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


Wednesday, January 31st

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Steiner, C., Bozzolan, F., Montagne, N., Maibeche, M. and Chertemps, T. (2017). Neofunctionalization of "Juvenile Hormone Esterase Duplication" in Drosophila as an odorant-degrading enzyme towards food odorants. Sci Rep 7(1): 12629. PubMed ID: 28974761
Odorant degrading enzymes (ODEs) are thought to be responsible, at least in part, for olfactory signal termination in the chemosensory system by rapid degradation of odorants in the vicinity of the receptors. A carboxylesterase, specifically expressed in Drosophila antennae, called "juvenile hormone esterase duplication (JHEdup)" has been previously reported to hydrolyse different fruit esters in vitro. This study functionally characterized JHEdup in vivo. The jhedup gene is highly expressed in large basiconic sensilla, housed in the the maxillary palps, that have been reported to detect several food esters. An electrophysiological analysis demonstrates that ab1A olfactory neurons of jhedup mutant flies exhibit an increased response to certain food acetates. Furthermore, mutant flies show a higher sensitivity towards the same odorants in behavioural assays. A phylogenetic analysis reveals that jhedup arose as a duplication of the juvenile hormone esterase gene during the evolution of Diptera, most likely in the ancestor of Schizophora, and has been conserved in all the 12 sequenced Drosophila species. Jhedup exhibits also an olfactory-predominant expression pattern in other Drosophila species. These results support the implication of JHEdup in the degradation of food odorants in D. melanogaster and propose a neofunctionalization of this enzyme as a bona fide ODE in Drosophilids.
Shimada, S., Oosaki, M., Takahashi, R., Uene, S., Yanagisawa, S., Tsukihara, T. and Shinzawa-Itoh, K. (2017). A unique respiratory adaptation in Drosophila independent of supercomplex formation. Biochim Biophys Acta. PubMed ID: 29191512
Large assemblies of respiratory chain complexes, known as supercomplexes, are present in the mitochondrial membrane. The formation of supercomplexes is thought to contribute to efficient electron transfer, stabilization of each enzyme complex, and inhibition of reactive oxygen species (ROS) generation. In this study, mitochondria from various organisms were solubilized with digitonin, and then the solubilized complexes were separated by blue native PAGE (BN-PAGE). The results revealed a supercomplex consisting of complexes I, III, and IV in mitochondria from bovine and porcine heart, and a supercomplex consisting primarily of complexes I and III in mitochondria from mouse heart and liver. However, supercomplexes were barely detectable in Drosophila flight-muscle mitochondria, and only dimeric complex V was present. Drosophila mitochondria exhibited the highest rates of oxygen consumption and NADH oxidation, and the concentrations of the electron carriers, cytochrome c and quinone were higher than in other species. Respiratory chain complexes were tightly packed in the mitochondrial membrane containing abundant phosphatidylethanol-amine with the fatty acid palmitoleic acid (C16:1), which is relatively high oxidation-resistant as compared to poly-unsaturated fatty acid. These properties presumably allow efficient electron transfer in Drosophila. These findings reveal the existence of a new mechanism of biological adaptation independent of supercomplex formation.
Musselman, L. P., Fink, J. L., Grant, A. R., Gatto, J. A., Tuthill, B. F., and Baranski, T. J. (2017). The relationship between immunity and metabolism in Drosophila diet-induced insulin resistance. Mol Cell Biol [Epub ahead of print]. PubMed ID: 29084810
This study analyzed insulin signaling in the fat body using loss- and gain-of-function. When expression of the sole Drosophila Insulin receptor (InR) was reduced in larval fat bodies, animals exhibited developmental delay and reduced size in a diet-dependent manner. Fat body InR knockdown also led to reduced survival on high-sugar diets. To look downstream of InR at potential mediators of insulin resistance, RNA-seq studies in insulin-resistant fat bodies revealed differential expression of genes, including those involved in innate immunity. Obesity-associated insulin resistance led to increased susceptibility of flies to infection, as in humans. Reduced innate immunity was dependent on fat body InR expression. The peptidoglycan recognition proteins (PGRPs) PGRP-SB2 and PGRP-SC2 were selected for further study based on differential expression studies. Downregulating PGRP-SB2 selectively in the fat body protected animals from the deleterious effects of overnutrition, whereas downregulating PGRP-SC2 produced InR-like phenotypes. These studies extend earlier work linking the immune and insulin signaling pathways and identify new targets of insulin signaling that could serve as potential drug targets to treat type 2 diabetes.
Park, J. and Carlson, J. R. (2017). Physiological responses of the Drosophila labellum to amino acids. J Neurogenet: 1-10. PubMed ID: 29191065
