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Current papers in developmental biology and gene function


Tuesday, March 31, 2020 - Chromatin

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Zraly, C. B., Zakkar, A., Perez, J. H., Ng, J., White, K. P., Slattery, M. and Dingwall, A. K. (2020). The Drosophila MLR COMPASS complex is essential for programming cis-regulatory information and maintaining epigenetic memory during development. Nucleic Acids Res. PubMed ID: 32052053
The MLR COMPASS complex monomethylates H3K4 that serves to epigenetically mark transcriptional enhancers to drive proper gene expression during animal development. Chromatin genrichment analyses of the Drosophila MLR complex reveals dynamic association with promoters and enhancers in embryos with late stage enrichments biased toward both active and poised enhancers. RNAi depletion of the Cmi (also known as Lpt) subunit that contains the chromatin binding PHD finger domains attenuates enhancer functions, but unexpectedly results in inappropriate enhancer activation during stages when hormone responsive enhancers are poised, revealing critical epigenetic roles involved in both the activation and repression of enhancers depending on developmental context. Cmi is necessary for robust H3K4 monomethylation and H3K27 acetylation that mark active enhancers, but not for the chromatin binding of Trr, the MLR methyltransferase. These data reveal two likely major regulatory modes of MLR function, contributions to enhancer commissioning in early embryogenesis and bookmarking enhancers to enable rapid transcriptional re-activation at subsequent developmental stages.
Nystrom, S. L., Niederhuber, M. J. and McKay, D. J. (2020). Expression of E93 provides an instructive cue to control dynamic enhancer activity and chromatin accessibility during development. Development 147(6). PubMed ID: 32094114
How temporal cues combine with spatial inputs to control gene expression during development is poorly understood. This study tested the hypothesis that the Drosophila transcription factor E93 controls temporal gene expression by regulating chromatin accessibility. Precocious expression of E93 early in wing development reveals that it can simultaneously activate and deactivate different target enhancers. Notably, the precocious patterns of enhancer activity resemble the wild-type patterns that occur later in development, suggesting that expression of E93 alters the competence of enhancers to respond to spatial cues. Genomic profiling reveals that precocious E93 expression is sufficient to regulate chromatin accessibility at a subset of its targets. These accessibility changes mimic those that normally occur later in development, indicating that precocious E93 accelerates the wild-type developmental program. Further, it was found that target enhancers that do not respond to precocious E93 in early wings become responsive after a developmental transition, suggesting that parallel temporal pathways work alongside E93. These findings support a model wherein E93 expression functions as an instructive cue that defines a broad window of developmental time through control of chromatin accessibility.
Walther, M., Schrahn, S., Krauss, V., Lein, S., Kessler, J., Jenuwein, T. and Reuter, G. (2020). Heterochromatin formation in Drosophila requires genome-wide histone deacetylation in cleavage chromatin before mid-blastula transition in early embryogenesis. Chromosoma 129(1): 83-98. PubMed ID: 31950239
Su(var) mutations define epigenetic factors controlling heterochromatin formation and gene silencing in Drosophila. This study identified SU(VAR)2-1 as a novel chromatin regulator that directs global histone deacetylation during the transition of cleavage chromatin into somatic blastoderm chromatin in early embryogenesis. SU(VAR)2-1 is heterochromatin-associated in blastoderm nuclei but not in later stages of development. In larval polytene chromosomes, SU(VAR)2-1 is a band-specific protein. SU(VAR)2-1 directs global histone deacetylation by recruiting the histone deacetylase RPD3. In Su(var)2-1 mutants H3K9, H3K27, H4K8 and H4K16 acetylation shows elevated levels genome-wide and heterochromatin displays aberrant histone hyper-acetylation. Whereas H3K9me2- and HP1a-binding appears unaltered, the heterochromatin-specific H3K9me2S10ph composite mark is impaired in heterochromatic chromocenters of larval salivary polytene chromosomes. SU(VAR)2-1 contains an NRF1/EWG domain and a C2HC zinc-finger motif. This study identifies SU(VAR)2-1 as a dosage-dependent, heterochromatin-initiating SU(VAR) factor, where the SU(VAR)2-1-mediated control of genome-wide histone deacetylation after cleavage and before mid-blastula transition (pre-MBT) is required to enable heterochromatin formation.
Ninova, M., Chen, Y. A., Godneeva, B., Rogers, A. K., Luo, Y., Fejes Toth, K. and Aravin, A. A. (2020). Su(var)2-10 and the SUMO Pathway Link piRNA-Guided Target Recognition to Chromatin Silencing. Mol Cell 77(3): 556-570. PubMed ID: 31901446
Regulation of transcription is the main mechanism responsible for precise control of gene expression. Whereas the majority of transcriptional regulation is mediated by DNA-binding transcription factors that bind to regulatory gene regions, an elegant alternative strategy employs small RNA guides, Piwi-interacting RNAs (piRNAs) to identify targets of transcriptional repression. This study shows that in Drosophila the small ubiquitin-like protein SUMO and the SUMO E3 ligase Su(var)2-10 are required for piRNA-guided deposition of repressive chromatin marks and transcriptional silencing of piRNA targets. Su(var)2-10 links the piRNA-guided target recognition complex to the silencing effector by binding the piRNA/Piwi complex and inducing SUMO-dependent recruitment of the SetDB1/Wde histone methyltransferase effector. It is proposed that in Drosophila, the nuclear piRNA pathway has co-opted a conserved mechanism of SUMO-dependent recruitment of the SetDB1/Wde chromatin modifier to confer repression of genomic parasites.
Loubiere, V., Papadopoulos, G. L., Szabo, Q., Martinez, A. M. and Cavalli, G. (2020). Widespread activation of developmental gene expression characterized by PRC1-dependent chromatin looping. Sci Adv 6(2): eaax4001. PubMed ID: 31950077
Polycomb repressive complexes 1 and 2 have been historically described as transcriptional repressors, but recent reports suggest that PRC1 might also support activation, although the underlying mechanisms remain elusive. This study shows that stage-specific PRC1 binding at a subset of active promoters and enhancers during Drosophila development coincides with the formation of three-dimensional (3D) loops, an increase in expression during development and repression in PRC1 mutants. Dissection of the dachshund locus indicates that PRC1-anchored loops are versatile architectural platforms that persist when surrounding genes are transcriptionally active and fine-tune their expression. The analysis of RING1B binding profiles and 3D contacts during neural differentiation in mice suggests that this role is conserved in mammals.
Jain, S., Maini, J., Narang, A., Maiti, S. and Brahmachari, V. (2020). The regulatory function of dIno80 correlates with its DNA binding activity. Gene 732: 144368. PubMed ID: 31954859
The INO80 complex, including the Ino80 protein, forms a highly conserved canonical complex that remodels chromatin in the context of multiple cellular functions. The Drosophila homologue, dIno80, is involved in homeotic gene regulation during development as a canonical Pho-dIno80 complex. Previously, it was found that dIno80 regulates homeotic genes by interacting with epigenetic regulators, such as polycomb and trithorax, suggesting the occurrence of non-canonical Ino80 complexes. Using spectroscopic methods and gel retardation assays, this study identified a set of consensus DNA sequences that DNA binding domain of dIno80 (DBINO) interacts with having differential affinity and high specificity. Testing these sequences in reporter assays, showed that this interaction can positively regulate transcription. These results suggest that, dIno80 has a sequence preference for interaction with DNA leading to transcriptional changes.

Monday, March 30th - Signaling

Wong, K. K. L., Liu, T. W., Parker, J. M., Sinclair, D. A. R., Chen, Y. Y., Khoo, K. H., Vocadlo, D. J. and Verheyen, E. M. (2020). The nutrient sensor OGT regulates Hipk stability and tumorigenic-like activities in Drosophila. Proc Natl Acad Sci U S A 117(4): 2004-2013. PubMed ID: 31932432
Environmental cues such as nutrients alter cellular behaviors by acting on a wide array of molecular sensors inside cells. Of emerging interest is the link observed between effects of dietary sugars on cancer proliferation. This study identified the requirements of hexosamine biosynthetic pathway (HBP) and O-GlcNAc transferase (OGT) for Drosophila homeodomain-interacting protein kinase (Hipk)-induced growth abnormalities in response to a high sugar diet. On a normal diet, OGT is both necessary and sufficient for inducing Hipk-mediated tumor-like growth. It was further shown that OGT maintains Hipk protein stability by blocking its proteasomal degradation and that Hipk is O-GlcNAcylated by OGT. In mammalian cells, human HIPK2 proteins accumulate posttranscriptionally upon OGT overexpression. Mass spectrometry analyses reveal that HIPK2 is at least O-GlcNAc modified at S852, T1009, and S1147 residues. Mutations of these residues reduce HIPK2 O-GlcNAcylation and stability. Together, these data demonstrate a conserved role of OGT in positively regulating the protein stability of HIPKs (fly Hipk and human HIPK2), which likely permits the nutritional responsiveness of HIPKs.
Yokoi, T., Nabe, T., Ishizuka, C., Hayashi, K., Ito-Harashima, S., Yagi, T., Nakagawa, Y. and Miyagawa, H. (2020). Transcription-inducing activity of natural and synthetic juvenile hormone agonists through the Drosophila Methoprene-tolerant protein. Pest Manag Sci. PubMed ID: 32003111
Juvenile hormones (JHs) are a class of sesquiterpenoids that play a pivotal role in insect growth and reproduction. Synthetic JH agonists (JHAs), including pyriproxyfen, have been widely used as insecticides to control agricultural pests and disease vectors. Recent studies revealed that the action of JHAs is mediated by their intracellular receptor, the heterodimer of Methoprene-tolerant (Met) and Taiman (Tai) proteins. Although a range of bioassay systems have been developed to detect the activity of JHAs, each of these systems has its own drawback(s), such as poor reproducibility, the use of radioactive ligands, or the effect of endogenous JH-signaling factors. To address these issues, a new luciferase reporter assay for JHAs was developed in mammalian HEK293T cells transiently transfected with the Drosophila Met and Tai genes. This reporter system gave highly reproducible results and showed nanomolar sensitivity to natural JHs. This reporter system was applied to structure-activity relationship (SAR) analyses of 14 natural and synthetic JHAs, leading to identification of the ligand structural factors important for the transcription-inducing activity. COBecause this reporter system is not affected by the signaling cascade downstream of the JH receptors, it is suitable to evaluate the intrinsic activity of JHAs. The SAR results obtained in this study therefore provide invaluable information on the rational design of novel JHA insecticides.
Schwartz, R., Guichard, A., Franc, N. C., Roy, S. and Bier, E. (2020). A Drosophila Model for Clostridium difficile Toxin CDT Reveals Interactions with Multiple Effector Pathways. iScience 23(2): 100865. PubMed ID: 32058973
Clostridium difficile infections (CDIs) cause severe and occasionally life-threatening diarrhea. Hyper-virulent strains produce CDT, a toxin that ADP-ribosylates actin monomers and inhibits actin polymerization. This study created transgenic Drosophila lines expressing the catalytic subunit CDTa to investigate its interaction with host signaling pathways in vivo. When expressed in the midgut, CDTa reduces body weight and fecal output and compromises survival, suggesting severe impairment of digestive functions. At the cellular level, CDTa induces F-actin network collapse, elimination of the intestinal brush border and disruption of intercellular junctions. Toxin-dependent re-distribution of Rab11 to enterocytes' apical surface occurs, and suppression was observed of CDTa phenotypes by a Dominant-Negative form of Rab11 or RNAi of the dedicated Rab11GEF Crag (DENND4). This study also reports that Calmodulin (Cam) is required to mediate CDTa activity. In parallel, chemical inhibition of the Cam/Calcineurin pathway by Cyclosporin A or FK506 also reduces CDTa phenotypes, potentially opening new avenues for treating CDIs.
Takemura, M., Noborn, F., Nilsson, J., Bowden, N., Nakato, E., Baker, S., Su, T. Y., Larson, G. and Nakato, H. (2020). Chondroitin sulfate proteoglycan Windpipe modulates Hedgehog signaling in Drosophila. Mol Biol Cell: mbcE19060327. PubMed ID: 32049582
Proteoglycans, a class of carbohydrate-modified proteins, often modulate growth factor signaling on the cell surface. However, the molecular mechanism by which proteoglycans regulate signal transduction is largely unknown. Using a recently-developed glycoproteomic method, this study found that Windpipe (Wdp) is a novel chondroitin sulfate proteoglycan (CSPG) in Drosophila. Wdp is a single-pass transmembrane protein with leucin-rich repeat (LRR) motifs and bears three CS sugar chain attachment sites in the extracellular domain. Wdp modulates the Hedgehog (Hh) pathway. In the wing disc, overexpression of wdp inhibits Hh signaling, which is dependent on its CS chains and the LRR motifs. wdp null mutant flies show a specific defect (supernumerary scutellar bristles) known to be caused by Hh overexpression. RNAi knockdown and mutant clone analyses showed that loss of wdp leads to the upregulation of Hh signaling. Altogether, this study demonstrates a novel role of CSPGs in regulating Hh signaling.
Yamamoto-Hino, M., Kawaguchi, K., Ono, M., Furukawa, K. and Goto, S. (2020). Lamin is essential for nuclear localization of the GPI synthesis enzyme PIG-B and GPI-AP production in Drosophila. J Cell Sci. PubMed ID: 32051283
Membrane lipid biosynthesis is a complex process that occurs in various intracellular compartments. In Drosophila, phosphatidylinositol glycan (PIG)-B (DPIG-B), which catalyzes addition of the third mannose in glycosylphosphatidylinositol (GPI), localizes to the nuclear envelope (NE). Although this NE localization is essential for Drosophila development, the underlying molecular mechanism remains unknown. To elucidate this mechanism, DPIG-B-interacting proteins were identified by performing immunoprecipitation followed by proteomic analysis. Which of these proteins are required for the NE localization of DPIG-B were identified. Knockdown of Lamin Dm0, a B-type lamin, led to mislocalization of DPIG-B from the NE to the endoplasmic reticulum. Lamin Dm0 associated with DPIG-B at the inner nuclear membrane, a process that required the tail domain of Lamin Dm0. Furthermore, GPI moieties were distributed abnormally in the Lamin Dm0 mutant. These data indicate that Lamin Dm0 is involved in the NE localization of DPIG-B and is required for proper GPI-anchor modification of proteins.
Yang, S., Weske, A., Du, Y., Valera, J. M., Jones, K. L. and Johnson, A. N. (2020). FGF signaling directs myotube guidance by regulating Rac activity. Development 147(3). PubMed ID: 31932350
Nascent myotubes undergo a dramatic morphological transformation during myogenesis, in which the myotubes elongate over several cell diameters and are directed to the correct muscle attachment sites. Although this process of myotube guidance is essential to pattern the musculoskeletal system, the mechanisms that control myotube guidance remain poorly understood. Using transcriptomics, this study found that components of the Fibroblast Growth Factor (FGF) signaling pathway were enriched in nascent myotubes in Drosophila embryos. Null mutations in the FGF receptor heartless (htl), or its ligands, caused significant myotube guidance defects. The FGF ligand Pyramus is expressed broadly in the ectoderm, and ectopic Pyramus expression disrupted muscle patterning. Mechanistically, Htl regulates the activity of Rho/Rac GTPases in nascent myotubes and effects changes in the actin cytoskeleton. FGF signals are thus essential regulators of myotube guidance that act through cytoskeletal regulatory proteins to pattern the musculoskeletal system.

Friday, March 27th - RNA and Transposons

Luo, S., Zhang, H., Duan, Y., Yao, X., Clark, A. G. and Lu, J. (2020). The evolutionary arms race between transposable elements and piRNAs in Drosophila melanogaster. BMC Evol Biol 20(1): 14. PubMed ID: 31992188
The piwi-interacting RNAs (piRNAs) are small non-coding RNAs that specifically repress transposable elements (TEs) in the germline of Drosophila. Despite expanding understanding of TE:piRNA interaction, whether there is an evolutionary arms race between TEs and piRNAs was unclear. This study examined the population genomics of TEs and piRNAs in the worldwide strains of D. melanogaster. By conducting a correlation analysis between TE contents and the abundance of piRNAs from ovaries of representative strains of D. melanogaster, positive correlations were found between TEs and piRNAs in six TE families. Simulations further highlight that TE activities and the strength of purifying selection against TEs are important factors shaping the interactions between TEs and piRNAs. These studies also suggest that the de novo generation of piRNAs is an important mechanism to repress the newly invaded TEs. These results revealed the existence of an evolutionary arms race between the copy numbers of TEs and the abundance of antisense piRNAs at the population level. Although the interactions between TEs and piRNAs are complex and many factors should be considered to impact their interaction dynamics, the results suggest the emergence, repression specificity and strength of piRNAs on TEs should be considered in studying the landscapes of TE insertions in Drosophila. These results deepen understanding of the interactions between piRNAs and TEs, and also provide novel insights into the nature of genomic conflicts of other forms.
Rahman, S., Modak, C., Akter, M. and Alam, M. S. (2020). Role of MicroRNA Genes miR-1000 and miR-375 in Forming Olfactory Conditional Memory in Drosophila melanogaster. Microrna. PubMed ID: 32013857
Learning and memory are basic aspects in neurogenetics as most of the neurological disorders start with dementia or memory loss. Several genes associated with memory formation have been discovered. MicroRNA genes miR-1000 and miR-375 were reported to be associated with neural integration and glucose homeostasis in some insects and vertebrates. However, neuronal function of these genes is yet to be established in D. melanogaster. Possible role of miR-1000 and miR-375 in learning and memory formation in this fly has been explored in the present study. Both appetitive and aversive olfactory conditional learning were tested in the miR-1000 and miR-375 knockout (KO) strains and compared with wild one. Five days old third instar larvae were trained by allowing them to be associated with an odor with reward (fructose) or punishment (salt). Then, the larvae were tested to calculate their preferences to the odor trained with. Learning index (LI) values and larval locomotion speed were calculated for all strains. No significant difference was observed for larval locomotion speed in mutant strains. Knockout strain of miR-1000 showed significant deficiency in both appetitive and aversive memory formation whereas miR-375 KO strain showed a significantly lower response only in appetitive one. The results of the present study indicate important role played by these two genes in forming short-term memory in D. melanogaster.
Martinez Corrales, G., Filer, D., Wenz, K. C., Rogan, A., Phillips, G., Li, M., Feseha, Y., Broughton, S. J. and Alic, N. (2020). Partial Inhibition of RNA Polymerase I Promotes Animal Health and Longevity. Cell Rep 30(6): 1661-1669. PubMed ID: 32049000
Health and survival in old age can be improved by changes in gene expression. RNA polymerase (Pol) I is the essential, conserved enzyme whose task is to generate the pre-ribosomal RNA (rRNA). Pol I is the fundamental structurally and functionally conserved eukaryotic enzyme that transcribes a single gene, ribosomal DNA (rDNA) (Vannini and Cramer, 2012). It generates the pre-rRNA that is processed into the mature 18S, 5.8S, and 28S rRNAs, the key structural and catalytic components of the ribosome. Reducing the levels of Pol I activity is sufficient to extend lifespan in the fruit fly. This effect can be recapitulated by partial, adult-restricted inhibition, with both enterocytes and stem cells of the adult midgut emerging as important cell types. In stem cells, Pol I appears to act in the same longevity pathway as Pol III, implicating rRNA synthesis in these cells as the key lifespan determinant. Importantly, reduction in Pol I activity delays broad, age-related impairment and pathology, improving the function of diverse organ systems. Hence, this study shows that Pol I activity in the adult drives systemic, age-related decline in animal health and anticipates mortality.
Yamaguchi, S., Oe, A., Nishida, K. M., Yamashita, K., Kajiya, A., Hirano, S., Matsumoto, N., Dohmae, N., Ishitani, R., Saito, K., Siomi, H., Nishimasu, H., Siomi, M. C. and Nureki, O. (2020). Crystal structure of Drosophila Piwi. Nat Commun 11(1): 858. PubMed ID: 32051406
PIWI-clade Argonaute proteins associate with PIWI-interacting RNAs (piRNAs), and silence transposons in animal gonads. This study reports the crystal structure of the Drosophila PIWI-clade Argonaute Piwi in complex with endogenous piRNAs, at 2.9 A resolution. A structural comparison of Piwi with other Argonautes highlights the PIWI-specific structural features, such as the overall domain arrangement and metal-dependent piRNA recognition. These structural and biochemical data reveal that, unlike other Argonautes including silkworm Siwi, Piwi has a non-canonical DVDK tetrad and lacks the RNA-guided RNA cleaving slicer activity. Furthermore, this study found that the Piwi mutant with the canonical DEDH catalytic tetrad exhibits the slicer activity and readily dissociates from less complementary RNA targets after the slicer-mediated cleavage, suggesting that the slicer activity could compromise the Piwi-mediated co-transcriptional silencing. It is thus proposed that Piwi lost the slicer activity during evolution to serve as an RNA-guided RNA-binding platform, thereby ensuring faithful co-transcriptional silencing of transposons.
Stolyarenko, A. D. (2020). Nuclear Argonaute Piwi Gene Mutation Affects rRNA by Inducing rRNA Fragment Accumulation, Antisense Expression, and Defective Processing in Drosophila Ovaries. Int J Mol Sci 21(3). PubMed ID: 32046213
Drosophila key nuclear piRNA silencing pathway protein Piwi of the Argonaute family has been classically studied as a factor controlling transposable elements and fertility. Piwi has been shown to concentrate in the nucleolus for reasons largely unknown. Ribosomal RNA is the main component of the nucleolus. In this work the effect of a piwi mutation on rRNA is described. This work led to three important conclusions: A mutation in piwi induces antisense 5S rRNA expression, a processing defect of 2S rRNA orthologous to the 3'-end of eukaryotic 5.8S rRNA, and accumulation of fragments of all five rRNAs in Drosophila melanogaster ovaries. Hypotheses to explain these phenomena are proposed, possibly involving the interaction of the components of the piRNA pathway with the RNA surveillance machinery.
Suh, Y. S., Yeom, E., Nam, J. W., Min, K. J., Lee, J. and Yu, K. (2020). Methionyl-tRNA Synthetase Regulates Lifespan in Drosophila. Mol Cells. PubMed ID: 31940717
Methionyl-tRNA synthetase (MRS) is essential for translation. MRS mutants reduce global translation, which usually increases lifespan in various genetic models. However, this study found that inhibition of MRS in Drosophila reduced lifespan despite of the reduced protein synthesis. Microarray analysis with MRS inhibited Drosophila revealed significant changes in inflammatory and immune response genes. Especially, the expression of anti-microbial peptides (AMPs) genes was reduced. When the expression levels of AMP genes during aging was measured, those were getting increased in the control flies but reduced in MRS inhibition flies age dependently. Interestingly, in the germ-free condition, the maximum lifespan was increased in MRS inhibition flies compared with that of the conventional condition. These findings suggest that the lifespan of MRS inhibition flies is reduced due to the down-regulated AMPs expression in Drosophila.

