile cells on the ventral side of the Drosophila embryo ensures robust tissue folding. Two network properties were identified that mechanically promote robustness. First, redundant supracellular cytoskeletal network paths ensure global connectivity, even with network degradation. By forming many more connections than are required, morphogenesis is not disrupted by local network damage, analogous to the way redundancy guarantees the large-scale function of vasculature and transportation networks. Second, directional stiffening of edges oriented orthogonal to the folding axis promotes furrow formation at lower contractility levels. Structural redundancy and directional network stiffening ensure robust tissue folding with proper orientation.
Sandate, C. R., Szyk, A., Zehr, E. A., Lander, G. C. and Roll-Mecak, A. (2019). An allosteric network in spastin couples multiple activities required for microtubule severing. Nat Struct Mol Biol. PubMed ID: 31285604
The AAA+ ATPase spastin remodels microtubule arrays through severing and its mutation is the most common cause of hereditary spastic paraplegias (HSP). Polyglutamylation of the tubulin C-terminal tail recruits spastin to microtubules and modulates severing activity. This paper present a ~3.2 A resolution cryo-EM structure of the Drosophila melanogaster spastin hexamer with a polyglutamate peptide bound in its central pore. Two electropositive loops arranged in a double-helical staircase coordinate the substrate sidechains. The structure reveals how concurrent nucleotide and substrate binding organizes the conserved spastin pore loops into an ordered network that is allosterically coupled to oligomerization, and suggests how tubulin tail engagement activates spastin for microtubule disassembly. This allosteric coupling may apply generally in organizing AAA+ protein translocases into their active conformations. This allosteric network is shown to be essential for severing and is a hotspot for HSP mutations.

Monday, September 9th - Signaling

Won, J. H., Kim, G. W., Kim, J. Y., Cho, D. G., Kwon, B., Bae, Y. K. and Cho, K. O. (2019). ADAMTS Sol narae cleaves extracellular Wingless to generate a novel active form that regulates cell proliferation in Drosophila. Cell Death Dis 10(8): 564. PubMed ID: 31332194
Wnt/ Wingless (Wg) is essential for embryonic development and adult homeostasis in all metazoans, but the mechanisms by which secreted Wnt/Wg is processed remain largely unknown. A Drosophila Sol narae (Sona) is a member of A Disintegrin And Metalloprotease with ThromboSpondin motif (ADAMTS) family, and positively regulates Wg signaling by promoting Wg secretion. This study reports that Sona and Wg are secreted by both conventional Golgi and exosomal transports, and Sona cleaves extracellular Wg at the two specific sites, leading to the generation of N-terminal domain (NTD) and C-terminal domain (CTD) fragments. The cleaved forms of extracellular Wg were detected in the extracellular region of fly wing discs, and its level was substantially reduced in sona mutants. Transient overexpression of Wg-CTD increased wing size while prolonged overexpression caused lethality and developmental defects. In contrast, Wg-NTD did not induce any phenotype. Moreover, the wing defects and lethality induced by sona RNAi were considerably rescued by Wg-CTD, indicating that a main function of extracellular Sona is the generation of Wg-CTD. Wg-CTD stabilized cytoplasmic Armadillo (Arm) and had genetic interactions with components of canonical Wg signaling. Wg-CTD also induced Wg downstream targets such as Distal-less (Dll) and Vestigial (Vg). Most importantly, Cyclin D (Cyc D) was induced by Wg-CTD but not by full-length Wg. Because Sona also induces Cyc D in a cell non-autonomous manner, Wg-CTD generated by Sona in the extracellular region activates a subset of Wg signaling whose major function is the regulation of cell proliferation.
Wolf, D., Smylla, T. K., Reichmuth, J., Hoffmeister, P., Kober, L., Zimmermann, M., Turkiewicz, A., Borggrefe, T., Nagel, A. C., Oswald, F., Preiss, A. and Maier, D. (2019). Nucleo-cytoplasmic shuttling of Drosophila Hairless/Su(H) heterodimer as a means of regulating Notch dependent transcription. Biochim Biophys Acta Mol Cell Res 1866(10): 1520-1532. PubMed ID: 31326540
Activation and repression of Notch target genes is mediated by transcription factor CSL, known as Suppressor of Hairless (Su(H)) in Drosophila and CBF1 or RBPJ in human. CSL associates either with co-activator Notch or with co-repressors such as Drosophila Hairless. The nuclear translocation of transcription factor CSL relies on co-factor association, both in mammals and in Drosophila. The Drosophila CSL orthologue Su(H) requires Hairless for repressor complex formation. Based on its role in transcriptional silencing, H protein would be expected to be strictly nuclear. However, H protein is also cytosolic, which may relate to its role in the stabilization and nuclear translocation of Su(H) protein. This study investigated the function of the predicted nuclear localization signals (NLS 1-3) and single nuclear export signal (NES) of co-repressor Hairless using GFP-fusion proteins, reporter assays and in vivo analyses using Hairless wild type and shuttling-defective Hairless mutants. NLS3 and NES were identified as being critical for Hairless function. In fact, H(NLS3) mutant flies match H null mutants, whereas H(NLS3NES) double mutants display weaker phenotypes in agreement with a crucial role for NES in H export. As expected for a transcriptional repressor, Notch target genes are deregulated in H(NLS3) mutant cells, demonstrating nuclear requirement for its activity. Importantly, it was revealed that Su(H) protein strictly follows Hairless protein localization. Together, it is proposed that shuttling between the nucleo-cytoplasmic compartments provides the possibility to fine tune the regulation of Notch target gene expression by balancing of Su(H) protein availability for Notch activation.