This study has systematically studied the physiological responses elicited by amino acids from the principal taste organ of the Drosophila head. Although the detection and coding of sugars and bitter compounds have been examined extensively in this organism, little attention has been paid to the physiology of amino acid taste. One class of sensilla, the labellar basiconic S sensilla, were found to yield the strongest responses to amino acids, although these responses were much weaker than the most robust responses to sugar or bitter compounds. S sensilla are heterogeneous in their amino acid responses and amino acids differ in the responses they elicit from individual sensilla. Tryptophan elicited relatively strong responses from S sensilla and these responses were eliminated when bitter-sensing neurons were ablated. Although tryptophan yielded little if any response in a behavioral paradigm, phenylalanine elicited a relatively strong response in the same paradigm and had a different physiological profile, supporting the notion that different amino acids are differentially encoded by the repertoire of taste neurons.
Stefana, M. I., Driscoll, P. C., Obata, F., Pengelly, A. R., Newell, C. L., MacRae, J. I. and Gould, A. P. (2017). Developmental diet regulates Drosophila lifespan via lipid autotoxins. Nat Commun 8(1): 1384. PubMed ID: 29123106
Early-life nourishment exerts long-term influences upon adult physiology and disease risk. These lasting effects of diet are well established but the underlying mechanisms are only partially understood. This study shows that restricting dietary yeast during Drosophila development can, depending upon the subsequent adult environment, more than double median lifespan. Developmental diet acts via a long-term influence upon the adult production of toxic molecules, which are termed autotoxins, that are shed into the environment and shorten the lifespan of both sexes. Autotoxins are synthesised by oenocytes and some of them correspond to alkene hydrocarbons that also act as pheromones. This study identifies a mechanism by which the developmental dietary history of an animal regulates its own longevity and that of its conspecific neighbours. It also has important implications for the design of lifespan experiments as autotoxins can influence the regulation of longevity by other factors including diet, sex, insulin signalling and population density.
Marxreiter, S. and Thummel, C. S. (2017). Adult functions for the Drosophila DHR78 nuclear receptor. Dev Dyn 247(2):315-32. PubMed ID: 29171103
The Testicular Receptors 2 and 4 (TR2, TR4) comprise a small subfamily of orphan nuclear receptors. Genetic studies in mouse models have identified roles for TR4 in developmental progression, fertility, brain development, and metabolism, as well as genetic redundancy with TR2. This paper describes a study the adult functions of the single Drosophila member of this subfamily, DHR78, with the goal of defining its ancestral functions in the absence of genetic redundancy. DHR78 mutants have a shortened lifespan, reduced motility, and mated DHR78 mutant females display a reduced feeding rate. Transcriptional profiling reveals a major role for DHR78 in promoting the expression of genes that are expressed in the midgut, suggesting that it contributes to nutrient uptake. Roles were identified for DHR78 in maintaining the expression of genes in the ecdysone and Notch signaling pathways. This study provides a new context for linking the molecular activity of the TR orphan nuclear receptors with their complex roles in adult physiology and lifespan.
Musso, P. Y., Lampin-Saint-Amaux, A., Tchenio, P. and Preat, T. (2017). Ingestion of artificial sweeteners leads to caloric frustration memory in Drosophila. Nat Commun 8(1): 1803. PubMed ID: 29180783
Non-caloric artificial sweeteners (NAS) are widely used in modern human food, raising the question about their health impact. This study asked whether NAS consumption is a neutral experience at neural and behavioral level, or if NAS can be interpreted and remembered as negative experience. Behavioral and imaging approaches were used to demonstrate that Drosophila melanogaster learn the non-caloric property of NAS through post-ingestion process. These results show that sweet taste is predictive of an energy value, and its absence leads to the formation of what we call Caloric Frustration Memory (CFM) that devalues the NAS or its caloric enantiomer. CFM formation involves activity of the associative memory brain structure, the mushroom bodies (MBs). In vivo calcium imaging of MB-input dopaminergic neurons that respond to sugar showed a reduced response to NAS after CFM formation. Altogether, these findings demonstrate that NAS are a negative experience for the brain.
Sujkowski, A., Ramesh, D., Brockmann, A. and Wessells, R. (2017). Octopamine drives endurance exercise adaptations in Drosophila. Cell Rep 21(7): 1809-1823. PubMed ID: 29141215
Endurance exercise is an effective therapeutic intervention with substantial pro-healthspan effects. Male Drosophila respond to a ramped daily program of exercise by inducing conserved physiological responses similar to those seen in mice and humans. Female flies respond to an exercise stimulus but do not experience the adaptive training response seen in males. This study used female flies as a model to demonstrate that differences in exercise response are mediated by differences in neuronal activity. The activity of octopaminergic neurons is specifically required to induce the conserved cellular and physiological changes seen following endurance training. Furthermore, either intermittent, scheduled activation of octopaminergic neurons or octopamine feeding is able to fully substitute for exercise, conferring a suite of pro-healthspan benefits to sedentary Drosophila. These experiments indicate that octopamine is a critical mediator of adaptation to endurance exercise in Drosophila.