Thursday, March 26th - Cell Cycle

Okazaki, R., Yamazoe, K. and Inoue, Y. H. (2020). Nuclear Export of Cyclin B Mediated by the Nup62 Complex Is Required for Meiotic Initiation in Drosophila Males. Cells 9(2). PubMed ID: 31979075
The central channel of the nuclear pore complex plays an important role in the selective transport of proteins between the nucleus and cytoplasm. Previous studies have demonstrated that the depletion of the Nup62 complex, constructing the nuclear pore channel in premeiotic Drosophila cells, resulted in the absence of meiotic cells. This study attempted to understand the mechanism underlying the cell cycle arrest before meiosis. dsRNAs was induced against the nucleoporin mRNAs using the Gal4/UAS system in Drosophila. The cell cycle of the Nup62-depleted cells was arrested before meiosis without CDK1 activation. The ectopic over-expression of CycB, but not constitutively active CDK1, resulted in partial rescue from the arrest. CycB continued to exist in the nuclei of Nup62-depleted cells and cells depleted of exportin encoded by emb. Protein complexes containing CycB, Emb, and Nup62 were observed in premeiotic spermatocytes. CycB, which had temporally entered the nucleus, was associated with Emb, and the complex was transported back to the cytoplasm through the central channel, interacting with the Nup62 complex. It is proposed that CycB is exported with Emb through the channel interacting with the Nup62 complex before the onset of meiosis. The nuclear export ensures the modification and formation of sufficient CycB-CDK1 in the cytoplasm.
Hylton, C. A., Hansen, K., Bourgeois, A. and Tomkiel Dean, J. E. (2020). Sex chromosome pairing mediated by euchromatic homology in Drosophila male meiosis. Genetics. PubMed ID: 31915134
Diploid germline cells must undergo two consecutive meiotic divisions before differentiating as haploid sex cells. During meiosis I, homologs pair and remain conjoined until segregation at anaphase. Drosophila melanogaster spermatocytes are unique in that the canonical events of meiosis I including synaptonemal complex (SC) formation, double-strand DNA breaks, and chiasmata are absent. Sex chromosomes pair at intergenic spacer sequences within the rDNA. Autosomes pair at numerous euchromatic homologies, but not at heterochromatin, suggesting that pairing may be limited to specific sequences. This study separately examined the capability of X euchromatin to pair and conjoin using an rDNA-deficient X and a series of Dp(1;Y) chromosomes. Genetic assays showed that duplicated X euchromatin can substitute for endogenous rDNA pairing sites. Segregation was not proportional to homology length, and pairing could be mapped to nonoverlapping sequences within a single Dp(1;Y). Using fluorescent in situ hybridization (FISH) to early prophase I spermatocytes, it was shown that pairing occurred with high fidelity at all homologies tested. Pairing was unaffected by the presence of X rDNA, nor could it be explained by rDNA magnification. By comparing genetic and cytological data, it was determined that centromere proximal pairings were best at segregation. Segregation was dependent on the conjunction protein Stromalin in Meiosis while the autosomal-specific Teflon was dispensable. Overall, these results suggest that pairing may occur at all homologies, but there may be sequence or positional requirements for conjunction.
Nano, M., Gemble, S., Simon, A., Pennetier, C., Fraisier, V., Marthiens, V. and Basto, R. (2019). Cell-cycle asynchrony generates DNA damage at mitotic entry in polyploid cells. Curr Biol 29(22): 3937-3945. PubMed ID: 31708395
Polyploidy arises from the gain of complete chromosome sets, and it is known to promote cancer genome evolution. Recent evidence suggests that a large proportion of human tumors experience whole-genome duplications (WGDs), which might favor the generation of highly abnormal karyotypes within a short time frame, rather than in a stepwise manner. However, the molecular mechanisms linking whole-genome duplication to genetic instability remain poorly understood. Using repeated cytokinesis failure to induce polyploidization of Drosophila neural stem cells (NSCs) (also called neuroblasts [NBs]), this study investigated the consequences of polyploidy in vivo. Surprisingly, DNA damage was found generated in a subset of nuclei of polyploid NBs during mitosis. Importantly, the observations in flies were confirmed in mouse NSCs (mNSCs) and human cancer cells after acute cytokinesis inhibition. Interestingly, DNA damage occurs in nuclei that were not ready to enter mitosis but were forced to do so when exposed to the mitotic environment of neighboring nuclei within the same cell. Additionally, it was found that polyploid cells are cell-cycle asynchronous and forcing cell-cycle synchronization was sufficient to lower the levels of DNA damage generated during mitosis. Overall, this work supports a model in which DNA damage at mitotic entry can generate DNA structural abnormalities that might contribute to the onset of genetic instability.
Barbosa, J., Martins, T., Bange, T., Tao, L., Conde, C. and Sunkel, C. (2019). Polo regulates Spindly to prevent premature stabilization of kinetochore-microtubule attachments. Embo j: e100789. PubMed ID: 31849090
Accurate chromosome segregation in mitosis requires sister kinetochores to bind to microtubules from opposite spindle poles. The stability of kinetochore-microtubule attachments is fine-tuned to prevent or correct erroneous attachments while preserving amphitelic interactions. Polo kinase has been implicated in both stabilizing and destabilizing kinetochore-microtubule attachments. However, the mechanism underlying Polo-destabilizing activity remains elusive. Resorting to an RNAi screen in Drosophila for suppressors of a constitutively active Polo mutant, this study has identified a strong genetic interaction between Polo and the Rod-ZW10-Zwilch (RZZ) complex, whose kinetochore accumulation has been shown to antagonize microtubule stability. Polo phosphorylates Spindly and impairs its ability to bind to Zwilch. This precludes dynein-mediated removal of the RZZ from kinetochores and consequently delays the formation of stable end-on attachments. It is proposed that high Polo-kinase activity following mitotic entry directs the RZZ complex to minimize premature stabilization of erroneous attachments, whereas a decrease in active Polo in later mitotic stages allows the formation of stable amphitelic spindle attachments. These findings demonstrate that Polo tightly regulates the RZZ-Spindly-dynein module during mitosis to ensure the fidelity of chromosome segregation.
Bonner, A. M., Hughes, S. E. and Hawley, R. S. (2020). Regulation of Polo Kinase by Matrimony Is Required for Cohesin Maintenance during Drosophila melanogaster Female Meiosis. Curr Biol. PubMed ID: 32008903
The Drosophila PLK Polo kinase (Polo) is inhibited by the female meiosis-specific protein Matrimony (Mtrm) in a stoichiometric manner. Drosophila Polo localizes strongly to kinetochores and to central spindle microtubules during prometaphase and metaphase I of female meiosis. Mtrm protein levels increase dramatically after nuclear envelope breakdown. This study shows that Mtrm is enriched along the meiotic spindle and that loss of mtrm results in mislocalization of the catalytically active form of Polo. The mtrm gene is haploinsufficient, and heterozygosity for mtrm results in high levels of achiasmate chromosome missegregation. In mtrm/(+) heterozygotes, there is a low level of sister centromere separation, as well as precocious loss of cohesion along the arms of achiasmate chromosomes. However, mtrm-null females are sterile, and sister chromatid cohesion is abolished on all chromosomes, leading to a failure to properly congress or orient chromosomes in metaphase I. These data demonstrate a requirement for the inhibition of Polo, perhaps by sequestering Polo to the microtubules during Drosophila melanogaster female meiosis and suggest that catalytically active Polo is a distinct subset of the total Polo population within the oocyte that requires its own regulation.
Warecki, B., Ling, X., Bast, I. and Sullivan, W. (2020). ESCRT-III-mediated membrane fusion drives chromosome fragments through nuclear envelope channels. J Cell Biol 219(3). PubMed ID: 32032426
Mitotic cells must form a single nucleus during telophase or exclude part of their genome as damage-prone micronuclei. While research has detailed how micronuclei arise from cells entering anaphase with lagging chromosomes, cellular mechanisms allowing late-segregating chromosomes to rejoin daughter nuclei remain underexplored. This study finds that late-segregating acentric chromosome fragments that rejoin daughter nuclei are associated with nuclear membrane but devoid of lamin and nuclear pore complexes in Drosophila melanogaster. Acentrics pass through membrane-, lamin-, and nuclear pore-based channels in the nuclear envelope that extend and retract as acentrics enter nuclei. Membrane encompassing the acentrics fuses with the nuclear membrane, facilitating integration of the acentrics into newly formed nuclei. Fusion, mediated by the membrane fusion protein Comt/NSF and ESCRT-III components Shrub/CHMP4B and CHMP2B, facilitates reintegration of acentrics into nuclei. These results suggest a previously unsuspected role for membrane fusion, similar to nuclear repair, in the formation of a single nucleus during mitotic exit and the maintenance of genomic integrity.

Wednesday, March25th - Apoptosis and Autophagy

Palu, R. A. S., Dalton, H. M. and Chow, C. Y. (2020). Decoupling of apoptosis from activation of the ER stress response by the Drosophila metallopeptidase superdeath. Genetics. PubMed ID: 32047096
Endoplasmic reticulum (ER) stress-induced apoptosis is a primary cause and modifier of degeneration in a number of genetic disorders. Understanding how genetic variation influences the ER stress response and subsequent activation of apoptosis could improve individualized therapies and predictions of outcomes for patients. This study finds that the uncharacterized, membrane-bound metallopeptidase CG14516 in Drosophila melanogaster, which was rename as SUPpressor of ER stress-induced DEATH (superdeath), plays a role in modifying ER stress-induced apoptosis. Loss of superdeath reduces apoptosis and degeneration in the Rh1(G69D) model of ER stress through the JNK signaling cascade. This effect on apoptosis occurs without altering the activation of the unfolded protein response (IRE1 and PERK), suggesting that the beneficial pro-survival effects of this response are intact. Furthermore, superdeath was was shown to function epistatically upstream of CDK5, a known JNK-activated pro-apoptotic factor in this model of ER stress. superdeath is not only a modifier of this particular model, but affects the general tolerance to ER stress, including ER stress-induced apoptosis. Finally, evidence is presented of Superdeath localization to the endoplasmic reticulum membrane. While similar in sequence to a number of human metallopeptidases found in the plasma membrane and ER membrane, its localization suggests that superdeath is orthologous to ERAP1/2 in humans. Together, this study provides evidence that superdeath is a link between stress in the ER and activation of cytosolic apoptotic pathways.
Khan, A., Paro, S., McGurk, L., Sambrani, N., Hogg, M. C., Brindle, J., Pennetta, G., Keegan, L. P. and O'Connell, M. A. (2020). Membrane and synaptic defects leading to neurodegeneration in Adar mutant Drosophila are rescued by increased autophagy. BMC Biol 18(1): 15. PubMed ID: 32059717
In fly brains, the Drosophila Adar (adenosine deaminase acting on RNA) enzyme edits hundreds of transcripts to generate edited isoforms of encoded proteins. It is unknown whether Drosophila Adar RNA editing events mediate some coherent physiological effect. To address this question, a genetic screen was performed for suppressors of Adar mutant defects. Adar5G1 null mutant flies are partially viable, severely locomotion defective, aberrantly accumulate axonal neurotransmitter pre-synaptic vesicles and associated proteins, and develop an age-dependent vacuolar brain neurodegeneration. A genetic screen revealed suppression of all Adar5G1 mutant phenotypes tested by reduced dosage of the Tor gene, which encodes a pro-growth kinase that increases translation and reduces autophagy in well-fed conditions. Suppression of Adar5G1) phenotypes by reduced Tor is due to increased autophagy; overexpression of Atg5, which increases canonical autophagy initiation, reduces aberrant accumulation of synaptic vesicle proteins and suppresses all Adar mutant phenotypes tested. Endosomal microautophagy (eMI) is another Tor-inhibited autophagy pathway involved in synaptic homeostasis in Drosophila. Increased expression of the key eMI protein Hsc70-4 also reduces aberrant accumulation of synaptic vesicle proteins and suppresses all Adar5G1 mutant phenotypes tested. These findings link Drosophila Adar mutant synaptic and neurotransmission defects to more general cellular defects in autophagy; presumably, edited isoforms of CNS proteins are required for optimum synaptic response capabilities in the brain during the behaviorally complex adult life stage.
Michel, M. and Dahmann, C. (2020). Tissue mechanical properties modulate cell extrusion in the Drosophila abdominal epidermis. Development 147(5). PubMed ID: 32161061
The replacement of cells is a common strategy during animal development. In the Drosophila pupal abdomen, larval epidermal cells (LECs) are replaced by adult progenitor cells (histoblasts). Previous work showed that interactions between histoblasts and LECs result in apoptotic extrusion of LECs during early pupal development. Extrusion of cells is closely preceded by caspase activation and is executed by contraction of a cortical actomyosin cable. This study identified a population of LECs that extrudes independently of the presence of histoblasts during late pupal development. Extrusion of these LECs is not closely preceded by caspase activation, involves a pulsatile medial actomyosin network, and correlates with a developmental time period when mechanical tension and E-cadherin turnover at adherens junctions is particularly high. This work reveals a developmental switch in the cell extrusion mechanism that correlates with changes in tissue mechanical properties.
Wagle, R. and Song, Y. H. (2019). Ionizing radiation reduces larval brain size by inducing premature differentiation of Drosophila neural stem cells. Biochem Biophys Res Commun. PubMed ID: 31864707
DNA damaging agents, such as ionizing radiation (IR), induce cell cycle arrest, senescence, differentiation, or cell death of stem cells, which may affect tissue homeostasis. Drosophila larval brain neuroblasts (NBs) and maintaining an appropriate number of NBs is critical to maintain brain size. Irradiation of larvae at early larval stage results in microcephaly, whereas the DNA damage response of NBs that could explain this small brain size is not clearly understood. This study observed that the irradiation of larvae in the second instar retarded brain growth, accompanied by fewer NBs. The IR-induced microcephaly does not seem to result from apoptosis since the irradiated larval brain was not stained with activated Caspase nor was the microcephaly affected by the ectopic expression of the apoptosis inhibitor. When analyzed for the percentage of mitotic cells, irradiated NBs recovered their proliferative potential within 6 h post-irradiation after transient cell cycle arrest. However, IR eventually reduced the proliferation of NBs at later time points and induced the premature differentiation of NBs. In summary, IR-induced microcephaly occurs by NB loss due to premature differentiation, rather than apoptotic cell death.
Kiss, V., Jipa, A., Varga, K., Takats, S., Maruzs, T., Lorincz, P., Simon-Vecsei, Z., Szikora, S., Foldi, I., Bajusz, C., Toth, D., Vilmos, P., Gaspar, I., Ronchi, P., Mihaly, J. and Juhasz, G. (2019). Drosophila Atg9 regulates the actin cytoskeleton via interactions with profilin and Ena. Cell Death Differ. PubMed ID: 31740789
Autophagy ensures the turnover of cytoplasm and requires the coordinated action of Atg proteins, some of which also have moonlighting functions in higher eukaryotes. This study shows that the transmembrane protein Atg9 is required for female fertility, and its loss leads to defects in actin cytoskeleton organization in the ovary and enhances filopodia formation in neurons in Drosophila. Atg9 localizes to the plasma membrane anchor points of actin cables and is also important for the integrity of the cortical actin network. Of note, such phenotypes are not seen in other Atg mutants, suggesting that these are independent of autophagy defects. Mechanistically, the known actin regulators profilin and Ena/VASP were identified as novel binding partners of Atg9 based on microscopy, biochemical, and genetic interactions. Accordingly, the localization of both profilin and Ena depends on Atg9. Taken together, these data identify a new and unexpected role for Atg9 in actin cytoskeleton regulation.
Li, Z., Wu, C., Ding, X., Li, W. and Xue, L. (2020). Toll signaling promotes JNK-dependent apoptosis in Drosophila. Cell Div 15: 7. PubMed ID: 32174999
Apoptosis plays pivotal roles in organ development and tissue homeostasis, with its major function to remove unhealthy cells that may compromise the fitness of the organism. Toll signaling, with the ancient evolutionary origin, regulates embryonic dorsal-ventral patterning, axon targeting and degeneration, and innate immunity. Using Drosophila as a genetic model, this study characterized the role of Toll signaling in apoptotic cell death. Toll signaling was found to trigger caspase-dependent cell death in development. In addition, JNK activity is required for Toll-induced cell death. Furthermore, ectopic Toll expression induces the activation of JNK pathway. Moreover, physiological activation of Toll signaling is sufficient to produce JNK-dependent cell death. Finally, Toll signaling activates JNK-mediated cell death through promoting ROS production. As Toll pathway has been evolutionarily conserved from Drosophila to human, this study may shed light on the mechanism of mammalian Toll-like receptors (TLRs) signaling in apoptotic cell death.

Tuesday, March 24th - Adult Neural Function

Serway, C. N., Dunkelberger, B. S., Del Padre, D., Nolan, N. W. C., Georges, S., Freer, S., Andres, A. J. and de Belle, J. S. (2020). Importin-alpha2 mediates brain development, learning and memory consolidation in Drosophila. J Neurogenet: 1-14. PubMed ID: 31965871
Neuronal development and memory consolidation are conserved processes that rely on nuclear-cytoplasmic transport of signaling molecules to regulate gene activity and initiate cascades of downstream cellular events. Surprisingly, few reports address and validate this widely accepted perspective. This study shows that Importin-alpha2 (Imp-alpha2), a soluble nuclear transporter that shuttles cargoes between the cytoplasm and nucleus, is vital for brain development, learning and persistent memory in Drosophila melanogaster. Mutations in importin-alpha2 (imp-alpha2, known as Pendulin or Pen and homologous with human KPNA2) are alleles of mushroom body miniature B (mbmB), a gene known to regulate aspects of brain development and influence adult behavior in flies. Mushroom bodies (MBs), paired associative centers in the brain, are smaller than normal due to defective proliferation of specific intrinsic Kenyon cell (KC) neurons in mbmB mutants. Extant KCs projecting to the MB beta-lobe terminate abnormally on the contralateral side of the brain. mbmB adults have impaired olfactory learning but normal memory decay in most respects, except that protein synthesis-dependent long-term memory (LTM) is abolished. This observation supports an alternative mechanism of persistent memory in which mutually exclusive protein-synthesis-dependent and -independent forms rely on opposing cellular mechanisms or circuits. A testable model of Imp-alpha2 and nuclear transport roles in brain development and conditioned behavior is proposed. Based on molecular characterization, it is suggested that mbmB is hereafter referred to as imp-alpha2(mbmB).
Silva, B., Niehage, C., Maglione, M., Hoflack, B., Sigrist, S. J., Wassmer, T., Pavlowsky, A. and Preat, T. (2020). Interactions between amyloid precursor protein-like (APPL) and MAGUK scaffolding proteins contribute to appetitive long-term memory in Drosophila melanogaster. J Neurogenet: 1-14. PubMed ID: 31965876
Amyloid precursor protein (APP), the precursor of amyloid beta peptide, plays a central role in Alzheimer's disease (AD), a pathology characterized by memory decline and synaptic loss upon aging. Understanding the physiological role of APP is fundamental in deciphering the progression of AD, and several studies suggest a synaptic function via protein-protein interactions. Nevertheless, it remains unclear whether and how these interactions contribute to memory. In Drosophila, previous work has shown that APP-like (APPL), the fly APP homolog, is required for aversive associative memory in the olfactory memory center, the mushroom body (MB). The present study shows that APPL is required for appetitive long-term memory (LTM), another form of associative memory, in a specific neuronal subpopulation of the MB, the alpha'/beta' Kenyon cells. Using a biochemical approach, this study identified the synaptic MAGUK (membrane-associated guanylate kinase) proteins X11, CASK, Dlgh2 and Dlgh4 as interactants of the APP intracellular domain (AICD). Next, this study shows that the Drosophila homologs CASK and Dlg are also required for appetitive LTM in the alpha'/beta' neurons. Finally, using a double RNAi approach, it was demonstrated that genetic interactions between APPL and CASK, as well as between APPL and Dlg, are critical for appetitive LTM. In summary, these results suggest that APPL contributes to associative long-term memory through its interactions with the main synaptic scaffolding proteins CASK and Dlg. This function should be conserved across species.
Vanderheyden, W. M., Van Dongen, H. P. A., Frank, M. G. and Gerstner, J. R. (2019). Sleep pressure regulates mushroom body neural-glial interactions in Drosophila. Matters Sel 2019. PubMed ID: 31938713
Sleep is a behavior that exists broadly across animal phyla, from flies to humans, and is necessary for normal brain function. Recent studies in both vertebrates and invertebrates have suggested a role for glial cells in sleep regulatory processes. Changes in neural-glial interactions have been shown to be critical for synaptic plasticity and circuit function. This study tested the hypothesis that changes in sleep pressure alters neural-glial interactions. In the fruit fly, Drosophila melanogaster, sleep is known to be regulated by mushroom body (MB) circuits. The technique GFP Reconstitution Across Synaptic Partners (GRASP) was used to test whether changes in sleep pressure affect neural-glial interactions between MB neurons and astrocytes, a specialized glial cell type known to regulate sleep in flies and mammals. The MB-astrocyte GRASP signal was reduced after 24 h of sleep deprivation, whereas the signal returned to baseline levels following 72 h of recovery. Social enrichment, which increases sleep drive, similarly reduced the MB-astrocyte GRASP signal. No changes were observed in the MB-astrocyte GRASP signal over time-of-day, or following paraquat exposure or starvation. These data suggest that changes in sleep pressure are linked to dynamic changes in neural-glial interactions between astrocytes and neuronal sleep circuits, which are not caused by normal rest-activity cycles or stressors.
Wang, H., Foquet, B., Dewell, R. B., Song, H., Dierick, H. A. and Gabbiani, F. (2020). Molecular characterization and distribution of the voltage-gated sodium channel, Para, in the brain of the grasshopper and vinegar fly. J Comp Physiol A Neuroethol Sens Neural Behav Physiol. PubMed ID: 31902005
Voltage-gated sodium (NaV) channels, encoded by the gene para, play a critical role in the rapid processing and propagation of visual information related to collision avoidance behaviors. This study investigated their localization by immunostaining the optic lobes and central brain of the grasshopper Schistocerca americana and the fruit fly Drosophila melanogaster with an antibody that recognizes the channel peptide domain responsible for fast inactivation gating. NaV channels were detected at high density at all stages of development. In the optic lobe, they revealed stereotypically repeating fascicles consistent with the regular structure of the eye. In the central brain, major axonal tracts were strongly labeled, particularly in the grasshopper olfactory system. The NaV channel sequence of Drosophila was used to identify an ortholog in the transcriptome of Schistocerca. The grasshopper, fruit fly, and human NaV channels exhibit a high degree of conservation at gating and ion selectivity domains. Comparison with three species evolutionarily close to Schistocerca identified splice variants of Para and their relation to those of Drosophila. The anatomical distribution of NaV channels molecularly analogous to those of humans in grasshoppers and vinegar flies provides a substrate for rapid signal propagation and visual processing in the context of visually-guided collision avoidance.
Shiozaki, H. M., Ohta, K. and Kazama, H. (2020). A multi-regional network encoding heading and steering maneuvers in Drosophila. Neuron. PubMed ID: 32023429
An internal sense of heading direction is computed from various cues, including steering maneuvers of the animal. Although neurons encoding heading and steering have been found in multiple brain regions, it is unclear whether and how they are organized into neural circuits. This study shows that, in flying Drosophila, heading and turning behaviors are encoded by population dynamics of specific cell types connecting the subregions of the central complex (CX), a brain structure implicated in navigation. Columnar neurons in the fan-shaped body (FB) of the CX exhibit circular dynamics that multiplex information about turning behavior and heading. These dynamics are coordinated with those in the ellipsoid body, another CX subregion containing a heading representation, although only FB neurons flip turn preference depending on the visual environment. Thus, the navigational system spans multiple subregions of the CX, where specific cell types show coordinated but distinct context-dependent dynamics.
Ping, Y., Shao, L., Li, M., Yang, L. and Zhang, J. (2020). Contribution of social influences through superposition of visual and olfactory inputs to circadian re-entrainment. iScience 23(2): 100856. PubMed ID: 32058967
Circadian patterns of locomotor activity are influenced by social interactions. Studies on insects highlight the importance of volatile odors and the olfactory system. Wild-type Drosophila exhibit immediate re-entrainment to new light:dark (LD) cycles, whereas cryb and jetc mutants show deficits in re-entrainability. This study found that both male mutants re-entrained faster to phase-shifted LD cycles when social interactions with WT female flies were promoted than the isolated males. In addition, accelerated re-entrainment mediated by social interactions was found to be depended on both visual and olfactory cues, and the effect of both cues presented jointly was nearly identical to the sum of the effects of the two cues presented separately. Moreover, re-entrainment deficits in period per expression-oscillation found in jetc mutants were partially restored by promoting social interactions. These results demonstrated that, in addition to olfaction, social interactions through the visual system also play important roles in clock entrainment.