Taylor, C. A. t., Cormier, K. W., Keenan, S. E., Earnest, S., Stippec, S., Wichaidit, C., Juang, Y. C., Wang, J., Shvartsman, S. Y., Goldsmith, E. J. and Cobb, M. H. (2019). Functional divergence caused by mutations in an energetic hotspot in ERK2. Proc Natl Acad Sci U S A 116(31): 15514-15523. PubMed ID: 31296562
The most frequent extracellular signal-regulated kinase 2 (ERK2; see Drosophila Rolled) mutation occurring in cancers is E322K (E-K). ERK2 E-K reverses a buried charge in the ERK2 common docking (CD) site, a region that binds activators, inhibitors, and substrates. Little is known about the cellular consequences associated with this mutation, other than apparent increases in tumor resistance to pathway inhibitors. ERK2 E-K, like the mutation of the preceding aspartate (ERK2 D321N [D-N]) known as the sevenmaker mutation, causes increased activity in cells and evades inactivation by dual-specificity phosphatases. As opposed to findings in cancer cells, in developmental assays in Drosophila, only ERK2 D-N displays a significant gain of function, revealing mutation-specific phenotypes. The crystal structure of ERK2 D-N is indistinguishable from that of wild-type protein, yet this mutant displays increased thermal stability. In contrast, the crystal structure of ERK2 E-K reveals profound structural changes, including disorder in the CD site and exposure of the activation loop phosphorylation sites, which likely account for the decreased thermal stability of the protein. These contiguous mutations in the CD site of ERK2 are both required for docking interactions but lead to unpredictably different functional outcomes. These results suggest that the CD site is in an energetically strained configuration, and this helps drive conformational changes at distal sites on ERK2 during docking interactions.
Xie, B., Morton, D. B. and Cook, T. A. (2019). Opposing transcriptional and post-transcriptional roles for Scalloped in binary Hippo-dependent neural fate decisions. Dev Biol. PubMed ID: 31265830
The Hippo tumor suppressor pathway plays many fundamental cell biological roles during animal development. Two central players in controlling Hippo-dependent gene expression are the TEAD transcription factor Scalloped (Sd) and its transcriptional co-activator Yorkie (Yki). Hippo signaling phosphorylates Yki, thereby blocking Yki-dependent transcriptional control. In post-mitotic Drosophila photoreceptors, a bistable negative feedback loop forms between the Hippo-dependent kinase Warts/Lats and Yki to lock in green vs blue-sensitive neuronal subtype choices, respectively. Previous experiments indicate that sd and yki mutants phenocopy each other's functions, both being required for promoting the expression of the blue photoreceptor fate determinant melted (melt) and the blue-sensitive opsin Rh5. This study demonstrates that Sd ensures the robustness of this neuronal fate decision via multiple antagonistic gene regulatory roles. In Hippo-positive (green) photoreceptors, Sd directly represses both melt and Rh5 gene expression through defined TEAD binding sites, a mechanism that is antagonized by Yki in Hippo-negative (blue) cells. Additionally, in blue photoreceptors, Sd is required to promote the translation of the Rh5 protein through a 3'UTR-dependent and microRNA-mediated process. Together, these studies reveal that Sd can drive context-dependent cell fate decisions through opposing transcriptional and post-transcriptional mechanisms.
Trylinski, M. and Schweisguth, F. (2019). Activation of Arp2/3 by WASp is essential for the endocytosis of Delta only during cytokinesis in Drosophila. Cell Rep 28(1): 1-10. PubMed ID: 31269431
The actin nucleator Arp2/3 generates pushing forces in response to signals integrated by SCAR and WASp. In Drosophila, the activation of Arp2/3 by WASp is specifically required for Notch signaling following asymmetric cell division. How WASp and Arp2/3 regulate Notch activity and why receptor activation requires WASp and Arp2/3 only in the context of intra-lineage fate decisions are unclear. This study found that WASp, but not SCAR, is required for Notch activation soon after division of the sensory organ precursor cell. Conversely, SCAR, but not WASp, is required to expand the cell-cell contact between the two SOP daughters. Thus, these two activities of Arp2/3 can be uncoupled. Using a time-resolved endocytosis assay, WASp and Arp2/3 were shown to be required for the endocytosis of Dl only during cytokinesis. It is proposed that WASp-Arp2/3 provides an extra pushing force that is specifically required for the efficient endocytosis of Dl during cytokinesis.