Tuesday, January 30th

Pinto, B. S. and Orr-Weaver, T. L. (2017). Drosophila protein phosphatases 2A B' Wdb and Wrd regulate meiotic centromere localization and function of the MEI-S332 Shugoshin. Proc Natl Acad Sci U S A. PubMed ID: 29158400
Proper segregation of chromosomes in meiosis is essential to prevent miscarriages and birth defects. This requires that sister chromatids maintain cohesion at the centromere as cohesion is released on the chromatid arms when the homologs segregate at anaphase I. The Shugoshin proteins preserve centromere cohesion by protecting the cohesin complex from cleavage, and this has been shown in yeasts to be mediated by recruitment of the protein phosphatase 2A B' (PP2A B'). In metazoans, delineation of the role of PP2A B' in meiosis has been hindered by its myriad of other essential roles. The Drosophila Shugoshin MEI-S332 can bind directly to both of the B' regulatory subunits of PP2A, Wdb and Wrd, in yeast two-hybrid experiments. Exploiting experimental advantages of Drosophila spermatogenesis, this study found that the Wdb subunit localizes first along chromosomes in meiosis I, becoming restricted to the centromere region as MEI-S332 binds. Wdb and MEI-S332 show colocalization at the centromere region until release of sister-chromatid cohesion at the metaphase II/anaphase II transition. MEI-S332 is necessary for Wdb localization, but, additionally, both Wdb and Wrd are required for MEI-S332 localization. Thus, rather than MEI-S332 being hierarchical to PP2A B', these proteins reciprocally ensure centromere localization of the complex. Functional relationships between MEI-S332 and the two forms of PP2A were analyzed by quantifying meiotic chromosome segregation defects in double or triple mutants. These studies revealed that both Wdb and Wrd contribute to MEI-S332's ability to ensure accurate segregation of sister chromatids, but, as in centromere localization, they do not act solely downstream of MEI-S332.
Nippe, O. M., Wade, A. R., Elliott, C. J. H. and Chawla, S. (2017). Circadian rhythms in visual responsiveness in the behaviorally arrhythmic Drosophila clock mutant Clk(Jrk). J Biol Rhythms 32(6):583-592. PubMed ID: 29172879
An organism's biological day is characterized by a pattern of anticipatory physiological and behavioral changes that are governed by circadian clocks to align with the 24-h cycling environment. This study used flash electroretinograms (ERGs) and steady-state visually evoked potentials (SSVEPs) to examine how visual responsiveness in wild-type Drosophila melanogaster and the circadian clock mutant ClkJrk varies over circadian time. The ERG parameters of wild-type flies vary over the circadian day, with a higher luminance response during the subjective night. The SSVEP response that assesses contrast sensitivity also showed a time-of-day dependence, including 2 prominent peaks within a 24-h period and a maximal response at the end of the subjective day, indicating a tradeoff between luminance and contrast sensitivity. Moreover, the behaviorally arrhythmic ClkJrk mutants maintained a circadian profile in both luminance and contrast sensitivity, but unlike the wild-types, which show bimodal profiles in their visual response, ClkJrk flies show a weakening of the bimodal character, with visual responsiveness tending to peak once a day. It is concluded that the ClkJrk mutation mainly affects 1 of 2 functionally coupled oscillators and that the visual system is partially separated from the locomotor circadian circuits that drive bouts of morning and evening activity. As light exposure is a major mechanism for entrainment, this work suggests that a detailed temporal analysis of electrophysiological responses is warranted to better identify the time window at which circadian rhythms are most receptive to light-induced phase shifting.
O'Farrell, F., Lobert, V. H., Sneeggen, M., Jain, A., Katheder, N. S., Wenzel, E. M., Schultz, S. W., Tan, K. W., Brech, A., Stenmark, H. and Rusten, T. E. (2017). Class III phosphatidylinositol-3-OH kinase controls epithelial integrity through endosomal LKB1 regulation. Nat Cell Biol 19(12): 1412-1423. PubMed ID: 29084199
The molecular mechanisms underlying the interdependence between intracellular trafficking and epithelial cell polarity are poorly understood. This study shows that inactivation of class III phosphatidylinositol-3-OH kinase (CIII-PI3K), which produces phosphatidylinositol-3-phosphate (PtdIns3P) on endosomes, disrupts epithelial organization. This is caused by dysregulation of endosomally localized LKB1, also known as STK11, which shows delocalized and increased activity accompanied by dysplasia-like growth and invasive behaviour of cells provoked by JNK pathway activation. CIII-PI3K inactivation cooperates with Ras(V12) to promote tumour growth in vivo in an LKB1-dependent manner. Strikingly, co-depletion of LKB1 reverts these phenotypes and restores epithelial integrity. The endosomal, but not autophagic, function of CIII-PI3K controls polarity. The CIII-PI3K effector, WD repeat and FYVE domain-containing 2 (WDFY2), was identified an LKB1 regulator in Drosophila tissues and human organoids. Thus, this study defines a CIII-PI3K-regulated endosomal signalling platform from which LKB1 directs epithelial polarity, the dysregulation of which endows LKB1 with tumour-promoting properties.
Rajan, A., Housden, B. E., Wirtz-Peitz, F., Holderbaum, L. and Perrimon, N. (2017). A mechanism coupling systemic energy sensing to adipokine secretion. Dev Cell 43(1): 83-98.e86. PubMed ID: 29017032
Adipocytes sense systemic nutrient status and systemically communicate this information by releasing adipokines. The mechanisms that couple nutritional state to adipokine release are unknown. This study investigated how Unpaired 2 (Upd2), a structural and functional ortholog of the primary human adipokine leptin, is released from Drosophila fat cells. Golgi reassembly stacking protein (GRASP), an unconventional secretion pathway component, is required for Upd2 secretion. In nutrient-rich fat cells, GRASP clusters in close proximity to the apical side of lipid droplets (LDs). During nutrient deprivation, glucagon-mediated increase in calcium (Ca(2+)) levels, via calmodulin kinase II (CaMKII) phosphorylation, inhibits proximal GRASP localization to LDs. Using a heterologous cell system, human leptin secretion was also shown to be regulated by Ca(2+) and CaMKII. In summary, a mechanism is described by which increased cytosolic Ca(2+) negatively regulates adipokine secretion and has uncovered an evolutionarily conserved molecular link between intracellular Ca(2+) levels and energy homeostasis.
Perez-Mockus, G., Mazouni, K., Roca, V., Corradi, G., Conte, V. and Schweisguth, F. (2017). Spatial regulation of contractility by Neuralized and Bearded during furrow invagination in Drosophila. Nat Commun 8(1): 1594. PubMed ID: 29150614
Embryo-scale morphogenesis arises from patterned mechanical forces. During Drosophila gastrulation, actomyosin contractility drives apical constriction in ventral cells, leading to furrow formation and mesoderm invagination. It remains unclear whether and how mechanical properties of the ectoderm influence this process. This study shows that Neuralized (Neur), an E3 ubiquitin ligase active in the mesoderm, regulates collective apical constriction and furrow formation. Conversely, the Bearded (Brd) proteins antagonize maternal Neur and lower medial-apical contractility in the ectoderm: in Brd-mutant embryos, the ventral furrow invaginates properly but rapidly unfolds as medial MyoII levels increase in the ectoderm. Increasing contractility in the ectoderm via activated Rho similarly triggers furrow unfolding whereas decreasing contractility restores furrow invagination in Brd-mutant embryos. Thus, the inhibition of Neur by Brd in the ectoderm differentiates the mechanics of the ectoderm from that of the mesoderm and patterns the activity of MyoII along the dorsal-ventral axis.
Ren, S., Huang, Z., Jiang, Y. and Wang, T. (2017). dTBC1D7 regulates systemic growth independently of TSC through insulin signaling. J Cell Biol [Epub ahead of print]. PubMed ID: 29187524
The insulin signaling pathway plays key roles in systemic growth. TBC1D7 has recently been identified as the third subunit of the tuberous sclerosis complex (TSC), a negative regulator of cell growth. This study used Drosophila as a model system to dissect the physiological function of TBC1D7 in vivo. In mutants lacking TBC1D7, cell and organ growth were promoted, and TBC1D7 limited cell growth in a cell-nonautonomous and TSC-independent manner. TBC1D7 is specifically expressed in insulin-producing cells in the fly brain and regulated biosynthesis and release of insulin-like peptide 2, leading to systemic growth. Furthermore, animals carrying the dTBC1D7 mutation were hypoglycemic, short-lived, and sensitive to oxidative stress. These findings provide new insights into the physiological function of TBC1D7 in the systemic control of growth, as well as insights into human disorders caused by TBC1D7 mutation.