Monday, March 23 - Disease Models

Diana, A., Collu, M., Casu, M. A., Mocci, I., Aguilar-Santelises, M. and Setzu, M. D. (2020). Improvements of motor performances in the Drosophila LRRK2 loss-of-function model of Parkinson's disease: Effects of dialyzed leucocyte Extracts from Human Serum. Brain Sci 10(1). PubMed ID: 31947539
Within neurodegenerative syndromes, Parkinson's disease (PD) is typically associated with its locomotor defects, sleep disturbances and related dopaminergic (DA) neuron loss. The fruit fly, Drosophila melanogaster, with leucine-rich repeat kinase 2 mutants (LRRK2) loss-of-function in the WD40 domain, provides mechanistic insights into corresponding human behaviour, possibly disclosing some physiopathologic features of PD in both genetic and sporadic forms. Moreover, several data support the boosting impact of innate and adaptive immunity pathways for driving the progression of PD. In this context, human dialyzable leukocyte extracts (DLE) have been extensively used to transfer antigen-specific information that influences the activity of various immune components, including inflammatory cytokines. Hence, the main goal of this study was to ascertain the therapeutic potential of DLE from male and female donors on D. melanogaster LRRK2 loss-of-function, as compared to D. melanogaster wild-type (WT), in terms of rescuing physiological parameters, such as motor and climbing activities, which are severely compromised in the mutant flies. Finally, in search of the anatomical structures responsible for restored functions in parkinsonian-like mutant flies, this study found a topographical correlation between improvement of locomotor performances and an increased number of dopaminergic neurons in selective areas of LRRK2 mutant brains.
Singh, M. D., Jensen, M., Lasser, M., Huber, E., Yusuff, T., Pizzo, L., Lifschutz, B., Desai, I., Kubina, A., Yennawar, S., Kim, S., Iyer, J., Rincon-Limas, D. E., Lowery, L. A. and Girirajan, S. (2020). NCBP2modulates neurodevelopmental defects of the 3q29 deletion in Drosophila and Xenopus laevis models. PLoS Genet 16(2): e1008590. PubMed ID: 32053595
The 1.6 Mbp deletion on chromosome 3q29 is associated with a range of neurodevelopmental disorders, including schizophrenia, autism, microcephaly, and intellectual disability. Despite its importance towards neurodevelopment, the role of individual genes, genetic interactions, and disrupted biological mechanisms underlying the deletion have not been thoroughly characterized. This study used quantitative methods to assay Drosophila melanogaster and Xenopus laevis models with tissue-specific individual and pairwise knockdown of 14 homologs of genes within the 3q29 region. Developmental, cellular, and neuronal phenotypes were identified for multiple homologs of 3q29 genes, potentially due to altered apoptosis and cell cycle mechanisms during development. Using the fly eye, screening was performed for 314 pairwise knockdowns of homologs of 3q29 genes and 44 interactions between pairs of homologs and 34 interactions with other neurodevelopmental genes were identified. Interestingly, NCBP2 homologs in Drosophila (Cbp20) and X. laevis (ncbp2) enhanced the phenotypes of homologs of the other 3q29 genes, leading to significant increases in apoptosis that disrupted cellular organization and brain morphology. These cellular and neuronal defects were rescued with overexpression of the apoptosis inhibitors Diap1 and xiap in both models, suggesting that apoptosis is one of several potential biological mechanisms disrupted by the deletion. NCBP2 was also highly connected to other 3q29 genes in a human brain-specific interaction network, providing support for the relevance of these results towards the human deletion. Overall, this study suggests that NCBP2-mediated genetic interactions within the 3q29 region disrupt apoptosis and cell cycle mechanisms during development.
Tonoki, A., Ogasawara, M., Yu, Z. and Itoh, M. (2020). Appetitive memory with survival benefit is robust across aging in Drosophila. J Neurosci. PubMed ID: 31992587
The formation of memory declines with advancing age. However, susceptibility to memory impairments depends on several factors, including the robustness of memory, the responsible neural circuits, and the internal state of aged individuals. How age-dependent changes in internal states and neural circuits affect memory formation remains unclear. This study showed in Drosophila melanogaster that aged flies of both sexes form robust appetitive memory conditioned with nutritious sugar, which suppresses their high mortality rates during starvation. In contrast, aging impairs the formation of appetitive memory conditioned with non-nutritious sugar that lacks survival benefits for the flies. Aging was found to enhanced the preference for nutritious sugar over non-nutritious sugar correlated with an age-dependent increase in the expression of Drosophila neuropeptide F, an ortholog of mammalian neuropeptide Y. Furthermore, a subset of dopaminergic neurons that signal the sweet taste of sugar decreases its function with aging, while a subset of dopaminergic neurons that signal the nutritional value of sugar maintains its function with age. These results suggest that aging impairs the ability to form memories without survival benefits; however, the ability to form memories with survival benefits is maintained through age-dependent changes in the neural circuits and neuropeptides.
Vernizzi, L., Paiardi, C., Licata, G., Vitali, T., Santarelli, S., Raneli, M., Manelli, V., Rizzetto, M., Gioria, M., Pasini, M. E., Grifoni, D., Vanoni, M. A., Gellera, C., Taroni, F. and Bellosta, P. (2020). Glutamine Synthetase 1 Increases Autophagy Lysosomal Degradation of Mutant Huntingtin Aggregates in Neurons, Ameliorating Motility in a Drosophila Model for Huntington's Disease. Cells 9(1). PubMed ID: 31941072
Glutamine Synthetase 1 (GS1) is a key enzyme that catalyzes the ATP-dependent synthesis of l-glutamine from l-glutamate and is also member of the Glutamate Glutamine Cycle, a complex physiological process between glia and neurons that controls glutamate homeostasis and is often found compromised in neurodegenerative diseases including Huntington's disease (HD). This study reports that the expression of GS1 in neurons ameliorates the motility defects induced by the expression of the mutant Htt, using a Drosophila model for HD. This phenotype is associated with the ability of GS1 to favor the autophagy that was associate with the presence of reduced Htt toxic protein aggregates in neurons expressing mutant Htt. Expression of GS1 prevents the TOR activation and phosphorylation of S6K, a mechanism that it associated with the reduced levels of essential amino acids, particularly of arginine and asparagine important for TOR activation. This study reveals a novel function for GS1 to ameliorate neuronal survival by changing amino acids' levels that induce a "starvation-like" condition responsible to induce autophagy. The identification of novel targets that inhibit TOR in neurons is of particular interest for the beneficial role that autophagy has in preserving physiological neuronal health and in the mechanisms that eliminate the formation of toxic aggregates in proteinopathies.
Parker, G. A., Kohn, N., Spirina, A., McMillen, A., Huang, W. and Mackay, T. F. C. (2020). Genetic Basis of Increased Lifespan and Postponed Senescence in Drosophila melanogaster. G3 (Bethesda). PubMed ID: 31969430
Limited lifespan and senescence are near-universal phenomena. These quantitative traits exhibit variation in natural populations due to the segregation of many interacting loci and from environmental effects. Due to the complexity of the genetic control of lifespan and senescence, understanding of the genetic basis of variation in these traits is incomplete. This study analyzed the pattern of genetic divergence between long-lived (O) Drosophila melanogaster lines selected for postponed reproductive senescence and unselected control (B) lines. The productivity of the O and B lines were quantified, and reproductive senescence was found to be maternally controlled. 57 candidate genes were selected that are expressed in ovaries, 49 of which have human orthologs, and the effects of RNA interference in ovaries and accessary glands on lifespan and reproduction were assessed. All but one candidate gene affected at least one life history trait in one sex or productivity week. In addition, 23 genes had antagonistic pleiotropic effects on lifespan and productivity. Identifying evolutionarily conserved genes affecting increased lifespan and delayed reproductive senescence is the first step towards understanding the evolutionary forces that maintain segregating variation at these loci in nature and may provide potential targets for therapeutic intervention to delay senescence while increasing lifespan.
Krishnaswamy, S., Huang, H. W., Marchal, I. S., Ryoo, H. D. and Sigurdsson, E. M. (2020). Neuronally expressed anti-tau scFv prevents tauopathy-induced phenotypes in Drosophila models. Neurobiol Dis 137: 104770. PubMed ID: 31982516

Single-chain variable fragments (scFv) have been derived from tau antibody hybridomas, and their promise as imaging diagnostic agents has been shown. This study examined the therapeutic potential of anti-tau scFv in transgenic Drosophila models that express in neurons wild-type (WT) human tau (htau) or the human tauopathy mutation R406W. scFv expressing flies were crossed with the tauopathy flies and analyzed. Overall, the survival curves differed significantly. Control flies not expressing htau survived the longest, whereas R406W expressing flies had the shortest lifespan, which was greatly prolonged by co-expressing the anti-tau scFv. Likewise, htau WT expressing flies had a moderately short lifespan, which was prolonged by co-expressing the anti-tau scFv. In addition, the htau expression impaired wing expansion after eclosion, and caused progressive abdomen expansion These features were more severe in htau R406W flies than in htau WT flies. Importantly, both phenotypes were prevented by co-expression of the anti-tau scFv. Lastly, brain analyses revealed scFv-mediated tau clearance, and its prevention of tau-mediated neurotoxicity. In summary, these findings support the therapeutic potential of an anti-tau scFv, including as gene therapies, and the use of Drosophila models for such screening.

Friday, March 20th - Cytoskeleton and junctions

Jonusaite, S., Beyenbach, K. W., Meyer, H., Paululat, A., Izumi, Y., Furuse, M. and Rodan, A. R. (2020). The septate junction protein Mesh is required for epithelial morphogenesis, ion transport and paracellular permeability in the Drosophila Malpighian tubule. Am J Physiol Cell Physiol. PubMed ID: 31913700
Septate junctions (SJs) are occluding cell-cell junctions that have roles in paracellular permeability and barrier function in the epithelia of invertebrates. Arthropods have two types of SJs, pleated SJs (pSJs) and smooth SJs (sSJs). In Drosophila melanogaster, sSJs are found in the midgut and Malpighian tubules, but the functions of sSJs and their protein components in the tubule epithelium are unknown. This study examined the role of the previously identified integral sSJ component, Mesh, in the Malpighian tubule. This study genetically manipulated mesh specifically in the principal cells of the tubule at different life stages. Tubules of flies with developmental mesh knockdown revealed defects in epithelial architecture, sSJ molecular and structural organization, and lack of urine production in basal and kinin-stimulated conditions, resulting in edema and early adult lethality. Knockdown of mesh during adulthood did not disrupt tubule epithelial and sSJ integrity, but decreased the transepithelial potential, diminished transepithelial fluid and ion transport, and decreased paracellular permeability to 4 kDa dextran. Drosophila kinin decreased transepithelial potential and increased chloride permeability, and stimulated fluid secretion in both control and adult mesh knockdown tubules, but had no effect on 4 kDa dextran flux. Together, these data indicate roles for Mesh in the developmental maturation of the Drosophila Malpighian tubule and in ion and macromolecular transport in the adult tubule.
Vajente, N., Norante, R., Redolfi, N., Daga, A., Pizzo, P. and Pendin, D. (2019). Microtubules Stabilization by Mutant Spastin Affects ER Morphology and Ca(2+) Handling. Front Physiol 10: 1544. PubMed ID: 31920731
The endoplasmic reticulum (ER) extends as a network of interconnected tubules and sheet-like structures in eukaryotic cells. ER tubules dynamically change their morphology and position within the cells in response to physiological stimuli and these network rearrangements depend on the microtubule (MT) cytoskeleton. Store-operated calcium entry (SOCE) relies on the repositioning of ER tubules to form specific ER-plasma membrane junctions. Indeed, the tips of polymerizing MTs are supposed to provide the anchor for ER tubules to move toward the plasma membrane, however the precise role of the cytoskeleton during SOCE has not been conclusively clarified. This study exploited an in vivo approach involving the manipulation of MT dynamics in Drosophila melanogaster by neuronal expression of a dominant-negative variant of the MT-severing protein spastin to induce MT hyper-stabilization. MT stabilization alters ER morphology, favoring an enrichment in ER sheets at the expense of tubules. Stabilizing MTs has a negative impact on the process of SOCE and results in a reduced ER Ca(2+) content, affecting the flight ability of the flies. Restoring proper MT organization by administering the MT-destabilizing drug vinblastine, chronically or acutely, rescues ER morphology, SOCE and flight ability, indicating that MT dynamics impairment is responsible for all the phenotypes observed.
Gonzalez-Morales, N., Xiao, Y. S., Schilling, M. A., Marescal, O., Liao, K. A. and Schock, F. (2019). Myofibril diameter is set by a finely tuned mechanism of protein oligomerization in Drosophila. Elife 8. PubMed ID: 31746737
Myofibrils are huge cytoskeletal assemblies embedded in the cytosol of muscle cells. They consist of arrays of sarcomeres, the smallest contractile unit of muscles. Within a muscle type, myofibril diameter is highly invariant and contributes to its physiological properties, yet little is known about the underlying mechanisms setting myofibril diameter. This study shows that the PDZ and LIM domain protein Zasp, a structural component of Z-discs, mediates Z-disc and thereby myofibril growth through protein oligomerization. Oligomerization is induced by an interaction of its ZM domain with LIM domains. Oligomerization is terminated upon upregulation of shorter Zasp isoforms which lack LIM domains at later developmental stages. The balance between these two isoforms, which were called growing and blocking isoforms sets the stereotyped diameter of myofibrils. If blocking isoforms dominate, myofibrils become smaller. If growing isoforms dominate, myofibrils and Z-discs enlarge, eventually resulting in large pathological aggregates that disrupt muscle function.
Aguilar-Aragon, M., Bonello, T. T., Bell, G. P., Fletcher, G. C. and Thompson, B. J. (2020). Adherens junction remodelling during mitotic rounding of pseudostratified epithelial cells. EMBO Rep: e49700. PubMed ID: 32030856
Epithelial cells undergo cortical rounding at the onset of mitosis to enable spindle orientation in the plane of the epithelium. In cuboidal epithelia in culture, the adherens junction protein E-cadherin recruits Pins/LGN/GPSM2 and Mud/NuMA to orient the mitotic spindle. In the pseudostratified columnar epithelial cells of Drosophila, septate junctions recruit Mud/NuMA to orient the spindle, while Pins/LGN/GPSM2 is surprisingly dispensable. This study shows that these pseudostratified epithelial cells downregulate E-cadherin as they round up for mitosis. Preventing cortical rounding by inhibiting Rho-kinase-mediated actomyosin contractility blocks downregulation of E-cadherin during mitosis. Mitotic activation of Rho-kinase depends on the RhoGEF ECT2/Pebble and its binding partners RacGAP1/MgcRacGAP/CYK4/Tum and MKLP1/KIF23/ZEN4/Pav. Cell cycle control of these Rho activators is mediated by the Aurora A and B kinases, which act redundantly during mitotic rounding. Thus, in Drosophila pseudostratified epithelia, disruption of adherens junctions during mitosis necessitates planar spindle orientation by septate junctions to maintain epithelial integrity (Aguilar-Aragon, 2020).
Palumbo, V., Tariq, A., Borgal, L., Metz, J., Brancaccio, M., Gatti, M., Wakefield, J. G. and Bonaccorsi, S. (2020). Drosophila Morgana is an Hsp90-interacting protein with a direct role in microtubule polymerisation. J Cell Sci 133(2). PubMed ID: 31907206
Morgana (Mora, also known as CHORD in flies) and its mammalian homologue, called CHORDC1 or CHP1, is a highly conserved cysteine and histidine-rich domain (CHORD)-containing protein that has been proposed to function as an Hsp90 co-chaperone. Morgana deregulation promotes carcinogenesis in both mice and humans while, in Drosophila, loss of mora causes lethality and a complex mitotic phenotype that is rescued by a human morgana transgene. This study shows that Drosophila Mora localises to mitotic spindles and co-purifies with the Hsp90-R2TP-TTT supercomplex and with additional well-known Hsp90 co-chaperones. Acute inhibition of Mora function in the early embryo results in a dramatic reduction in centrosomal microtubule stability, leading to small spindles nucleated from mitotic chromatin. Purified Mora binds to microtubules directly and promotes microtubule polymerisation in vitro, suggesting that Mora directly regulates spindle dynamics independently of its Hsp90 co-chaperone role.
Edwards-Jorquera, S. S., Bosveld, F., Bellaiche, Y. A., Lennon-Dumenil, A. M. and Glavic, A. (2020). Trpml controls actomyosin contractility and couples migration to phagocytosis in fly macrophages. J Cell Biol 219(3). PubMed ID: 31940424
Phagocytes use their actomyosin cytoskeleton to migrate as well as to probe their environment by phagocytosis or macropinocytosis. Although migration and extracellular material uptake have been shown to be coupled in some immune cells, the mechanisms involved in such coupling are largely unknown. By combining time-lapse imaging with genetics, this study identified the lysosomal Ca2+ channel Trpml as an essential player in the coupling of cell locomotion and phagocytosis in hemocytes, the Drosophila macrophage-like immune cells. Trpml is needed for both hemocyte migration and phagocytic processing at distinct subcellular localizations: Trpml regulates hemocyte migration by controlling actomyosin contractility at the cell rear, whereas its role in phagocytic processing lies near the phagocytic cup in a myosin-independent fashion. This study further highlighted that Vamp7 also regulates phagocytic processing and locomotion but uses pathways distinct from those of Trpml. These results suggest that multiple mechanisms may have emerged during evolution to couple phagocytic processing to cell migration and facilitate space exploration by immune cells.

