What's hot today: Current papers in developmental biology and gene function Follow @interactivefly Tweet

Tuesday, December 15th

Khan, M.R., Li, L., Pérez-Sánchez, C., Saraf, A., Florens, L., Slaughter, B.D., Unruh, J.R. and Si, K. (2015). Amyloidogenic oligomerization transforms Drosophila Orb2 from a translation repressor to an activator. Cell 163: 1468-1483. PubMed ID: 26638074 Summary: Memories are thought to be formed in response to transient experiences, in part through changes in local protein synthesis at synapses. In Drosophila, the amyloidogenic (prion-like) state of the RNA binding protein Orb2 has been implicated in long-term memory, but how conformational conversion of Orb2 promotes memory formation is unclear. Combining in vitro and in vivo studies, this study finds that the monomeric form of Orb2 represses translation and removes mRNA poly(A) tails, while the oligomeric form enhances translation and elongates the poly(A) tails and imparts its translational state to the monomer. The CG13928 protein, which binds only to monomeric Orb2, promotes deadenylation, whereas the putative poly(A) binding protein CG4612 promotes oligomeric Orb2-dependent translation. These data support a model in which monomeric Orb2 keeps target mRNA in a translationally dormant state and experience-dependent conversion to the amyloidogenic state activates translation, resulting in persistent alteration of synaptic activity and stabilization of memory.

Lin, T., et al. (2015). Spindle-F is the central mediator of Ik2 kinase-dependent dendrite pruning in Drosophila sensory neurons. PLoS Genet 11: e1005642. PubMed ID: 26540204 Summary: During development, certain Drosophila sensory neurons undergo dendrite pruning that selectively eliminates their dendrites but leaves the axons intact. How these neurons regulate pruning activity in the dendrites remains unknown. This study identifies a coiled-coil protein Spindle-F (Spn-F) that is required for dendrite pruning in Drosophila sensory neurons. Spn-F acts downstream of IKK-related kinase Ik2 in the same pathway for dendrite pruning. Spn-F exhibits a punctate pattern in larval neurons, whereas these Spn-F puncta become redistributed in pupal neurons, a step that is essential for dendrite pruning. The redistribution of Spn-F from puncta in pupal neurons requires the phosphorylation of Spn-F by Ik2 kinase to decrease Spn-F self-association, and depends on the function of microtubule motor dynein complex. Spn-F is a key component to link Ik2 kinase to dynein motor complex, and the formation of Ik2/Spn-F/dynein complex is critical for Spn-F redistribution and for dendrite pruning. These findings reveal a novel regulatory mechanism for dendrite pruning achieved by temporal activation of Ik2 kinase and dynein-mediated redistribution of Ik2/Spn-F complex in neurons.

Cattenoz, P. B., Popkova, A., Southall, T., Aiello, G., Brand, A. and Giangrande, A. (2015). . Functional conservation of the Glide/Gcm regulatory network controlling glia, hemocyte and tendon cell differentiation in Drosophila. Genetics [Epub ahead of print] . PubMed ID: 26567182 Summary: This study shows the results and the validation of a DamID genome-wide screen that identifies the direct targets of Gcm, a potent transcription factor that controls glia, hemocyte and tendon cell differentiation in Drosophila. The screen highlights three major signaling pathways interacting with Gcm: Notch, Hedgehog and JAK/STAT, which all involve feedback loops. Furthermore, the screen identifies effector molecules that are necessary for cell-cell interactions during late developmental processes and/or in ontogeny. Typically, Immunoglobulin domain-containing proteins control cell adhesion and axonal navigation. This shows that early and transiently expressed fate determinants not only control other transcription factors that in turn implement a specific developmental program but also directly affect late developmental events and cell function. This study also provides the first evidence for the conservation of Gcm direct targets in humans.

Sitaraman, D., Aso, Y., Rubin, G.M. and Nitabach, M.N. (2015). Control of sleep by dopaminergic inputs to the Drosophila mushroom body. Front Neural Circuits 9: 73. PubMed ID: 26617493 Summary: The Drosophila mushroom body (MB) is an associative learning network that is important for the control of sleep. Particular intrinsic MB Kenyon cell (KC) classes have been identified that regulate sleep through synaptic activation of particular MB output neurons (MBONs) whose axons convey sleep control signals out of the MB to downstream target regions. Specifically, it was found that sleep-promoting KCs increase sleep by preferentially activating cholinergic sleep-promoting MBONs, while wake-promoting KCs decrease sleep by preferentially activating glutamatergic wake-promoting MBONs. By using a combination of genetic and physiological approaches to identify wake-promoting dopaminergic neurons (DANs) that innervate the MB, it was shown that they activate wake-promoting MBONs. These studies reveal a dopaminergic sleep control mechanism that likely operates by modulation of KC-MBON microcircuits.

Monday, December 14th

Liu, C., Haynes, P. R., Donelson, N. C., Aharon, S. and Griffith, L. C. (2015). Sleep in populations of Drosophila melanogaster. eNeuro 2 [Epub ahead of print]. PubMed ID: 26465005 Summary: The fruit fly Drosophila melanogaster is a diurnal insect active during the day with consolidated sleep at night. Social interactions between pairs of flies have been shown to affect locomotor activity patterns, but effects on locomotion and sleep patterns have not been assessed for larger populations. This study used a commercially available locomotor activity monitor (LAM25H) system to record and analyze sleep behavior. Surprisingly, it was found that same-sex populations of flies synchronize their sleep/wake activity, resulting in a population sleep pattern, which is similar but not identical to that of isolated individuals. Like individual flies, groups of flies show circadian and homeostatic regulation of sleep, as well as sexual dimorphism in sleep pattern and sensitivity to starvation and a known sleep-disrupting mutation (amnesiac). Populations of flies, however, exhibit distinct sleep characteristics from individuals. Differences in sleep appear to be due to olfaction-dependent social interactions and change with population size and sex ratio. These data support the idea that it is possible to investigate neural mechanisms underlying the effects of population behaviors on sleep by directly looking at a large number of animals in laboratory conditions.

Chen, C., Buhl, E., Xu, M., Croset, V., Rees, J. S., Lilley, K. S., Benton, R., Hodge, J. J. and Stanewsky, R. (2015). Drosophila Ionotropic Receptor 25a mediates circadian clock resetting by temperature. Nature 527: 516-520. PubMed ID: 26580016 Summary: Circadian clocks are endogenous timers adjusting behaviour and physiology with the solar day. Visual and non-visual photoreceptors are responsible for synchronizing circadian clocks to light, but clock-resetting is also achieved by alternating day and night temperatures with only 2-4 degrees C difference. This study shows that Drosophila Ionotropic Receptor 25a (IR25a) is required for behavioural synchronization to low-amplitude temperature cycles. This channel is expressed in sensory neurons of internal stretch receptors previously implicated in temperature synchronization of the circadian clock. IR25a is required for temperature-synchronized clock protein oscillations in subsets of central clock neurons. Extracellular leg nerve recordings reveal temperature- and IR25a-dependent sensory responses, and IR25a misexpression confers temperature-dependent firing of heterologous neurons. It is proposed that IR25a is part of an input pathway to the circadian clock that detects small temperature differences. This pathway operates in the absence of known 'hot' and 'cold' sensors in the Drosophila antenna, revealing the existence of novel periphery-to-brain temperature signalling channels.

Shim, J., Lee, Y., Jeong, Y. T., Kim, Y., Lee, M. G., Montell, C. and Moon, S. J. (2015). The full repertoire of Drosophila gustatory receptors for detecting an aversive compound. Nat Commun 6: 8867. PubMed ID: 26568264 Summary: The ability to detect toxic compounds in foods is essential for animal survival. However, the minimal subunit composition of gustatory receptors required for sensing aversive chemicals in Drosophila is unknown. This study reports that three gustatory receptors, GR8a, GR66a and GR98b function together in the detection of L-canavanine, a plant-derived insecticide. Ectopic co-expression of Gr8a and Gr98b in Gr66a-expressing, bitter-sensing gustatory receptor neurons (GRNs) confers responsiveness to L-canavanine. Furthermore, misexpression of all three Grs enables salt- or sweet-sensing GRNs to respond to L-canavanine. Introduction of these Grs in sweet-sensing GRNs switches L-canavanine from an aversive to an attractive compound. Co-expression of GR8a, GR66a and GR98b in Drosophila S2 cells induces an L-canavanine-activated nonselective cation conductance. It is concluded that three GRs collaborate to produce a functional L-canavanine receptor. Thus, these results clarify the full set of GRs underlying the detection of a toxic tastant that drives avoidance behaviour in an insect.

Kallman, B. R., Kim, H. and Scott, K. (2015). Excitation and inhibition onto central courtship neurons biases mate choice. Elife 4 [Epub ahead of print]. PubMed ID: 26568316 Summary: The ability to distinguish males from females is essential for productive mate selection and species propagation. Recent studies in Drosophila have identified different classes of contact chemosensory neurons that detect female or male pheromones and influence courtship decisions. This study examined central neural pathways in the male brain that process female and male pheromones using anatomical, calcium imaging, optogenetic, and behavioral studies. Sensory neurons were found that detect female pheromones, but not male pheromones, activate a novel class of neurons in the ventral nerve cord to cause activation of P1 neurons, male-specific command neurons that trigger courtship. In addition, sensory neurons that detect male pheromones, as well as those that detect female pheromones, activate central mAL neurons to inhibit P1. These studies demonstrate that the balance of excitatory and inhibitory drives onto central courtship-promoting neurons controls mating decisions.

Kidd, P. B., Young, M. W. and Siggia, E. D. (2015). Temperature compensation and temperature sensation in the circadian clock. Proc Natl Acad Sci U S A 112: E6284-6292. PubMed ID: 26578788 Summary: All known circadian clocks have an endogenous period that is remarkably insensitive to temperature, a property known as temperature compensation, while at the same time being readily entrained by a diurnal temperature oscillation. Although temperature compensation and entrainment are defining features of circadian clocks, their mechanisms remain poorly understood. Most models presume that multiple steps in the circadian cycle are temperature-dependent, thus facilitating temperature entrainment, but then insist that the effect of changes around the cycle sums to zero to enforce temperature compensation. An alternative theory proposes that the circadian oscillator evolved from an adaptive temperature sensor: a gene circuit that responds only to temperature changes. This theory implies that temperature changes should linearly rescale the amplitudes of clock component oscillations but leave phase relationships and shapes unchanged. This study shows using timeless luciferase reporter measurements and Western blots against Timeless protein that this prediction is satisfied by the Drosophila circadian clock. Evidence is reviewed for pathways that couple temperature to the circadian clock; previously unidentified evidence is shown for coupling between the Drosophila clock and the heat-shock pathway.