Wong, K. K. L., Liao, J. Z. and Verheyen, E. M. (2019). A positive feedback loop between Myc and aerobic glycolysis sustains tumor growth in a Drosophila tumor model. Elife 8. PubMed ID: 31259690
Cancer cells usually exhibit aberrant cell signaling and metabolic reprogramming. However, mechanisms of crosstalk between these processes remain elusive. This study shows that in an in vivo tumor model expressing oncogenic Drosophila Homeodomain-interacting protein kinase (Hipk), tumor cells display elevated aerobic glycolysis. Mechanistically, elevated Hipk drives transcriptional upregulation of Drosophila Myc (dMyc; MYC in vertebrates) likely through convergence of multiple perturbed signaling cascades. dMyc induces robust expression of pfk2 (encoding 6-Phosphofructo-2-kinase/fructose-2,6-bisphosphatase; PFKFB in vertebrates) among other glycolytic genes. Pfk2 catalyzes the synthesis of fructose-2,6-bisphosphate, which acts as a potent allosteric activator of Phosphofructokinase (Pfk) and thus stimulates glycolysis. Pfk2 and Pfk in turn are required to sustain dMyc protein accumulation post-transcriptionally, establishing a positive feedback loop. Disruption of the loop abrogates tumorous growth. Together, this study demonstrates a reciprocal stimulation of Myc and aerobic glycolysis and identifies the Pfk2-Pfk governed committed step of glycolysis as a metabolic vulnerability during tumorigenesis.

Friday, September 6th - Evolution

King, T. D., Leonard, C. J., Cooper, J. C., Nguyen, S., Joyce, E. F. and Phadnis, N. (2019). Recurrent losses and rapid evolution of the condensin II complex in insects. Mol Biol Evol. PubMed ID: 31270536
Condensins play a crucial role in the organization of genetic material by compacting and disentangling chromosomes. Based on studies in a few model organisms, the condensin I and condensin II complexes are considered to have distinct functions, with the condensin II complex playing a role in meiosis and somatic pairing of homologous chromosomes in Drosophila. Intriguingly, the Cap-G2 subunit of condensin II is absent in Drosophila melanogaster, and this loss may be related to the high levels of chromosome pairing seen in flies. This study finds that all three non-SMC subunits of condensin II (Cap-G2, Cap-D3, and Cap-H2) have been repeatedly and independently lost in taxa representing multiple insect orders, with some taxa lacking all three. All non-Dipteran insects display near-uniform low pairing levels regardless of their condensin II complex composition, suggesting that some key aspects of genome organization are robust to condensin II subunit losses. Finally, consistent signatures of positive selection were found in condensin subunits across flies and mammals. These findings suggest that these ancient complexes are far more evolutionarily labile than previously suspected, and are at the crossroads of several forms of genomic conflicts. These results raise fundamental questions about the specific functions of the two condensin complexes in taxa that have experienced subunit losses, and open the door to further investigations to elucidate the diversity of molecular mechanisms that underlie genome organization across various life forms.
Panara, V., Budd, G. E. and Janssen, R. (2019). Phylogenetic analysis and embryonic expression of panarthropod Dmrt genes. Front Zool 16: 23. PubMed ID: 31303887
One set of the developmentally important Doublesex and Male-abnormal-3 Related Transcription factors (Dmrt) is subject of intense research, because of their role in sex-determination and sexual differentiation. This likely non-monophyletic group of Dmrt genes is represented by the Drosophila melanogaster gene Doublesex (Dsx), the Caenorhabditis elegans Male-abnormal-3 (Mab-3) gene, and vertebrate Dmrt1 genes. However, other members of the Dmrt family are much less well studied, and in arthropods, including the model organism Drosophila melanogaster, data on these genes are virtually absent with respect to their embryonic expression and function. This study investigate the complete set of Dmrt genes in members of all main groups of Arthropoda and a member of Onychophora, extending the data to Panarthropoda as a whole. The presence of at least four families of Dmrt genes (including Dsx-like genes) in Panarthropoda is confirmed, and their expression profiles during embryogenesis were examined. This work shows that the expression patterns of Dmrt11E, Dmrt93B, and Dmrt99B orthologs are highly conserved among panarthropods. Embryonic expression of Dsx-like genes, however, is more derived, likely as a result of neo-functionalization after duplication. These data suggest deep homology of most of the panarthropod Dmrt genes with respect to their function that likely dates back to their last common ancestor. The function of Dsx and Dsx-like genes which are critical for sexual differentiation in animals, however, appears to be much less conserved.
Kopp, A., Barmina, O. and Prigent, S. R. (2019). Phylogenetic position of the Drosophila fima and dentissima lineages, and the status of the D. melanogaster species group. Mol Phylogenet Evol 139: 106543. PubMed ID: 31247309
The subgenus Sophophora of Drosophila, which includes D. melanogaster, is an important model for the study of molecular evolution, comparative genomics, and evolutionary developmental biology. Numerous phylogenetic studies have examined species relationships in the well-known melanogaster, obscura, willistoni, and saltans species groups, as well as the relationships among these clades. In contrast, other species groups of Sophophora have been relatively neglected and have not been subjected to molecular phylogenetic analysis. This study focuses on the endemic African Drosophila fima and dentissima lineages. Both these clades fall within the broadly defined melanogaster species group, but are otherwise distantly related to each other. The new phylogeny supports pervasive divergent and convergent evolution of male-specific grasping structures (sex combs). The implications of these results are discussed for defining the boundaries of the melanogaster species group, and weigh the relative merits of "splitting" and "lumping" approaches to the taxonomy of this key model system.