Monday, January 29th

Seugnet, L., Dissel, S., Thimgan, M., Cao, L. and Shaw, P. J. (2017). Identification of genes that maintain behavioral and structural plasticity during sleep loss. Front Neural Circuits 11: 79. PubMed ID: 29109678
Although patients with primary insomnia experience sleep disruption, they are able to maintain normal performance on a variety of cognitive tasks. This observation suggests that insomnia may be a condition where predisposing factors simultaneously increase the risk for insomnia and also mitigate against the deleterious consequences of waking. To gain insight into processes that might regulate sleep and buffer neuronal circuits during sleep loss, three genes, fat facet (faf), highwire (hiw) and the GABA receptor Resistance to dieldrin (Rdl), were manipulated that were differentially modulated in a Drosophila model of insomnia. The results indicate that increasing faf and decreasing hiw or Rdl within wake-promoting large ventral lateral clock neurons (lLNvs) induces sleep loss. As expected, sleep loss induced by decreasing hiw in the lLNvs results in deficits in short-term memory and increases of synaptic growth. However, sleep loss induced by knocking down Rdl in the lLNvs protects flies from sleep-loss induced deficits in short-term memory and increases in synaptic markers. Surprisingly, decreasing hiw and Rdl within the Mushroom Bodies (MBs) protects against the negative effects of sleep deprivation (SD) as indicated by the absence of a subsequent homeostatic response, or deficits in short-term memory. Together these results indicate that specific genes are able to disrupt sleep and protect against the negative consequences of waking in a circuit dependent manner.
Song, Q., Feng, G., Zhang, J., Xia, X., Ji, M., Lv, L. and Ping, Y. (2017). NMDA receptor-mediated Ca2+ influx in the absence of Mg2+ block disrupts rest:activity rhythms in Drosophila. Sleep 40(12). PubMed ID: 29029290
The correlated activation of pre- and postsynaptic neurons is essential for the NMDA receptor-mediated Ca2+ influx by removing Mg2+ from block site and NMDA receptors have been implicated in phase resetting of circadian clocks. So this study assessed rest:activity rhythms in Mg2+ block defective animals. Using Drosophila locomotor monitoring system, circadian rest:activity rhythms of different mutants were checked under constant darkness (DD) and light:dark (LD) conditions. Mg2+ block defective mutant flies were found to exhibit completely arrhythmic under DD. To further understand the role of Mg2+ block in daily circadian rest:activity, the mutant flies were observed under LD cycles, and severely reduced morning anticipation and advanced evening peak compared to control flies. Tissue-specific expression of Mg2+ block defective NMDA receptors was used, and pigment-dispersing factor receptor (PDFR) expressing circadian neurons were implicated in mediating the circadian rest:activity deficits. Endogenous functional NMDA receptors are expressed in most Drosophila neurons, including in a subgroup of dorsal neurons (DN1s). Subsequently, it was determined that the uncorrelated extra Ca2+ influx may act in part through Ca2+/Calmodulin (CaM)-stimulated PDE1c pathway leading to morning behavior phenotypes. These results demonstrate that Mg2+ block of NMDA receptors at resting potential is essential for the daily circadian rest:activity and it is proposes that Mg2+ block functions to suppress CaM-stimulated PDE1c activation at resting potential, thus regulating Ca2+ and cAMP oscillations in circadian and sleep circuits.
Saxena, N., Natesan, D. and Sane, S. P. (2017). Odor source localization in complex visual environments by fruit flies. J Exp Biol. PubMed ID: 29146771
Flying insects routinely forage in complex and cluttered sensory environments. Their search for a food or a pheromone source typically begins with a whiff of odor, which triggers a flight response, eventually bringing the insect near the odor source. However, pinpointing the precise location of an odor source requires use of both visual and olfactory modalities, aided by odor plumes. This study investigated odor-tracking behavior in fruit flies (Drosophila melanogaster) presented with low- or high-contrast visual landmarks, either paired with or separate from an attractive odor cue. These experiments were conducted either in a gentle air stream which generated laminar odor plumes, or in still air in which odor dissipates uniformly in all directions. Trajectories of flies revealed several novel features of their odor-tracking behavior in addition to those previously documented. First, in both moving and still air, odor-seeking flies rely on co-occurrence of visual landmarks with olfactory cues to guide them to odorant objects. Second, flies abruptly decelerate upon encountering an odor plume, thereafter steering towards nearest visual objects that had no inherent salience in the absence of odor. Thus, interception of an attractive odor increases their salience to nearby high-contrast visual landmarks. Third, flies adopt distinct odor tracking strategies during flight in moving vs. still air. Whereas they weave in and out of plumes towards an odor source in airflow, their approach is more incremental in still air. Both strategies are robust and flexible, and enable flies to reliably find odor sources under diverse visual and airflow environments.
Ryglewski, S., Duch, C. and Altenhein, B. (2017). Tyramine actions on Drosophila flight behavior are affected by a glial dehydrogenase/reductase. Front Syst Neurosci 11: 68. PubMed ID: 29021745
The biogenic amines octopamine (OA) and tyramine (TA) modulate insect motor behavior in an antagonistic manner. OA generally enhances locomotor behaviors such as Drosophila larval crawling and flight, whereas TA decreases locomotor activity. However, the mechanisms and cellular targets of TA modulation of locomotor activity are incompletely understood. This study combines immunocytochemistry, genetics and flight behavioral assays in the Drosophila model system to test the role of a candidate enzyme for TA catabolism, named Nazgul (Naz), in flight motor behavioral control. It is hypothesized that the dehydrogenase/reductase Naz represents a critical step in TA catabolism. Immunocytochemistry reveals that Naz is localized to a subset of Repo positive glial cells with cell bodies along the motor neuropil borders and numerous positive Naz arborizations extending into the synaptic flight motor neuropil. RNAi knock down of Naz in Repo positive glial cells reduces Naz protein level below detection level by Western blotting. The resulting consequence is a reduction in flight durations, thus mimicking known motor behavioral phenotypes as resulting from increased TA levels. In accord with the interpretation that reduced TA degradation by Naz results in increased TA levels in the flight motor neuropil, the motor behavioral phenotype can be rescued by blocking TA receptors. These findings indicate that TA modulates flight motor behavior by acting on central circuitry and that TA is normally taken up from the central motor neuropil by Repo-positive glial cells, desaminated and further degraded by Naz.
Shahandeh, M. P., Pischedda, A. and Turner, T. L. (2017). Male mate choice via cuticular hydrocarbon pheromones drives reproductive isolation between Drosophila species. Evolution 72(1):123-135. PubMed ID: 29098691
Mate discrimination is a key mechanism restricting gene flow between species. While studied extensively with respect to female mate choice, mechanisms of male mate choice between species are far less studied. Thus, there is little knowledge of the relative frequency, importance, or overall contribution of male mate discrimination to reproductive isolation. This study estimated the relative contributions of male and female choice to reproductive isolation between Drosophila simulans and D. sechellia, and showed that male mate discrimination accounts for the majority of the current isolation between these species. It was further demonstrate that males discriminate based on female cuticular hydrocarbon pheromones, and collect evidence supporting the hypothesis that male mate discrimination may alleviate the costs associated with heterospecific courtship and mating. These findings highlight the potentially significant contribution of male mate choice to the formation of reproductive isolating barriers, and thus the speciation process.
Lee, M. J., Sung, H. Y., Jo, H., Kim, H. W., Choi, M. S., Kwon, J. Y. and Kang, K. (2017). Ionotropic Receptor 76b is required for gustatory aversion to excessive Na+ in Drosophila. Mol Cells 40(10): 787-795. PubMed ID: 29081083
Avoiding ingestion of excessively salty food is essential for cation homeostasis that underlies various physiological processes in organisms. The molecular and cellular basis of the aversive salt taste, however, remains elusive. Through a behavioral reverse genetic screening, feeding suppression by Na(+)-rich food was found to require Ionotropic Receptor 76b (Ir76b) in Drosophila labellar gustatory receptor neurons (GRNs). Concentrated sodium solutions with various anions caused feeding suppression dependent on Ir76b. Feeding aversion to caffeine and high concentrations of divalent cations and sorbitol was unimpaired in Ir76b-deficient animals, indicating sensory specificity of Ir76b-dependent Na(+) detection and the irrelevance of hyperosmolarity-driven mechanosensation to Ir76b-mediated feeding aversion. Ir76b-dependent Na(+)-sensing GRNs in both L- and s-bristles are required for repulsion as opposed to the previous report where the L-bristle GRNs direct only low-Na(+) attraction. This work extends the physiological implications of Ir76b from low-Na(+) attraction to high-Na(+) aversion, prompting further investigation of the physiological mechanisms that modulate two competing components of Na(+)-evoked gustation coded in heterogeneous Ir76b-positive GRNs.