Gardner, B., Strus, E., Meng, Q. C., Coradetti, T., Naidoo, N. N., Kelz, M. B. and Williams, J. A. (2015). Sleep homeostasis and general anesthesia: Are fruit flies well rested after emergence from Propofol?. Anesthesiology [Epub ahead of print]. PubMed ID: 26556728 Summary: Shared neurophysiologic features between sleep and anesthetic-induced hypnosis indicate a potential overlap in neuronal circuitry underlying both states. The authors explored the hypothesis that propofol anesthesia also dispels sleep pressure in the fruit fly. Propofol was administered by transferring flies onto food containing various doses of propofol. High-performance liquid chromatography was used to measure the tissue concentrations of ingested propofol. To determine whether propofol anesthesia substitutes for natural sleep, the flies were subjected to 10-h sleep deprivation (SD), followed by 6-h propofol exposure, and monitored for subsequent sleep. Oral propofol treatment was shown to cause anesthesia in flies as indicated by a dose-dependent reduction in locomotor activity and increased arousal threshold. Recovery sleep in flies fed propofol after SD was delayed until after flies had emerged from anesthesia. SD was also associated with a significant increase in mortality in propofol-fed flies. Together, these data indicate that fruit flies are effectively anesthetized by ingestion of propofol and suggest that homologous molecular and neuronal targets of propofol are conserved in Drosophila. However, behavioral measurements indicate that propofol anesthesia does not satisfy the homeostatic need for sleep and may compromise the restorative properties of sleep.

Sunday, December 13th

Costechareyre, D., Capo, F., Fabre, A., Chaduli, D., Kellenberger, C., Roussel, A., Charroux, B. and Royet, J. (2015). Tissue-specific regulation of Drosophila NF-κB pathway activation by peptidoglycan recognition protein SC. J Innate Immun [Epub ahead of print]. PubMed ID: 26513145 Summary: In Drosophila, peptidoglycan (PGN) is detected by PGN recognition proteins (PGRPs) that act as pattern recognition receptors. Some PGRPs such as PGRP-LB or PGRP-SCs are able to cleave PGN, therefore reducing the amount of immune elicitors and dampening immune deficiency (IMD) pathway activation. By generating PGRP-SC-specific mutants, this study reevaluated the roles of PGRP-LB, PGRP-SC1 and PGRP-SC2 during immune responses. These genes were shown to be expressed in different gut domains, and they follow distinct transcriptional regulation. Loss-of-function mutant analysis indicates that PGRP-LB is playing a major role in IMD pathway activation and bacterial load regulation in the gut, although PGRP-SCs are expressed at high levels in this organ. PGRP-SC2 is the main negative regulator of IMD pathway activation in the fat body. Accordingly, mutants for either PGRP-LB or PGRP-SC2 displayed a distinct susceptibility to bacteria depending on the infection route. Lastly, PGRP-SC1 and PGRP-SC2 are required in vivo for full Toll pathway activation by Gram-positive bacteria.

Sansone, C. L., Cohen, J., Yasunaga, A., Xu, J., Osborn, G., Subramanian, H., Gold, B., Buchon, N. and Cherry, S. (2015). Microbiota-dependent priming of antiviral intestinal immunity in Drosophila. Cell Host Microbe 18: 571-581. PubMed ID: 26567510 Summary: Enteric pathogens must overcome intestinal defenses to establish infection. In Drosophila, the ERK signaling pathway inhibits enteric virus infection. The intestinal microflora also impacts immunity but its role in enteric viral infection is unknown. This study shows that two signals are required to activate antiviral ERK signaling in the intestinal epithelium. One signal depends on recognition of peptidoglycan from the microbiota, particularly from the commensal Acetobacter pomorum, which primes the NF-κB-dependent induction of a secreted factor, Pvf2. However, the microbiota is not sufficient to induce this pathway; a second virus-initiated signaling event involving release of transcriptional paused genes mediated by the kinase Cdk9 is also required for Pvf2 production. Pvf2 stimulates antiviral immunity by binding to the receptor tyrosine kinase PVR, which is necessary and sufficient for intestinal ERK responses. These findings demonstrate that sensing of specific commensals primes inflammatory signaling required for epithelial responses that restrict enteric viral infections.

Helenius, I. T., Haake, R. J., Kwon, Y. J., Hu, J. A., Krupinski, T., Casalino-Matsuda, S. M., Sporn, P. H., Sznajder, J. I. and Beitel, G. J. (2015). Identification of Drosophila Zfh2 as a mediator of hypercapnic immune regulation by a Genome-Wide RNA interference screen. J Immunol. PubMed ID: 26643480 Summary: Hypercapnia, elevated partial pressure of CO2 in blood and tissue, develops in many patients with chronic severe obstructive pulmonary disease and other advanced lung disorders. Patients with advanced disease frequently develop bacterial lung infections. Hypercapnia has been shown to suppress induction of NF-κB-regulated innate immune response genes required for host defense, and it increases mortality from bacterial infections. However, the molecular mediators of hypercapnic immune suppression are undefined. This study reports a genome-wide RNA interference screen in Drosophila S2* cells stimulated with bacterial peptidoglycan. The screen identified 16 genes with human orthologs whose knockdown reduced hypercapnic suppression of the gene encoding the antimicrobial peptide Diptericin (Dipt), but did not increase Dipt mRNA levels in air. In vivo tests of one of the strongest screen hits, zinc finger homeodomain 2 (Zfh2; mammalian orthologs ZFHX3/ATBF1 and ZFHX4), demonstrate that reducing zfh2 function improves survival of flies exposed to elevated CO2 and infected with Staphylococcus aureus. Tissue-specific knockdown of zfh2 in the fat body mitigates hypercapnia-induced reductions in Dipt and improves resistance of CO2-exposed flies to infection. Zfh2 mutations also partially rescue hypercapnia-induced delays in egg hatching, suggesting that Zfh2's role in mediating responses to hypercapnia extends beyond the immune system. Taken together these results identify Zfh2 as the first in vivo mediator of hypercapnic immune suppression.

Gutzwiller, F., Carmo, C. R., Miller, D. E., Rice, D. W., Newton, I. L., Hawley, R. S., Teixeira, L. and Bergman, C. M. (2015). Dynamics of Wolbachia pipientis gene expression across the Drosophila melanogaster life cycle. G3 (Bethesda) [Epub ahead of print]. PubMed ID: 26497146 Summary: Symbiotic interactions between microbes and their multicellular hosts have manifold biological consequences. To better understand how bacteria maintain symbiotic associations with animal hosts, this study analyzed genome-wide gene expression for the endosymbiotic alpha-proteobacteria Wolbachia pipientis across the entire life cycle of Drosophila. The majority of Wolbachia genes are expressed stably across the D. melanogaster life cycle, but 7.8% of Wolbachia genes exhibit robust stage- or sex-specific expression differences when studied in the whole-organism context. Differentially-expressed Wolbachia genes are typically up-regulated after Drosophila embryogenesis and include many bacterial membrane, secretion system, and ankyrin-repeat containing proteins. Sex-biased genes are often organized as small operons of uncharacterized genes and are mainly up-regulated in adult Drosophila males in an age-dependent manner. Expression levels of previously reported candidate genes thought to be involved in host-microbe interaction were systematically investigate, including those in the WO-A and WO-B prophages and in the Octomom region that has been implicated in regulating bacterial titre and pathogenicity. This work provides comprehensive insight into the developmental dynamics of gene expression for a widespread endosymbiont in its natural host context, and shows that public gene expression data harbour rich resources to probe the functional basis of the Wolbachia-Drosophila symbiosis and annotate the transcriptional outputs of the Wolbachia genome.

Saturday, December 12th

Bozicevic, V., Hutter, S., Stephan, W. and Wollstein, A. (2015). Population genetic evidence for cold adaptation in European Drosophila melanogaster populations. Mol Ecol [Epub ahead of print]. PubMed ID: 26558479 Summary: Drosophila melanogaster populations from Europe (the Netherlands and France) and Africa (Rwanda and Zambia) were studied to uncover genetic evidence of adaptation to cold. This study presents four lines of evidence for genes involved in cold adaptation from four perspectives: (1) the frequency of SNPs at genes previously known to be associated with chill-coma recovery time (CCRT), startle reflex (SR), and resistance to starvation stress (RSS) vary along environmental gradients and therefore among populations; (2) SNPs of genes that correlate significantly with latitude and altitude in African and European populations overlap with SNPs that correlate with a latitudinal cline from North America; (3) at the genome-wide level, the top candidate genes are enriched in gene ontology (GO) terms that are related to cold tolerance; (4) GO enriched terms from North American clinal genes overlap significantly with those from Africa and Europe. Each SNP was tested in 10 independent runs of Bayenv2, using the median Bayes factors to ascertain candidate genes. None of the candidate genes were found close to the breakpoints of cosmopolitan inversions, and only four candidate genes were linked to QTLs related to CCRT. To overcome the limitation that only four populations were used to test correlations with environmental gradients, simulations were performed to estimate the power of this approach for detecting selection. Based on the results a novel network of genes is proposed that is involved in cold adaptation.

Bergland, A. O., Tobler, R., Gonzalez, J., Schmidt, P. and Petrov, D. (2015). Secondary contact and local adaptation contribute to genome-wide patterns of clinal variation in Drosophila melanogaster. Mol Ecol [Epub ahead of print]. PubMed ID: 26547394 Summary: Populations arrayed along broad latitudinal gradients often show patterns of clinal variation in phenotype and genotype. Such population differentiation can be generated and maintained by both historical demographic events and local adaptation. These evolutionary forces are not mutually exclusive and can in some cases produce nearly identical patterns of genetic differentiation among populations. This study investigated the evolutionary forces that generated and maintain clinal variation genome-wide among populations of Drosophila melanogaster sampled in North America and Australia. Patterns of clinal variation in these continents were contrasted with patterns of differentiation among ancestral European and African populations. Using established and novel methods derive in this study, it was shown that recently derived North America and Australia populations were likely founded by both European and African lineages and that this hybridization event likely contributed to genome-wide patterns of parallel clinal variation between continents. The pervasive effects of admixture means that differentiation at only several hundred loci can be attributed to the operation of spatially varying selection using an FST outlier approach. These results provide novel insight into the well-studied system of clinal differentiation in D. melanogaster and provide a context for future studies seeking to identify loci contributing to local adaptation in a wide variety of organisms, including other invasive species as well as temperate endemics.

Poyet, M., Le Roux, V., Gibert, P., Meirland, A., Prevost, G., Eslin, P. and Chabrerie, O. (2015). The wide potential trophic niche of the asiatic fruit fly Drosophila suzukii: The key of its invasion success in temperate Europe?. PLoS One 10: e0142785. PubMed ID: 26581101 Summary: The Asiatic fruit fly Drosophila suzukii has recently invaded Europe and North and South America, causing severe damage to fruit production systems. Although agronomic host plants of that fly are now well documented, little is known about the suitability of wild and ornamental hosts in its exotic area. In order to study the potential trophic niche of D. suzukii with relation to fruit characteristics, fleshy fruits from 67 plant species were sampled in natural and anthropic ecosystems (forests, hedgerows, grasslands, coastal areas, gardens and urban areas) of the north of France and submitted to experimental infestations. A set of fruit traits (structure, colour, shape, skin texture, diameter and weight, phenology) potentially interacting with oviposition choices and development success of D. suzukii was measured. Almost half of the tested plant species belonging to 17 plant families allowed the full development of D. suzukii. This suggests that the extreme polyphagy of the fly and the very large reservoir of hosts producing fruits all year round ensure temporal continuity in resource availability and contribute to the persistence and the exceptional invasion success of D. suzukii in natural habitats and neighbouring cultivated systems. Nevertheless, this very plastic trophic niche is not systematically beneficial to the fly. Some of the tested plants attractive to D. suzukii gravid females stimulate oviposition but do not allow full larval development. Planted near sensitive crops, these "trap plants" may attract and lure D. suzukii, therefore contributing to the control of the invasive fly.