Liu, Y., Ramos-Womack, M., Han, C., Reilly, P., Brackett, K. L., Rogers, W., Williams, T. M., Andolfatto, P., Stern, D. L. and Rebeiz, M. (2019). Changes throughout a genetic network mask the contribution of Hox gene evolution. Curr Biol 29(13): 2157-2166. PubMed ID: 31257142
Hox genes pattern the anterior-posterior axis of animals and are posited to drive animal body plan evolution, yet their precise role in evolution has been difficult to determine. This study identified evolutionary modifications in the Hox gene Abd-B that dramatically altered its expression along the body plan of Drosophila santomea. Abd-B is required for pigmentation in Drosophila yakuba, the sister species of D. santomea, and changes to Abd-B expression would be predicted to make large contributions to the loss of body pigmentation in D. santomea. However, manipulating Abd-B expression in current-day D. santomea does not affect pigmentation. This is attributed to epistatic interaction to four other genes within the D. santomea pigmentation network, three of which have evolved expression patterns that do not respond to Abd-B. These results demonstrate how body plans may evolve through small evolutionary steps distributed throughout Hox-regulated networks. Polygenicity and epistasis may hinder efforts to identify genes and mechanisms underlying macroevolutionary traits.

Thursday, September 5th - Enhancers and Transcriptional Regulation

Kovalenko, E. V., Mazina, M. Y., Krasnov, A. N. and Vorobyeva, N. E. (2019). The Drosophila nuclear receptors EcR and ERR jointly regulate the expression of genes involved in carbohydrate metabolism. Insect Biochem Mol Biol 112: 103184. PubMed ID: 31295549
The rate of carbohydrate metabolism is tightly coordinated with developmental transitions in Drosophila, and fluctuates depending on the requirements of a particular developmental stage. These successive metabolic switches result from changes in the expression levels of genes encoding glycolytic, tricarboxylic acid cycle (TCA), and oxidative phosphorylation enzymes. This report describes a repressive action of ecdysone signaling on the expression of glycolytic genes and enzymes of glycogen metabolism in Drosophila development. The basis of this effect is an interaction between the ecdysone receptor (EcR) and the estrogen-related receptor (ERR), a specific regulator of the Drosophila glycolysis. An overlapping DNA-binding pattern was found for the EcR and ERR in the Drosophila S2 cells. EcR was detected at a subset of the ERR target genes responsible for carbohydrate metabolism. The 20-hydroxyecdysone treatment of both the Drosophila larvae and the S2 cells decreased transcriptional levels of ERR targets. A joint action mode is proposed for both the EcR and ERR, for at least a subset of the glycolytic genes. Both receptors bind to the same regulatory regions and may form or be part of a joint transcriptional regulatory complex in the Drosophila S2 cells.
Fritsch, C., et al. (2019). Multilevel regulation of the glass locus during Drosophila eye development. PLoS Genet 15(7): e1008269. PubMed ID: 31299050
Development of eye tissue is initiated by a conserved set of transcription factors termed retinal determination network (RDN). In Drosophila, the zinc-finger transcription factor Glass acts directly downstream of the RDN to control identity of photoreceptor as well as non-photoreceptor cells. The molecular mechanisms that control expression of glass, however, remain largely unknown. All information to recapitulate glass expression are contained in a compact 5.2 kb cis-acting genomic element by combining different cell-type specific and general enhancers with repressor elements. Moreover, the immature RNA of the locus contains an alternative small open reading frame (smORF) upstream of the actual glass translation start, resulting in a small peptide instead of the three possible Glass protein isoforms. CRISPR/Cas9-based mutagenesis shows that the smORF is not required for the formation of functioning photoreceptors, but is able to attenuate effects of glass misexpression. Furthermore, editing the genome to generate glass loci eliminating either one or two isoforms shows that only one of the three proteins is critical for formation of functioning photoreceptors, while removing the two other isoforms did not cause defects in developmental or photoreceptor function. These results show that eye development and function is largely unaffected by targeted manipulations of critical features of the glass transcript, suggesting a strong selection pressure to allow the formation of a functioning eye.
Koromila, T. and Stathopoulos, A. (2019). Distinct roles of broadly expressed repressors support dynamic enhancer action and change in time. Cell Rep 28(4): 855-863. PubMed ID: 31340149
How broadly expressed repressors regulate gene expression is incompletely understood. To gain insight, this study investigated how Suppressor of Hairless-Su(H)-and Runt regulate expression of bone morphogenetic protein (BMP) antagonist short-gastrulation via the sog_Distal enhancer. A live imaging protocol was optimized to capture this enhancer's spatiotemporal output throughout the early Drosophila embryo, finding in this context that Runt regulates transcription initiation, Su(H) regulates transcription rate, and both factors control spatial expression. Furthermore, whereas Su(H) functions as a dedicated repressor, Runt temporally switches from repressor to activator. These results demonstrate that broad repressors play temporally distinct roles and contribute to dynamic gene expression. Both Run and Su(H)'s ability to influence the spatiotemporal domains of gene expression may serve to counterbalance activators and function in this manner as important regulators of the maternal-to-zygotic transition in early embryos.