Thursday, March 19th - Immune Response

Cohen, L. B., Lindsay, S. A., Xu, Y., Lin, S. J. H. and Wasserman, S. A. (2020). The Daisho Peptides Mediate Drosophila Defense Against a Subset of Filamentous Fungi. Front Immunol 11: 9. PubMed ID: 32038657
Fungal infections, widespread throughout the world, affect a broad range of life forms, including agriculturally relevant plants, humans, and insects. In defending against fungal infections, the fruit fly Drosophila melanogaster employs the Toll pathway to induce a large number of immune peptides. Some have been investigated, such as the antimicrobial peptides (AMPs) and Bomanins (Boms); many, however, remain uncharacterized. This study examined the role in innate immunity of two related peptides, Daisho1 and Daisho2 (formerly IM4 and IM14, respectively), found in hemolymph following Toll pathway activation. By generating a CRISPR/Cas9 knockout of both genes, Deltadaisho, this study found that the Daisho peptides are required for defense against a subset of filamentous fungi, including Fusarium oxysporum, but not other Toll-inducible pathogens, such as Enterococcus faecalis and Candida glabrata. Analysis of null alleles and transgenes revealed that the two daisho genes are each required for defense, although their functions partially overlap. Generating and assaying a genomic epitope-tagged Daisho2 construct, interaction was detected in vitro of Daisho2 peptide in hemolymph with the hyphae of F. oxysporum. Together, these results identify the Daisho peptides as a new class of innate immune effectors with humoral activity against a select set of filamentous fungi.
Lin, S. J. H., Fulzele, A., Cohen, L. B., Bennett, E. J. and Wasserman, S. A. (2019). Bombardier Enables Delivery of Short-Form Bomanins in the Drosophila Toll Response. Front Immunol 10: 3040. PubMed ID: 31998316
Toll mediates a robust and effective innate immune response across vertebrates and invertebrates. In Drosophila melanogaster, activation of Toll by systemic infection drives the accumulation of a rich repertoire of immune effectors in hemolymph, including the recently characterized Bomanins, as well as the classical antimicrobial peptides (AMPs). This study report the functional characterization of a Toll-induced hemolymph protein encoded by the bombardier (CG18067) gene. Using the CRISPR/Cas9 system to generate a precise deletion of the bombardier transcriptional unit, this study found that Bombardier is required for Toll-mediated defense against fungi and Gram-positive bacteria. Assaying cell-free hemolymph, it was found that the Bomanin-dependent candidacidal activity is also dependent on Bombardier, but is independent of the antifungal AMPs Drosomycin and Metchnikowin. Using mass spectrometry, it was demonstrated that deletion of bombardier results in the specific absence of short-form Bomanins from hemolymph. In addition, flies lacking Bombardier exhibited a defect in pathogen tolerance that was traced to an aberrant condition triggered by Toll activation. These results leadn to a model in which the presence of Bombardier in wild-type flies enables the proper folding, secretion, or intermolecular associations of short-form Bomanins, and the absence of Bombardier disrupts one or more of these steps, resulting in defects in both immune resistance and tolerance.
Layton, E. M., On, J., Perlmutter, J. I., Bordenstein, S. R. and Shropshire, J. D. (2019). Paternal grandmother age affects the strength of Wolbachia-induced cytoplasmic incompatibility in Drosophila melanogaster. MBio 10(6). PubMed ID: 31690673
Wolbachia are obligate intracellular bacteria that are globally distributed in half of all arthropod species. As the most abundant maternally inherited microbe in animals, Wolbachia manipulate host reproduction via reproductive parasitism strategies, including cytoplasmic incompatibility (CI). CI manifests as embryonic death when Wolbachia-modified sperm fertilize uninfected eggs but not maternally infected eggs. Thus, CI can provide a relative fitness advantage to Wolbachia-infected females and drive the infection through a population. In the genetic model Drosophila melanogaster, the Wolbachia strain wMel induces variable CI, making mechanistic studies in D. melanogaster cumbersome. This study demonstrates that sons of older paternal D. melanogaster grandmothers induce stronger CI than sons of younger paternal grandmothers, and this was termed relationship the "paternal grandmother age effect" (PGAE). Moreover, the embryos and adult sons of older D. melanogaster grandmothers have higher Wolbachia densities, correlating with their ability to induce stronger CI. In addition, it is reported that Wolbachia density positively correlates with female age and decreases after mating, suggesting that females transmit Wolbachia loads that are proportional to their own titers. These findings reveal a transgenerational impact of age on wMel-induced CI, elucidate Wolbachia density dynamics in D. melanogaster, and provide a methodological advance to studies aimed at understanding wMel-induced CI in the D. melanogaster model.
Iwashita, S., Suzuki, H., Goto, A., Oyama, T., Kanoh, H., Kuraishi, T., Fuse, N., Yano, T., Oshima, Y., Dow, J. A. T., Davies, S. A. and Kurata, S. (2020). A Receptor Guanylate Cyclase, Gyc76C, Mediates Humoral, and Cellular Responses in Distinct Ways in Drosophila Immunity. Front Immunol 11: 35. PubMed ID: 32063902
Innate immunity is an evolutionarily conserved host defense system against infections. The fruit fly Drosophila relies solely on innate immunity for infection defense, and the conservation of innate immunity makes Drosophila an ideal model for understanding the principles of innate immunity, which comprises both humoral and cellular responses. The mechanisms underlying the coordination of humoral and cellular responses, however, has remained unclear. Previous work has identified Gyc76C, a receptor-type guanylate cyclase that produces cyclic guanosine monophosphate (cGMP), as an immune receptor in Drosophila. Gyc76C mediates the induction of antimicrobial peptides for humoral responses by a novel cGMP pathway including a membrane-localized cGMP-dependent protein kinase, DG2, through downstream components of the Toll receptor such as dMyD88. This study shows that Gyc76C is also required for the proliferation of blood cells (hemocytes) for cellular responses to bacterial infections. In contrast to Gyc76C-dependent antimicrobial peptide induction, Gyc76C-dependent hemocyte proliferation is meditated by a small GTPase, Ras85D, and not by DG2 or dMyD88, indicating that Gyc76C mediates the cellular and humoral immune responses in distinct ways.
Sanchez Bosch, P., Makhijani, K., Herboso, L., Gold, K. S., Baginsky, R., Woodcock, K. J., Alexander, B., Kukar, K., Corcoran, S., Jacobs, T., Ouyang, D., Wong, C., Ramond, E. J. V., Rhiner, C., Moreno, E., Lemaitre, B., Geissmann, F. and Bruckner, K. (2019). Adult Drosophila lack hematopoiesis but rely on a blood cell reservoir at the respiratory epithelia to relay infection signals to surrounding tissues. Dev Cell. PubMed ID: 31735669
The use of adult Drosophila melanogaster as a model for hematopoiesis or organismal immunity has been debated. Addressing this question, an extensive reservoir of blood cells (hemocytes) was identified at the respiratory epithelia (tracheal air sacs) of the thorax and head. Lineage tracing and functional analyses demonstrate that the majority of adult hemocytes are phagocytic macrophages (plasmatocytes) from the embryonic lineage that parallels vertebrate tissue macrophages. Surprisingly, no sign of adult hemocyte expansion was observed. Instead, hemocytes play a role in relaying an innate immune response to the blood cell reservoir: through Imd signaling and the Jak/Stat pathway ligand Upd3, hemocytes act as sentinels of bacterial infection, inducing expression of the antimicrobial peptide Drosocin in respiratory epithelia and colocalizing fat body domains. Drosocin expression in turn promotes animal survival after infection. This work identifies a multi-signal relay of organismal humoral immunity, establishing adult Drosophila as model for inter-organ immunity.
Cattenoz, P. B., Sakr, R., Pavlidaki, A., Delaporte, C., Riba, A., Molina, N., Hariharan, N., Mukherjee, T. and Giangrande, A. (2020). Temporal specificity and heterogeneity of Drosophila immune cells. EMBO J: e104486. PubMed ID: 32162708
Immune cells provide defense against non-self and have recently been shown to also play key roles in diverse processes such as development, metabolism, and tumor progression. The heterogeneity of Drosophila immune cells (hemocytes) remains an open question. Using bulk RNA sequencing, this study found that the hemocytes display distinct features in the embryo, a closed and rapidly developing system, compared to the larva, which is exposed to environmental and metabolic challenges. Through single-cell RNA sequencing, fourteen hemocyte clusters were identified present in unchallenged larvae and associated with distinct processes, e.g., proliferation, phagocytosis, metabolic homeostasis, and humoral response. Finally, this study characterizes the changes occurring in the hemocyte clusters upon wasp infestation, which triggers the differentiation of a novel hemocyte type, the lamellocyte. This first molecular atlas of hemocytes provides insights and paves the way to study the biology of the Drosophila immune cells in physiological and pathological conditions.

Wednesday, March 18th - Gonads

Rastegari, E., Kajal, K., Tan, B. S., Huang, F., Chen, R. H., Hsieh, T. S. and Hsu, H. J. (2020). WD40 protein Wuho controls germline homeostasis via TRIM-NHL tumor suppressor Mei-p26 in Drosophila. Development 147(2). PubMed ID: 31941704
WD40 proteins control many cellular processes via protein interactions. Drosophila Wuho (Wh, a WD40 protein) controls fertility, although the involved mechanisms are unclear. This study shows that Wh promotion of Mei-p26 (a human TRIM32 ortholog) function maintains ovarian germ cell homeostasis. Wh and Mei-p26 are epistatically linked, with wh and mei-p26 mutants showing nearly identical phenotypes, including germline stem cell (GSC) loss, stem-cyst formation due to incomplete cytokinesis between GSCs and daughter cells, and overproliferation of GSC progeny. Mechanistically, Wh interacts with Mei-p26 in different cellular contexts to induce cell type-specific effects. In GSCs, Wh and Mei-p26 promote BMP stemness signaling for proper GSC division and maintenance. In GSC progeny, Wh and Mei-p26 silence nanos translation, downregulate a subset of microRNAs involved in germ cell differentiation and suppress ribosomal biogenesis via dMyc to limit germ cell mitosis. The human ortholog of Wh (WDR4) interacts with TRIM32 in human cells. These results show that Wh is a regulator of Mei-p26 in Drosophila germ cells and suggest that the WD40-TRIM interaction may also control tissue homeostasis in other stem cell systems.
Sokolova, O. A., Mikhaleva, E. A., Kharitonov, S. L., Abramov, Y. A., Gvozdev, V. A. and Klenov, M. S. (2020). Special vulnerability of somatic niche cells to transposable element activation in Drosophila larval ovaries. Sci Rep 10(1): 1076. PubMed ID: 31974416
In the Drosophila ovary, somatic escort cells (ECs) form a niche that promotes differentiation of germline stem cell (GSC) progeny. The piRNA (Piwi-interacting RNA) pathway, which represses transposable elements (TEs), is required in ECs to prevent the accumulation of undifferentiated germ cells (germline tumor phenotype). The soma-specific piRNA cluster flamenco (flam) produces a substantial part of somatic piRNAs. This study characterized the biological effects of somatic TE activation on germ cell differentiation in flam mutants. The choice between normal and tumorous phenotypes of flam mutant ovaries depends on the number of persisting ECs, which is determined at the larval stage. Accordingly, much more frequent DNA breaks were found in somatic cells of flam larval ovaries than in adult ECs. The absence of Chk2 or ATM checkpoint kinases dramatically enhanced oogenesis defects of flam mutants, in contrast to the germline TE-induced defects that are known to be mostly suppressed by chk2 mutation. These results demonstrate a crucial role of checkpoint kinases in protecting niche cells against deleterious TE activation and suggest substantial differences between DNA damage responses in ovarian somatic and germ cells.
Senos Demarco, R., Uyemura, B. S. and Jones, D. L. (2020). EGFR Signaling Stimulates Autophagy to Regulate Stem Cell Maintenance and Lipid Homeostasis in the Drosophila Testis. Cell Rep 30(4): 1101-1116.e1105. PubMed ID: 31995752
Although typically upregulated upon cellular stress, autophagy can also be utilized under homeostatic conditions as a quality control mechanism or in response to developmental cues. This study reports that autophagy is required for the maintenance of somatic cyst stem cells (CySCs) in the Drosophila testis. Disruption of autophagy in CySCs and early cyst cells (CCs) by the depletion of autophagy-related (Atg) genes reduced early CC numbers and affected CC function, resembling decreased epidermal growth factor receptor (EGFR) signaling. Indeed, the data indicate that EGFR acts to stimulate autophagy to preserve early CC function, whereas target of rapamycin (TOR) negatively regulates autophagy in the differentiating CCs. Finally, this study shows that the EGFR-mediated stimulation of autophagy regulates lipid levels in CySCs and CCs. These results demonstrate a key role for autophagy in regulating somatic stem cell behavior and tissue homeostasis by integrating cues from both the EGFR and TOR signaling pathways to control lipid metabolism.
Shen, W. and Sun, J. (2020). Different modes of Notch activation and strength regulation in the spermathecal secretory lineage. Development 147(3). PubMed ID: 31988187
The strength of Notch signaling contributes to pleiotropic actions of Notch; however, there is as yet no a full understanding of the molecular regulation of Notch-signaling strength. This study investigated the mode of Notch activation in binary fate specification in the Drosophila spermathecal linage, where Notch is asymmetrically activated across three divisions to specify different cell fates. Using clonal analysis, this study shows that Delta (Dl) serves as the ligand for Notch in the first and second divisions. Dl and Serrate (Ser) function redundantly in the third division. Compared with the third division, cell-fate decision in the second division requires a lower level of Suppressor of Hairless protein, and, consequently, a lower level of Notch signaling. Several Notch endosomal trafficking regulators differentially regulate Notch signaling between the second and third divisions. This study demonstrate that cell differentiation in spermathecae involves different Notch-activation modes, Notch-signaling strengths and Notch-trafficking regulations. Thus, the Drosophila spermathecal lineage is an exciting model for probing the molecular mechanisms that modulate the Notch signaling pathway.
Slaidina, M., Banisch, T. U., Gupta, S. and Lehmann, R. (2020). A single-cell atlas of the developing Drosophila ovary identifies follicle stem cell progenitors. Genes Dev 34(3-4): 239-249. PubMed ID: 31919193
Addressing the complexity of organogenesis at a system-wide level requires a complete understanding of adult cell types, their origin, and precursor relationships. The Drosophila ovary has been a model to study how coordinated stem cell units, germline, and somatic follicle stem cells maintain and renew an organ. However, lack of cell type-specific tools have limited the ability to study the origin of individual cell types and stem cell units. This study used a single-cell RNA sequencing approach to uncover all known cell types of the developing ovary, reveal transcriptional signatures, and identify cell type-specific markers for lineage tracing. This study identifies a novel cell type corresponding to the elusive follicle stem cell precursors and predicts subtypes of known cell types. Altogether, this study reveals a previously unanticipated complexity of the developing ovary and provide a comprehensive resource for the systematic analysis of ovary morphogenesis.
Persico, V., Migliorini, M., Callaini, G. and Riparbelli, M. G. (2020). The Singularity of the Drosophila Male Germ Cell Centriole: The Asymmetric Distribution of Sas4 and Sas6. Cells 9(1). PubMed ID: 31947732
Drosophila spermatocytes have giant centrioles that display unique properties. Both the parent centrioles maintain a distinct cartwheel and nucleate a cilium-like region that persists during the meiotic divisions and organizes a structured sperm axoneme. Moreover, the parent centrioles are morphologically undistinguishable, unlike vertebrate cells in which mother and daughter centrioles have distinct structural features. However, immunofluorescence analysis of the parent centrioles in mature primary spermatocytes revealed an asymmetric accumulation of the typical Sas4 and Sas6 proteins. Notably, the fluorescence intensity of Sas4 and Sas6 at the daughter centrioles is greater than the intensity found at the mother ones. In contrast, the centrioles of wing imaginal disc cells display an opposite condition in which the loading of Sas4 and Sas6 at the mother centrioles is greater. These data underlie a subtle asymmetry among the parent centrioles and point to a cell type diversity of the localization of the Sas4 and Sas6 proteins (Persico, 2020).

Wednesday March 17th - Adult neural development and function

Okamoto, N. and Yamanaka, N. (2020). Steroid Hormone Entry into the Brain Requires a Membrane Transporter in Drosophila. Curr Biol 30(2): 359-366. PubMed ID: 31928869
Steroid hormones control various aspects of brain development and behavior in metazoans, but how they enter the central nervous system (CNS) through the blood-brain barrier (BBB) remains poorly understood. It is generally believed that steroid hormones freely diffuse through the plasma membrane of the BBB cells to reach the brain, because of the predominant "simple diffusion" model of steroid hormone transport across cell membranes. Recently, however, the simple diffusion model was challenged by showing that a Drosophila organic anion-transporting polypeptide (OATP), which was named Ecdysone Importer (EcI), is required for cellular uptake of the primary insect steroid hormone ecdysone. As ecdysone is first secreted into the hemolymph before reaching the CNS, this finding raised the question of how ecdysone enters the CNS through the BBB to exert its diverse role in Drosophila brain development. This study demonstrates in the Drosophila BBB that EcI is indispensable for ecdysone entry into the CNS to facilitate brain development. EcI is highly expressed in surface glial cells that form the BBB, and EcI knockdown in the BBB suppresses ecdysone signaling within the CNS and blocks ecdysone-mediated neuronal events during development. In an ex vivo culture system, the CNS requires EcI in the BBB to incorporate ecdysone from the culture medium. The results suggest a transporter-mediated mechanism of steroid hormone entry into the CNS, which may provide important implications in controlling brain development and behavior by regulating steroid hormone permeability across the BBB.
Noyes, N. C., Walkinshaw, E. and Davis, R. L. (2020). Ras acts as a molecular switch between two forms of consolidated memory in Drosophila. Proc Natl Acad Sci U S A 117(4): 2133-2139. PubMed ID: 31932418
Long-lasting, consolidated memories require not only positive biological processes that facilitate long-term memories (LTM) but also the suppression of inhibitory processes that prevent them. The mushroom body neurons (MBn) in Drosophila melanogaster store protein synthesis-dependent LTM (PSD-LTM) as well as protein synthesis-independent, anesthesia-resistant memory (ARM). The formation of ARM inhibits PSD-LTM but the underlying molecular processes that mediate this interaction remain unknown. This study demonstrate that the Ras-->Raf-->rho kinase (ROCK) pathway in MBn suppresses ARM consolidation, allowing the formation of PSD-LTM. The initial results revealed that the effects of Ras on memory are due to postacquisition processes. Ras knockdown enhanced memory expression but had no effect on acquisition. Additionally, increasing Ras activity optogenetically after, but not before, acquisition impaired memory performance. The elevated memory produced by Ras knockdown is a result of increased ARM. While Ras knockdown enhanced the consolidation of ARM, it eliminated PSD-LTM. These effects are mediated by the downstream kinase Raf. Similar to Ras, knockdown of Raf enhanced ARM consolidation and impaired PSD-LTM. Surprisingly, knockdown of the canonical downstream extracellular signal-regulated kinase did not reproduce the phenotypes observed with Ras and Raf knockdown. Rather, Ras/Raf inhibition of ROCK was found to be responsible for suppressing ARM. Constitutively active ROCK enhanced ARM and impaired PSD-LTM, while decreasing ROCK activity rescued the enhanced ARM produced by Ras knockdown. It is concluded that MBn Ras/Raf inhibition of ROCK suppresses the consolidation of ARM, which permits the formation of PSD-LTM.
Philpott, J. M., Narasimamurthy, R., Ricci, C. G., Freeberg, A. M., Hunt, S. R., Yee, L. E., Pelofsky, R. S., Tripathi, S., Virshup, D. M. and Partch, C. L. (2020). Casein kinase 1 dynamics underlie substrate selectivity and the PER2 circadian phosphoswitch. Elife 9. PubMed ID: 32043967
Post-translational control of PERIOD stability by Casein Kinase 1delta/epsilon (CK1: Doubletime) plays a key regulatory role in metazoan circadian rhythms. Despite the deep evolutionary conservation of CK1 in eukaryotes, little is known about its regulation and the factors that influence substrate selectivity on functionally antagonistic sites in PERIOD that directly control circadian period. This study describes a molecular switch involving a highly conserved anion binding site in CK1. This switch controls conformation of the kinase activation loop and determines which sites on mammalian PER2 are preferentially phosphorylated, thereby directly regulating PER2 stability. Integrated experimental and computational studies shed light on the allosteric linkage between two anion binding sites that dynamically regulate kinase activity. Period-altering kinase mutations from humans to Drosophila differentially modulate this activation loop switch to elicit predictable changes in PER2 stability, providing a foundation to understand and further manipulate CK1 regulation of circadian rhythms.
Ozer, I. and Carle, T. (2020). Back to the light, coevolution between vision and olfaction in the "Dark-flies" (Drosophila melanogaster). PLoS One 15(2): e0228939. PubMed ID: 32045466
Trade-off between vision and olfaction, the fact that investment in one correlates with decreased investment in the other, has been demonstrated by a wealth of comparative studies. However, there is still no empirical evidence suggesting how these two sensory systems coevolve, i.e. simultaneously or alternatively. The "Dark-flies" (Drosophila melanogaster) constitute a unique model to investigate such relation since they have been reared in the dark since 1954, approximately 60 years (~1500 generations). To observe how vision and olfaction evolve, populations of Dark-flies were reared in normal lighting conditions for 1 (DF1G) and 65 (DF65G) generations. The sizes of the visual (optic lobes, OLs) and olfactory (antennal lobes, ALs) primary centres, as well as the rest of the brain, and compared the results with the original and its genetically most similar strain (Oregon flies). Whereas the ALs decreased in size, the OLs (together with the brain) increased in size in the Dark-flies returned back to the light, both in the DF1G and DF65G. These results experimentally show that trade-off between vision and olfaction occurs simultaneously, and suggests that there are possible genetic and epigenetic processes regulating the size of both optic and antennal lobes. Furthermore, although the Dark-flies were able to mate and survive in the dark with a reduced neural investment, individuals being returned to the light seem to have been selected with reinvestment in visual capabilities despite a potential higher energetic cost.
Mishra, A., Cronley, P., Ganesan, M., Schulz, D. J. and Zars, T. (2020). Dopaminergic neurons can influence heat-box place learning in Drosophila. J Neurogenet: 1-8. PubMed ID: 31997669
Dopamine provides crucial neuromodulatory functions in several insect and rodent learning and memory paradigms. However, an early study suggested that dopamine may be dispensable for aversive place memory in Drosophila. This study tested the involvement of particular dopaminergic neurons in place learning and memory. The thermogenetic tool Gr28bD was used to activate protocerebral anterior medial (PAM) cluster and non-PAM dopaminergic neurons in an operant way in heat-box place learning. Activation of PAM neurons influences performance during place learning, but not during memory testing. These findings provide a gateway to explore how dopamine influences place learning.
McKinney, H., Sherer, L. M., Williams, J. L., Certel, S. J. and Stowers, R. S. (2020). Characterization of Drosophila Octopamine receptor neuronal expression using MiMIC-converted Gal4 lines. J Comp Neurol. PubMed ID: 32060912
Octopamine, the invertebrate analog of norepinephrine, is known to modulate a large variety of behaviors in Drosophila including feeding initiation, locomotion, aggression, and courtship, among many others. Significantly less is known about the identity of the neurons that receive octopamine input and how they mediate octopamine-regulated behaviors. This study characterized adult neuronal expression of MiMIC-converted Trojan-Gal4 lines for each of the five Drosophila octopamine receptors. Broad neuronal expression was observed for all five octopamine receptors, yet distinct differences among them were also apparent. Use of immunostaining for the octopamine neurotransmitter synthesis enzyme Tdc2, along with a novel genome-edited conditional Tdc2-LexA driver, revealed all five octopamine receptors express in Tdc2/octopamine neurons to varying degrees. This suggests autoreception may be an important circuit mechanism by which octopamine modulates behavior.