Kain, J. S., Zhang, S., Akhund-Zade, J., Samuel, A. D., Klein, M. and de Bivort, B. L. (2015). Variability in thermal and phototactic preferences in Drosophila may reflect an adaptive bet-hedging strategy. Evolution [Epub ahead of print]. PubMed ID: 26531165 Summary: Organisms use various strategies to cope with fluctuating environmental conditions. In diversified bet-hedging, a single genotype exhibits phenotypic heterogeneity with the expectation that some individuals will survive transient selective pressures. To date, empirical evidence for bet-hedging is scarce. This study observed that individual Drosophila melanogaster flies exhibit striking variation in light- and temperature-preference behaviors. With a modeling approach that combines real world weather and climate data to simulate temperature preference-dependent survival and reproduction, this study found that a bet-hedging strategy may underlie the observed inter-individual behavioral diversity. Specifically, bet-hedging outcompetes strategies in which individual thermal preferences are heritable. Animals employing bet-hedging refrain from adapting to the coolness of spring with increased warm-seeking that inevitably becomes counterproductive in the hot summer. This strategy is particularly valuable when mean seasonal temperatures are typical, or when there is considerable fluctuation in temperature within the season. The model predicts, and and this study experimentally verified, that the behaviors of individual flies are not heritable. Finally, the effects of historical weather data, climate change, and geographic seasonal variation on the optimal strategies underlying behavioral variation between individuals was modeled, characterizing the regimes in which bet-hedging is advantageous.

Giraldo-Perez, P., Herrera, P., Campbell, A., Taylor, M. L., Skeats, A., Aggio, R., Wedell, N. and Price, T. A. (2015). Winter is coming: hibernation reverses the outcome of sperm competition in a fly. J Evol Biol [Epub ahead of print]. PubMed ID: 26565889 Summary: Sperm commonly compete within females to fertilise ova, but research has focused on short term sperm storage: sperm that are maintained in a female for only a few days or weeks before use. In nature, females of many species store sperm for months or years, often during periods of environmental stress, such as cold winters. This study examined the outcome of sperm competition in the fruit fly Drosophila pseudoobscura, simulating the conditions in which females survive winter. Females were mated to two males, and then the female was stored for up to 120 days at 4 degrees C. It was found that the outcome of sperm competition was consistent when sperm from two males was stored for 0, 1 or 30 days, with the last male to mate fathering most of the offspring. However, when females were stored in the cold for 120 days, the last male to mate fathered less than 5% of the offspring. Moreover, when sperm were stored long term the first male fathered almost all offspring even when he carried a meiotic driving sex chromosome that drastically reduces sperm competitive success under short term storage conditions. This suggests that long-term sperm storage can radically alter the outcome of sperm competition.

Yassin, A. and David, J. R. (2015). Within species reproductive costs affect the asymmetry of satyrisation in Drosophila. J Evol Biol [Epub ahead of print]. PubMed ID: 26538290 Summary: Understanding how species interactions influence their distribution and evolution is a fundamental question in evolutionary biology. Theory suggests that asymmetric reproductive interference, in which one species induces higher reproductive costs on another species, may be more important in delimiting species boundaries than interspecific competition over resources. However, the underlying mechanisms of such asymmetry remains unclear. This study tested whether differences in within species reproductive costs determine the between species asymmetry of costs using three allopatric Drosophila species belonging to the melanogaster subgroup. The results support this hypothesis, especially in a pair of insular species. Males of one species that induce costs to their conspecific females led to a 5-fold increase of heterospecific females mortality with dead flies bearing spectacular large melanised wounds on their genitalia. Males of the other species were harmful neither to their conspecific nor heterospecific females. Comparative studies of within species reproductive costs may therefore be a valuable tool for predicting between species interactions and community structures.

Friday, December 11th

Spellberg, M. J. and Marr, M. T. (2015). FOXO regulates RNA interference in Drosophila and protects from RNA virus infection. Proc Natl Acad Sci U S A 112: 14587-14592. PubMed ID: 26553999 Summary: Small RNA pathways are important players in posttranscriptional regulation of gene expression. These pathways play important roles in all aspects of cellular physiology from development to fertility to innate immunity. However, almost nothing is known about the regulation of the central genes in these pathways. The forkhead box O (FOXO) family of transcription factors is a conserved family of DNA-binding proteins that responds to a diverse set of cellular signals. FOXOs are crucial regulators of cellular homeostasis that have a conserved role in modulating organismal aging and fitness. This study shows that Drosophila FOXO (dFOXO) regulates the expression of core small RNA pathway genes. In addition, increased dFOXO activity results in an increase in RNA interference (RNAi) efficacy, establishing a direct link between cellular physiology and RNAi. Consistent with these findings, dFOXO activity is stimulated by viral infection and is required for effective innate immune response to RNA virus infection. This study reveals an unanticipated connection among dFOXO, stress responses, and the efficacy of small RNA-mediated gene silencing and suggests that organisms can tune their gene silencing in response to environmental and metabolic conditions.

Brewer-Jensen, P., Wilson, C. B., Abernethy, J., Mollison, L., Card, S. and Searles, L. L. (2015). Suppressor of sable [Su(s)] and Wdr82 down-regulate RNA from heat-shock-inducible repetitive elements by a mechanism that involves transcription termination. RNA [Epub ahead of print]. PubMed ID: 26577379 Summary: Although RNA polymerase II (Pol II) productively transcribes very long genes in vivo, transcription through extragenic sequences often terminates in the promoter-proximal region and the nascent RNA is degraded. Mechanisms that induce early termination and RNA degradation are not well understood in multicellular organisms. This study presents evidence that the suppressor of sable [su(s)] regulatory pathway of Drosophila plays a role in this process. Previous studies have shown that Su(s) promotes exosome-mediated degradation of transcripts from endogenous repeated elements at an Hsp70 locus (Hsp70-αβ elements). This report identifies Wdr82 as a component of this process and shows that it works with Su(s) to inhibit Pol II elongation through Hsp70-αβ elements. Furthermore, the unstable transcripts produced during this process are shown to be polyadenylated at heterogeneous sites that lack canonical polyadenylation signals. Two distinct regions were defined that mediate this regulation. These results indicate that the Su(s) pathway promotes RNA degradation and transcription termination through a novel mechanism.

Xie, X. and Dubrovsky, E. B. (2015). Knockout of Drosophila RNase ZL impairs mitochondrial transcript processing, respiration and cell cycle progression. Nucleic Acids Res [Epub ahead of print]. PubMed ID: 26553808 Summary: RNase ZL is a highly conserved tRNA 3'-end processing endoribonuclease. Similar to its mammalian counterpart, Drosophila RNase ZL (dRNaseZ) has a mitochondria targeting signal (MTS) flanked by two methionines at the N-terminus. Alternative translation initiation yields two protein forms: the long one is mitochondrial, and the short one may localize in the nucleus or cytosol. This study has generated a mitochondria specific knockout of the dRNaseZ gene. In this in vivo model, cells deprived of dRNaseZ activity display impaired mitochondrial polycistronic transcript processing, increased reactive oxygen species (ROS) and a switch to aerobic glycolysis compensating for cellular ATP. Damaged mitochondria impose a cell cycle delay at the G2 phase disrupting cell proliferation without affecting cell viability. Antioxidants attenuate genotoxic stress and rescue cell proliferation, implying a critical role for ROS. It is suggested that under a low-stress condition, ROS activate tumor suppressor p53, which modulates cell cycle progression and promotes cell survival. Transcriptional profiling of p53 targets confirms upregulation of antioxidant and cycB-Cdk1 inhibitor genes without induction of apoptotic genes. This study implicates Drosophila RNase ZL in a novel retrograde signaling pathway initiated by the damage in mitochondria and manifested in a cell cycle delay before the mitotic entry.

Novak, S. M., Joardar, A., Gregorio, C. C. and Zarnescu, D. C. (2015). Regulation of heart rate in Drosophila via Fragile X mental retardation protein. PLoS One 10: e0142836. PubMed ID: 26571124 Summary: RNA binding proteins play a pivotal role in post-transcriptional gene expression regulation, however little is understood about their role in cardiac function. Alterations in the levels of Fragile X Related 1 protein, FXR1, the predominant FraX member expressed in vertebrate striated muscle, have been linked to structural and functional defects in mice and zebrafish models. FraX proteins are established regulators of translation and are known to regulate specific targets in different tissues. To decipher the direct role of FraX proteins in the heart in vivo, Drosophila, which harbors a sole, functionally conserved and ubiquitously expressed FraX protein, dFmr1, was investigated. Using classical loss of function alleles as well as muscle specific RNAi knockdown, dFmr1 was shown to be required for proper heart rate during development. Functional analyses in the context of cardiac-specific dFmr1 knockdown by RNAi demonstrate that dFmr1 is required cell autonomously in cardiac cells for regulating heart rate. Interestingly, these functional defects are not accompanied by any obvious structural abnormalities, suggesting that dFmr1 may regulate a different repertoire of targets in Drosophila than in vertebrates. Taken together, these findings support the hypothesis that dFmr1 protein is essential for proper cardiac function and establish the fly as a new model for studying the role(s) of FraX proteins in the heart.

Thursday, December 10th

Yamanaka, N., Marques, G. and O'Connor, M. B. (2015). Vesicle-mediated steroid hormone secretion in Drosophila melanogaster. Cell 163: 907-919. PubMed ID: 26544939 Summary: Steroid hormones are a large family of cholesterol derivatives regulating development and physiology in both the animal and plant kingdoms, but little is known concerning mechanisms of their secretion from steroidogenic tissues. This study presents evidence that in Drosophila, endocrine release of the steroid hormone ecdysone is mediated through a regulated vesicular trafficking mechanism. Inhibition of calcium signaling in the steroidogenic prothoracic gland results in the accumulation of unreleased ecdysone, and the knockdown of calcium-mediated vesicle exocytosis components in the gland caused developmental defects due to deficiency of ecdysone. Accumulation of synaptotagmin-labeled vesicles in the gland is observed when calcium signaling is disrupted, and these vesicles contain an ABC transporter that functions as an ecdysone pump to fill vesicles. It is proposed that trafficking of steroid hormones out of endocrine cells is not always through a simple diffusion mechanism as presently thought, but instead can involve a regulated vesicle-mediated release process.

Winther, A. M., Vorontsova, O., Rees, K. A., Nareoja, T., Sopova, E., Jiao, W. and Shupliakov, O. (2015). An endocytic scaffolding protein together with Synapsin regulates synaptic vesicle clustering in the Drosophila neuromuscular junction. J Neurosci 35: 14756-14770. PubMed ID: 26538647 Summary: Many endocytic proteins accumulate in the reserve pool of synaptic vesicles (SVs) in synapses and relocalize to the endocytic periactive zone during neurotransmitter release. Currently little is known about their functions outside the periactive zone. This study shows that in the Drosophila neuromuscular junction (NMJ), the endocytic scaffolding protein Dap160 colocalizes during the SV cycle and forms a functional complex with the SV-associated phosphoprotein Synapsin, previously implicated in SV clustering. This direct interaction is essential for proper localization of synapsin at NMJs. In a dap160 rescue mutant lacking the interaction between Dap160 and synapsin, perturbed reclustering of SVs during synaptic activity is observed. The data indicate that in addition to the function in endocytosis, Dap160 is a component of a network of protein-protein interactions that serves for clustering of SVs in conjunction with synapsin. During the SV cycle, Dap160 interacts with synapsin dispersed from SVs and helps direct synapsin back to vesicles.