Heist, T., Fukaya, T. and Levine, M. (2019). Large distances separate coregulated genes in living Drosophila embryos. Proc Natl Acad Sci U S A 116(30): 15062-15067. PubMed ID: 31285341
There is extensive evidence that remote enhancers are brought into proximity with their target promoters via long-range looping interactions. However, the exact physical distances of these enhancer-promoter interactions remain uncertain. This study employed high-resolution imaging of living Drosophila embryos to visualize the distances separating linked genes that are coregulated by a shared enhancer. Cotransvection assays (linked genes on separate homologs) suggest a surprisingly large distance during transcriptional activity: at least 100-200 nm. Similar distances were observed when a shared enhancer was placed into close proximity with linked reporter genes in cis. These observations are consistent with the occurrence of "transcription hubs," whereby clusters (or condensates) of multiple RNA polymerase II complexes and associated cofactors are periodically recruited to active promoters. The dynamics of this process might be responsible for rapid fluctuations in the distances separating the transcription of coregulated reporter genes during transvection. It is proposed that enhancer-promoter communication depends on a combination of classical looping and linking models.
Rice, G. R., Barmina, O., Luecke, D., Hu, K., Arbeitman, M. and Kopp, A. (2019). Modular tissue-specific regulation of doublesex underpins sexually dimorphic development in Drosophila. Development 146(14). PubMed ID: 31285355
The ability of a single genome to produce distinct and often dramatically different male and female forms is one of the wonders of animal development. In Drosophila melanogaster, most sexually dimorphic traits are controlled by sex-specific isoforms of the doublesex (dsx) transcription factor, and dsx expression is mostly limited to cells that give rise to sexually dimorphic traits. However, it is unknown how this mosaic of sexually dimorphic and monomorphic organs arises. This study characterized the cis-regulatory sequences that control dsx expression in the foreleg, which contains multiple types of sex-specific sensory organs. Separate modular enhancers were found to be responsible for dsx expression in each sexually dimorphic organ. Expression of dsx in the sex comb is co-regulated by two enhancers with distinct spatial and temporal specificities that are separated by a genitalia-specific enhancer. The sex comb-specific enhancer from D. willistoni, a species that primitively lacks sex combs, is not active in the foreleg. Thus, the mosaic of sexually dimorphic and monomorphic organs depends on modular regulation of dsx transcription by dedicated cell type-specific enhancers.
Tsai, A., Alves, M. R. and Crocker, J. (2019). Multi-enhancer transcriptional hubs confer phenotypic robustness. Elife 8. PubMed ID: 31294690
Previous work has shown in Drosophila melanogaster embryos that low-affinity Ultrabithorax (Ubx)-responsive shavenbaby (svb) enhancers drive expression using localized transcriptional environments and that active svb enhancers on different chromosomes tended to colocalize. This study tested the hypothesis that these multi-enhancer 'hubs' improve phenotypic resilience to stress by buffering against decreases in transcription factor concentrations and transcriptional output. Deleting a redundant enhancer from the svb locus led to reduced trichome numbers in embryos raised at elevated temperatures. Using high-resolution fluorescence microscopy, lower Ubx concentration and transcriptional output were observed in this deletion allele. Transcription sites of the full svb cis-regulatory region inserted into a different chromosome colocalized with the svb locus, increasing Ubx concentration, the transcriptional output of svb, and partially rescuing the phenotype. Thus, multiple enhancers could reinforce a local transcriptional hub to buffer against environmental stresses and genetic perturbations, providing a mechanism for phenotypical robustness.

Wednesday, September 4th - Signaling

Ozakman, Y. and Eleftherianos, I. (2019). TGF-beta signaling interferes With the Drosophila innate immune and metabolic response to parasitic nematode infection. Front Physiol 10: 716. PubMed ID: 31316388
The common fruit fly, Drosophila melanogaster, is an outstanding model to study the molecular basis of anti-pathogen immunity. The parasitic nematode Heterorhabditis gerrardi, together with its mutualistic bacteria Photorhabdus asymbiotica, infects a wide range of insects, including D. melanogaster. Transforming growth factor-beta (TGF-ss) signaling in D. melanogaster has been shown to regulate in response to parasitic nematode infection. This study investigated the contribution of two TGF-ss signaling branches, the activin and the bone morphogenetic protein (BMP), to D. melanogaster immune function against H. gerrardi. D. melanogaster larvae were used carrying mutations in the genes coding for the TGF-ss extracellular ligands daw and dpp. The number of circulating hemocytes in uninfected daw and dpp mutants decreases twofold compared to background controls, yet no significant changes in hemocyte numbers and survival of the TGF-ss mutants are observed upon nematode infection. However, nematode-infected daw mutants express Dual oxidase at higher levels and phenoloxidase activity at lower levels compared to their background controls. To elucidate the contribution of TGF-ss signaling in the metabolic response of D. melanogaster to parasitic nematodes, lipid and carbohydrate levels were estimated in daw and dpp mutant larvae infected with H. gerrardi. Both nematode-infected mutants contain lipid droplets of larger size, with daw mutant larvae also containing elevated glycogen levels. Overall, these findings indicate that the regulation of activin and BMP branches of TGF-ss signaling can alter the immune and metabolic processes in D. melanogaster during response to parasitic nematode infection. Results from this study shed light on the molecular signaling pathways insects activate to regulate mechanisms for fighting potent nematode parasites, which could lead to the identification of novel management strategies for the control of damaging pests.