Monday March 16th - Disease models and signaling

Elvira, R., Cha, S. J., Noh, G. M., Kim, K. and Han, J. (2020). PERK-mediated eIF2alpha phosphorylation contributes to the protection of dopaminergic neurons from chronic heat stress in Drosophila. Int J Mol Sci 21(3). PubMed ID: 32013014
Environmental high-temperature heat exposure is linked to physiological stress such as disturbed protein homeostasis caused by endoplasmic reticulum (ER) stress. Abnormal proteostasis in neuronal cells is a common pathological factor of Parkinson's disease (PD). Chronic heat stress is thought to induce neuronal cell death during the onset and progression of PD, but the exact role and mechanism of ER stress and the activation of the unfolded protein response (UPR) remains unclear. This study showed that chronic heat exposure induces ER stress mediated by the PKR-like eukaryotic initiation factor 2alpha kinase (PERK)/eIF2alpha phosphorylation signaling pathway in Drosophila neurons. Chronic heat-induced eIF2alpha phosphorylation was regulated by PERK activation and required for neuroprotection from chronic heat stress. Moreover, the attenuated protein synthesis by eIF2alpha phosphorylation was a critical factor for neuronal cell survival during chronic heat stress. Genetic downregulation of PERK, specifically in dopaminergic (DA) neurons, impaired motor activity and led to DA neuron loss. Therefore, these findings provide in vivo evidence demonstrating that chronic heat exposure may be a critical risk factor in the onset of PD, and eIF2alpha phosphorylation mediated by PERK may contribute to the protection of DA neurons against chronic heat stress in Drosophila.
Kwon, S. Y., Massey, K., Watson, M. A., Hussain, T., Volpe, G., Buckley, C. D., Nicolaou, A. and Badenhorst, P. (2020). Oxidised metabolites of the omega-6 fatty acid linoleic acid activate dFOXO. Life Sci Alliance 3(2). PubMed ID: 31992650
Obesity-induced inflammation, or meta-inflammation, plays key roles in metabolic syndrome and is a significant risk factor in diabetes and cardiovascular disease. To investigate causal links between obesity, meta-inflammation, and insulin signaling a Drosophila model was established to determine how elevated dietary fat and changes in the levels and balance of saturated fatty acids (SFAs) and polyunsaturated fatty acids (PUFAs) influence inflammation. Negligible effect of saturated fatty acid on inflammation was observed but marked enhancement or suppression by omega-6 and omega-3 PUFAs, respectively. Using combined lipidomic and genetic analysis, omega-6 polyunsaturated fatty acid was shown to enhance meta-inflammation by producing linoleic acid-derived lipid mediator 9-hydroxy-octadecadienoic acid (9-HODE). Transcriptome analysis reveals 9-HODE functions by regulating FOXO family transcription factors. 9-HODE activates JNK, triggering FOXO nuclear localisation and chromatin binding. FOXO TFs are important transducers of the insulin signaling pathway that are normally down-regulated by insulin. By activating FOXO, 9-HODE could antagonise insulin signaling providing a molecular conduit linking changes in dietary fatty acid balance, meta-inflammation, and insulin resistance.
Ham, S. J., Lee, D., Yoo, H., Jun, K., Shin, H. and Chung, J. (2020). Decision between mitophagy and apoptosis by Parkin via VDAC1 ubiquitination. Proc Natl Acad Sci U S A. PubMed ID: 32047033
VDAC1 is a critical substrate of Parkin responsible for the regulation of mitophagy and apoptosis. This study demonstrates that VDAC1 can be either mono- or polyubiquitinated by Parkin in a PINK1-dependent manner. VDAC1 deficient with polyubiquitination (VDAC1 Poly-KR) hampers mitophagy, but VDAC1 deficient with monoubiquitination (VDAC1 K274R) promotes apoptosis by augmenting the mitochondrial calcium uptake through the mitochondrial calcium uniporter (MCU) channel. The transgenic flies expressing Drosophila Porin K273R, corresponding to human VDAC1 K274R, show Parkinson disease (PD)-related phenotypes including locomotive dysfunction and degenerated dopaminergic neurons, which are relieved by suppressing MCU and mitochondrial calcium uptake. To further confirm the relevance of these findings in PD, a missense mutation of Parkin was discovered in PD patients, T415N, which lacks the ability to induce VDAC1 monoubiquitination but still maintains polyubiquitination. Interestingly, Drosophila Parkin T433N, corresponding to human Parkin T415N, fails to rescue the PD-related phenotypes of Parkin-null flies. Taken together, these results suggest that VDAC1 monoubiquitination plays important roles in the pathologies of PD by controlling apoptosis.
Hwang, S. H., Bang, S., Kim, W. and Chung, J. (2020). Von Hippel-Lindau tumor suppressor (VHL) stimulates TOR signaling by interacting with phosphoinositide 3-kinase (PI3K). J Biol Chem. PubMed ID: 31959630
Cell growth is positively controlled by phosphoinositide 3-kinase (PI3K)-TOR signaling pathway under conditions of abundant growth factors and nutrients. To discover additional mechanisms that regulate cell growth, an RNAi-based mosaic analyses was performed in the Drosophila fat body, the primary metabolic organ in the fly. Unexpectedly, the knockdown of the Drosophila von Hippel-Lindau (VHL) gene markedly decreased cell size and body size. These cell growth phenotypes induced by VHL loss-of-function were recovered by activation of TOR signaling in Drosophila. Consistent with the genetic interactions between VHL and the signaling components of PI3K-TOR pathway in Drosophila, it was observed that VHL loss-of-function in mammalian cells causes decreased phosphorylation of ribosomal protein S6 kinase (S6K) and Akt, which represent the main activities of this pathway. It was further demonstrated that VHL activates TOR signaling by directly interacting with the p110 catalytic subunit of PI3K. On the basis of the evolutionarily conserved regulation of PI3K-TOR signaling by VHL observed in this study, it is proposed that VHL plays an important role in the regulation and maintenance of proper cell growth in metazoans.
Lee, J. J., Andreazza, S. and Whitworth, A. J. (2020). The STING pathway does not contribute to behavioural or mitochondrial phenotypes in Drosophila Pink1/parkin or mtDNA mutator models. Sci Rep 10(1): 2693. PubMed ID: 32060339
Mutations in PINK1 and Parkin/PRKN cause the degeneration of dopaminergic neurons in familial forms of Parkinson's disease but the precise pathogenic mechanisms are unknown. The PINK1/Parkin pathway has been described to play a central role in mitochondrial homeostasis by signalling the targeted destruction of damaged mitochondria, however, how disrupting this process leads to neuronal death was unclear until recently. An elegant study in mice revealed that the loss of Pink1 or Prkn coupled with an additional mitochondrial stress resulted in the aberrant activation of the innate immune signalling, mediated via the cGAS/STING pathway, causing degeneration of dopaminergic neurons and motor impairment. Genetic knockout of Sting was sufficient to completely prevent neurodegeneration and accompanying motor deficits. To determine whether Sting plays a conserved role in Pink1/parkin related pathology, tests were performed for genetic interactions between Sting and Pink1/parkin in Drosophila. Surprisingly, loss of Sting, or its downstream effector Relish, was insufficient to suppress the behavioural deficits or mitochondria disruption in the Pink1/parkin mutants. Thus, it is concluded that phenotypes associated with loss of Pink1/parkin are not universally due to aberrant activation of the STING pathway.
Norkett, R., Del Castillo, U., Lu, W. and Gelfand, V. I. (2020). Ser/Thr kinase Trc controls neurite outgrowth in Drosophila by modulating microtubule-microtubule sliding. Elife 9. PubMed ID: 32022690
Correct neuronal development requires tailored neurite outgrowth. Neurite outgrowth is driven in part by microtubule-sliding - the transport of microtubules along each other. It has been recently demonstrated that a 'mitotic' kinesin-6 (Pavarotti in Drosophila) effectively inhibits microtubule-sliding and neurite outgrowth. However, mechanisms regulating Pavarotti itself in interphase cells and specifically in neurite outgrowth are unknown. This study used a combination of live imaging and biochemical methods to show that the inhibition of microtubule-sliding by Pavarotti is controlled by phosphorylation. The Ser/Thr NDR kinase Tricornered (Trc) as a Pavarotti-dependent regulator of microtubule sliding in neurons. Further, Trc-mediated phosphorylation of Pavarotti was shown to promote its interaction with 14-3-3 proteins. Loss of 14-3-3 prevents Pavarotti from associating with microtubules. Thus, a pathway is proposed by which microtubule-sliding can be up- or downregulated in neurons to control neurite outgrowth, and establish parallels between microtubule-sliding in mitosis and post-mitotic neurons.

Friday, March 13th - Gonads

McCambridge, A., Solanki, D., Olchawa, N., Govani, N., Trinidad, J. C. and Gao, M. (2020). Comparative Proteomics Reveal Me31B's Interactome Dynamics, Expression Regulation, and Assembly Mechanism into Germ Granules during Drosophila Germline Development. Sci Rep 10(1): 564. PubMed ID: 31953495
Me31B is a protein component of Drosophila germ granules and plays an important role in germline development by interacting with other proteins and RNAs. To understand the dynamic changes that the Me31B interactome undergoes from oogenesis to early embryogenesis, this study characterized the early embryo Me31B interactome and compared it to the known ovary interactome. The two interactomes shared RNA regulation proteins, glycolytic enzymes, and cytoskeleton/motor proteins, but the core germ plasm proteins Vas, Tud, and Aub were significantly decreased in the embryo interactome. Follow-up on two RNA regulations proteins present in both interactomes, Tral and Cup, revealed that they colocalize with Me31B in nuage granules, P-bodies/sponge bodies, and possibly in germ plasm granules. It was further shown that Tral and Cup are both needed for maintaining Me31B protein level and mRNA stability, with Tral's effect being more specific. In addition, evidence is provided that Me31B likely colocalizes and interacts with germ plasm marker Vas in the ovaries and early embryo germ granules. Finally, it was shown that Me31B's localization in germ plasm is likely independent of the Osk-Vas-Tud-Aub germ plasm assembly pathway although its proper enrichment in the germ plasm may still rely on certain conserved germ plasm proteins.
McGeary, M. K. and Findlay, G. D. (2020). Molecular evolution of the sex peptide network in Drosophila. J Evol Biol. PubMed ID: 31991034
This study investigated the molecular evolution of the 'sex peptide network' of Drosophila melanogaster, a functionally well-characterized reproductive protein network. In this species, the peptide hormone sex peptide (SP) and its interacting proteins cause major changes in female physiology and behaviour after mating. In contrast, females of more distantly related Drosophila species do not respond to SP. In spite of these phenotypic differences, orthologs of all network proteins were detected across 22 diverse Drosophila species, and evidence was found that most orthologs likely function in reproduction throughout the genus. Within SP-responsive species, the recurrent, adaptive evolution was detected of several network proteins, consistent with sexual selection acting to continually refine network function. Some evidence was found for adaptive evolution of several proteins along two specific phylogenetic lineages that correspond with increased expression of the SP receptor in female reproductive tracts or increased sperm length, respectively. Finally, gene expression profiling was used to examine the likely degree of functional conservation of the paralogs of an SP network protein that arose via gene duplication. These results suggest a dynamic history for the SP network in which network members arose before the onset of robust SP-mediated responses and then were shaped by both purifying and positive selection.
Lu, W., Lakonishok, M., Liu, R., Billington, N., Rich, A., Glotzer, M., Sellers, J. R. and Gelfand, V. I. (2020). Competition between kinesin-1 and myosin-V defines Drosophila posterior determination. Elife 9. PubMed ID: 32057294
Local accumulation of oskar (osk) mRNA in the Drosophila oocyte determines the posterior pole of the future embryo. Two major cytoskeletal components, microtubules and actin filaments, together with a microtubule motor, kinesin-1, and an actin motor, myosin-V, are essential for osk mRNA posterior localization. This study used Staufen, an RNA-binding protein that colocalizes with osk mRNA, as a proxy for osk mRNA. Posterior localization of osk/Staufen was shown to be determined by competition between kinesin-1 and myosin-V. While kinesin-1 removes osk/Staufen from the cortex along microtubules, myosin-V anchors osk/Staufen at the cortex. Myosin-V wins over kinesin-1 at the posterior pole due to low microtubule density at this site, while kinesin-1 wins at anterior and lateral positions because they have high density of cortically-anchored microtubules. As a result, posterior determinants are removed from the anterior and lateral cortex but retained at the posterior pole. Thus, posterior determination of Drosophila oocytes is defined by kinesin-myosin competition, whose outcome is primarily determined by cortical microtubule density.
Lee, Y. M., Chiang, P. H., Cheng, J. H., Shen, W. H., Chen, C. H., Wu, M. L., Tian, Y. L., Ni, C. H., Wang, T. F., Lin, M. D. and Chou, T. B. (2020). Drosophila decapping protein 2 modulates the formation of cortical F-actin for germ plasm assembly. Dev Biol. PubMed ID: 32007453
In Drosophila, the deposition of the germ plasm at the posterior pole of the oocyte is essential for the abdomen and germ cell formation during embryogenesis. To assemble the germ plasm, oskar (osk) mRNA, produced by nurse cells, should be localized and anchored on the posterior cortex of the oocyte. Processing bodies (P-bodies) are cytoplasmic RNA granules responsible for the 5'->3' mRNA degradation. Evidence suggests that the components of P-bodies, such as Drosophila decapping protein 1 and Ge-1, are involved in the posterior localization of osk. However, whether the decapping core enzyme, Drosophila decapping protein 2 (dDcp2), is also involved remains unclear. This study generated a dDcp2 null allele and showed that dDcp2 was required for the posterior localization of germ plasm components including osk. dDcp2 was distributed on the oocyte cortex and was localized posterior to the osk. In the posterior pole of dDcp2 mutant oocytes, osk was mislocalized and colocalized with F-actin detached from the cortex; moreover, considerably fewer F-actin projections were observed. Using the F-actin cosedimentation assay, dDcp2 was shown to interact with F-actin through its middle region. In conclusion, these findings explored a novel function of dDcp2 in assisting osk localization by modulating the formation of F-actin projections on the posterior cortex.
Kovacs, L., Nagy, A., Pal, M. and Deak, P. (2020). Usp14 is required for spermatogenesis and ubiquitin stress responses in Drosophila melanogaster. J Cell Sci 133(2). PubMed ID: 31974276
Deubiquitylating (DUB) enzymes free covalently linked ubiquitin moieties from ubiquitin-ubiquitin and ubiquitin-protein conjugates, and thereby maintain the equilibrium between free and conjugated ubiquitin moieties and regulate ubiquitin-mediated cellular processes. This study performed genetic analyses of mutant phenotypes in Drosophila melanogaster and demonstrates that loss of Usp14 function results in male sterility, with defects in spermatid individualization and reduced testicular free monoubiquitin levels. These phenotypes were rescued by germline-specific overexpression of wild-type Usp14. Synergistic genetic interactions with Ubi-p63E and cycloheximide sensitivity suggest that ubiquitin shortage is a primary cause of male sterility. In addition, Usp14 is predominantly expressed in testes in Drosophila, indicating a higher demand for this DUB in testes that is also reflected by testis-specific loss-of-function Usp14 phenotypes. Collectively, these results suggest a major role of Usp14 in maintaining normal steady state free monoubiquitin levels during the later stages of Drosophila spermatogenesis.
Eichler, C. E., Hakes, A. C., Hull, B. and Gavis, E. R. (2020). Compartmentalized oskar degradation in the germ plasm safeguards germline development. Elife 9. PubMed ID: 31909715
Partitioning of mRNAs into ribonucleoprotein (RNP) granules supports diverse regulatory programs within the crowded cytoplasm. At least two types of RNP granules populate the germ plasm, a cytoplasmic domain at the posterior of the Drosophila oocyte and embryo. Germ granules deliver mRNAs required for germline development to pole cells, the germ cell progenitors. A second type of RNP granule, here named founder granules, contains oskar mRNA, which encodes the germ plasm organizer. Whereas oskar mRNA is essential for germ plasm assembly during oogenesis, this study shows that it is toxic to pole cells. Founder granules mediate compartmentalized degradation of oskar during embryogenesis to minimize its inheritance by pole cells. Degradation of oskar in founder granules is temporally and mechanistically distinct from degradation of oskar and other mRNAs during the maternal-to-zygotic transition. These results show how compartmentalization in RNP granules differentially controls fates of mRNAs localized within the same cytoplasmic domain.

Thursday, March 12 - Adult neural development and function

Suzuki, Y., Schenk, J. E., Tan, H. and Gaudry, Q. (2020). A Population of Interneurons Signals Changes in the Basal Concentration of Serotonin and Mediates Gain Control in the Drosophila Antennal Lobe. Curr Biol. PubMed ID: 32142699
Serotonin (5-HT) represents a quintessential neuromodulator, having been identified in nearly all animal species where it functions in cognition, motor control, and sensory processing. In the olfactory circuits of flies and mice, serotonin indirectly inhibits odor responses in olfactory receptor neurons (ORNs) via GABAergic local interneurons (LNs). However, the effects of 5-HT in olfaction are likely complicated, because multiple receptor subtypes are distributed throughout the olfactory bulb (OB) and antennal lobe (AL), the first layers of olfactory neuropil in mammals and insects, respectively. For example, serotonin has a non-monotonic effect on odor responses in Drosophila projection neurons (PNs), where low concentrations suppress odor-evoked activity and higher concentrations boost PN responses. Serotonin reaches the AL via the diffusion of paracrine 5-HT through the fly hemolymph and by activation of the contralaterally projecting serotonin-immunoreactive deuterocerebral interneurons (CSDns): the only serotonergic cells that innervate the AL. Concentration-dependent effects could arise by either the expression of multiple 5-HT receptors (5-HTRs) on the same cells or by populations of neurons dedicated to detecting serotonin at different concentrations. This study identify a population of LNs that express 5-HT7Rs exclusively to detect basal concentrations of 5-HT. These LNs inhibit PNs via GABAB receptors and mediate subtractive gain control. LNs expressing 5-HT7Rs are broadly tuned to odors and target every glomerulus in the antennal lobe. These results demonstrate that serotonergic modulation at low concentrations targets a specific population of LNs to globally downregulate PN odor responses in the AL.
Maierbrugger, K. T., Sousa-Nunes, R. and Bateman, J. M. (2020). The mTOR pathway component Unkempt regulates neural stem cell and neural progenitor cell cycle in the Drosophila central nervous system. Dev Biol. PubMed ID: 31978396
The formation of a complex nervous system requires the coordinated action of progenitor cell proliferation, differentiation and maturation. The Drosophila postembryonic central nervous system provides a powerful model for dissecting the cellular and molecular mechanisms underpinning neurogenesis. The conserved zinc finger/RING protein Unkempt (Unk) has been identified as a key temporal regulator of neuronal differentiation in the Drosophila developing eye. Unk acts downstream of the mechanistic target of rapamycin (mTOR) pathway together with its binding partner Headcase (Hdc). This study investigated the role of Unk in Drosophila postembryonic thoracic neurogenesis. The Drosophila central nervous system contains neural stem cells, called neuroblasts, and neural progenitors, known as ganglion mother cells (GMCs). Unk is highly expressed in the central brain and ventral nerve cord but is not required to maintain neuroblast numbers or for the regulation of temporal series factor expression in neuroblasts. However, loss of Unk increases the number of neuroblasts and GMCs in S-phase of the cell cycle, resulting in the overproduction of neurons.Unk interacts with Hdc through its zinc finger domain. The zinc finger domain is required for the synergistic activity of Unk with Hdc during eye development but is not necessary for the activity of Unk in thoracic neurogenesis. Overall, this study shows that Unk and Hdc are novel negative regulators of neurogenesis in Drosophila and indicates a conserved role of mTOR signalling in nervous system development.
Ketkar, M. D., Sporar, K., Gur, B., Ramos-Traslosheros, G., Seifert, M. and Silies, M. (2020). Luminance Information Is Required for the Accurate Estimation of Contrast in Rapidly Changing Visual Contexts. Curr Biol. PubMed ID: 32008904
Visual perception scales with changes in the visual stimulus, or contrast, irrespective of background illumination. However, visual perception is challenged when adaptation is not fast enough to deal with sudden declines in overall illumination, for example, when gaze follows a moving object from bright sunlight into a shaded area. This study shows that the visual system of the fly employs a solution by propagating a corrective luminance-sensitive signal. In vivo 2-photon imaging and behavioral analyses were used to demonstrate that distinct OFF-pathway inputs encode contrast and luminance. Predictions of contrast-sensitive neuronal responses show that contrast information alone cannot explain behavioral responses in sudden dim light. The luminance-sensitive pathway via the L3 neuron is required for visual processing in such rapidly changing light conditions, ensuring contrast constancy when pure contrast sensitivity underestimates a stimulus. Thus, retaining a peripheral feature, luminance, in visual processing is required for robust behavioral responses.
Kondo, S., Takahashi, T., Yamagata, N., Imanishi, Y., Katow, H., Hiramatsu, S., Lynn, K., Abe, A., Kumaraswamy, A. and Tanimoto, H. (2020). Neurochemical Organization of the Drosophila Brain Visualized by Endogenously Tagged Neurotransmitter Receptors. Cell Rep 30(1): 284-297. PubMed ID: 31914394
Neurotransmitters often have multiple receptors that induce distinct responses in receiving cells. Expression and localization of neurotransmitter receptors in individual neurons are therefore critical for understanding the operation of neural circuits. This study describes a comprehensive library of reporter strains in which a convertible T2A-GAL4 cassette is inserted into endogenous neurotransmitter receptor genes of Drosophila. Using this library, the expression of 75 neurotransmitter receptors was profiled in the brain. Cluster analysis reveals neurochemical segmentation of the brain, distinguishing higher brain centers from the rest. By recombinase-mediated cassette exchange, T2A-GAL4 was converted into split-GFP and Tango to visualize subcellular localization and activation of dopamine receptors in specific cell types. This reveals striking differences in their subcellular localization, which may underlie the distinct cellular responses to dopamine in different behavioral contexts. These resources thus provide a versatile toolkit for dissecting the cellular organization and function of neurotransmitter systems in the fly brain.
Masson, J. B., Laurent, F., Cardona, A., Barre, C., Skatchkovsky, N., Zlatic, M. and Jovanic, T. (2020). Identifying neural substrates of competitive interactions and sequence transitions during mechanosensory responses in Drosophila. PLoS Genet 16(2): e1008589. PubMed ID: 32059010
Nervous systems have the ability to select appropriate actions and action sequences in response to sensory cues. The circuit mechanisms by which nervous systems achieve choice, stability and transitions between behaviors are still incompletely understood. To identify neurons and brain areas involved in controlling these processes, a large-scale neuronal inactivation screen was combined with automated action detection in response to a mechanosensory cue in Drosophila larva. Behaviors were analyzed from 2.9x105 larvae, and 66 candidate lines were identified for mechanosensory responses out of which 25 for competitive interactions between actions. The neurons in these lines were further characterized in detail and their connectivity was analyzed using electron microscopy. The neurons in the mechanosensory network were found to be located in different regions of the nervous system consistent with a distributed model of sensorimotor decision-making. These findings provide the basis for understanding how selection and transition between behaviors are controlled by the nervous system.
Matulis, C. A., Chen, J., Gonzalez-Suarez, A. D., Behnia, R. and Clark, D. A. (2020). Heterogeneous Temporal Contrast Adaptation in Drosophila Direction-Selective Circuits. Curr Biol 30(2): 222-236. PubMed ID: 31928874
In visual systems, neurons adapt both to the mean light level and to the range of light levels, or the contrast. Contrast adaptation has been studied extensively, but it remains unclear how it is distributed among neurons in connected circuits, and how early adaptation affects subsequent computations. This study investigated temporal contrast adaptation in neurons across Drosophila's visual motion circuitry. Several ON-pathway neurons showed strong adaptation to changes in contrast over time. One of these neurons, Mi1, showed almost complete adaptation on fast timescales, and experiments ruled out several potential mechanisms for its adaptive properties. When contrast adaptation reduced the gain in ON-pathway cells, it was accompanied by decreased motion responses in downstream direction-selective cells. Simulations show that contrast adaptation can substantially improve motion estimates in natural scenes. The benefits are larger for ON-pathway adaptation, which helps explain the heterogeneous distribution of contrast adaptation in these circuits.