Ukken, F. P., Bruckner, J. J., Weir, K. L., Hope, S. J., Sison, S. L., Birschbach, R. M., Hicks, L., Taylor, K. L., Dent, E. W., Gonsalvez, G. B. and O'Connor-Giles, K. M. (2015). BAR-SH3 Sorting nexins are conserved Nervous wreck interactors that organize synapses and promote neurotransmission. J Cell Sci [Epub ahead of print]. PubMed ID: 26567222 Summary: Nervous wreck (Nwk) is a conserved F-BAR protein that attenuates synaptic growth and promotes synaptic function in Drosophila. In an effort to understand how Nwk carries out its dual roles, this study isolated interacting proteins through mass spectrometry. A conserved interaction is reported between Nwk proteins and BAR-SH3 Sorting nexins, a family of membrane-binding proteins implicated in diverse intracellular trafficking processes. In mammalian cells, BAR-SH3 Sorting nexins induce plasma membrane tubules that colocalize Nwk2, consistent with a possible functional interaction during early stages of endocytic trafficking. To study the role of BAR-SH3 Sorting nexins in vivo, advantage was taken of the lack of genetic redundancy in Drosophila, and CRISPR-based genome engineering was employed to generate null and endogenously tagged alleles of SH3PX1. SH3PX1 localizes to neuromuscular junctions where it regulates synaptic ultrastructure, but not synapse number. Consistently, neurotransmitter release is significantly diminished in SH3PX1 mutants. Double mutant and tissue-specific rescue experiments indicate that SH3PX1 promotes neurotransmitter release presynaptically, at least in part through functional interactions with Nwk, and may act to distinguish Nwk's roles in regulating synaptic growth and function.

Gireud, M., Sirisaengtaksin, N., Tsunoda, S. and Bean, A. J. (2015). Cell-free reconstitution of multivesicular body (MVB) cargo sorting. Journal-Methods Mol Biol 1270: 115-124. PubMed ID: 25702113 Summary: The signaling activity of cell surface localized membrane proteins occurs primarily while these proteins are located on the plasma membrane but is, in some cases, not terminated until the proteins are degraded. Following internalization and movement through the endocytic pathway en route to lysosomes, membrane proteins transit a late endosomal organelle called the multivesicular body (MVB). MVBs are formed by invagination of the limiting membrane of endosomes, resulting in an organelle possessing a limiting membrane and containing internal vesicles. The molecular machinery that regulates the separation of membrane proteins destined for degradation from those resulting in surface expression is not well understood. This study reconstituted an endosomal sorting event under cell-free conditions. Advantage was taken of the itinerary of a prototypical membrane protein, the Epidermal growth factor receptor (EGFR) and a biochemical monitor was designed for cargo movement into internal MVB vesicles that is generally modifiable for other membrane proteins. Since is it not known how internal vesicle formation is related to cargo sorting, morphological examination using transmission electron microscopy (TEM) allows separate monitoring of vesicle formation. This study determined that MVB sorting is dependent on cytosolic components, adenosine triphosphate (ATP), time, temperature, and an intact proton gradient. This assay reconstitutes the maturation of late endosomes and allows the morphological and biochemical examination of vesicle formation and membrane protein sorting.

Wednesday December 9th

Harmansa, S., Hamaratoglu, F., Affolter, M. and Caussinus, E. (2015). Dpp spreading is required for medial but not for lateral wing disc growth. Nature 527: 317-322. PubMed ID: 26550827 Summary: Drosophila Decapentaplegic has served as a paradigm to study morphogen-dependent growth control. However, the role of a Dpp gradient in tissue growth remains highly controversial. Two fundamentally different models have been proposed: the 'temporal rule' model suggests that all cells of the wing imaginal disc divide upon a 50% increase in Dpp signalling, whereas the 'growth equalization model' suggests that Dpp is only essential for proliferation control of the central cells. To discriminate between these two models, morphotrap, a membrane-tethered anti-green fluorescent protein (GFP) nanobody was generated and used, that enables immobilization of enhanced (e)GFP::Dpp on the cell surface, thereby abolishing Dpp gradient formation. In the absence of Dpp spreading, wing disc patterning is lost; however, lateral cells still divide at normal rates. These data are consistent with the growth equalization model, but do not fit a global temporal rule model in the wing imaginal disc.

Okamoto, N. and Nishimura, T. (2015). Signaling from glia and cholinergic neurons controls nutrient-dependent production of an insulin-like peptide for Drosophila body growth. Dev Cell 35: 295-310. PubMed ID: 26555050 Summary: The insulin-like peptide (ILP) family plays key biological roles in the control of body growth. Although the functions of ILPs are well understood, the mechanisms by which organisms sense their nutrient status and thereby control ILP production remain largely unknown. This study shows that signaling relay and feedback mechanisms control the nutrient-dependent expression of Drosophila ILP5 (Dilp5). The expression of dilp5 in brain insulin-producing cells (IPCs) is negatively regulated by the transcription factor FoxO. Glia-derived Dilp6 remotely regulates the FoxO activity in IPCs, primarily through Jeb secreted by cholinergic neurons. Dilp6 production by surface glia is amplified by cellular response to circulating Dilps derived from IPCs, in concert with amino acid signals. The induction of dilp5 is critical for sustaining body growth under restricted food conditions. These results provide a molecular framework that explains how the production of an endocrine hormone in a specific tissue is coordinated with environmental conditions.

Akiyama, T. and Gibson, M. C. (2015). Decapentaplegic and growth control in the developing Drosophila wing. Nature 527: 375-378. PubMed ID: 26550824 Summary: As a central model for morphogen action during animal development, the bone morphogenetic protein 2/4 (BMP2/4)-like ligand Decapentaplegic is proposed to form a long-range signalling gradient that directs both growth and pattern formation during Drosophila wing disc development. While the patterning role of Dpp secreted from a stripe of cells along the anterior-posterior compartmental boundary is well established, the mechanism by which a Dpp gradient directs uniform cell proliferation remains controversial and poorly understood. To determine the precise spatiotemporal requirements for Dpp during wing disc development, this study used CRISPR-Cas9-mediated genome editing to generate a flippase recognition target (FRT)-dependent conditional null allele. By genetically removing Dpp from its endogenous stripe domain, the requirement was confirmed of Dpp for the activation of a downstream phospho-Mothers against dpp gradient and the regulation of the patterning targets spalt, optomotor blind and brinker. Surprisingly, however, third-instar wing blade primordia devoid of compartmental dpp expression maintain relatively normal rates of cell proliferation and exhibit only mild defects in growth. These results indicate that during the latter half of larval development, the Dpp morphogen gradient emanating from the anterior-posterior compartment boundary is not directly required for wing disc growth.

Pahi, Z., Kiss, Z., Komonyi, O., Borsos, B. N., Tora, L., Boros, I. M. and Pankotai, T. (2015). dTAF10- and dTAF10b-containing complexes are required for ecdysone-driven larval-pupal morphogenesis in Drosophila melanogaster. PLoS One 10: e0142226. PubMed ID: 26556600 Summary: In eukaryotes the TFIID complex is required for preinitiation complex assembly which positions RNA polymerase II around transcription start sites. Histone acetyltransferase complexes including SAGA and ATAC, modulate transcription at several steps through modification of specific core histone residues. This study investigated the function of Drosophila proteins TAF10 and TAF10b, which are subunits of dTFIID and dSAGA, respectively. The simultaneous deletion of both dTaf10 genes impaired the recruitment of the dTFIID subunit dTAF5 to polytene chromosomes, while binding of other TFIID subunits, RNAPII was not affected. The lack of both dTAF10 proteins resulted in failures in the larval-pupal transition during metamorphosis and in transcriptional reprogramming at this developmental stage. Importantly, the phenotype resulting from dTaf10+dTaf10b mutation could be rescued by ectopically added ecdysone, suggesting that dTAF10- and/or dTAF10b-containing complexes are involved in the expression of ecdysone biosynthetic genes. These data support the idea that the presence of dTAF10 proteins in dTFIID and/or dSAGA is required only at specific developmental steps. It is proposed that distinct forms of dTFIID and/or dSAGA exist during Drosophila metamorphosis, wherein different TAF compositions serve to target RNAPII at different developmental stages and tissues.

Tuesday, December 8th

Bhadra, U., Mondal, T., Bag, I., Mukhopadhyay, D., Das, P., Parida, B. B., Mainkar, P. S., Reddy, C. R. and Bhadra, M. P. (2015). HDAC inhibitor misprocesses bantam oncomiRNA, but stimulates hid induced apoptotic pathway. Sci Rep 5: 14747. PubMed ID: 26442596 Summary: Apoptosis or programmed cell death is critical for embryogenesis and tissue homeostasis. Uncontrolled apoptosis leads to different human disorders including immunodeficiency, autoimmune disorder and cancer. Several small molecules that control apoptosis have been identified. This study has shown the functional role of triazole derivative (DCPTN-PT) that acts as a potent HDAC inhibitor (reducing the level of Rpd3/HDAC1 in Drosophila) and leads to the misexpression of proto onco microRNA (miRNA) bantam. To further understanding the mechanism of action of the molecule in the apoptotic pathway, a series of experiments were also performed in Drosophila, a well known model organism in which the nature of human apoptosis is very analogous. DCPTN-PT mis processes bantam microRNA and alters its down regulatory target hid function and cleavage of Caspase-3 which in turn influence components of the mitochondrial apoptotic pathway in Drosophila. However regulatory microRNAs in other pro-apoptotic genes are not altered. Simultaneously, treatment of same molecule also affects the mitochondrial regulatory pathway in human tumour cell lines suggesting its conservative nature between fly and human. It is reasonable to propose that triazole derivative (DCPTN-PT) controls bantam oncomiRNA and increases hid induced apoptosis and is also able to influence mitochondrial apoptotic pathway.

Wu, Y., Lindblad, J. L., Garnett, J., Kamber Kaya, H. E., Xu, D., Zhao, Y., Flores, E. R., Hardy, J. and Bergmann, A. (2015). Genetic characterization of two gain-of-function alleles of the effector caspase DrICE in Drosophila. Cell Death Differ . PubMed ID: 26542461 Summary: Caspases are the executioners of apoptosis. Although much is known about their physiological roles and structures, detailed analyses of missense mutations of caspases are lacking. As mutations within caspases are identified in various human diseases, the study of caspase mutants will help to elucidate how caspases interact with other components of the apoptosis pathway and how they may contribute to disease. DrICE is the major effector caspase in Drosophila required for developmental and stress-induced cell death. This study reports the isolation and characterization of six de novo drICE mutants, all of which carry point mutations affecting amino acids conserved among caspases in various species. These six mutants behave as recessive loss-of-function mutants in a homozygous condition. Surprisingly, however, two of the newly isolated drICE alleles are gain-of-function mutants in a heterozygous condition, although they are loss-of-function mutants homozygously. Interestingly, they only behave as gain-of-function mutants in the presence of an apoptotic signal. These two alleles carry missense mutations affecting conserved amino acids in close proximity to the catalytic cysteine residue. This result provides a significant exception to the expectation that mutations of conserved amino acids always abolish the pro-apoptotic activity of caspases.