Madan, E. et al., (2019). Flower isoforms promote competitive growth in cancer. Nature. PubMed ID: 31341286
In humans, the adaptive immune system uses the exchange of information between cells to detect and eliminate foreign or damaged cells; however, the removal of unwanted cells does not always require an adaptive immune system. For example, cell selection in Drosophila uses a cell selection mechanism based on 'fitness fingerprints', which allow it to delay ageing, prevent developmental malformations and replace old tissues during regeneration. At the molecular level, these fitness fingerprints consist of combinations of Flower membrane proteins. Proteins that indicate reduced fitness are called Flower-Lose, because they are expressed in cells marked to be eliminated. However, the presence of Flower-Lose isoforms at a cell's membrane does not always lead to elimination, because if neighbouring cells have similar levels of Lose proteins, the cell will not be killed. Humans could benefit from the capability to recognize unfit cells, because accumulation of damaged but viable cells during development and ageing causes organ dysfunction and disease. However, in Drosophila this mechanism is hijacked by premalignant cells to gain a competitive growth advantage. This would be undesirable for humans because it might make tumours more aggressive. It is unknown whether a similar mechanism of cell-fitness comparison is present in humans. This study shows that two human Flower isoforms (hFWE1 and hFWE3) behave as Flower-Lose proteins, whereas the other two isoforms (hFWE2 and hFWE4) behave as Flower-Win proteins. The latter give cells a competitive advantage over cells expressing Lose isoforms, but Lose-expressing cells are not eliminated if their neighbours express similar levels of Lose isoforms; these proteins therefore act as fitness fingerprints. Moreover, human cancer cells show increased Win isoform expression and proliferate in the presence of Lose-expressing stroma, which confers a competitive growth advantage on the cancer cells. Inhibition of the expression of Flower proteins reduces tumour growth and metastasis, and induces sensitivity to chemotherapy. These results show that ancient mechanisms of cell recognition and selection are active in humans and affect oncogenic growth.
Pfeifle, I., Bohnekamp, J., Volkhardt, A., Kirsten, H., Rohwedder, A., Thum, A., Magin, T. M. and Kunz, M. (2019). MEK inhibitor cobimetinib rescues a dRaf mutant lethal phenotype in Drosophila melanogaster. Exp Dermatol. PubMed ID: 31338879
Since Drosophila melanogaster has proven to be a useful model system to study phenotypes of oncogenic mutations and to identify new anti-cancer drugs, human BRAF(V600E) homologous dRaf mutant (dRaf(A572E)) Drosophila melanogaster strains to use these for characterization of mutant phenotypes and exploit these phenotypes for drug testing. For mutant gene expression, the GAL4/UAS expression system was used. dRaf (A572E) was expressed tissue-specific in the eye, epidermis, heart, wings, secretory glands and in the whole animal. Expression of dRaf (A572E) under the control of an eye-specific driver led to semi-lethality and a rough eye phenotype. The vast majority of other tissue-specific and ubiquitous drivers led to a lethal phenotype only. The rough eye phenotype was used to test BRAF inhibitor vemurafenib and MEK1/2 inhibitor cobimetinib. There was no phenotype rescue by this treatment. However, a significant rescue of the lethal phenotype was observed under a gut-specific driver. Here, MEK1/2 inhibitor cobimetinib rescued Drosophila larvae to reach pupal stage in 37% of cases as compared to 1% in control experiments. Taken together, the BRAF(V600E) homolog dRaf (A572E) exerts mostly lethal effects in Drosophila. Gut-specific dRaf (A572E) expression might in future be developed further for drug testing.
Spracklen, A. J., Thornton-Kolbe, E. M., Bonner, A. N., Florea, A., Compton, P. J., Fernandez-Gonzalez, R. and Peifer, M. (2019). The Crk adapter protein is essential for Drosophila embryogenesis, where it regulates multiple actin-dependent morphogenic events. Mol Biol Cell: mbcE19050302. PubMed ID: 31318326
Small SH2/SH3 adapter proteins regulate cell fate and behavior by mediating interactions between cell surface receptors and downstream signaling effectors in many signal transduction pathways. The Crk family has tissue-specific roles in phagocytosis, cell migration and neuronal development, and mediates oncogenic signaling in pathways like that of Abelson kinase. However, redundancy among the two mammalian family members and the position of the Drosophila gene on the fourth chromosome precluded assessment of Crk's full role in embryogenesis. These limitations were circumvented with shRNA and CRISPR technology to assess Crk's function in Drosophila morphogenesis. Crk was found to be essential beginning in the first few hours of development, where it ensures accurate mitosis by regulating orchestrated dynamics of the actin cytoskeleton to keep mitotic spindles in syncytial embryos from colliding. In this role, it positively regulates cortical localization of the Arp2/3 complex, its regulator SCAR, and F-actin to actin caps and pseudocleavage furrows. Crk loss leads to loss of nuclei and formation of multinucleate cells. Roles were found for Crk in embryonic wound healing and in axon patterning in the nervous system, where it localizes to the axons and midline glia. Thus, Crk regulates diverse events in embryogenesis that require orchestrated cytoskeletal dynamics.