Wednesday, March 11th - Neural development

Gonsior, M. and Ismat, A. (2019). sli is required for proper morphology and migration of sensory neurons in the Drosophila PNS. Neural Dev 14(1): 10. PubMed ID: 31651354
Neurons and glial cells coordinate with each other in many different aspects of nervous system development. Both types of cells are receiving multiple guidance cues to guide the neurons and glial cells to their proper final position. The lateral chordotonal organs (lch5) of the Drosophila peripheral nervous system (PNS) are composed of five sensory neurons surrounded by four different glial cells, scolopale cells, cap cells, attachment cells and ligament cells. During embryogenesis, the lch5 neurons go through a rotation and ventral migration to reach their final position in the lateral region of the abdomen. This study shows that the extracellular ligand sli is required for the proper ventral migration and morphology of the lch5 neurons. It was further shown that mutations in the Sli receptors Robo and Robo2 also display similar defects as loss of sli, suggesting a role for Slit-Robo signaling in lch5 migration and positioning. Additionally, it was demonstrated that the scolopale, cap and attachment cells follow the mis-migrated lch5 neurons in sli mutants, while the ventral stretching of the ligament cells seems to be independent of the lch5 neurons. This study sheds light on the role of Slit-Robo signaling in sensory neuron development.
Lin, T., Kao, H. H., Chou, C. H., Chou, C. Y., Liao, Y. C. and Lee, H. H. (2020). Rab11 activation by Ik2 kinase is required for dendrite pruning in Drosophila sensory neurons. PLoS Genet 16(2): e1008626. PubMed ID: 32059017
Neuronal pruning is a commonly observed phenomenon for the developing nervous systems to ensure precise wiring of neural circuits. The function of Ik2 kinase and its downstream mediator, Spindle-F (Spn-F), are essential for dendrite pruning of Drosophila sensory neurons during development. However, little is known about how Ik2/Spn-F signaling is transduced in neurons and ultimately results in dendrite pruning. Genetic analyses and rescue experiments have demonstrated that the small GTPase Rab11, especially the active GTP-bound form, is required for dendrite pruning. It was also found that Rab11 shows genetic interactions with spn-F and ik2 on pruning. Live imaging of single neurons and antibody staining reveal normal Ik2 kinase activation in Rab11 mutant neurons, suggesting that Rab11 could have a functional connection downstream of and/or parallel to the Ik2 kinase signaling. Moreover, biochemical evidence is provided that both the Ik2 kinase activity and the formation of Ik2/Spn-F/Rab11 complexes are central to promote Rab11 activation in cells. Together, these studies reveal that a critical role of Ik2/Spn-F signaling in neuronal pruning is to promote Rab11 activation, which is crucial for dendrite pruning in neurons.
Arefin, B., Parvin, F., Bahrampour, S., Stadler, C. B. and Thor, S. (2019). Drosophila neuroblast selection is gated by Notch, Snail, SoxB, and EMT bene interplay. Cell Rep 29(11): 3636-3651. PubMed ID: 31825841
In the developing Drosophila central nervous system (CNS), neural progenitor (neuroblast [NB]) selection is gated by lateral inhibition, controlled by Notch signaling and proneural genes. However, proneural mutants still generate many NBs, indicating the existence of additional proneural genes. Moreover, recent studies reveal involvement of key epithelial-mesenchymal transition (EMT) genes in NB selection, but the regulatory interplay between Notch signaling and the EMT machinery is unclear. This study finds that SoxNeuro (SoxB family) and worniu (Snail family) are integrated with the Notch pathway, and constitute the missing proneural genes. Notch signaling, the proneural, SoxNeuro, and worniu genes regulate key EMT genes to orchestrate the NB selection process. Hence, this study has uncovered an expanded lateral inhibition network for NB selection and demonstrate its link to key players in the EMT machinery. The evolutionary conservation of the genes involved suggests that the Notch-SoxB-Snail-EMT network may control neural progenitor selection in many other systems.
Li, J., Han, S., Li, H., Udeshi, N. D., Svinkina, T., Mani, D. R., Xu, C., Guajardo, R., Xie, Q., Li, T., Luginbuhl, D. J., Wu, B., McLaughlin, C. N., Xie, A., Kaewsapsak, P., Quake, S. R., Carr, S. A., Ting, A. Y. and Luo, L. (2020). Cell-Surface Proteomic Profiling in the Fly Brain Uncovers Wiring Regulators. Cell 180(2): 373-386. PubMed ID: 31955847
Molecular interactions at the cellular interface mediate organized assembly of single cells into tissues and, thus, govern the development and physiology of multicellular organisms. This study developed a cell-type-specific, spatiotemporally resolved approach to profile cell-surface proteomes in intact tissues. Quantitative profiling of cell-surface proteomes of Drosophila olfactory projection neurons (PNs) in pupae and adults revealed global downregulation of wiring molecules and upregulation of synaptic molecules in the transition from developing to mature PNs. A proteome-instructed in vivo screen identified 20 cell-surface molecules regulating neural circuit assembly, many of which belong to evolutionarily conserved protein families not previously linked to neural development. Genetic analysis further revealed that the lipoprotein receptor LRP1 cell-autonomously controls PN dendrite targeting, contributing to the formation of a precise olfactory map. These findings highlight the power of temporally resolved in situ cell-surface proteomic profiling in discovering regulators of brain wiring.
Seroka, A. Q. and Doe, C. Q. (2019). The Hunchback temporal transcription factor determines motor neuron axon and dendrite targeting in Drosophila. Development. PubMed ID: 30890568
The generation of neuronal diversity is essential for circuit formation and behavior. Morphological differences in sequentially born neurons could be due to intrinsic molecular identity specified by temporal transcription factors (henceforth called intrinsic temporal identity) or due to changing extrinsic cues. This study used the Drosophila NB7-1 lineage to address this question. NB7-1 generates the U1-U5 motor neurons sequentially; each has a distinct intrinsic temporal identity due to inheritance of different temporal transcription factors at its time of birth. This study shows that the U1-U5 neurons project axons sequentially, followed by sequential dendrite extension. The earliest temporal transcription factor, Hunchback, was misexpressed to create "ectopic" U1 neurons with an early intrinsic temporal identity but later birth-order. These ectopic U1 neurons have axon muscle targeting and dendrite neuropil targeting consistent with U1 intrinsic temporal identity, rather than their time of birth or differentiation. It is concluded that intrinsic temporal identity plays a major role in establishing both motor axon muscle targeting and dendritic arbor targeting, which are required for proper motor circuit development.
Gorla, M., Santiago, C., Chaudhari, K., Layman, A. A. K., Oliver, P. M. and Bashaw, G. J. (2019). Ndfip proteins target Robo receptors for degradation and allow commissural axons to cross the midline in the developing spinal cord. Cell Rep 26(12): 3298-3312. PubMed ID: 30893602
Commissural axons initially respond to attractive signals at the midline, but once they cross, they become sensitive to repulsive cues. This switch prevents axons from re-entering the midline. In insects and mammals, negative regulation of Roundabout (Robo) receptors prevents premature response to the midline repellant Slit. In Drosophila, the endosomal protein Commissureless (Comm) prevents Robo1 surface expression before midline crossing by diverting Robo1 into late endosomes. Notably, Comm is not conserved in vertebrates. This study identified two Nedd-4-interacting proteins, Ndfip1 and Ndfip2, that act analogously to Comm to localize Robo1 to endosomes. Ndfip proteins recruit Nedd4 E3 ubiquitin ligases to promote Robo1 ubiquitylation and degradation. Ndfip proteins are expressed in commissural axons in the developing spinal cord and removal of Ndfip proteins results in increased Robo1 expression and reduced midline crossing. The results define a conserved Robo1 intracellular sorting mechanism between flies and mammals to avoid premature responsiveness to Slit.

Tuesday, March 10th - Synapse and Vesicles

Coombes, C. E., Saunders, H. A. J., Mannava, A. G., Johnson-Schlitz, D. M., Reid, T. A., Parmar, S., McClellan, M., Yan, C., Rogers, S. L., Parrish, J. Z., Wagenbach, M., Wordeman, L., Wildonger, J. and Gardner, M. K. (2020). Non-enzymatic activity of the alpha-Tubulin acetyltransferase alphaTAT limits synaptic bouton growth in neurons. Curr Biol. PubMed ID: 31928876
Neuronal axons terminate as synaptic boutons that form stable yet plastic connections with their targets. Synaptic bouton development relies on an underlying network of both long-lived and dynamic microtubules that provide structural stability for the boutons while also allowing for their growth and remodeling. However, a molecular-scale mechanism that explains how neurons appropriately balance these two microtubule populations remains a mystery. It was hypothesized that alpha-tubulin acetyltransferase (alphaTAT), which both stabilizes long-lived microtubules against mechanical stress via acetylation and has been implicated in promoting microtubule dynamics, could play a role in this process. Using the Drosophila neuromuscular junction as a model, this study found that non-enzymatic αTAT activity limits the growth of synaptic boutons by affecting dynamic, but not stable, microtubules. Loss of αTAT results in the formation of ectopic boutons. These ectopic boutons can be similarly suppressed by resupplying enzyme-inactive αTAT or by treatment with a low concentration of the microtubule-targeting agent vinblastine, which acts to suppress microtubule dynamics. Biophysical reconstitution experiments revealed that non-enzymatic alphaTAT1 activity destabilizes dynamic microtubules but does not substantially impact the stability of long-lived microtubules. Further, during microtubule growth, non-enzymatic αTAT activity results in increasingly extended tip structures, consistent with an increased rate of acceleration of catastrophe frequency with microtubule age, perhaps via tip structure remodeling. Through these mechanisms, αTAT enriches for stable microtubules at the expense of dynamic ones. It is proposed that the specific suppression of dynamic microtubules by non-enzymatic αTAT activity regulates the remodeling of microtubule networks during synaptic bouton development.
Kopke, D. L., Leahy, S. N., Vita, D. J., Lima, S. C., Newman, Z. L. and Broadie, K. (2020). Carrier of Wingless (Cow) Regulation of Drosophila Neuromuscular Junction Development. eNeuro. PubMed ID: 32024666
The first Wnt signaling ligand discovered, Drosophila Wingless (Wg; Wnt1 in mammals), plays critical roles in neuromuscular junction (NMJ) development, regulating synaptic architecture and function. Heparan sulfate proteoglycans (HSPGs), consisting of a core protein with heparan sulfate (HS) glycosaminoglycan (GAG) chains, bind to Wg ligands to control both extracellular distribution and intercellular signaling function. Drosophila HSPGs previously shown to regulate Wg trans-synaptic signaling at the NMJ include the glypican Dally-like Protein (Dlp) and perlecan Terribly Reduced Optic Lobes (Trol). This study investigated synaptogenic functions of the most recently described Drosophila HSPG, secreted Carrier of Wingless (Cow), which directly binds Wg in the extracellular space. At the glutamatergic NMJ, it was found that Cow secreted from the presynaptic motor neuron acts to limit synaptic architecture and neurotransmission strength. In cow null mutants, this study found increased synaptic bouton number and elevated excitatory current amplitudes, phenocopying presynaptic Wg overexpression. cow null mutants exhibit an increased number of glutamatergic synapses and increased synaptic vesicle (SV) fusion frequency based both on GCaMP imaging and electrophysiology recording. Membrane-tethered Wg prevents cow null defects in NMJ development, indicating that Cow mediates secreted Wg signaling. It has been shown previously that the secreted Wg deacylase Notum restricts Wg signaling at the NMJ, and this study shows here that Cow and Notum work through the same pathway to limit synaptic development. It is concluded Cow acts cooperatively with Notum to coordinate neuromuscular synapse structural and functional differentiation via negative regulation of Wg trans-synaptic signaling within the extracellular synaptomatrix.
Lamaze, A., Jepson, J. E. C., Akpoghiran, O. and Koh, K. (2020). Antagonistic Regulation of Circadian Output and Synaptic Development by JETLAG and the DYSCHRONIC-SLOWPOKE Complex. iScience 23(2): 100845. PubMed ID: 32058958
Circadian output genes act downstream of the clock to promote rhythmic changes in behavior and physiology, yet their molecular and cellular functions are not well understood. This study characterized an interaction between regulators of circadian entrainment, output, and synaptic development in Drosophila that influences clock-driven anticipatory increases in morning and evening activity. Previous work has shown the JETLAG (JET) E3 ubiquitin ligase resets the clock upon light exposure, whereas the PDZ protein DYSCHRONIC (DYSC) regulates circadian locomotor output and synaptic development. Surprisingly, this study found that JET and DYSC antagonistically regulate synaptic development at the larval neuromuscular junction, and reduced JET activity rescues arrhythmicity of dysc mutants. Consistent with the prior finding that DYSC regulates SLOWPOKE (SLO) potassium channel expression, jet mutations also rescue circadian and synaptic phenotypes in slo mutants. Collectively, these data suggest that JET, DYSC, and SLO promote circadian output in part by regulating synaptic morphology.
Laiouar, S., Berns, N., Brech, A. and Riechmann, V. (2020). RabX1 Organizes a Late Endosomal Compartment that Forms Tubular Connections to Lysosomes Consistent with a "Kiss and Run" Mechanism. Curr Biol. PubMed ID: 32059769
Degradation of endocytosed proteins involves the formation of transient connections between late endosomes and lysosomes in a process called "kiss and run." Genes and proteins controlling this mechanism are unknown. This study identify the small guanosine triphosphatase (GTPase) RabX1 as an organizer of a late endosomal compartment that forms dynamic tubular connections to lysosomes. By analyzing trafficking of the adhesion protein Fasciclin2 in the Drosophila follicular epithelium, this study shows that a reduction of RabX1 function leads to defects in Fasciclin2 degradation. RabX1 mutants fail to form normal lysosomes and accumulate Fasciclin2 in a swelling late-endosomal compartment. RabX1 protein localizes to late endosomes, where it induces the formation of tubular connections to lysosomes. It is proposed that these tubules facilitate influx of lysosomal content into late endosomes and that this influx leads to the formation of endolysosomes, in which Fasciclin2 is degraded. The formation of RabX1 tubules is dependent on the V-ATPase proton pump. Moreover, evidence that V-ATPase activity is upregulated during epithelial differentiation. This upregulation intensifies RabX1 tubulation and thereby boosts the capacity of the endolysosomal pathway. Enhanced endolysosomal capacity is required for the removal of Fasciclin2 from the epithelium, which is part of a developmental program promoting epithelial morphogenesis.
Fujii, S., Kurokawa, K., Inaba, R., Hiramatsu, N., Tago, T., Nakamura, Y., Nakano, A., Satoh, T. and Satoh, A. K. (2020). Recycling endosomes attach to the trans-side of Golgi stacks in Drosophila and mammalian cells. J Cell Sci. PubMed ID: 31974113
Historically, the trans-Golgi network (TGN) has been recognized as a sorting center of newly synthesized proteins, whereas recycling endosome (RE) is a compartment where endocytosed materials transit before being recycled to the plasma membrane. However, recent findings revealed that both the TGN and RE connect endocytosis and exocytosis, and thus are functionally overlapping. This study reports, in both Drosophila and microtubule-disrupted HeLa cells, that REs are interconvertible between two distinct states, namely Golgi-associated REs and free REs. Detachment and reattachment of REs and Golgi stacks were often observed. These two types of REs were in the route of Glycosylphosphatidylinositol-anchored cargo protein released from the endoplasmic reticulum, but not in that of Vesicular stomatitis virus G protein. In plants, it has been established that there are two types of TGNs: the Golgi-associated TGN and Golgi-independent TGN. Dynamics of REs in both Drosophila and mammalian cells revealed strong similarity to plant TGNs. Together with the molecular-level similarity, these results indicate that fly/mammalian REs are equivalent organelles to TGNs in plants, and evoke reconsideration of identities and functional relationships between REs and TGNs.
Kim, Y. and Cho, K. O. (2020). POU domain motif3 (Pdm3) induces wingless (wg) transcription and is essential for development of larval neuromuscular junctions in Drosophila. Sci Rep 10(1): 517. PubMed ID: 31949274
Wnt is a conserved family of secreted proteins that play diverse roles in tissue growth and differentiation. Identification of transcription factors that regulate wnt expression is pivotal for understanding tissue-specific signaling pathways regulated by Wnt. This study identified pdm3m7, a new allele of the pdm3 gene encoding a POU family transcription factor, in a lethality-based genetic screen for modifiers of Wingless (Wg) signaling in Drosophila. Interestingly, pdm3m7 larvae showed slow locomotion, implying neuromuscular defects. Analysis of larval neuromuscular junctions (NMJs) revealed decreased bouton number with enlarged bouton in pdm3 mutants. pdm3 NMJs also had fewer branches at axon terminals than wild-type NMJs. Consistent with pdm3m7 being a candidate wg modifier, NMJ phenotypes in pdm3 mutants were similar to those of wg mutants, implying a functional link between these two genes. Indeed, lethality caused by Pdm3 overexpression in motor neurons was completely rescued by knockdown of wg, indicating that Pdm3 acts upstream to Wg. Furthermore, transient expression of Pdm3 induced ectopic expression of wg-LacZ reporter and Wg effector proteins in wing discs. It is proposed that Pdm3 expressed in presynaptic NMJ neurons regulates wg transcription for growth and development of both presynaptic neurons and postsynaptic muscles.