Jin, A., Neufeld, T. P. and Choe, J. (2015). Kibra and aPKC regulate starvation-induced autophagy in Drosophila.Biochem Biophys Res Commun 468: 1-7. PubMed ID: 26551466 Summary: Autophagy is a bulk degradation system that functions in response to cellular stresses such as metabolic stress, endoplasmic reticulum stress, oxidative stress, and developmental processes. During autophagy, cytoplasmic components are captured in double-membrane vesicles called autophagosomes. The autophagosome fuses with the lysosome, producing a vacuole known as an autolysosome. The cellular components are degraded by lysosomal proteases and recycled. Autophagy is important for maintaining cellular homeostasis, and the process is evolutionarily conserved. Kibra is an upstream regulator of the hippo signaling pathway, which controls organ size by affecting cell growth, proliferation, and apoptosis. Kibra is mainly localized in the apical membrane domain of epithelial cells and acts as a scaffold protein. This study found that Kibra is required for autophagy to function properly. The absence of Kibra caused defects in the formation of autophagic vesicles and autophagic degradation. It was also found that the well-known cell polarity protein aPKC interacts with Kibra, and its activity affects autophagy upstream of Kibra. Constitutively active aPKC decreased autophagic vesicle formation and autophagic degradation. The interaction between aPKC and Kibra was confirmed in S2 cells and Drosophila larva. Taken together, these data suggest that Kibra and aPKC are essential for regulating starvation-induced autophagy.

Joffre, C., et al. (2015). The pro-apoptotic STK38 kinase is a new Beclin1 partner positively regulating autophagy. Curr Biol 25: 2479-2492. PubMed ID: 26387716 Summary: Autophagy plays key roles in development, oncogenesis, cardiovascular, metabolic, and neurodegenerative diseases. This study describes the STK38 protein kinase (also termed NDR1 and Tricornered in Drosophila) as a conserved regulator of autophagy. STK38 was discovered as a novel binding partner of Beclin1 (Atg6; see Drosophila Atg6), a key regulator of autophagy. STK38 promotes autophagosome formation in human cells and in Drosophila. Upon autophagy induction, STK38-depleted cells display impaired LC3B-II conversion; reduced ATG14L, ATG12, and WIPI-1 puncta formation; and significantly decreased Vps34 (Pi3K59F in Drosophila) activity, as judged by PI3P formation. Furthermore, it was observed that STK38 supports the interaction of the exocyst component Exo84 with Beclin1 and RalB, which is required to initiate autophagosome formation. Upon studying the activation of STK38 during autophagy induction, STK38 was found to be stimulated in a MOB1- and exocyst-dependent manner. In contrast, RalB depletion triggers hyperactivation of STK38, resulting in STK38-dependent apoptosis under prolonged autophagy conditions. Together, these data establish STK38 as a conserved regulator of autophagy in human cells and flies. Evidence is also provided demonstrating that STK38 and RalB assist the coordination between autophagic and apoptotic events upon autophagy induction, hence further proposing a role for STK38 in determining cellular fate in response to autophagic conditions.

Monday, December 7th

Russo, J., Harrington, A. W. and Steiniger, M. (2015). Antisense transcription of retrotransposons in Drosophila: The origin of endogenous small interfering RNA precursors. Genetics [Epub ahead of print]. PubMed ID: 26534950 Summary: To repress transposons and combat genomic instability, eukaryotes have evolved several small RNA mediated defense mechanisms. Specifically, in Drosophila somatic cells, endogenous small interfering (esi)RNAs suppress retrotransposon mobility. EsiRNAs are produced by Dicer-2 processing of double-stranded RNA precursors, yet the origins of these precursors are unknown. This study shows that most transposon families are transcribed in both the sense and antisense direction. LTR retrotransposons are generated from intra element transcription start sites with canonical RNA polymerase II promoters. Retrotransposon antisense transcripts were shown to be less polyadenylated than sense transcripts, which may promote nuclear retention of antisense transcripts and the double-stranded RNAs they form. Dicer-2 RNAi-depletion causes a decrease in the number of esiRNAs mapping to retrotransposons. These data support a model in which double-stranded RNA precursors are derived from convergent transcription and processed by Dicer-2 into esiRNAs that silence both sense and antisense retrotransposon transcripts. Reduction of sense retrotransposon transcripts potentially lowers element specific protein levels to prevent transposition. This mechanism preserves genomic integrity and is especially important for Drosophila fitness because mobile genetic elements are highly active.

Loedige, I., Jakob, L., Treiber, T., Ray, D., Stotz, M., Treiber, N., Hennig, J., Cook, K. B., Morris, Q., Hughes, T. R., Engelmann, J. C., Krahn, M. P. and Meister, G. (2015). The crystal structure of the NHL domain in complex with RNA reveals the molecular basis of Drosophila Brain-tumor-mediated gene regulation. Cell Rep 13: 1206-1220. PubMed ID: 26527002 Summary: TRIM-NHL proteins are conserved among metazoans and control cell fate decisions in various stem cell linages. The Drosophila TRIM-NHL protein Brain tumor (Brat) directs differentiation of neuronal stem cells by suppressing self-renewal factors. Brat is an RNA-binding protein and functions as a translational repressor. However, it is unknown which RNAs Brat regulates and how RNA-binding specificity is achieved. Using RNA immunoprecipitation and RNAcompete, this study identified Brat-bound mRNAs in Drosophila embryos and defined consensus binding motifs for Brat as well as a number of additional TRIM-NHL proteins, indicating that TRIM-NHL proteins are conserved, sequence-specific RNA-binding proteins. Brat-mediated repression and direct RNA-binding depend on the identified motif and show that binding of the localization factor Miranda to the Brat-NHL domain inhibits Brat activity. Finally, to unravel the sequence specificity of the NHL domain, the Brat-NHL domain in complex was crystallize with RNA, and a high-resolution protein-RNA structure of this fold is presented.

Prabu, J. R., Muller, M., Thomae, A. W., Schussler, S., Bonneau, F., Becker, P. B. and Conti, E. (2015). Structure of the RNA helicase MLE reveals the molecular mechanisms for uridine specificity and RNA-ATP coupling. Mol Cell 60: 487-499. PubMed ID: 26545078 Summary: The MLE helicase remodels the roX lncRNAs, enabling the lncRNA-mediated assembly of the Drosophila dosage compensation complex. This study identified a stable MLE core comprising the DExH helicase module and two auxiliary domains: a dsRBD and an OB-like fold. MLEcore is an unusual DExH helicase that can unwind blunt-ended RNA duplexes and has specificity for uridine nucleotides.The 2.1 A resolution structure of MLEcore was determined bound to a U10 RNA and ADP-AlF4. The OB-like and dsRBD folds bind the DExH module and contribute to form the entrance of the helicase channel. Four uridine nucleotides engage in base-specific interactions, rationalizing the conservation of uridine-rich sequences in critical roX substrates. roX2 binding is orchestrated by MLE's auxiliary domains, which is prerequisite for MLE localization to the male X chromosome. The structure visualizes a transition-state mimic of the reaction and suggests how eukaryotic DEAH/RHA helicases couple ATP hydrolysis to RNA translocation.

Yue, Y., Yang, Y., Dai, L., Cao, G., Chen, R., Hong, W., Liu, B., Shi, Y., Meng, Y., Shi, F., Xiao, M. and Jin, Y. (2015). Long-range RNA pairings contribute to mutually exclusive splicing. RNA [Epub ahead of print]. PubMed ID: 26554032 Summary: Mutually exclusive splicing is an important means of increasing the protein repertoire, by which the Down's syndrome cell adhesion molecule (Dscam) gene potentially generates 38,016 different isoforms in Drosophila melanogaster. However, the regulatory mechanisms remain obscure due to the complexity of the Dscam exon cluster. This study reveal a molecular model for the regulation of the mutually exclusive splicing of the serpent pre-mRNA based on competition between upstream and downstream RNA pairings. Such dual RNA pairings confer fine tuning of the inclusion of alternative exons. Moreover, the splicing outcome of alternative exons is mediated in relative pairing strength-correlated mode. Combined comparative genomics analysis and experimental evidence revealed similar bidirectional structural architectures in exon clusters 4 and 9 of the Dscam gene. These findings provide a novel mechanistic framework for the regulation of mutually exclusive splicing and may offer potentially applicable insights into long-range RNA-RNA interactions in gene regulatory networks.

Sunday, December 6th

Schoborg, T., Zajac, A.L., Fagerstrom, C.J., Guillen, R.X. and Rusan, N.M. (2015). An Asp-CaM complex is required for centrosome-pole cohesion and centrosome inheritance in neural stem cells. J Cell Biol [Epub ahead of print]. PubMed ID: 26620907 Summary: The interaction between centrosomes and mitotic spindle poles is important for efficient spindle formation, orientation, and cell polarity. However, our understanding of the dynamics of this relationship and implications for tissue homeostasis remains poorly understood. This study reports that Drosophila melanogaster calmodulin (CaM) regulates the ability of the microcephaly-associated protein, abnormal spindle (Asp), to cross-link spindle microtubules. Both proteins colocalize on spindles and move toward spindle poles, suggesting that they form a complex. Binding and structure-function analysis support this hypothesis. Disruption of the Asp-CaM interaction alone leads to unfocused spindle poles and centrosome detachment. This behavior leads to randomly inherited centrosomes after neuroblast division. It was further shown that spindle polarity is maintained in neuroblasts despite centrosome detachment, with the poles remaining stably associated with the cell cortex. Finally, CaM is required for Asp's spindle function; however, it is completely dispensable for Asp's role in microcephaly suppression.

Hall, E. T. and Verheyen, E. M. (2015). Ras-activated Dsor1 promotes Wnt signaling in Drosophila development. J Cell Sci. [Epub ahead of print]. PubMed ID: 26542023 Summary: Wnt/Wingless (Wg) and Ras/MAPK signaling both play fundamental roles in growth, cell-fate determination, and when dysregulated, can lead to tumorigenesis. Several conflicting modes of interaction between Ras/MAPK and Wnt signaling have been identified in specific cellular contexts, causing synergistic or antagonistic effects on target genes. This study found novel evidence that the dual specificity kinase Downstream of Raf1 (Dsor1), also known as MEK. is required for Wnt signaling. Knockdown of Dsor1 results in loss of Wingless target gene expression, as well as reductions in stabilized Armadillo (Arm; Drosophila beta-catenin). A close physical interaction was found between Dsor1 and Arm; catalytically inactive Dsor1 causes a reduction inactive Arm. These results suggest that Dsor1 normally counteracts the Axin-mediated destruction of Arm. Ras-Dsor1 activity is independent of upstream activation by EGFR, rather it appears to be activated by the insulin-like growth factor receptor to promote Wg signaling. Together our results suggest novel crosstalk between Insulin and Wg signaling via Dsor1.