Friday, January 26th

Penke, T. J. R., McKay, D. J., Strahl, B. D., Matera, A. G. and Duronio, R. J. (2018). Functional redundancy of variant and canonical histone H3 Lysine 9 modification in Drosophila. Genetics 208(1):229-244. PubMed ID: 29133298
Histone post-translational modifications (PTMs) and differential incorporation of variant and canonical histones into chromatin are central modes of epigenetic regulation. Despite similar protein sequences, histone variants are enriched for different suites of PTMs compared to their canonical counterparts. For example, variant histone H3.3 occurs primarily in transcribed regions and is enriched for "active" histone PTMs like Lys9 acetylation (H3.3K9ac), whereas the canonical histone H3 is enriched for Lys9 methylation (H3K9me), which is found in transcriptionally silent heterochromatin. To determine the functions of K9 modification on variant versus canonical H3, the phenotypes caused by engineering H3.3(K9R) and H3(K9R) mutant genotypes in Drosophila melanogaster were compared. Whereas most H3.3(K9R) and a small number of H3(K9R) mutant animals are capable of completing development and do not have substantially altered protein coding transcriptomes, all H3.3(K9R) H3(K9R) combined mutants die soon after embryogenesis and display decreased expression of genes enriched for K9ac. These data suggest that the role of K9ac in gene activation during development can be provided by either H3 or H3.3. Conversely, it was found that H3.3K9 is methylated at telomeric transposons, and this mark contributes to repressive chromatin architecture, supporting a role for H3.3 in heterochromatin that is distinct from that of H3. Thus, these genetic and molecular analyses demonstrate that K9 modification of variant and canonical H3 have overlapping roles in development and transcriptional regulation, though to differing extents in euchromatin and heterochromatin.
Rickels, R., Herz, H. M., Sze, C. C., Cao, K., Morgan, M. A., Collings, C. K., Gause, M., Takahashi, Y. H., Wang, L., Rendleman, E. J., Marshall, S. A., Krueger, A., Bartom, E. T., Piunti, A., Smith, E. R., Abshiru, N. A., Kelleher, N. L., Dorsett, D. and Shilatifard, A. (2017). Histone H3K4 monomethylation catalyzed by Trr and mammalian COMPASS-like proteins at enhancers is dispensable for development and viability. Nat Genet 49(11): 1647-1653. PubMed ID: 28967912
Histone H3 lysine 4 monomethylation (H3K4me1) is an evolutionarily conserved feature of enhancer chromatin catalyzed by the COMPASS-like methyltransferase family, which includes Trr in Drosophila melanogaster and MLL3 (encoded by KMT2C) and MLL4 (encoded by KMT2D) in mammals. This study demonstrates that Drosophila embryos expressing catalytically deficient Trr eclose and develop to productive adulthood. Parallel experiments with a trr allele that augments enzyme product specificity show that conversion of H3K4me1 at enhancers to H3K4me2 and H3K4me3 is also compatible with life and results in minimal changes in gene expression. Similarly, loss of the catalytic SET domains of MLL3 and MLL4 in mouse embryonic stem cells (mESCs) does not disrupt self-renewal. Drosophila embryos with trr alleles encoding catalytic mutants manifest subtle developmental abnormalities when subjected to temperature stress or altered cohesin levels. Collectively, these findings suggest that animal development can occur in the context of Trr or mammalian COMPASS-like proteins deficient in H3K4 monomethylation activity and point to a possible role for H3K4me1 on cis-regulatory elements in specific settings to fine-tune transcriptional regulation in response to environmental stress.
Shimaji, K., Tanaka, R., Maeda, T., Ozaki, M., Yoshida, H., Ohkawa, Y., Sato, T., Suyama, M. and Yamaguchi, M. (2017). Histone methyltransferase G9a is a key regulator of the starvation-induced behaviors in Drosophila melanogaster. Sci Rep 7(1): 14763. PubMed ID: 29116191
Organisms have developed behavioral strategies to defend themselves from starvation stress. Despite of their importance in nature, the underlying mechanisms have been poorly understood. This study shows that Drosophila G9a (dG9a), one of the histone H3 Lys 9-specific histone methyltransferases, functions as a key regulator for the starvation-induced behaviors. RNA-sequencing analyses utilizing dG9a null mutant flies revealed that the expression of some genes relating to gustatory perception are regulated by dG9a under starvation conditions. Reverse transcription quantitative-PCR analyses showed that the expression of gustatory receptor genes for sensing sugar are up-regulated in starved dG9a null mutant. Consistent with this, proboscis extension reflex tests indicated that dG9a depletion increased the sensitivity to sucrose under starvation conditions. Furthermore, the locomotion activity was promoted in starved dG9a null mutant. It was also found that dG9a depletion downregulates the expression of insulin-like peptide genes that are required for the suppression of starvation-induced hyperactivity. Furthermore, refeeding of wild type flies after starvation conditions restores the hyperactivity and increased sensitivity to sucrose as well as dG9a expression level. These data suggest that dG9a functions as a key regulator for the decision of behavioral strategies under starvation conditions.
Posukh, O. V., Maksimov, D. A., Laktionov, P. P., Koryakov, D. E. and Belyakin, S. N. (2017). Functional dissection of Drosophila melanogaster SUUR protein influence on H3K27me3 profile. Epigenetics Chromatin 10(1): 56. PubMed ID: 29191233
In eukaryotes, heterochromatin replicates late in S phase of the cell cycle and contains specific covalent modifications of histones. SuUR mutation found in Drosophila makes heterochromatin replicate earlier than in wild type and reduces the level of repressive histone modifications. SUUR protein was shown to be associated with moving replication forks, apparently through the interaction with PCNA. The biological process underlying the effects of SUUR on replication and composition of heterochromatin remains unknown. This study performed a functional dissection of SUUR protein effects on H3K27me3 level. Using hidden Markow model-based algorithm SuUR-sensitive chromosomal regions were revealed that demonstrated unusual characteristics: They do not contain Polycomb and require SUUR function to sustain H3K27me3 level. This study tested the role of SUUR protein in the mechanisms that could affect H3K27me3 histone levels in these regions. SUUR did not affect the initial H3K27me3 pattern formation in embryogenesis or Polycomb distribution in the chromosomes. The possible effect of SUUR on histone genes expression and its involvement in DSB repair were also ruled out. These results support the idea that SUUR protein contributes to the heterochromatin maintenance during the chromosome replication.
Stadler, M. R., Haines, J. E. and Eisen, M. (2017). Convergence of topological domain boundaries, insulators, and polytene interbands revealed by high-resolution mapping of chromatin contacts in the early Drosophila melanogaster embryo. Elife 6. PubMed ID: 29148971
High-throughput assays of three-dimensional interactions of chromosomes have shed considerable light on the structure of animal chromatin. Despite this progress, the precise physical nature of observed structures and the forces that govern their establishment remain poorly understood. This study presents high resolution Hi-C data from early Drosophila embryos. Boundaries between topological domains of various sizes were shown to map to DNA elements that resemble classical insulator elements: short genomic regions sensitive to DNase digestion that are strongly bound by known insulator proteins and are frequently located between divergent promoters. Further, a striking correspondence was shown between these elements and the locations of mapped polytene interband regions. It is likely this relationship between insulators, topological boundaries, and polytene interbands extends across the genome, and a model is proposed in which decompaction of boundary-insulator-interband regions drives the organization of interphase chromosomes by creating stable physical separation between adjacent domains.
Morgan, M. A. J., Rickels, R. A., Collings, C. K., He, X., Cao, K., Herz, H. M., Cozzolino, K. A., Abshiru, N. A., Marshall, S. A., Rendleman, E. J., Sze, C. C., Piunti, A., Kelleher, N. L., Savas, J. N. and Shilatifard, A. (2017). A cryptic Tudor domain links BRWD2/PHIP to COMPASS-mediated histone H3K4 methylation. Genes Dev 31(19): 2003-2014. PubMed ID: 29089422
Histone H3 Lys4 (H3K4) methylation is a chromatin feature enriched at gene cis-regulatory sequences such as promoters and enhancers. This study identified an evolutionarily conserved factor, BRWD2/PHIP, which colocalizes with histone H3K4 methylation genome-wide in human cells, mouse embryonic stem cells, and Drosophila. Biochemical analysis of BRWD2 demonstrated an association with the Cullin-4-RING ubiquitin E3 ligase-4 (CRL4) complex, nucleosomes, and chromatin remodelers. BRWD2/PHIP binds directly to H3K4 methylation through a previously unidentified chromatin-binding module related to Royal Family Tudor domains, which has been named the CryptoTudor domain. Using CRISPR-Cas9 genetic knockouts, it was demonstrated that COMPASS H3K4 methyltransferase family members differentially regulate BRWD2/PHIP chromatin occupancy. Finally, it was demonstrated that depletion of the single Drosophila homolog dBRWD3 results in altered gene expression and aberrant patterns of histone H3 Lys27 acetylation at enhancers and promoters, suggesting a cross-talk between these chromatin modifications and transcription through the BRWD protein family.