Monday, March 9th - Adult and larval physiology

Andersen, M. K. and Overgaard, J. (2020). Maintenance of hindgut reabsorption during cold exposure is a key adaptation for Drosophila cold tolerance. J Exp Biol. PubMed ID: 31953360
Maintaining extracellular osmotic and ionic homeostasis is crucial for organismal function. In insects, hemolymph volume and ion content is regulated by the secretory Malpighian tubules and reabsorptive hindgut. When exposed to stressful cold, homeostasis is gradually disrupted, characterized by a debilitating increase in extracellular K(+) concentration (hyperkalemia). Accordingly, studies have found a strong link between the species-specific cold tolerance and their ability to maintain ion and water homeostasis at low temperature. This is also true for drosophilids where inter- and intra-specific differences in cold tolerance are linked to the secretory capacity of Malpighian tubules. There is, however, little information on the reabsorptive capacity of the hindgut in Drosophila To address this, a novel method was developed that permits continued measurements of hindgut ion and fluid reabsorption in Drosophila. This assay is temporally stable (approximately 2 hours) and responsive to cAMP stimulation and pharmacological intervention in accordance with the current insect hindgut reabsorption model. Then, how cold acclimation or cold adaptation affected hindgut reabsorption at benign (24 degrees C) and low temperature (3 degrees C) was investigated. Cold tolerant Drosophila species and cold-acclimated D. melanogaster maintain superior fluid and Na(+) reabsorption at low temperature. Furthermore, cold adaptation and acclimation caused a relative reduction in K(+) reabsorption at low temperature. These characteristic responses of cold adaptation/ acclimation will promote maintenance of ion and water homeostasis at low temperature. This study of hindgut function therefore provides evidence to suggest that adaptations in osmoregulatory capacity of insects are critical for their ability to tolerate cold.
Cao, T., Sujkowski, A., Cobb, T., Wessells, R. J. and Jin, J. P. (2020). The glutamic acid-rich long C-terminal extension of troponin T has a critical role in insect muscle functions. J Biol Chem. PubMed ID: 32024695
The troponin complex regulates the Ca2+-activation of myofilaments during striated muscle contraction and relaxation. Troponin genes emerged 500-700 million years ago during early animal evolution. Troponin T (TnT) is the thin filament-anchoring subunit of troponin. Vertebrate and invertebrate TnTs have conserved core structures, reflecting conserved functions in regulating muscle contraction, and also contain significantly diverged structures, reflecting muscle type- and species-specific adaptations. TnT in insects contains a highly diverged structure consisting of a long glutamic acid-rich C-terminal extension of ~70 residues with unknown function. C-terminally truncated Drosophila TnT (TpnT-CD70) retains binding of tropomyosin, troponin I, and troponin C, indicating a preserved core structure of TnT. However, the mutant TpnTCD70 gene residing on the X chromosome resulted in lethality in male flies. This X-linked mutation produces dominant-negative phenotypes, including decreased flying and climbing abilities, in heterozygous female flies. Immunoblot quantification with a TpnT-specific monoclonal antibody indicated expression of TpnT-CD70 in vivo and normal stoichiometry of total TnT in myofilaments of heterozygous female flies. Light and electron microscopy examinations revealed primarily normal sarcomere structures in female heterozygous animals, whereas Z-band streaming could be observed in the jump muscle of these flies. Although TpnT-CD70-expressing flies exhibited lower resistance to cardiac stress, their hearts were significantly more tolerant to Ca2+ overloading induced by high-frequency electrical pacing. These findings suggest that the Glu-rich long C-terminal extension of insect TnT functions as a myofilament Ca2+buffer/reservoir, potentially critical to the high-frequency asynchronous contraction of flight muscles.
Cabrero, P., Terhzaz, S., Dornan, A. J., Ghimire, S., Holmes, H. L., Turin, D. R., Romero, M. F., Davies, S. A. and Dow, J. A. T. (2020). Specialized stellate cells offer a privileged route for rapid water flux in Drosophila renal tubule. Proc Natl Acad Sci U S A 117(3): 1779-1787. PubMed ID: 31907321
Insects are highly successful, in part through an excellent ability to osmoregulate. The renal (Malpighian) tubules can secrete fluid faster on a per-cell basis than any other epithelium, but the route for these remarkable water fluxes has not been established. In Drosophila melanogaster, four genes of the major intrinsic protein family are expressed at a very high level in the fly renal tissue: the aquaporins (AQPs) Drip and Prip and the aquaglyceroporins Eglp2 and Eglp4 As predicted from their structure, and by their transport function by expressing these proteins in Xenopus oocytes, Drip, Prip, and Eglp2 show significant and specific water permeability, whereas Eglp2 and Eglp4 show very high permeability to glycerol and urea. Knockdowns of any of these genes result in impaired hormone-induced fluid secretion. The Drosophila tubule has 2 main secretory cell types: active cation-transporting principal cells, wherein the aquaglyceroporins localize to opposite plasma membranes, and small stellate cells, the site of the chloride shunt conductance, with these AQPs localizing to opposite plasma membranes. This suggests a model in which osmotically obliged water flows through the stellate cells. Consistent with this model, fluorescently labeled dextran, an in vivo marker of membrane water permeability, is trapped in the basal infoldings of the stellate cells after kinin diuretic peptide stimulation, confirming that these cells provide the major route for transepithelial water flux. The spatial segregation of these components of epithelial water transport may help to explain the unique success of the higher insects in regulating their internal environments.
Gutorov, R., Peters, M., Katz, B., Brandwine, T., Barbera, N. A., Levitan, I. and Minke, B. (2019). Modulation of Transient receptor potential C channel activity by cholesterol. Front Pharmacol 10: 1487. PubMed ID: 31920669
Changes of cholesterol level in the plasma membrane of cells have been shown to modulate ion channel function. The proposed mechanisms underlying these modulations include association of cholesterol to a single binding site at a single channel conformation, association to a highly flexible cholesterol binding site adopting multiple poses, and perturbation of lipid rafts. These perturbations have been shown to induce reversible targeting of mammalian transient receptor potential C (TRPC) channels to the cholesterol-rich membrane environment of lipid rafts. Thus, the observed inhibition of TRPC channels by methyl-beta-cyclodextrin (MbetaCD), which induces cholesterol efflux from the plasma membrane, may result from disruption of lipid rafts. This perturbation was also shown to disrupt multimolecular signaling complexes containing TRPC channels. The Drosophila TRP and TRP-like (TRPL) channels belong to the TRPC channel subfamily. When the Drosophila TRPL channel was expressed in S2 or HEK293 cells and perfused with MbetaCD, the TRPL current was abolished in less than 100 s, fitting well the fast kinetic phase of cholesterol sequestration experiments in cells. It was thus suggested that the fast kinetics of TRPL channel suppression by MbetaCD arise from disruption of lipid rafts. Accordingly, lipid raft perturbation by cholesterol sequestration could give clues to the function of lipid environment in TRPC channel activity and its mechanism.
Kierdorf, K., Hersperger, F., Sharrock, J., Vincent, C. M., Ustaoglu, P., Dou, J., Gyoergy, A., Gross, O., Siekhaus, D. E. and Dionne, M. S. (2020). Muscle function and homeostasis require cytokine inhibition of AKT activity in Drosophila. Elife 9. PubMed ID: 31944178
Unpaired ligands are secreted signals that act via a GP130-like receptor, Domeless, to activate JAK/STAT signalling in Drosophila. Like many mammalian cytokines, unpaireds can be activated by infection and other stresses and can promote insulin resistance in target tissues. However, the importance of this effect in non-inflammatory physiology is unknown. This study identified a requirement for unpaired-JAK signalling as a metabolic regulator in healthy adult Drosophila muscle. Adult muscles show basal JAK-STAT signalling activity in the absence of any immune challenge. Plasmatocytes (Drosophila macrophages) are an important source of this tonic signal. Loss of the dome receptor on adult muscles significantly reduces lifespan and causes local and systemic metabolic pathology. These pathologies result from hyperactivation of AKT and consequent deregulation of metabolism. Thus, this study identify a cytokine signal that must be received in muscle to control AKT activity and metabolic homeostasis.
Guo, R. and Reinhardt, K. (2020). Dietary polyunsaturated fatty acids affect volume and metabolism of Drosophila melanogaster sperm. J Evol Biol. PubMed ID: 31961473
Dietary fatty acids can accumulate in sperm and affect their function in vertebrates. There is currently no clear prediction as to how dietary fatty acids affect the sperm of D. melanogaster. This study manipulated the amount and identity of dietary polyunsaturated fatty acids (PUFA) in the food of D. melanogaster males (a treatment known to affect membrane fluidity) and measured changes in sperm parameters. The following foundings were made: (a) males reared on food containing PUFA-rich, plant-derived lipids showed a slower increase in sperm volume over male age compared to males reared on yeast-derived lipid food which is richer in saturated fatty acids. (b) The resistance of sperm membrane integrity to osmotic stress was not altered by dietary lipid treatment, but (c) food containing yeast-derived lipids induced a 46% higher in situ rate of production of reactive oxygen species in sperm cells. These findings show that dietary lipids have similar effects on sperm parameters in Drosophila as in vertebrates, affect some, but not all, sperm parameters and modulate male reproductive ageing. In concert with recent findings of sex-specific seasonal variation of diet choice in the wild, these results suggest a substantial dietary impact on the dynamics of male reproduction in the wild.

Friday, March 6th - Neural Development

Hernandez, E., MacNamee, S. E., Kaplan, L. R., Lance, K., Garcia-Verdugo, H. D., Farhadi, D. S., Deer, C., Lee, S. W. and Oland, L. A. (2020). The astrocyte network in the ventral nerve cord neuropil of the Drosophila third-instar larva. J Comp Neurol. PubMed ID: 31909826
Understanding neuronal function at the local and circuit level requires understanding astrocyte function. This study provides a detailed analysis of astrocyte morphology and territory in the Drosophila third-instar ventral nerve cord where there already exists considerable understanding of the neuronal network. Astrocyte shape varies more than previously reported; many have bilaterally symmetrical partners, many have a high percentage of their arborization in adjacent segments, and many have branches that follow structural features. Taken together, these data are consistent with, but not fully explained by, a model of a developmental growth process dominated by competitive or repulsive interactions between astrocytes. The data suggest that the model should also include cell-autonomous aspects, as well as the use of structural features for growth. Variation in location of arborization territory for identified astrocytes was great enough that a standardized scheme of neuropil division among the six astrocytes that populate each hemi-segment is not possible at the third instar. The arborizations of the astrocytes can extend across neuronal functional domains. The ventral astrocyte in particular, whose territory can extend well into the proprioceptive region of the neuropil, has no obvious branching pattern that correlates with domains of particular sensory modalities, suggesting that the astrocyte would respond to neuronal activity in any of the sensory modalities, perhaps integrating across them. This study sets the stage for future studies that will generate a robust, functionally oriented connectome that includes both partners in neuronal circuits-the neurons and the glial cells, providing the foundation necessary for studies to elucidate neuron-glia interactions in this neuropil.
Hu, C., Kanellopoulos, A. K., Richter, M., Petersen, M., Konietzny, A., Tenedini, F., Hoyer, N., Cheng, L., Poon, C. L. C., Harvey, K. F., Windhorst, S., Parrish, J. Z., Mikhaylova, M., Bagni, C., Calderon de Anda, F. and Soba, P. (2020). Conserved Tao kinase activity regulates dendritic arborization, cytoskeletal dynamics and sensory function in Drosophila. J Neurosci. PubMed ID: 31964717
Dendritic arborization is highly regulated and requires tight control of dendritic growth, branching, cytoskeletal dynamics and ion channel expression to ensure proper function. Abnormal dendritic development can result in altered network connectivity, which has been linked to neurodevelopmental disorders including Autism Spectrum Disorders (ASDs). How neuronal growth control programs tune dendritic arborization to ensure function is still not fully understood. Using Drosophila dendritic arborization (da) neurons as a model, this study identified the conserved Ste20-like kinase Tao as a negative regulator of dendritic arborization. Tao kinase activity regulates cytoskeletal dynamics and sensory channel localization required for proper sensory function in both, male and female flies. Evidence is provided for functional conservation of Tao kinase showing that its ASD-linked human orthologue, Tao kinase 2 (Taok2), could replace Drosophila Tao and rescue dendritic branching, dynamic microtubule alterations and behavioral defects. However, several ASD-linked Taok2 variants displayed impaired rescue activity suggesting Tao/Taok2 mutations can disrupt sensory neuron development and function. Consistently, it was shown that Tao kinase activity is required in developing and as well as adult stages for maintaining normal dendritic arborization and sensory function to regulate escape and social behavior. These data suggest an important role for Tao kinase signaling in cytoskeletal organization to maintain proper dendritic arborization and sensory function, providing a strong link between developmental sensory aberrations and behavioral abnormalities relevant for Taok2-dependent ASDs.
Ishikawa, Y., Fujiwara, M., Wong, J., Ura, A. and Kamikouchi, A. (2019). Stereotyped combination of hearing and wind/gravity-sensing neurons in the Johnston's organ of Drosophila. Front Physiol 10: 1552. PubMed ID: 31969834
The antennal ear of the fruit fly, called the Johnston's organ (JO), detects a wide variety of mechanosensory stimuli, including sound, wind, and gravity. Like many sensory cells in insect, JO neurons are compartmentalized in a sensory unit (i.e., scolopidium). To understand how different subgroups of JO neurons are organized in each scolopidial compartment, this study visualized individual JO neurons by labeling various subgroups of JO neurons in different combinations. It was found that vibration-sensitive (or deflection-sensitive) neurons rarely grouped together in a single scolopidial compartment. This finding suggests that JO neurons are grouped in stereotypical combinations each with a distinct response property in a scolopidium.
Kato, K., Orihara-Ono, M. and Awasaki, T. (2020). Multiple lineages enable robust development of the neuropil-glia architecture in adult Drosophila. Development. PubMed ID: 32051172
Neural remodeling is essential for the development of a functional nervous system and has been extensively studied in the metamorphosis of Drosophila Despite the crucial roles of glial cells in brain functions, including learning and behavior, little is known of how adult glial cells develop in the context of neural remodeling. This study shows that the architecture of neuropil-glia in the adult Drosophila brain, composed of astrocyte-like glia (ALG) and ensheathing glia (EG), robustly develops from two different populations in the larva: the larval EG and glial cell missing-positive (gcm +) cells. While larval EG dedifferentiate, proliferate, and redifferentiate into adult ALG and EG, gcm + cells also generate the same glial subtypes. Each glial lineage occupies a certain brain area complementary to the other, and together they form the adult neuropil-glia architecture. Both lineages require the FGF receptor Heartless to proliferate, and the homeoprotein Prospero to differentiate into ALG. Lineage-specific inhibition of gliogenesis revealed that each lineage compensates for deficiency in the proliferation of the other. Together, the lineages ensure the robust development of adult neuropil-glia, thereby ensuring a functional brain.
Inami, S., Sato, S., Kondo, S., Tanimoto, H., Kitamoto, T. and Sakai, T. (2020). Environmental light is required for maintenance of long-term memory in Drosophila. J Neurosci 40(7): 1427-1439. PubMed ID: 31932417
Long-term memory (LTM) is stored as functional modifications of relevant neural circuits in the brain. A large body of evidence indicates that the initial establishment of such modifications through the process known as memory consolidation requires learning-dependent transcriptional activation and de novo protein synthesis. However, it remains poorly understood how the consolidated memory is maintained for a long period in the brain, despite constant turnover of molecular substrates. Using the Drosophila courtship conditioning assay of adult males as a memory paradigm, this study shows that in Drosophila, environmental light plays a critical role in LTM maintenance. LTM is impaired when flies are kept in constant darkness (DD) during the memory maintenance phase. Because light activates the brain neurons expressing the neuropeptide pigment-dispersing factor (Pdf), the possible involvement of Pdf neurons in LTM maintenance was examined. Temporal activation of Pdf neurons compensated for the DD-dependent LTM impairment, whereas temporal knockdown of Pdf during the memory maintenance phase impaired LTM in light/dark cycles. Furthermore, it was demonstrated that the transcription factor cAMP response element-binding protein (CREB) is required in the memory center, namely, the mushroom bodies (MBs), for LTM maintenance, and Pdf signaling regulates light-dependent transcription via CREB. These results demonstrate for the first time that universally available environmental light plays a critical role in LTM maintenance by activating the evolutionarily conserved memory modulator CREB in MBs via the Pdf signaling pathway.
Karunanithi, S., Cylinder, D., Ertekin, D., Zalucki, O. H., Marin, L., Lavidis, N. A., Atwood, H. L. and van Swinderen, B. (2020). Proportional downscaling of glutamatergic release sites by the general anesthetic propofol at Drosophila motor nerve terminals. eNeuro. PubMed ID: 32019872
Propofol is the most common general anesthetic used for surgery in humans, yet its complete mechanism of action remains elusive. In addition to potentiating inhibitory synapses in the brain, propofol also impairs excitatory neurotransmission. This study used electrophysiological recordings from individual glutamatergic boutons in male and female larval Drosophila melanogaster motor nerve terminals to characterize this effect. Recording was performed from two bouton types, which have distinct presynaptic physiology and different average numbers of release sites or active zones. A clinically relevant dose of propofol (3muM) impairs neurotransmitter release similarly at both bouton types, by decreasing the number of active release sites by half, without affecting release probability. In contrast, an analog of propofol has no effect on glutamate release. Co-expressing a truncated syntaxin1A protein in presynaptic boutons completely blocked this effect of propofol. Overexpressing wild-type syntaxin1A in boutons also conferred a level of resistance, by increasing the number of active release sites to a physiological ceiling set by the number of active zones or T-bars, and in this way counteracting the effect of propofol. These results point to the presynaptic release machinery as a target for the general anesthetic. Proportionally equivalent effects of propofol on the number of active release sites across the different bouton types suggests that smaller glutamatergic boutons with fewer release sites may be more vulnerable to the presynaptic effects of the drug.

Thursday, March 5th - Evolution

Kanippayoor, R. L., Alpern, J. H. M. and Moehring, A. J. (2020). A common suite of cellular abnormalities and spermatogenetic errors in sterile hybrid males in Drosophila. Proc Biol Sci 287(1919): 20192291. PubMed ID: 31964309
When two species interbreed, the resulting hybrid offspring are often sterile, with the heterogametic (e.g. XY) hybrid usually being more severely affected. The prevailing theory for this pattern of sterility evokes divergent changes in separate lineages having maladaptive interactions when placed together in a hybrid individual, with recessive factors on the sex chromosome interacting with dominant factors on the autosomes. The effect of these interactions on gametogenesis should not be uniform across species pairs unless genetic divergence follows the same paths in different lineages or if a specific stage of gametogenesis is more susceptible to detrimental genetic interactions. A detailed cellular characterization of hybrid male sterility was performed across three recently diverged species pairs of Drosophila. Across all three pairs, sterile hybrid sperm are alive but exhibit rapid nuclear de-condensation with age, with active, but non-differentiated, mitochondria. Surprisingly, all three sets of interspecies hybrids produce half of the number of sperm per round of spermatogenesis, with each sperm cell containing two tails. Non-disjunction failures during meiosis I were identified as the likely cause. Thus, errors during meiosis I may be a general phenomenon underlying Drosophila male sterility, indicating either a heightened sensitivity of this spermatogenic stage to failure, or a basis to sterility other than the prevailing model.
Nagarajan-Radha, V., Aitkenhead, I., Clancy, D. J., Chown, S. L. and Dowling, D. K. (2020). Sex-specific effects of mitochondrial haplotype on metabolic rate in Drosophila melanogaster support predictions of the Mother's Curse hypothesis. Philos Trans R Soc Lond B Biol Sci 375(1790): 20190178. PubMed ID: 31787038
Evolutionary theory proposes that maternal inheritance of mitochondria will facilitate the accumulation of mitochondrial DNA (mtDNA) mutations that are harmful to males but benign or beneficial to females. It is predicted that the genetic variation which delineates distinct mtDNA haplotypes of a given species should confer larger phenotypic effects on males than females (reflecting mtDNA mutations that are male-harming, but female-benign), or sexually antagonistic effects (reflecting mutations that are male-harming, but female-benefitting). This study explored whether similar signatures of male-bias or sexual antagonism extend to a key physiological trait-metabolic rate. The effects of mitochondrial haplotypes on the amount of carbon dioxide produced by individual flies, controlling for mass and activity, was measured across 13 strains of D. melanogaster that differed only in their mtDNA haplotype. The effects of mtDNA haplotype on metabolic rate were larger in males than females. Furthermore, a negative intersexual correlation was observed across the haplotypes for metabolic rate. Finally, a male-specific negative correlation, across haplotypes, was uncovered between metabolic rate and longevity. These results are consistent with the hypothesis that maternal mitochondrial inheritance has led to the accumulation of a sex-specific genetic load within the mitochondrial genome, which affects metabolic rate and that may have consequences for the evolution of sex differences in life history.
Machado, H. E., Lawrie, D. S. and Petrov, D. A. (2019). Pervasive strong selection at the level of codon usage bias in Drosophila melanogaster. Genetics. PubMed ID: 31871131
Codon usage bias (CUB), where certain codons are used more frequently than expected by chance, is a ubiquitous phenomenon and occurs across the tree of life. The dominant paradigm is that the proportion of preferred codons is set by weak selection. While experimental changes in codon usage have at times shown large phenotypic effects in contrast to this paradigm, genome-wide population genetic estimates have supported the weak selection model. This study used deep genomic population sequencing of two Drosophila melanogaster populations to measure selection on synonymous sites in a way that allowed estimation of the prevalence of both weak and strong purifying selection. Selection in favor of preferred codons was fpimd tp range from weak (|Nes| approximately 1) to strong (|Nes| > 10), with strong selection acting on 10-20% of synonymous sites in preferred codons. While previous studies indicated that selection at synonymous sites could be strong, this is the first study to detect and quantify strong selection specifically at the level of CUB. Further, it was found that CUB-associated polymorphism accounts for the majority of strong selection on synonymous sites, with secondary contributions of splicing (selection on alternatively spliced genes, splice junctions and spliceosome-bound sites) and transcription factor binding. These findings support a new model of CUB and indicate that the functional importance of CUB, as well as synonymous sites in general, have been underestimated.
Gu, L., Reilly, P. F., Lewis, J. J., Reed, R. D., Andolfatto, P. and Walters, J. R. (2019). Dichotomy of Dosage Compensation along the Neo Z Chromosome of the Monarch Butterfly. . Curr Biol 29(23): 4071-4077. PubMed ID: 31735674
Mechanisms of sex chromosome dosage compensation (SCDC) differ strikingly among animals. In Drosophila flies, chromosome-wide transcription is doubled from the single X chromosome in hemizygous (XY) males, whereas in Caenorhabditis nematodes, expression is halved for both X copies in homozygous (XX) females. Unlike other female-heterogametic (WZ female and ZZ male) animals, moths and butterflies exhibit sex chromosome dosage compensation patterns typically seen only in male-heterogametic species. The monarch butterfly carries a newly derived Z chromosome segment that arose from an autosomal fusion with the ancestral Z. Using a highly contiguous genome assembly, this study shows that gene expression is balanced between sexes along the entire Z chromosome but with distinct modes of compensation on the two segments. On the ancestral Z segment, depletion of H4K16ac corresponds to nearly halving of biallelic transcription in males, a pattern convergent to nematodes. Conversely, the newly derived Z segment shows a Drosophila-like mode of compensation, with enriched H4K16ac levels corresponding to doubled monoallelic transcription in females. This work reveals that, contrary to the expectation of co-opting regulatory mechanisms readily in place, the evolution of plural modes of dosage compensation is also possible along a single sex chromosome within a species.
Matute, D. R., Comeault, A. A., Earley, E., Serrato-Capuchina, A., Peede, D., Monroy-Eklund, A., Huang, W., Jones, C. D., Mackay, T. F. C. and Coyne, J. A. (2019). Rapid and predictable evolution of admixed populations between two Drosophila species pairs. Genetics. PubMed ID: 31767631
The consequences of hybridization are varied, ranging from the origin of new lineages, introgression of some genes between species, to the extinction of one of the hybridizing species. This study generated replicate admixed populations between two pairs of sister species of Drosophila: D. simulans and D. mauritiana; and D. yakuba and D. santomea. Each pair consisted of a continental species and an island endemic. The admixed populations were maintained by random mating in discrete generations for over 20 generations. Morphological, behavioral, and fitness-related traits from each replicate population were assessed periodically, and genomic DNA was sequenced from the populations at generation 20. For both pairs of species, species-specific traits and their genomes regressed to those of the continental species. A few alleles from the island species persisted, but they tended to be proportionally rare among all sites in the genome and were rarely fixed within the populations. This paucity of alleles from the island species was particularly pronounced on the X-chromosome. These results indicate that nearly all foreign genes were quickly eliminated after hybridization and that selection against the minor species genome might be similar across experimental replicates.
Luo, Y., Zhang, Y., Farine, J. P., Ferveur, J. F., Ramirez, S. and Kopp, A. (2019). Evolution of sexually dimorphic pheromone profiles coincides with increased number of male-specific chemosensory organs in Drosophila prolongata. Ecol Evol 9(23): 13608-13618. PubMed ID: 31871670
Binary communication systems that involve sex-specific signaling and sex-specific signal perception play a key role in sexual selection and in the evolution of sexually dimorphic traits. The driving forces and genetic changes underlying such traits can be investigated in systems where sex-specific signaling and perception have emerged recently and show evidence of potential coevolution. A promising model is found in Drosophila prolongata, which exhibits a species-specific increase in the number of male chemosensory bristles. This transition is shown to coincides with recent evolutionary changes in cuticular hydrocarbon (CHC) profiles. Long-chain CHCs that are sexually monomorphic in the closest relatives of D. prolongata (D. rhopaloa, D. carrolli, D. kurseongensis, and D. fuyamai) are strongly male-biased in this species. An intraspecific female-limited polymorphism, where some females have male-like CHC profiles, was also identifid. Both the origin of sexually dimorphic CHC profiles and the female-limited polymorphism in D. prolongata involve changes in the relative amounts of three mono-alkene homologs, 9-tricosene, 9-pentacosene, and 9-heptacosene, all of which share a common biosynthetic origin and point to a potentially simple genetic change underlying these traits. These results suggest that pheromone synthesis may have coevolved with chemosensory perception and open the way for reconstructing the origin of sexual dimorphism in this communication system.