Zhou, Y., Chang, R., Ji, W., Wang, N., Qi, M., Xu, Y., Guo, J. and Zhan, L. (2015). Loss of Scribble promotes Snail translation through translocation of HuR and enhances cancer drug resistance. J Biol Chem [Epub ahead of print]. PubMed ID: 26527679 Summary: Drug resistance of cancer cells to various therapeutic agents and molecular targets is a major problem facing current cancer research. The tumor suppressor gene Scribble encodes a polarity protein that is conserved between Drosophila and mammals; loss of the locus disrupts cell polarity, inhibits apoptosis, and mediates cancer process. However, the role of Scribble in drug resistance remains unknown. This study shows that knockdown of Scribble enhances drug resistance by permitting accumulation of Snail, which functions as a transcription factor during the epithelial-mesenchymal transition. Then, loss of Scribble activates the mRNA binding protein HuR (ELAV in Drosophila) by facilitating translocation of HuR from the nucleus to the cytoplasm. Furthermore, HuR can recognize AU-rich elements (AREs) of the Snail-encoding mRNA, thereby regulating Snail translation. Moreover, Loss of Scribble induced HuR translocation mediates the accumulation of Snail via activation of the p38 MAPK pathway. Thus, this work clarifies the role of polarity protein Scribble, which is directly implicated in the regulation of developmental transcription factor Snail, and suggesting a mechanism for Scribble mediating cancer drug resistance.

Lin, Y. H., Currinn, H., Pocha, S. M., Rothnie, A., Wassmer, T. and Knust, E. (2015). AP-2 complex-mediated endocytosis of Drosophila Crumbs regulates polarity via antagonizing Stardust. J Cell Sci [Epub ahead of print]. PubMed ID: 26527400 Summary: Maintenance of epithelial polarity depends on the correct localization and levels of polarity determinants. The evolutionarily conserved transmembrane protein Crumbs is crucial for the size and identity of the apical membrane, yet little is known about the molecular mechanisms controlling the amount of Crumbs at the surface. This study shows that Crumbs levels on the apical membrane depend on a well-balanced state of endocytosis and stabilization. The Adaptor Protein 2 (AP-2) complex binds to a motif in the cytoplasmic tail of Crumbs that overlaps with the binding site of Stardust, a protein known to stabilize Crumbs on the surface. Preventing endocytosis by mutations in AP-2 causes expansion of the Crumbs-positive plasma membrane and polarity defects, which can be partially rescued by removing one copy of crumbs. Strikingly, knocking-down both AP-2 and Stardust retains Crumbs on the membrane. This study provides evidence for a molecular mechanism, based on stabilization and endocytosis, to adjust surface levels of Crumbs, which are essential for maintaining epithelial polarity.

Saturday, December 5th

Talsma, A. D., Chaves, J. F., LaMonaca, A., Wieczorek, E. D. and Palladino, M. J. (2014).. Genome-wide screen for modifiers of Na+/K+ ATPase alleles identifies critical genetic loci. Mol Brain 7: 89. PubMed ID: 25476251 Summary: Mutations affecting the Na+/ K+ ATPase (a.k.a. the sodium-potassium pump) genes cause conditional locomotor phenotypes in flies and three distinct complex neurological diseases in humans. More than 50 mutations have been identified affecting the human ATP1A2 and ATP1A3 genes that are known to cause rapid-onset Dystonia Parkinsonism, familial hemiplegic migraine, alternating hemiplegia of childhood, and variants of familial hemiplegic migraine with neurological complications including seizures and various mood disorders. In flies, mutations affecting the Na pump α subunit gene have dramatic phenotypes including altered longevity, neural dysfunction, neurodegeneration, myodegeneration, and striking locomotor impairment. Locomotor defects can manifest as conditional bang-sensitive (BS) or temperature-sensitive (TS) paralysis: phenotypes well-suited for genetic screening. This study performed a genome-wide deficiency screen using three distinct missense alleles of ATPα and conditional locomotor function assays to identify novel modifier loci. A secondary screen confirmed allele-specificity of the interactions and many of the interactions were mapped to single genes and subsequently validated. Sixty-four modifier loci were identified, and classical mutations and RNAi were used to confirm 50 single gene interactions. The genes identified include those with known function, several with unknown function or that were otherwise uncharacterized, and many loci with no described association with locomotor or Na+/K+ ATPase function. In conclusion, this study has identified many critical regions and narrowed several of these to single genes. These data demonstrate there are many loci capable of modifying ATPα dysfunction, which may provide the basis for modifying migraine, locomotor and seizure dysfunction in animals.

Wu, Y., Gause, M., Xu, D., Misulovin, Z., Schaaf, C. A., Mosarla, R. C., Mannino, E., Shannon, M., Jones, E., Shi, M., Chen, W. F., Katz, O. L., Sehgal, A., Jongens, T. A., Krantz, I. D. and Dorsett, D. (2015). Drosophila Nipped-B mutants model Cornelia de Lange Syndrome in growth and behavior. PLoS Genet 11: e1005655. PubMed ID: 26544867 Summary: Individuals with Cornelia de Lange Syndrome (CdLS) display diverse developmental deficits, including slow growth, multiple limb and organ abnormalities, and intellectual disabilities. Severely-affected individuals most often have dominant loss-of-function mutations in the Nipped-B-Like (NIPBL) gene, and milder cases often have missense or in-frame deletion mutations in genes encoding subunits of the cohesin complex. Cohesin mediates sister chromatid cohesion to facilitate accurate chromosome segregation, and NIPBL is required for cohesin to bind to chromosomes. Individuals with CdLS, however, do not display overt cohesion or segregation defects. Rather, studies in human cells and model organisms indicate that modest decreases in NIPBL and cohesin activity alter the transcription of many genes that regulate growth and development. Sister chromatid cohesion factors, including the Nipped-B ortholog of human NIPBL, are also critical for gene expression and development in Drosophila melanogaster. This study describes how a modest reduction in Nipped-B activity alters growth and neurological function in Drosophila. These studies reveal that Nipped-B heterozygous mutant Drosophila show reduced growth, learning, and memory, and altered circadian rhythms. Importantly, the growth deficits are not caused by changes in systemic growth controls, but reductions in cell number and size attributable in part to reduced expression of myc (diminutive) and other growth control genes. The learning, memory and circadian deficits are accompanied by morphological abnormalities in brain structure. These studies confirm that Drosophila Nipped-B mutants provide a useful model for understanding CdLS, and provide new insights into the origins of birth defects.

Ruan, K., Zhu, Y., Li, C., Brazill, J.M. and Zhai, R.G. (2015). Alternative splicing of Drosophila Nmnat functions as a switch to enhance neuroprotection under stress. Nat Commun 6: 10057. PubMed ID: 26616331 Summary: Nicotinamide mononucleotide adenylyltransferase (NMNAT) is a conserved enzyme in the NAD synthetic pathway. It has also been identified as an effective and versatile neuroprotective factor. However, it remains unclear how healthy neurons regulate the dual functions of NMNAT and achieve self-protection under stress. This study shows that Drosophila Nmnat (DmNmnat) is alternatively spliced into two mRNA variants, RA and RB, which translate to protein isoforms with divergent neuroprotective capacities against spinocerebellar ataxia 1-induced neurodegeneration. Isoform PA/PC translated from RA is nuclear-localized with minimal neuroprotective ability, and isoform PB/PD translated from RB is cytoplasmic and has robust neuroprotective capacity. Under stress, RB is preferably spliced in neurons to produce the neuroprotective PB/PD isoforms. These results indicate that alternative splicing functions as a switch that regulates the expression of functionally distinct DmNmnat variants. Neurons respond to stress by driving the splicing switch to produce the neuroprotective variant and therefore achieve self-protection.

Wang, X., Perumalsamy, H., Kwon, H. W., Na, Y. E. and Ahn, Y. J. (2015). Effects and possible mechanisms of action of acacetin on the behavior and eye morphology of Drosophila models of Alzheimer's disease. Sci Rep 5: 16127. PubMed ID: 26530776 Summary: The human beta-amyloid (Aβ; see Drosophila Appl) cleaving enzyme (BACE-1) is a target for Alzheimer's disease (AD) treatments. This study was conducted to determine if acacetin extracted from the whole Agastache rugosa plant had anti-BACE-1 and behavioral activities in Drosophila AD models and to determine acacetin's mechanism of action. Acacetin rescued APP/BACE1-expressing flies and kept them from developing both eye morphology and behavioral defects. The rtPCR analysis revealed that acacetin reduced both the human APP and BACE-1 mRNA levels in the transgenic flies, suggesting that it plays an important role in the transcriptional regulation of human BACE-1 and APP. Western blot analysis revealed that acacetin reduced Aβ production by interfering with BACE-1 activity and APP synthesis, resulting in a decrease in the levels of the APP carboxy-terminal fragments and the APP intracellular domain. Therefore, the protective effect of acacetin on Aβ production is mediated by transcriptional regulation of BACE-1 and APP, resulting in decreased APP protein expression and BACE-1 activity. Acacetin also inhibited APP synthesis, resulting in a decrease in the number of amyloid plaques.

Chakraborty, M., Selma-Soriano, E., Magny, E., Couso, J. P., Perez-Alonso, M., Charlet-Berguerand, N., Artero, R. and Llamusi, B. (2015). Pentamidine rescues contractility and rhythmicity in a Drosophila model of myotonic dystrophy heart dysfunction. Dis Model Mech. PubMed ID: 26515653 Summary: Up to 80% of myotonic dystrophy type 1 (DM1) patients will develop cardiac abnormalities at some point during the progression of their disease. Despite its importance, very few animal model studies have focused on the heart dysfunction in DM1. This study describes the characterization of the heart phenotype in a Drosophila model expressing pure expanded CUG repeats under the control of the cardiomyocyte-specific driver GMH5-Gal4. Morphologically, expression of 250 CUG repeats caused abnormalities in the parallel alignment of the spiral myofibrils in dissected fly hearts revealed by phalloidin staining. Moreover, combined immunofluorescence and in situ hybridization of splice factor Muscleblind and CUG repeats, respectively, confirmed detectable ribonuclear foci and Muscleblind sequestration, characteristic features of DM1, exclusively in flies expressing the expanded CTG repeats. Similarly to what has been reported in human DM1 patients, heart-specific expression of toxic RNA resulted in reduced survival, increased arrhythmia, altered diastolic and systolic function and reduced heart tube diameters and contractility in the model flies. As a proof of concept that the fly heart model can be used for in vivo testing of promising therapeutic compounds, flies were fed with pentamidine, a compound previously described to improve DM1 phenotypes. Pentamidine not only released Muscleblind and reduced ribonuclear formation in the Drosophila heart but rescued heart arrhythmicity and contractility, and improved fly survival in animals expressing 250 CUG repeats.