Thursday, July 25th

Myachina, F., Bosshardt, F., Bischof, J., Kirschmann, M. and Lehner, C. F. (2017). Drosophila beta-tubulin 97EF is upregulated at low temperature and stabilizes microtubules. Development 144(24):4573-4587. PubMed ID: 29084803
Cells in ectotherms function normally within an often wide temperature range. As temperature dependence is not uniform across all the distinct biological processes, acclimation presumably requires complex regulation. The molecular mechanisms coping with the disruptive effects of temperature variation are still poorly understood. Interestingly, one of five different beta-tubulin paralogs, betaTub97EF, was among the genes up-regulated at low temperature in cultured Drosophila cells. As microtubules are known to be cold-sensitive, whether betaTub97EF protects microtubules at low temperatures was analyzed. During development at the optimal temperature (25 ° C), betaTub97EF was expressed in a tissue-specific pattern primarily in the gut. There, as well as in hemocytes, expression was increased at low temperature (14 ° C). While betaTub97EF mutants were viable and fertile at 25 ° C, their sensitivity within the well-tolerated range was slightly enhanced during embryogenesis specifically at low temperatures. Changing beta-tubulin isoform ratios in hemocytes demonstrated that beta-Tubulin 97EF has a pronounced microtubule stabilizing effect. Moreover, betaTub97EF is required for normal microtubule stability in the gut. These results suggest that betaTub97EF up-regulation at low temperature contributes to acclimation by stabilizing microtubules.
Jenkins, B. V., Saunders, H. A. J., Record, H. L., Johnson-Schlitz, D. M. and Wildonger, J. (2017). Effects of mutating alpha-tubulin lysine 40 on sensory dendrite development. J Cell Sci 130(24):4120-4131. PubMed ID: 29122984
Microtubules are essential to neuronal structure and function. Axonal and dendritic microtubules are enriched in post-translational modifications that impact microtubule dynamics, transport, and microtubule-associated proteins. Acetylation of alpha-tubulin lysine 40 (K40) is a prominent, conserved modification of neuronal microtubules. However, the cellular role of microtubule acetylation remains controversial. To resolve how microtubule acetylation might affect neuronal morphogenesis, endogenous alpha-tubulin was mutated in vivo using a new fly strain that facilitates the rapid knock-in of designer alpha-tubulin alleles. Leveraging this new strain, it was found that microtubule acetylation, as well as polyglutamylation and (de)tyrosination, is not essential for survival. However, it was found that dendrite branch refinement in sensory neurons relies on alpha-tubulin K40. Mutagenesis of K40 reveals moderate yet significant changes in dendritic lysosome transport, microtubule polymerization, and Futsch distribution in dendrites but not axons. These studies point to an unappreciated role for alpha-tubulin K40 and acetylation in dendrite morphogenesis. While the results are consistent with the idea that microtubule acetylation patterns microtubule function within neurons, they also suggest there may be an acetylation-independent requirement for alpha-tubulin K40.
Samarasekera, G. and Auld, V. J. (2017). C-terminal Src kinase (Csk) regulates the tricellular junction protein Gliotactin independent of Src. Mol Biol Cell 29(2):123-136. PubMed ID: 29167383
Tricellular junctions (TCJs) are uniquely placed permeability barriers formed at the corners of polarized epithelia where tight junctions (TJ) in vertebrates or septate junctions (SJ) in invertebrates from three cells converge. Gliotactin is a Drosophila TCJ protein and loss of Gliotactin results in SJ and TCJ breakdown, and permeability barrier loss. When overexpressed, Gliotactin spreads away from the TCJs resulting in disrupted epithelial architecture including over-proliferation, cell delamination and migration. Gliotactin levels are tightly controlled at the mRNA level and at the protein level through endocytosis and degradation triggered by tyrosine phosphorylation. This study identified C-terminal Src kinase (Csk) as a tyrosine kinase responsible for regulating Gliotactin endocytosis. Increased Csk suppresses the Gliotactin overexpression phenotypes by increasing endocytosis. Loss of Csk causes Gliotactin to spread away from the TCJ. Although Csk is known as a negative regulator of Src kinases, the effects of Csk on Gliotactin are independent of Src, and likely occurs through an adherens junction (AJ) associated complex. Overall, this study identified a new Src-independent role for Csk in the control of Gliotactin, a key tricellular junction protein.
Nieuwburg, R., Nashchekin, D., Jakobs, M., Carter, A. P., Khuc Trong, P., Goldstein, R. E. and St Johnston, D. (2017). Localised dynactin protects growing microtubules to deliver oskar mRNA to the posterior cortex of the Drosophila oocyte. Elife 6. PubMed ID: 29035202
The localisation of oskar mRNA to the posterior of the Drosophila oocyte defines where the abdomen and germ cells form in the embryo. Kinesin 1 transports oskar mRNA to the oocyte posterior along a polarised microtubule cytoskeleton that grows from non-centrosomal microtubule organising centres (ncMTOCs) along the anterior/lateral cortex. This study shows that the formation of this polarised microtubule network also requires the posterior regulation of microtubule growth. A missense mutation in the dynactin Arp1 subunit causes most oskar mRNA to localise in the posterior cytoplasm rather than cortically. oskar mRNA transport and anchoring are normal in this mutant, but the microtubules fail to reach the posterior pole. Thus, dynactin acts as an anti-catastrophe factor that extends microtubule growth posteriorly. Kinesin 1 transports dynactin to the oocyte posterior, creating a positive feedback loop that increases the length and persistence of the posterior microtubules that deliver oskar mRNA to the cortex.
Devambez, I., van Dijk, J., Benlefki, S., Layalle, S., Grau, Y., Rogowski, K., Parmentier, M. L. and Soustelle, L. (2017). Identification of DmTTLL5 as a major Tubulin glutamylase in the Drosophila nervous system. Sci Rep 7(1): 16254. PubMed ID: 29176602
Microtubules (MTs) play crucial roles during neuronal life. They are formed by heterodimers of alpha and beta-tubulins, which are subjected to several post-translational modifications (PTMs). Amongst them, glutamylation consists in the reversible addition of a variable number of glutamate residues to the C-terminal tails of tubulins. Glutamylation is the most abundant MT PTM in the mammalian adult brain, suggesting that it plays an important role in the nervous system (NS). This study shows that the previously uncharacterized CG31108 gene encodes an alpha-tubulin glutamylase acting in the Drosophila NS. This glutamylase, which was named DmTTLL5, initiates MT glutamylation specifically on alpha-tubulin, the only glutamylated tubulin in the Drosophila brain. In DmTTLL5 mutants, MT glutamylation was not detected in the NS, allowing for determining its potential function. DmTTLL5 mutants are viable and no defect was found in vesicular axonal transport, synapse morphology and larval locomotion. Moreover, DmTTLL5 mutant flies display normal negative geotaxis behavior and their lifespan is not altered. Thus, this work identifies DmTTLL5 as the major enzyme responsible for initiating neuronal MT glutamylation specifically on alpha-tubulin, and the absence of MT glutamylation was not detrimental for Drosophila NS function.
Patel, A. A., Oztug Durer, Z. A., van Loon, A. P., Bremer, K. V. and Quinlan, M. E. (2017). Drosophila and human FHOD family formins nucleate actin filaments. J Biol Chem 293(2):532-540. PubMed ID: 29127202
Formins are a conserved group of proteins that nucleate and processively elongate actin filaments. Among them, the formin homology domain-containing protein (FHOD) family of formins contributes to contractility of striated muscle and cell motility in several contexts. However, the mechanisms by which they carry out these functions remain poorly understood. Mammalian FHOD proteins were reported not to accelerate actin assembly in vitro; instead, they were proposed to act as barbed end cappers or filament bundlers. This study shows that purified Drosophila Fhod (also known as Fhos or knittrig) and human FHOD1 both accelerate actin assembly by nucleation. FHOD1's nucleation activity is restricted to cytoplasmic actin, whereas Drosophila Fhod potently nucleates both cytoplasmic and sarcomeric actin isoforms. Drosophila Fhod binds tightly to barbed ends, where it slows elongation in the absence of profilin and allows, but does not accelerate, elongation in the presence of profilin. Fhod antagonizes capping protein, but dissociates from barbed ends relatively quickly. Finally, it was determined that Fhod binds the sides of and bundles actin filaments. This work establishes that Fhod shares the capacity of other formins to nucleate and bundle actin filaments, but is notably less effective at processively elongating barbed ends than most well studied formins.