Wednesday, March 4th - Enhancers and Gene Regulaton

Nakamura, S., Hira, S., Fujiwara, M., Miyagata, N., Tsuji, T., Kondo, A., Kimura, H., Shinozuka, Y., Hayashi, M., Kobayashi, S. and Mukai, M. (2019). A truncated form of a transcription factor Mamo activates vasa in Drosophila embryos. Commun Biol 2: 422. PubMed ID: 31799425
Expression of the vasa gene is associated with germline establishment. Therefore, identification of vasa activator(s) should provide insights into germline development. However, the genes sufficient for vasa activation remain unknown. Previous work showed that the BTB/POZ-Zn-finger protein Mamo is necessary for vasa expression in Drosophila. This study showed that the truncated Mamo lacking the BTB/POZ domain (MamoAF) is a potent vasa activator. Overexpression of MamoAF was sufficient to induce vasa expression in both primordial germ cells and brain. Indeed, Mamo mRNA encoding a truncated Mamo isoform, which is similar to MamoAF, was predominantly expressed in primordial germ cells. The results of these genetic and biochemical studies showed that MamoAF, together with CBP, epigenetically activates vasa expression. Furthermore, MamoAF and the germline transcriptional activator OvoB exhibited synergy in activating vasa transcription. It is proposed that a Mamo-mediated network of epigenetic and transcriptional regulators activates vasa expression.
Hong, Y. G., Kang, B., Lee, S., Lee, Y., Ju, B. G. and Jeong, S. (2020). Identification of cis-regulatory region controlling Semaphorin-1a expression in the Drosophila embryonic nervous system. Mol Cells. PubMed ID: 32024353
The Drosophila transmembrane semaphorin Sema-1a mediates forward and reverse signaling that plays an essential role in motor and central nervous system (CNS) axon pathfinding during embryonic neural development. Previous immunohistochemical analysis revealed that Sema-1a is expressed on most commissural and longitudinal axons in the CNS and five motor nerve branches in the peripheral nervous system (PNS). This study uncovered three cis -regulatory elements (CREs), R34A03, R32H10, and R33F06, that robustly drove reporter expression in a large subset of neurons in the CNS. In the transgenic lines R34A03 and R32H10 reporter expression was consistently observed on both ISNb and SNa nerve branches, whereas in the line R33F06 reporter expression was irregularly detected on ISNb or SNa nerve branches in small subsets of abdominal hemisegments. Through complementation test with a Sema1a loss-of-function allele, it was found that neuronal expression of Sema-1a driven by each of R34A03 and R32H10 restores robustly the CNS and PNS motor axon guidance defects observed in Sema-1a homozygous mutants. The results suggest that in a redundant manner, the CREs, R34A03, R32H10, and R33F06 govern the Sema-1a expression required for the axon guidance function of Sema-1a during embryonic neural development.
Bandodkar, P. U., Al Asafen, H. and Reeves, G. T. (2020). Spatiotemporal control of gene expression boundaries using a feedforward loop. Dev Dyn. PubMed ID: 31925874
A feedforward loop (FFL) is commonly observed in several biological networks. The FFL network motif has been mostly studied with respect to variation of the input signal in time, with only a few studies of FFL activity in a spatially distributed system such as morphogen-mediated tissue patterning. However, most morphogen gradients also evolve in time. The spatiotemporal behavior of a coherent FFL in two contexts was studied: (a) a generic, oscillating morphogen gradient and (b) the dorsal-ventral patterning of the early Drosophila embryo by a gradient of the NF-kappaB homolog Dorsal with its early target Twist. In both models, features in the dynamics of the intermediate node-phase difference and noise filtering-that were found to be largely independent of the parameterization of the models, and thus were functions of the structure of the FFL itself. In the Dorsal gradient model, it was also found that proper target gene expression was not possible without including the effect of maternal pioneer factor Zelda. CONCLUSIONS: An FFL buffers fluctuation to changes in the morphogen signal ensuring stable gene expression boundaries.
Blanco, J., Cooper, J. C. and Baker, N. E. (2020). Roles of C/EBP class bZip proteins in the growth and cell competition of Rp ('Minute') mutants in Drosophila. Elife 9. PubMed ID: 31909714
Reduced copy number of ribosomal protein (Rp) genes adversely affects both flies and mammals. Xrp1 encodes a reportedly Drosophila-specific AT-hook, bZIP protein responsible for many of the effects including the elimination of Rp mutant cells by competition with wild type cells. Irbp18, an evolutionarily conserved bZIP gene, heterodimerizes with Xrp1 and with another bZip protein, dATF4. This study shows that Irbp18 is required for the effects of Xrp1, whereas dATF4 does not share the same phenotype, indicating that Xrp1/Irbp18 is the complex active in Rp mutant cells, independently of other complexes that share Irbp18. Xrp1 and Irbp18 transcripts and proteins are upregulated in Rp mutant cells by auto-regulatory expression that depends on the Xrp1 DNA binding domains and is necessary for cell competition. Xrp1 is conserved beyond Drosophila, although under positive selection for rapid evolution, and that at least one human bZip protein can similarly affect Drosophila development.
Castellanos, M., Mothi, N. and Munoz, V. (2020). Eukaryotic transcription factors can track and control their target genes using DNA antennas. Nat Commun 11(1): 540. PubMed ID: 31992709
Eukaryotic transcription factors (TF) function by binding to short 6-10 bp DNA recognition sites located near their target genes, which are scattered through vast genomes. Such process surmounts enormous specificity, efficiency and celerity challenges using a molecular mechanism that remains poorly understood. Combining biophysical experiments, theory and bioinformatics, this study dissected the interplay between the DNA-binding domain of Engrailed, a Drosophila TF, and the regulatory regions of its target genes. Engrailed binding affinity is strongly amplified by the DNA regions flanking the recognition site, which contain long tracts of degenerate recognition-site repeats. Such DNA organization operates as an antenna that attracts TF molecules in a promiscuous exchange among myriads of intermediate affinity binding sites. The antenna ensures a local TF supply, enables gene tracking and fine control of the target site's basal occupancy. This mechanism illuminates puzzling gene expression data and suggests novel engineering strategies to control gene expression.
Sajwan, S. and Mannervik, M. (2019). Gene activation by dCas9-CBP and the SAM system differ in target preference. Sci Rep 9(1): 18104. PubMed ID: 31792240
Gene overexpression through the targeting of transcription activation domains to regulatory DNA via catalytically defective Cas9 (dCas9) represents a powerful approach to investigate gene function as well as the mechanisms of gene control. To date, the most efficient dCas9-based activator is the Synergistic Activation Mediator (SAM) system whereby transcription activation domains are directly fused to dCas9 as well as tethered through MS2 loops engineered into the gRNA. This study shows that dCas9 fused to the catalytic domain of the histone acetyltransferase CBP is a more potent activator than the SAM system at some loci, but less efficient at other locations in Drosophila cells. These results suggest that different rate-limiting steps in the transcription cycle are affected by dCas9-CBP and the SAM system, and that comparing these activators may be useful for mechanistic studies of transcription as well as for increasing the number of hits in genome-wide overexpression screens.

Wednesday, March 3rd - Behavior

Brown, E., Layne, J. E., Elchert, A. R. and Rollmann, S. M. (2020). Behavioral and Transcriptional Response to Selection for Olfactory Behavior in Drosophila. G3 (Bethesda). PubMed ID: 32024668
The detection, discrimination, and behavioral responses to chemical cues in the environment can have marked effects on organismal survival and reproduction, eliciting attractive or aversive behavior. To gain insight into mechanisms mediating this hedonic valence, this study applied thirty generations of divergent artificial selection for Drosophila melanogaster olfactory behavior. Positive and negative behavioral responses were independently selected for two ecologically relevant chemical compounds: 2,3-butanedione and cyclohexanone. The correlated responses were also subjected to selection by testing behavioral responses to other odorants and life history traits. Measurements of behavioral responses of the selected lines and unselected controls to additional odorants showed that the mechanisms underlying responses to these odorants are, in some cases, differentially affected by selection regime and generalization of the response to other odorants was only detected in the 2,3-butanedione selection lines. Food consumption and lifespan varied with selection regime and, at times, sex. An analysis of gene expression of both selection regimes identified multiple differentially expressed genes. New genes and genes previously identified in mediating olfactory behavior were identified. In particular, functional enrichment was found of several gene ontology terms, including cell-cell adhesion and sulfur compound metabolic process, the latter including genes belonging to the glutathione S-transferase family. These findings highlight a potential role for glutathione S-transferases in the evolution of hedonic valence to ecologically relevant volatile compounds and set the stage for a detailed investigation into mechanisms by which these genes mediate attraction and aversion.
Brunner, B., Saumweber, J., Samur, M., Weber, D., Schumann, I., Mahishi, D., Rohwedder, A. and Thum, A. S. (2020). Food restriction reconfigures naive and learned choice behavior in Drosophila larvae. J Neurogenet: 1-10. PubMed ID: 31975653
In many animals, the establishment and expression of food-related memory is limited by the presence of food and promoted by its absence, implying that this behavior is driven by motivation. In the past, this has already been demonstrated in various insects including honeybees and adult Drosophila. For Drosophila larvae, which are characterized by an immense growth and the resulting need for constant food intake, however, knowledge is rather limited. Accordingly, this study has analyzed whether starvation modulates larval memory formation or expression after appetitive classical olfactory conditioning, in which an odor is associated with a sugar reward. Odor-sugar memory of starved larvae was shown to last longer than in fed larvae, although the initial performance is comparable. 80 minutes after odor fructose conditioning, only starved but not fed larvae show a reliable odor-fructose memory. This is likely due to a specific increase in the stability of anesthesia-resistant memory (ARM). Furthermore, it was observe that starved larvae, in contrast to fed ones, prefer sugars that offer a nutritional benefit in addition to their sweetness. Taken together this work shows that Drosophila larvae adjust the expression of learned and naive choice behaviors in the absence of food. These effects are only short-lasting probably due to their lifestyle and their higher internal motivation to feed. In the future, the extensive use of established genetic tools will allow identification of development-specific differences arising at the neuronal and molecular level.
Filice, D. C. S., Bhargava, R. and Dukas, R. (2020). Plasticity in male mating behavior modulates female life history in fruit flies. Evolution 74(2): 365-376. PubMed ID: 31925958
In many species, intense male-male competition for the opportunity to sire offspring has led to the evolution of selfish reproductive traits that are harmful to the females they mate with. In the fruit fly, Drosophila melanogaster, males modulate their reproductive behavior based on the perceived intensity of competition in their premating environment. Specifically, males housed with other males subsequently transfer a larger ejaculate during a longer mating compared to males housed alone. Although the potential fitness benefits to males from such plasticity are clear, its effects on females are mostly unknown. Hence, this study tested the long-term consequences to females from mating with males with distinct social experiences. First, it was verified that competitive experience influences male mating behavior and it was found that males housed with rivals subsequently have shorter mating latencies and longer mating durations. Then, females were exposed every other day for 20 days to males that were either housed alone or with rivals, and subsequently their fitness was measured. Females mated to males housed with rivals were found to produce more offspring early in life but fewer offspring later in life and have shorter lifespans but similar intrinsic population growth rates. These results indicate that plasticity in male mating behavior can influence female life histories by altering females' relative allocation to early versus late investment in reproduction and survival.
He, J., Hommen, F., Lauer, N., Balmert, S. and Scholz, H. (2020). Serotonin transporter dependent modulation of food-seeking behavior. PLoS One 15(1): e0227554. PubMed ID: 31978073
The olfactory pathway integrates the odor information required to generate correct behavioral responses. To address how changes of serotonin signaling in two contralaterally projecting, serotonin-immunoreactive deutocerebral neurons impacts key odorant attraction in Drosophila melanogaster, this study selectively altered serotonin signaling using the serotonin transporter with mutated serotonin binding sites in these neurons, and the consequence on odorant-guided food seeking was analyzed. The expression of the mutated serotonin transporter selectively changed the odorant attraction in an odorant-specific manner. The shift in attraction was not influenced by more up-stream serotonergic mechanisms mediating behavioral inhibition. The expression of the mutated serotonin transporter in CSD neurons did not influence other behaviors associated with food seeking such as olfactory learning and memory or food consumption. Evidence is provided that the change in the attraction by serotonin transporter function might be achieved by increased serotonin signaling and by different serotonin receptors. The 5-HT1B receptor positively regulated the attraction to low and negatively regulated the attraction to high concentrations of acetic acid. In contrast, 5-HT1A and 5-HT2A receptors negatively regulated the attraction in projection neurons to high acetic acid concentrations. These results provide insights into how serotonin signaling in two serotonergic neurons selectively regulates the behavioral response to key odorants during food seeking.
Rihani, K., Fraichard, S., Chauvel, I., Poirier, N., Delompre, T., Neiers, F., Tanimura, T., Ferveur, J. F. and Briand, L. (2019). A conserved odorant binding protein is required for essential amino acid detection in Drosophila. Commun Biol 2: 425. PubMed ID: 31799428
Animals need to detect in the food essential amino acids that they cannot synthesize. This study found that the odorant binding protein OBP19b, which is highly expressed in Drosophila melanogaster taste sensilla, is necessary for the detection of several amino acids including the essential l-phenylalanine. The recombinant OBP19b protein was produced and characterized for its binding properties: it stereoselectively binds to several amino acids. Using a feeding-choice assay, it was found that OBP19b is necessary for detecting l-phenylalanine and l-glutamine, but not l-alanine or D-phenylalanine. The cells expressing OBP19b were mapped, and the electrophysiological responses of a single taste sensillum were compared to several amino acids: OBP19b mutant flies showed a reduced response compared to control flies when tested to preferred amino acids, but not to the other ones. OBP19b is well conserved in phylogenetically distant species suggesting that this protein is necessary for detection of specific amino acids in insects.
Heys, C., Lize, A., Lewis, Z. and Price, T. A. R. (2020). Drosophila sexual attractiveness in older males is mediated by their microbiota. Microorganisms 8(2). PubMed ID: 31991698
Age is well known to be a basis for female preference of males. However, the mechanisms underlying age-based choices are not well understood, with several competing theories and little consensus. The idea that the microbiota can affect host mate choice is gaining traction, and in this study it was examine whether the male microbiota influences female preference for older individuals in the fruit fly Drosophila pseudoobscura. An intact microbiota was found to be a key component of attractiveness in older males. However, no evidence was found that this decrease in older male attractiveness was simply due to impaired microbiota generally reducing male quality. Instead, it is suggested that the microbiota underlies an honest signal used by females to assess male age, and that impaired microbiota disrupt this signal. This suggests that age-based preferences may break down in environments where the microbiota is impaired, for example when individuals are exposed to naturally occurring antibiotics, extreme temperatures, or in animals reared in laboratories on antibiotic supplemented diet.

Monday, March 2nd - Adult neural function

Heath, S. L., Christenson, M. P., Oriol, E., Saavedra-Weisenhaus, M., Kohn, J. R. and Behnia, R. (2020). Circuit mechanisms underlying chromatic encoding in Drosophila photoreceptors. Curr Biol 30(2): 264-275. PubMed ID: 31928878
Spectral information is commonly processed in the brain through generation of antagonistic responses to different wavelengths. In many species, these color opponent signals arise as early as photoreceptor terminals.This study measured the spectral tuning of photoreceptors in Drosophila. In addition to a previously described pathway comparing wavelengths at each point in space, A horizontal-cell-mediated pathway similar to that was found found in mammals. This pathway enables additional spectral comparisons through lateral inhibition, expanding the range of chromatic encoding in the fly. Together, these two pathways enable efficient decorrelation and dimensionality reduction of photoreceptor signals while retaining maximal chromatic information. A biologically constrained model accounts for the findings and predicts a spatio-chromatic receptive field for fly photoreceptor outputs, with a color opponent center and broadband surround. This dual mechanism combines motifs of both an insect-specific visual circuit and an evolutionarily convergent circuit architecture, endowing flies with the ability to extract chromatic information at distinct spatial resolutions.
Del Valle Rodriguez, A., Cera, M. and Portillo, J. R. (2020). A network approach to analyze neuronal lineage and layer innervation in the Drosophila optic lobes. PLoS One 15(2): e0227897. PubMed ID: 32023281
The optic lobes of the fruit fly Drosophila melanogaster form a highly wired neural network composed of roughly 130.000 neurons of more than 80 different types. This study use the optic lobe of the fruit fly as a paradigm to understand how neuroblasts generate multiple neuron types. A large-scale lineage bioinformatics analysis was performed using the graph theory. A large collection of cell clones were generated that genetically label the progeny of neuroblasts, and a database was built to draw graphs showing the lineage relationships between cell types. By establishing biological criteria that measures the strength of the neuronal relationships and applying community detection tools, eight clusters of neurons were identified. Each cluster contains different cell types that are posed to be the product of eight distinct classes of neuroblasts. Three of these clusters match the available lineage data, supporting the predictive value of the analysis. Finally, it was shown that the neuronal progeny of a neuroblast do not have preferential innervation patterns, but instead become part of different layers and neuropils. This study established a new methodology that helps understanding the logic of Drosophila brain development and can be applied to the more complex vertebrate brains.
Elkahlah, N. A., Rogow, J. A., Ahmed, M. and Clowney, E. J. (2020). Presynaptic developmental plasticity allows robust sparse wiring of the Drosophila mushroom body. Elife 9. PubMed ID: 31913123
In order to represent complex stimuli, principle neurons of associative learning regions receive combinatorial sensory inputs. Density of combinatorial innervation is theorized to determine the number of distinct stimuli that can be represented and distinguished from one another, with sparse innervation thought to optimize the complexity of representations in networks of limited size. How the convergence of combinatorial inputs to principle neurons of associative brain regions is established during development is unknown. This study explored the developmental patterning of sparse olfactory inputs to Kenyon cells of the Drosophila melanogaster mushroom body. By manipulating the ratio between pre- and post-synaptic cells, it was found that postsynaptic Kenyon cells set convergence ratio: Kenyon cells produce fixed distributions of dendritic claws while presynaptic processes are plastic. Moreover, this study showed that sparse odor responses are preserved in mushroom bodies with reduced cellular repertoires, suggesting that developmental specification of convergence ratio allows functional robustness.
Drews, M. S., Leonhardt, A., Pirogova, N., Richter, F. G., Schuetzenberger, A., Braun, L., Serbe, E. and Borst, A. (2020). Dynamic signal compression for robust motion vision in flies. Curr Biol. 30(2):209-221. PubMed ID: 31928873
Sensory systems need to reliably extract information from highly variable natural signals. Flies, for instance, use optic flow to guide their course and are remarkably adept at estimating image velocity regardless of image statistics. Current circuit models, however, cannot account for this robustness. This study demonstrates that the Drosophila visual system reduces input variability by rapidly adjusting its sensitivity to local contrast conditions. Functional properties were exhaustively mapped of neurons in the motion detection circuit; local responses were found to be compressed by surround contrast. The compressive signal is fast, integrates spatially, and derives from neural feedback. Training convolutional neural networks on estimating the velocity of natural stimuli shows that this dynamic signal compression can close the performance gap between model and organism. Overall, this work represents a comprehensive mechanistic account of how neural systems attain the robustness to carry out survival-critical tasks in challenging real-world environments.
Dissel, S., Morgan, E., Duong, V., Chan, D., van Swinderen, B., Shaw, P. and Zars, T. (2020). Sleep restores place learning to the adenylyl cyclase mutant rutabaga. J Neurogenet: 1-9. PubMed ID: 31997683
Sleep plays an important role in regulating plasticity. In Drosophila, the relationship between sleep and learning and memory has primarily focused on mushroom body dependent operant-learning assays such as aversive phototaxic suppression and courtship conditioning. In this study, sleep was increased in the classic mutant rutabaga (rut2080) and dunce (dnc1) by feeding them the GABA-A agonist gaboxadol (Gab). Performance was evaluated in each mutant in response to social enrichment and place learning, tasks that do not require the mushroom body. Gab-induced sleep did not restore behavioral plasticity to either rut2080 or dnc1 mutants following social enrichment. However, increased sleep restored place learning to rut2080 mutants. These data extend the positive effects of enhanced sleep to place learning and highlight the utility of Gab for elucidating the beneficial effects of sleep on brain functioning.
Davis, F. P., Nern, A., Picard, S., Reiser, M. B., Rubin, G. M., Eddy, S. R. and Henry, G. L. (2020). A genetic, genomic, and computational resource for exploring neural circuit function. Elife 9. PubMed ID: 31939737
The anatomy of many neural circuits is being characterized with increasing resolution, but their molecular properties remain mostly unknown. This study characterized gene expression patterns in distinct neural cell types of the Drosophila visual system using genetic lines to access individual cell types, the TAPIN-seq method to measure their transcriptomes, and a probabilistic method to interpret these measurements. These tools were used to build a resource of high-resolution transcriptomes for 100 driver lines covering 67 cell types. Combining these transcriptomes with recently reported connectomes helps characterize how information is transmitted and processed across a range of scales, from individual synapses to circuit pathways. Examples are described that include identifying neurotransmitters, including cases of apparent co-release, generating functional hypotheses based on receptor expression, as well as identifying strong commonalities between different cell types.
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