Landry, G. M., Hirata, T., Anderson, J. B., Cabrero, P., Gallo, C. J., Dow, J. A. and Romero, M. F. (2015). Sulfate and thiosulfate inhibit oxalate transport via a dPrestin(mSlc26a6)-dependent mechanism in an insect model of calcium oxalate nephrolithiasis. Am J Physiol Renal Physiol: ajprenal 00406 02015. PubMed ID: 26538444 Summary: Nephrolithiasis is one of the most common urinary tract disorders with the majority of kidney stones composed of calcium oxalate (CaOx). Given its prevalence (US occurrence 10%), it is still poorly understood, lacking progress in identifying new therapies because of its complex etiology. Drosophila is a recently developed model of CaOx nephrolithiasis. Effects of sulfate and thiosulfate on crystal formation were investigated using the Drosophila model, as well as electrophysiological effects on both Drosophila (Slc26a5/6; dprestin) and mouse (mSlc26a6) oxalate transporters utilizing the Xenopus oocyte heterologous expression system. Results indicate that both transport thiosulfate with a much higher affinity than sulfate. Additionally, both compounds were effective at decreasing CaOx crystallization when added to the diet. However, these results were not observed when compounds were applied to MTs ex vivo. Neither compound affected CaOx crystallization in dPrestin knock down animals indicating a role for principal cell-specific dPrestin in luminal oxalate transport. Furthermore,thiosulfate has a higher affinity for dPrestin and mSlc26a6 compared to oxalate These data indicate that thiosulfate's ability to act as a competitive inhibitor of oxalate via dPrestin, can explain the decrease in CaOx crystallization seen in the presence of thiosulfate, but not sulfate. Overall, these findings predict that thiosulfate or oxalate-mimics may be effective as therapeutic competitive inhibitors of CaOx crystallization.

Friday, December 4th

Ulianov, S. V., et al. (2015). Active chromatin and transcription play a key role in chromosome partitioning into topologically associating domains. Genome Res [Epub ahead of print]. PubMed ID: 26518482 Summary: Recent advances enabled by the Hi-C technique have unraveled many principles of chromosomal folding that were subsequently linked to disease and gene regulation. In particular, Hi-C revealed that chromosomes of animals are organized into topologically associating domains (TADs), evolutionary conserved compact chromatin domains that influence gene expression. Mechanisms that underlie partitioning of the genome into TADs remain poorly understood. To explore principles of TAD folding in Drosophila melanogaster, Hi-C and poly(A)+ RNA-seq were performed in four cell lines of various origins (S2, Kc167, DmBG3-c2, and OSC). Contrary to previous studies, regions between TADs (i.e., the inter-TADs and TAD boundaries) in Drosophila were found to be only weakly enriched with the insulator protein dCTCF, while another insulator protein Su(Hw) was found to be preferentially present within TADs. However, Drosophila inter-TADs harbor active chromatin and constitutively transcribed (housekeeping) genes. Accordingly, binding of insulator proteins dCTCF and Su(Hw) was found to predict TAD boundaries much worse than active chromatin marks do. Interestingly, inter-TADs correspond to decompacted inter-bands of polytene chromosomes, whereas TADs mostly correspond to densely packed bands. Collectively, these results suggest that TADs are condensed chromatin domains depleted in active chromatin marks, separated by regions of active chromatin. The mechanism of TAD self-assembly is proposed based on the ability of nucleosomes from inactive chromatin to aggregate, and on the lack of this ability in acetylated nucleosomal arrays. Finally, this hypothesis was tested by polymer simulations and it was found that TAD partitioning may be explained by different modes of inter-nucleosomal interactions for active and inactive chromatin.

Zielke, T., Glotov, A. and Saumweber, H. (2015). High-resolution in situ hybridization analysis on the chromosomal interval 61C7-61C8 of Drosophila melanogaster reveals interbands as open chromatin domains. Chromosoma [Epub ahead of print]. PubMed ID: 26520107 Summary: Eukaryotic chromatin is organized in contiguous domains that differ in protein binding, histone modifications, transcriptional activity, and in their degree of compaction. Genome-wide comparisons suggest that, overall, the chromatin organization is similar in different cells within an organism. This study compared the structure and activity of the 61C7-61C8 interval in polytene and diploid cells of Drosophila. By in situ hybridization on polytene chromosomes combined with high-resolution microscopy, the boundaries were mapped of the 61C7-8 interband and of the 61C7 and C8 band regions, respectively. The results demonstrate that the 61C7-8 interband is significantly larger than estimated previously. This interband extends over 20 kbp and is in the range of the flanking band domains. It contains several active genes and therefore can be considered as an open chromatin domain. Comparing the 61C7-8 structure of Drosophila S2 cells and polytene salivary gland cells by ChIP for chromatin protein binding and histone modifications, a highly consistent domain structure was observed for the proximal 13 kbp of the domain in both cell types. However, the distal 7 kbp of the open domain differs in protein binding and histone modification between both tissues. The domain contains four protein-coding genes in the proximal part and two noncoding transcripts in the distal part. The differential transcriptional activity of one of the noncoding transcripts correlates with the observed differences in the chromatin structure between both tissues.

Eagen, K. P., Hartl, T. A. and Kornberg, R. D. (2015). Stable Chromosome Condensation Revealed by Chromosome Conformation Capture. Cell 163: 934-946. PubMed ID: 26544940 Summary: Chemical cross-linking and DNA sequencing have revealed regions of intra-chromosomal interaction, referred to as topologically associating domains (TADs), interspersed with regions of little or no interaction, in interphase nuclei. TADs and the regions between them were found to correspond with the bands and interbands of polytene chromosomes of Drosophila. Further, the conservation of TADs between polytene and diploid cells of Drosophila was established. From direct measurements on light micrographs of polytene chromosomes, the states of chromatin folding in the diploid cell nucleus was deduced. Two states of folding, fully extended fibers containing regulatory regions and promoters, and fibers condensed up to 10-fold containing coding regions of active genes, constitute the euchromatin of the nuclear interior. Chromatin fibers condensed up to 30-fold, containing coding regions of inactive genes, represent the heterochromatin of the nuclear periphery. A convergence of molecular analysis with direct observation thus reveals the architecture of interphase chromosomes.

Panikar, C. S., Rajpathak, S. N., Abhyankar, V., Deshmukh, S. and Deobagkar, D. D. (2015). Presence of DNA methyltransferase activity and CpC methylation in Drosophila melanogaster. Mol Biol Rep 42: 1615-1621. PubMed ID: 26547851 Summary: Flies lack DNMT1/DNMT3 based methylation machinery. Despite recent reports confirming the presence of low DNA methylation in Drosophila; little is known about the methyltransferase. This study investigated the possible functioning of DNA methyltransferase in Drosophila. The 14 K oligo microarray slide was incubated with native cell extract from adult Drosophila to check the presence of the methyltransferase activity. The methylated oligo sequences were then identified by the binding of anti 5-methylcytosine monoclonal antibody. Methylation sensitive restriction enzyme mediated PCR was used to assess the methylation at a few selected loci identified on the array. A few of the total oligos were seen to be methylated under the assay conditions. Evidence is provided for the presence of de novo methyltransferase activity and identification of its sequence specificity. Presence of CpC methylation was detected in the selected genomic regions. This study provides an innovative approach to investigate methylation specificity of a native methyltransferase.

Thursday, December 3rd

Jiang, T., McKinley, R. F., McGill, M. A., Angers, S. and Harris, T. J. (2015). A Par-1-Par-3-centrosome cell polarity pathway and its tuning for isotropic cell adhesion. Curr Biol 25: 2701-2708. PubMed ID: 26455305 Summary: To form regulated barriers between body compartments, epithelial cells polarize into apical and basolateral domains and assemble adherens junctions (AJs). Despite close links with polarity networks that generate single polarized domains, AJs distribute isotropically around the cell circumference for adhesion with all neighboring cells. How AJs avoid the influence of polarity networks to maintain their isotropy has been unclear. In established epithelia, trans cadherin interactions could maintain AJ isotropy, but AJs are dynamic during epithelial development and remodeling, and thus specific mechanisms may control their isotropy. In Drosophila, aPKC prevents hyper-polarization of junctions as epithelia develop from cellularization to gastrulation. This study shows that aPKC does so by inhibiting a positive feedback loop between Bazooka (Baz)/Par-3, a junctional organizer, and centrosomes. Without aPKC, Baz and centrosomes lose their isotropic distributions and recruit each other to single plasma membrane (PM) domains. Surprisingly, loss- and gain-of-function analyses show that the Baz-centrosome positive feedback loop is driven by Par-1, a kinase known to phosphorylate Baz and inhibit its basolateral localization. This study found that Par-1 promotes the positive feedback loop through both centrosome microtubule effects and Baz phosphorylation. Normally, aPKC attenuates the circuit by expelling Par-1 from the apical domain at gastrulation. The combination of local activation and global inhibition is a common polarization strategy. Par-1 seems to couple both effects for a potent Baz polarization mechanism that is regulated for the isotropy of Baz and AJs around the cell circumference.

Tiwari, P., Kumar, A., Das, R. N., Malhotra, V. and VijayRaghavan, K. (2015). A tendon cell specific RNAi screen reveals novel candidates essential for muscle tendon interaction. PLoS One 10: e0140976. PubMed ID: 26488612 Summary: Tendons are fibrous connective tissue which connect muscles to the skeletal elements thus acting as passive transmitters of force during locomotion and provide appropriate body posture. Tendon-derived cues, albeit poorly understood, are necessary for proper muscle guidance and attachment during development. This study used dorsal longitudinal muscles of Drosophila and their tendon attachment sites to unravel the molecular nature of interactions between muscles and tendons. A genetic screen using RNAi-mediated knockdown in tendon cells was performed to find out molecular players involved in the formation and maintenance of myotendinous junction; 21 candidates were found out of 2507 RNAi lines screened. Of these, 19 were novel molecules in context of myotendinous system. Integrin-βPS and Talin, picked as candidates in this screen, are known to play important role in the cell-cell interaction and myotendinous junction formation validating the screen. Candidates were found with enzymatic function, transcription activity, cell adhesion, protein folding and intracellular transport function. Tango1, an ER exit protein involved in collagen secretion was identified as a candidate molecule involved in the formation of myotendinous junction. Tango1 knockdown was found to affect development of muscle attachment sites and formation of myotendinous junction. Tango1 was also found to be involved in secretion of Viking (Collagen type IV) and BM-40 from hemocytes and fat cells.

Hosono, C., Matsuda, R., Adryan, B. and Samakovlis, C. (2015). Transient junction anisotropies orient annular cell polarization in the Drosophila airway tubes. Nat Cell Biol 17: 1569-1576. PubMed ID: 26551273 Summary: Tubular organs exhibit a striking orientation of landmarks according to the physical anisotropy of the 3D shape, in addition to planar cell polarization. However, the influence of 3D tissue topography on the constituting cells remains underexplored. This study identified a regulatory network polarizing cellular biochemistry according to the physical anisotropy of the 3D tube geometry (tube cell polarization) by a genome-wide, tissue-specific RNAi screen. During Drosophila airway remodelling, each apical cellular junction is equipotent to establish perpendicular actomyosin cables, irrespective of the longitudinal or transverse tube axis. A dynamic transverse enrichment of atypical protein kinase C (aPKC) shifts the balance and transiently targets activated small GTPase RhoA, myosin phosphorylation and Rab11 vesicle trafficking to longitudinal junctions. It is proposed that the PAR complex translates tube physical anisotropy into longitudinal junctional anisotropy, where cell-cell communication aligns the contractile cytoskeleton of neighbouring cells.