Wednesday, January 24th

Matsubayashi, Y., Louani, A., Dragu, A., Sanchez-Sanchez, B. J., Serna-Morales, E., Yolland, L., Gyoergy, A., Vizcay, G., Fleck, R. A., Heddleston, J. M., Chew, T. L., Siekhaus, D. E. and Stramer, B. M. (2017). A moving source of matrix components is essential for de novo basement membrane formation. Curr Biol 27(22): 3526-3534. PubMed ID: 29129537
The basement membrane (BM) is a thin layer of extracellular matrix (ECM) beneath nearly all epithelial cell types that is critical for cellular and tissue function. It is composed of numerous components conserved among all bilaterians; however, it is unknown how all of these components are generated and subsequently constructed to form a fully mature BM in the living animal. Although BM formation is thought to simply involve a process of self-assembly, this concept suffers from a number of logistical issues when considering its construction in vivo. First, incorporation of BM components, including Col IV, Perl and LanA appears to be hierarchical, yet it is unclear whether their production during embryogenesis must also be regulated in a temporal fashion. Second, many BM proteins are produced not only by the cells residing on the BM but also by surrounding cell types, and it is unclear how large, possibly insoluble protein complexes are delivered into the matrix. This study exploited the ability to live image and genetically dissect de novo BM formation during Drosophila development. This reveals that there is a temporal hierarchy of BM protein production that is essential for proper component incorporation. Furthermore, it was shown that BM components require secretion by migrating macrophages (hemocytes) during their developmental dispersal, which is critical for embryogenesis. Indeed, hemocyte migration is essential to deliver a subset of ECM components evenly throughout the embryo. This reveals that de novo BM construction requires a combination of both production and distribution logistics allowing for the timely delivery of core components.
Mohamad Ishak, N. S., Nong, Q. D., Matsuura, T., Kato, Y. and Watanabe, H. (2017). Co-option of the bZIP transcription factor Vrille as the activator of Doublesex1 in environmental sex determination of the crustacean Daphnia magna. PLoS Genet 13(11): e1006953. PubMed ID: 29095827
Evolutionary Homolog Study
Divergence of upstream regulatory pathways of the transcription factor Doublesex (Dsx) serves as a basis for evolution of sex-determining mechanisms in animals. However, little is known about the regulation of Dsx in environmental sex determination. In the crustacean Daphnia magna, environmental sex determination is implemented by male-specific expression of the Dsx ortholog, Dsx1. Transcriptional regulation of Dsx1 comprises at least three phases during embryogenesis: non-sex-specific initiation, male-specific up-regulation, and its maintenance. This study demonstrates that the male-specific up-regulation is controlled by the bZIP transcription factor, Vrille (Vri), an ortholog of the circadian clock genes-Drosophila Vri and mammalian E4BP4/NFIL3. Sequence analysis of the Dsx1 promoter/enhancer revealed a conserved element among two Daphnia species (D. magna and D. pulex), which contains a potential enhancer harboring a consensus Vri binding site overlapped with a consensus Dsx binding site. Besides non-sex-specific expression of Vri in late embryos, male-specific expression was found in early gastrula before the Dsx1 up-regulation phase begins. Knockdown of Vri in male embryos showed reduction of Dsx1 expression. In addition, transient overexpression of Vri in early female embryos up-regulated the expression of Dsx1 and induced male-specific trait. Targeted mutagenesis using CRISPR/Cas9 disrupted the enhancer in males, which led to the reduction of Dsx1 expression. These results indicate that Vri was co-opted as a transcriptional activator of Dsx1 in environmental sex determination of D. magna. The data suggests the remarkably plastic nature of gene regulatory network in sex determination.
Nakao, H. (2017). A Bombyx homolog of ovo is a segmentation gene that acts downstream of Bm-wnt1(Bombyx wnt1 homolog). Gene Expr Patterns 27: 1-7. PubMed ID: 28988845
Evolutionary Homolog Study

Insect embryogenesis is divided into long and short/intermediate germ types. The long germ type may exhibit Drosophila-like hierarchical segmentation mechanisms, whereas the short/intermediate type assumes some repeating mechanisms that are considered to be ancestral. Embryogenesis in Bombyx mori possesses both characteristics. In this study, Bombyx ovo homolog (Bm-ovo) was identified as a gene involved in segmentation. Ovo is a Drosophila gene that encodes a zinc finger transcription factor and studies on its homolog functions in other systems have suggested that it acts as a switch to enable the initiation of differentiation from a progenitor cell state. This is the first description for ovo homologs being involved in insect segmentation. Bm-ovo is expressed dynamically during embryogenesis in a pattern that resembles that of gap and pair-rule genes. In Bm-ovo RNAi knockdown embryos, posterior segmentation does not proceed. In addition, defects in anterior segments are observed. In Bm-wnt1 knockdown embryos, the Bm-ovo expression pattern was changed, suggesting that Bm-wnt1 is an upstream regulator of Bm-ovo. The involvement of Bm-ovo may represent a novel ancestral step under the control of wnt genes in insect segmentation: this step may resemble those operating in cell differentiation processes.