Murcia, L., Clemente-Ruiz, M., Pierre-Elies, P., Royou, A. and Milan, M. (2019). Selective killing of RAS-malignant tissues by exploiting oncogene-induced DNA damage. Cell Rep 28(1): 119-131. PubMed ID: 31269434
Several oncogenes induce untimely entry into S phase and alter replication timing and progression, thereby generating replicative stress, a well-known source of genomic instability and a hallmark of cancer. Using an epithelial model in Drosophila, this study shows that the RAS oncogene, which triggers G1/S transition, induces DNA damage and, at the same time, silences the DNA damage response pathway. RAS compromises ATR-mediated phosphorylation of the histone variant H2Av and ATR-mediated cell-cycle arrest in G2 and blocks, through ERK, Dp53-dependent induction of cell death. ERK is also activated in normal tissues by an exogenous source of damage, and this activation is necessary to dampen the pro-apoptotic role of Dp53. This study exploits the pro-survival role of ERK activation upon endogenous and exogenous sources of DNA damage to present evidence that its genetic or chemical inhibition can be used as a therapeutic opportunity to selectively eliminate RAS-malignant tissues.
Otsuka, Y., Satoh, T., Nakayama, N., Inaba, R., Yamashita, H. and Satoh, A. K. (2019). Parcas is the predominant Rab11GEF for rhodopsin transport in Drosophila photoreceptors. J Cell Sci. PubMed ID: 31296556
Rab11 is essential for polarized post-Golgi vesicle trafficking to photosensitive membrane rhabdomeres in Drosophila photoreceptors. This study found that Parcas (Pcs), recently shown to have guanine-nucleotide-exchange (GEF) activity toward Rab11, co-localizes with Rab11 on the trans-side of Golgi units and post-Golgi vesicles at the base of the rhabdomeres in pupal photoreceptors. Pcs fused with the EM-tag APEX2 localizes on 150-300 nm vesicles at the trans-side of Golgi units, which are presumably fly recycling endosomes. Loss of Pcs impairs Rab11 localization on the trans-side of Golgi units and induces the cytoplasmic accumulation of post-Golgi vesicles bearing rhabdomere proteins, as observed in Rab11-deficiency. In contrast, loss of the specific subunits of TRAPPII, another known Rab11-GEF, does not cause any defects on the eye development nor the transport of rhabdomere proteins, however, simultaneous loss of TRAPPII and Pcs shows severe defects on eye development. These results indicated that both TRAPPII and Pcs are required for eye development, but Pcs functions as the predominant Rab11-GEF for post-Golgi transport to photosensitive membrane rhabdomeres.

Tuesday, September 3rd - Gonads

Chen, X., Luan, X., Zheng, Q., Qiao, C., Chen, W., Wang, M., Yan, Y., Xie, B., Shen, C., He, Z., Zhang, J., Liu, M., Hu, X., Li, H., Zheng, B., Fang, J. and Yu, J. (2019). Precursor RNA processing 3 is required for male fertility, and germline stem cell self-renewal and differentiation via regulating spliceosome function in Drosophila testes. Sci Rep 9(1): 9988. PubMed ID: 31292463
The nuclear pre-mRNA spliceosome is a large complex containing five small nuclear ribonucleoprotein particles (snRNPs) and many splicing factors. Messenger RNAs (mRNAs) are generated from pre-mRNAs by the process of RNA splicing, which is conserved in eukaryotes. Precursor RNA processing 3 (Prp3) is a U4/U6-associated snRNP whose function remains largely unknown. In the present study, using genetic manipulation of a Drosophila melanogaster testis model, Prp3 was demonstrated to be essential for male fertility in Drosophila. Prp3 deficiency in germline stem cells (GSCs) and early cyst cells resulted in abnormal structure of testes and maintenance defects of GSCs and cyst stem cells. Knockdown of Prp3 in spermatogonia and early cyst cells mediated tumor formation caused by differentiation defects. Using an in vitro assay, knockdown of Prp3 decreased proliferation and increased cell death, and controlled the spliceosome function via regulating spliceosome subunits expression in Drosophila S2 cells. Two other splicing factors were identified in the Prp complex (Prp19 and Prp8), which mimicked the phenotype of Prp3 in the Drosophila stem cell niche. The results revealed a significant role of precursor RNA processing factors in male testes, indicating that Prp3, a key spliceosome component in the Prp complex, is essential for male fertility, and germline stem cell self-renewal and differentiation, via regulating the spliceosome function in Drosophila testes.
Romero-Pozuelo, J., Foronda, D., Martin, P., Hudry, B., Merabet, S., Graba, Y. and Sanchez-Herrero, E. (2019). Cooperation of axial and sex specific information controls Drosophila female genitalia growth by regulating the Decapentaplegic pathway. Dev Biol. PubMed ID: 31251896
The specification and morphogenesis of an organ requires the coordinate deployment and integration of regulatory information, including sex specific information when the organ is sex specific. Only a few gene networks controlling size and pattern development have been deciphered, which limits the emergence of principles, general or not, underlying the organ-specifying gene networks. This study elucidates the genetic and molecular network determining the control of size in the Drosophila abdominal A9 primordium, contributing to the female genitalia. This network requires axial regulatory information provided by the Hox protein Abdominal-BR (Abd-BR), the Hox cofactors Extradenticle (Exd) and Homothorax (Hth), and the sex specific transcription factor Doublesex Female (DsxF). These factors synergize to control size in the female A9 by the coordinate regulation of the Decapentaplegic (Dpp) growth pathway. Molecular dissection of the dpp regulatory region and in vivo protein interaction experiments suggest that Abd-BR, Exd, Hth and DsxF coordinately regulate a short dpp enhancer to repress dpp expression and restrict female A9 size. The same regulators can also suppress dpp expression in the A8, but this requires the absence of the Abd-BM isoform, which specifies A8. These results delineate the network controlling female A9 growth in Drosophila.