Flores-Benitez, D. and Knust, E. (2015). Crumbs is an essential regulator of cytoskeletal dynamics and cell-cell adhesion during dorsal closure in Drosophila. Elife 4 [Epub ahead of print]. PubMed ID: 26544546 Summary: The evolutionarily conserved Crumbs protein is required for epithelial polarity and morphogenesis. This study identified a novel role of Crumbs as a negative regulator of actomyosin dynamics during dorsal closure in the Drosophila embryo. Embryos carrying a mutation in the FERM (protein 4.1/ezrin/radixin/moesin) domain-binding motif of Crumbs die due to an overactive actomyosin network associated with disrupted adherens junctions. This phenotype is restricted to the amnioserosa and does not affect other embryonic epithelia. This function of Crumbs requires DMoesin, the Rho1-GTPase, class-I p21-activated kinases and the Arp2/3 complex. Data presented here point to a critical role of Crumbs in regulating actomyosin dynamics, cell junctions and morphogenesis.

Wednesday, December 2nd

Nie, Y., Li, Q., Amcheslavsky, A., Duhart, J. C., Veraksa, A., Stocker, H., Raftery, L. A. and Ip, Y. T. (2015). Bunched and Madm function downstream of Tuberous sclerosis complex to regulate the growth of intestinal stem cells in Drosophila. Stem Cell Rev. PubMed ID: 26323255 Summary: The Drosophila adult midgut contains intestinal stem cells that support homeostasis and repair. This study shows that the leucine zipper protein Bunched and the adaptor protein Madm are novel regulators of intestinal stem cells. MARCM mutant clonal analysis and cell type specific RNAi revealed that Bunched and Madm were required within intestinal stem cells for proliferation. Transgenic expression of a tagged Bunched showed a cytoplasmic localization in midgut precursors, and the addition of a nuclear localization signal to Bunched reduced its function to cooperate with Madm to increase intestinal stem cell proliferation. Furthermore, the elevated cell growth and 4EBP phosphorylation phenotypes induced by loss of Tuberous Sclerosis Complex or overexpression of Rheb were suppressed by the loss of Bunched or Madm. Therefore, while the mammalian homolog of Bunched, TSC-22, is able to regulate transcription and suppress cancer cell proliferation, these data suggest the model that Bunched and Madm functionally interact with the TOR pathway in the cytoplasm to regulate the growth and subsequent division of intestinal stem cells.

Bailey, A. P., Koster, G., Guillermier, C., Hirst, E. M., MacRae, J. I., Lechene, C. P., Postle, A. D. and Gould, A. P. (2015). Antioxidant Role for lipid droplets in a stem cell niche of Drosophila. Cell 163: 340-353. PubMed ID: 26451484 Summary: Stem cells reside in specialized microenvironments known as niches. During Drosophila development, glial cells provide a niche that sustains the proliferation of neural stem cells (neuroblasts) during starvation. This study finds that the glial cell niche also preserves neuroblast proliferation under conditions of hypoxia and oxidative stress. Lipid droplets that form in niche glia during oxidative stress limit the levels of reactive oxygen species (ROS) and inhibit the oxidation of polyunsaturated fatty acids (PUFAs). These droplets protect glia and also neuroblasts from peroxidation chain reactions that can damage many types of macromolecules. The underlying antioxidant mechanism involves diverting PUFAs, including diet-derived linoleic acid, away from membranes to the core of lipid droplets, where they are less vulnerable to peroxidation. This study reveals an antioxidant role for lipid droplets that could be relevant in many different biological contexts.

Liu, Z., Zhong, G., Chai, P. C., Luo, L., Liu, S., Yang, Y., Baeg, G. H. and Cai, Y. (2015). Coordinated niche-associated signals promote germline homeostasis in the Drosophila ovary. J Cell Biol 211: 469-484. PubMed ID: 26504174 Summary: Stem cell niches provide localized signaling molecules to promote stem cell fate and to suppress differentiation. The Drosophila melanogaster ovarian niche is established by several types of stromal cells, including terminal filament cells, cap cells, and escort cells (ECs). This study shows that, in addition to its well-known function as a niche factor expressed in cap cells, the Drosophila transforming growth factor β molecule Decapentaplegic (Dpp) is expressed at a low level in ECs to maintain a pool of partially differentiated germline cells that may dedifferentiate to replenish germline stem cells upon their depletion under normal and stress conditions. This study further reveals that the Dpp level in ECs is modulated by Hedgehog (Hh) ligands, which originate from both cap cells and ECs. Hh signaling exerts its function by suppressing Janus kinase/signal transducer activity, which promotes Dpp expression in ECs. Collectively, these data suggest a complex interplay of niche-associated signals that controls the development of a stem cell lineage.

Fu, Z., Geng, C., Wang, H., Yang, Z., Weng, C., Li, H., Deng, L., Liu, L., Liu, N., Ni, J. and Xie, T. (2015). Twin promotes the maintenance and differentiation of germline stem cell lineage through modulation of multiple pathways. Cell Rep 13: 1366-1379. PubMed ID: 26549449 Summary: The central question in stem cell regulation is how the balance between self-renewal and differentiation is controlled at the molecular level. This study uses germline stem cells (GSCs) in the Drosophila ovary to demonstrate that the Drosophila CCR4 homolog Twin is required intrinsically to promote both GSC self-renewal and progeny differentiation. Twin/CCR4 is one of the two catalytic subunits in the highly conserved CCR4-NOT mRNA deadenylase complex. Twin works within the CCR4-NOT complex to intrinsically maintain GSC self-renewal, at least partly by sustaining E-cadherin-mediated GSC-niche interaction and preventing transposable element-induced DNA damage. It promotes GSC progeny differentiation by forming protein complexes with differentiation factors Bam and Bgcn independently of other CCR4-NOT components. Interestingly, Bam can competitively inhibit the association of Twin with Pop2 in the CCR4-NOT complex. Therefore, this study demonstrates that Twin has important intrinsic roles in promoting GSC self-renewal and progeny differentiation by functioning in different protein complexes.

Tuesday, December 1st

Seidner, G., Robinson, J. E., Wu, M., Worden, K., Masek, P., Roberts, S. W., Keene, A. C. and Joiner, W. J. (2015). Identification of neurons with a privileged role in sleep homeostasis in Drosophila melanogaster. Curr Biol 25: 2928-2938. PubMed ID: 26526372 Summary: Sleep is thought to be controlled by two main processes: a circadian clock that primarily regulates sleep timing and a homeostatic mechanism that detects and responds to sleep need. Whereas abundant experimental evidence suggests that sleep need increases with time spent awake, the contributions of different brain arousal systems have not been assessed independently of each other to determine whether certain neural circuits, rather than waking per se, selectively contribute to sleep homeostasis. This study found that flies sustained thermogenetic activation of three independent neurotransmitter systems promoted nighttime wakefulness. However, only sleep deprivation resulting from activation of cholinergic neurons was sufficient to elicit subsequent homeostatic recovery sleep, as assessed by multiple behavioral criteria. In contrast, sleep deprivation resulting from activation of octopaminergic neurons suppressed homeostatic recovery sleep, indicating that wakefulness can be dissociated from accrual of sleep need. Neurons that promote sleep homeostasis were found to innervate the central brain and motor control regions of the thoracic ganglion. Blocking activity of these neurons suppressed recovery sleep but did not alter baseline sleep, further differentiating between neural control of sleep homeostasis and daily fluctuations in the sleep/wake cycle. Importantly, selective activation of wake-promoting neurons without engaging the sleep homeostat impaired subsequent short-term memory, thus providing evidence that neural circuits that regulate sleep homeostasis are important for behavioral plasticity. Together, these data suggest a neural circuit model involving distinct populations of wake-promoting neurons, some of which are involved in homeostatic control of sleep and cognition.

Takemura, S. Y., et al. (2015). Synaptic circuits and their variations within different columns in the visual system of Drosophila. Journal-Proc Natl Acad Sci U S A 112: 13711-13716. PubMed ID: 26483464 Summary: This study reconstructed the synaptic circuits of seven columns in the second neuropil or medulla behind the fly's compound eye. These neurons embody some of the most stereotyped circuits in one of the most miniaturized of animal brains. The reconstructions allow study of variations between circuits in the medulla's neighboring columns. This variation in the number of synapses and the types of their synaptic partners has previously been little addressed because methods that visualize multiple circuits have not resolved detailed connections, and existing connectomic studies, which can see such connections, have not so far examined multiple reconstructions of the same circuit. This study addresses the omission by comparing the circuits common to all seven columns to assess variation in their connection strengths and the resultant rates of several different and distinct types of connection error. Error rates reveal that, overall, :lt;1% of contacts are not part of a consensus circuit, and those contacts that supplement (E+) or are missing from it (E-) were classified. Autapses, in which the same cell is both presynaptic and postsynaptic at the same synapse, are occasionally seen; two cells in particular, Dm9 and Mi1, form >/=20-fold more autapses than do other neurons. These results delimit the accuracy of developmental events that establish and normally maintain synaptic circuits with such precision, and thereby address the operation of such circuits. They also establish a precedent for error rates that will be required in the new science of connectomics.

Stevens, C. F. (2015). What the fly's nose tells the fly's brain. Proc Natl Acad Sci U S A 112: 9460-9465. PubMed ID: 26150492 Summary: The fly olfactory system has a three-layer architecture: The fly's olfactory receptor neurons send odor information to the first layer (the encoder) where this information is formatted as combinatorial odor code, one which is maximally informative, with the most informative neurons firing fastest. This first layer then sends the encoded odor information to the second layer (decoder), which consists of about 2,000 neurons that receive the odor information and "break" the code. For each odor, the amplitude of the synaptic odor input to the 2,000 second-layer neurons is approximately normally distributed across the population, which means that only a very small fraction of neurons receive a large input. Each odor, however, activates its own population of large-input neurons and so a small subset of the 2,000 neurons serves as a unique tag for the odor. Strong inhibition prevents most of the second-stage neurons from firing spikes, and therefore spikes from only the small population of large-input neurons is relayed to the third stage. This selected population provides the third stage (the user) with an odor label that can be used to direct behavior based on what odor is present (Stevens, 2015).

Kim, H., Choi, M. S., Kang, K. and Kwon, J. Y. (2015). Behavioral analysis of bitter taste perception in Drosophila larvae. Chem Senses [Epub ahead of print]. PubMed ID: 26512069 Summary: Insect larvae, which recognize food sources through chemosensory cues, are a major source of global agricultural loss. Gustation is an important factor that determines feeding behavior, and the gustatory receptors (Grs) act as molecular receptors that recognize diverse chemicals in gustatory receptor neurons (GRNs). The behavior of Drosophila larvae is relatively simpler than the adult fly, and a gustatory receptor-to-neuron map was established in a previous study of the major external larval head sensory organs. This study extensively examined The bitter taste responses of larvae using 2-choice behavioral assays. First, a panel of 23 candidate bitter compounds was tested to compare the behavioral responses of larvae and adults. Nine bitter compounds were tested, which elicit aversive behavior in a dose-dependent manner. A functional map of the larval GRNs was constructed with the use of Gr-GAL4 lines that drive expression of UAS-tetanus toxin and UAS-VR1 in specific gustatory neurons to identify bitter tastants-GRN combinations by suppressing and activating discrete subsets of taste neurons, respectively. The results suggest that many gustatory neurons act cooperatively in larval bitter sensing, and that these neurons have different degrees of responsiveness to different bitter compounds.

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