Primus, S., Pozmanter, C., Baxter, K. and Van Doren, M. (2019). Tudor-domain containing protein 5-like promotes male sexual identity in the Drosophila germline and is repressed in females by Sex lethal. PLoS Genet 15(7): e1007617. PubMed ID: 31329582
For sexually reproducing organisms, production of male or female gametes depends on specifying the correct sexual identity in the germline. In D. melanogaster, Sex lethal (Sxl) is the key gene that controls sex determination in both the soma and the germline, but how it does so in the germline is unknown, other than that it is different than in the soma. An RNA expression profiling experiment was conducted to identify direct and indirect germline targets of Sxl specifically in the undifferentiated germline. In these cells, Sxl loss does not lead to a global masculinization observed at the whole-genome level. In contrast, Sxl appears to affect a discrete set of genes required in the male germline, such as Phf7. Tudor domain containing protein 5-like (Tdrd5l) was identified as a target for Sxl regulation that is important for male germline identity. Tdrd5l is repressed by Sxl in female germ cells, but is highly expressed in male germ cells where it promotes proper male fertility and germline differentiation. Additionally, Tdrd5l localizes to cytoplasmic granules with some characteristics of RNA Processing (P-) Bodies, suggesting that it promotes male identity in the germline by regulating post-transcriptional gene expression.
Li, M., Hu, X., Zhang, S., Ho, M. S., Wu, G. and Zhang, L. (2019). Traffic jam regulates the function of the ovarian germline stem cell progeny differentiation niche during pre-adult stage in Drosophila. Sci Rep 9(1): 10124. PubMed ID: 31300663
Stem cell self-renewal and the daughter cell differentiation are tightly regulated by the respective niches, which produce extrinsic cues to support the proper development. In Drosophila ovary, Dpp is secreted from germline stem cell (GSC) niche and activates the BMP signaling in GSCs for their self-renewal. Escort cells (ECs) in differentiation niche restrict Dpp outside the GSC niche and extend protrusions to help with proper differentiation of the GSC daughter cells. This study provides evidence that loss of large Maf transcriptional factor Traffic jam (Tj) blocks GSC progeny differentiation. Spatio-temporal specific knockdown experiments indicate that Tj is required in pre-adult EC lineage for germline differentiation control. Further molecular and genetic analyses suggest that the defective germline differentiation caused by tj-depletion is partly attributed to the elevated dpp in the differentiation niche. Moreover, this study reveals that tj-depletion induces ectopic En expression outside the GSC niche, which contributes to the upregulated dpp expression in ECs as well as GSC progeny differentiation defect. Alternatively, loss of EC protrusions and decreased EC number elicited by tj-depletion may also partially contribute to the germline differentiation defect. Collectively, these findings suggest that Tj in ECs regulates germline differentiation by controlling the differentiation niche characteristics.
Dhiman, N., Shweta, K., Tendulkar, S., Deshpande, G., Ratnaparkhi, G. S. and Ratnaparkhi, A. (2019). Drosophila Mon1 constitutes a novel node in the brain-gonad axis that is essential for female germline maturation. Development 146(13). PubMed ID: 31292144
Monensin-sensitive 1 (Mon1) is an endocytic regulator that participates in the conversion of Rab5-positive early endosomes to Rab7-positive late endosomes. In Drosophila, loss of mon1 leads to sterility as the mon1 mutant females have extremely small ovaries with complete absence of late stage egg chambers - a phenotype reminiscent of mutations in the insulin pathway genes. This study shows that expression of many Drosophila insulin-like peptides (ILPs) is reduced in mon1 mutants and feeding mon1 adults an insulin-rich diet can rescue the ovarian defects. Surprisingly, however, mon1 functions in the tyramine/octopaminergic neurons (OPNs) and not in the ovaries or the insulin-producing cells (IPCs). Consistently, knockdown of mon1 in only the OPNs is sufficient to mimic the ovarian phenotype, while expression of the gene in the OPNs alone can 'rescue' the mutant defect. Last, ilp3 and ilp5 were assessessed as critical targets of mon1. This study thus identifies mon1 as a novel molecular player in the brain-gonad axis and underscores the significance of inter-organ systemic communication during development.
Persico, V., Callaini, G. and Riparbelli, M. G. (2019). The male stem cell niche of Drosophila melanogaster: Interactions between the germline stem cells and the hub. Exp Cell Res: 111489. PubMed ID: 31278899
The Drosophila male stem cell niche is a well characterized structure in which a small cluster of somatic cells send self-renewal signals to neighbouring germ cells. Although the molecular information involved in the stem cell fate have been identified, much less is understand on the mechanisms driving their short-range specific release. Ultrastructural analysis reveals distinct protrusions of the stem cell plasma membrane that interdigitate with membrane protrusions of the facing hub cells. Some of these protrusions are very elongated and extend into the hub and could correspond to the Mt-Nanotubes. Therefore, the interface between the stem cells and the hub appears more complex than previously reported and the membrane protrusions of the stem cells might represent specialized surface areas involved in the niche-stem cell communication. The presence was noticed of clathrin-coated vesicles in the germline plasma membrane that might be also involved in delivering information from the hub.
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