Rupprecht, J. F., Ong, K. H., Yin, J., Huang, A., Dinh, H. H., Singh, A. P., Zhang, S., Yu, W. and Saunders, T. E. (2017). Geometric constraints alter cell arrangements within curved epithelial tissues. Mol Biol Cell 28(25): 3582-3594. PubMed ID: 28978739
Organ and tissue formation are complex three-dimensional processes involving cell division, growth, migration, and rearrangement, all of which occur within physically constrained regions. However, analyzing such processes in three dimensions in vivo is challenging. This study focused on the process of cellularization in the anterior pole of the early Drosophila embryo to explore how cells compete for space under geometric constraints. Using microfluidics combined with fluorescence microscopy, quantitative information was extracted on the three-dimensional epithelial cell morphology. A cellular membrane rearrangement was observed in which cells exchange neighbors along the apical-basal axis. Such apical-to-basal neighbor exchanges were observed more frequently in the anterior pole than in the embryo trunk. Furthermore, cells within the anterior pole skewed toward the trunk along their long axis relative to the embryo surface, with maximum skew on the ventral side. A vertex model was constructed for cells in a curved environment. The observed cellular skew was reproduced in both wild-type embryos and embryos with distorted morphology. Further, such modeling showed that cell rearrangements were more likely in ellipsoidal, compared with cylindrical, geometry. Overall, it was demonstrated that geometric constraints can influence three-dimensional cell morphology and packing within epithelial tissues.
Carrell, S. N., O'Connell, M. D., Jacobsen, T., Pomeroy, A. E., Hayes, S. M. and Reeves, G. T. (2017). A facilitated diffusion mechanism establishes the Drosophila Dorsal gradient. Development 144(23): 4450-4461. PubMed ID: 29097443
The transcription factor NF-kappaB plays an important role in the immune system, apoptosis and inflammation. Dorsal, a Drosophila homolog of NF-kappaB, patterns the dorsal-ventral axis in the blastoderm embryo. During this stage, Dorsal is sequestered outside the nucleus by the IkappaB homolog Cactus. Toll signaling on the ventral side breaks the Dorsal/Cactus complex, allowing Dorsal to enter the nucleus to regulate target genes. Fluorescent data show that Dorsal accumulates on the ventral side of the syncytial blastoderm. This study used modeling and experimental studies to show that this accumulation is caused by facilitated diffusion, or shuttling, of the Dorsal/Cactus complex. Active Toll receptors are limiting in wild-type embryos, which is a key factor in explaining global Dorsal gradient formation. These results suggest that shuttling is necessary for viability of embryos from mothers with compromised dorsal levels. Therefore, Cactus not only has the primary role of regulating Dorsal nuclear import, but also has a secondary role in shuttling. Given that this mechanism has been found in other, independent, systems, it is suggested that it might be more prevalent than previously thought.
Mir, M., Reimer, A., Haines, J. E., Li, X. Y., Stadler, M., Garcia, H., Eisen, M. B. and Darzacq, X. (2017). Dense Bicoid hubs accentuate binding along the morphogen gradient. Genes Dev 31(17): 1784-1794. PubMed ID: 28982761
Morphogen gradients direct the spatial patterning of developing embryos; however, the mechanisms by which these gradients are interpreted remain elusive. This study used lattice light-sheet microscopy to perform in vivo single-molecule imaging in early Drosophila melanogaster embryos of the transcription factor Bicoid that forms a gradient and initiates patterning along the anteroposterior axis. In contrast to canonical models, it was observed that Bicoid binds to DNA with a rapid off rate throughout the embryo such that its average occupancy at target loci is on-rate-dependent. Bicoid was observed forming transient "hubs" of locally high density that facilitate binding as factor levels drop, including in the posterior, where Bicoid binding was observed despite vanishingly low protein levels. It is proposed that localized modulation of transcription factor on rates via clustering provides a general mechanism to facilitate binding to low-affinity targets and that this may be a prevalent feature of other developmental transcription factors.

Tuesday, January 23rd

Lan, Q., Cao, M., Kollipara, R. K., Rosa, J. B., Kittler, R. and Jiang, H. (2017). FoxA transcription factor Fork head maintains the intestinal stem/progenitor cell identities in Drosophila. Dev Biol 433(2):324-343. PubMed ID: 29108672
Understanding how somatic stem cells respond to tissue needs is important, since aberrant somatic stem cell behaviors may lead to tissue degeneration or tumorigenesis. From an in vivo RNAi screen targeting transcription factors that regulate intestinal regeneration, this study uncovered a requirement for the Drosophila FoxA transcription factor Fork head (Fkh) in the maintenance of intestinal stem/progenitor cell identities. FoxA/Fkh maintains the expressions of stem/progenitor cell markers and is required for stem cell proliferation during intestinal homeostasis and regeneration. Furthermore, FoxA/Fkh prevents the intestinal stem/progenitor cells from precocious differentiation into the Enterocyte lineage, likely in cooperation with the transcription factor bHLH/Daughterless (Da). In addition, loss of FoxA/Fkh suppresses the intestinal tumorigenesis caused by Notch pathway inactivation. To reveal the gene program underlying stem/progenitor cell identities, the genome-wide chromatin binding sites of transcription factors Fkh and Da were profiled, and interestingly, around half of Fkh binding regions are shared by Da, further suggesting their collaborative roles. Finally, the genes were identified associated with their shared binding regions. This comprehensive gene list may contain stem/progenitor maintenance factors functioning downstream of Fkh and Da, and would be helpful for future gene discoveries in the Drosophila intestinal stem cell lineage.
Suong, D. N. A., Shimaji, K., Pyo, J. H., Park, J. S., Yoshida, H., Yoo, M. A. and Yamaguchi, M. (2017). Overexpression of dJmj differentially affects intestinal stem cells and differentiated enterocytes. Cell Signal 42: 194-210. PubMed ID: 29102770
Jumonji (Jmj)/Jarid2 is a DNA-binding transcriptional repressor mediated via histone methylation. Nevertheless, the well-known function of Jmj is as a scaffold for the recruitment of various complexes including Polycomb repressive complex 2 (PRC2), and is required for mouse embryonic stem cell development. However, PRC2 independent function is suggested for Drosophila Jumonji (dJmj). To clarify the function of dJmj during cell differentiation, the Drosophila adult intestinal stem cell system was used that allows following of stem cell behaviors in vivo. Overexpression of dJmj in intestinal stem cells/enteroblasts (ISCs/EBs) induces cell-autonomous ISC proliferation followed by differentiation, that is controlled by the Notch and EGFR pathway. In contrast, overexpression of dJmj in enterocytes (ECs) resulted in activation of the JNK pathway in ECs followed by the induction of apoptosis. Activated JNK increased the level of Yorkie in ECs and induced the reduction of Upd proteins and EGFR ligands, which activated the JAK/STAT and EGFR pathway in both ISCs and EBs to promote ISC proliferation. The Notch signaling pathway appears to be highly activated to support the differentiation of EBs to ECs. Thus, the combination of these signaling pathways caused by ECs-specific dJmj-overexpression induced non-cell-autonomous ISC proliferation and differentiation. Surprisingly, these effects did not relate to H3K27me3 status, likely represented PRC2 activity, in cells that overexpressed dJmj. Instead of this, the disappearance of H3K27me3 in ISC/EB-specific overexpressed dJmj suggested a possible PRC2-independent role of dJmj in regulating chromatin structure.
Levings, D. C. and Nakato, H. (2017). Loss of heparan sulfate in the niche leads to tumor-like germ cell growth in the Drosophila testis. Glycobiology 28(1):32-41. PubMed ID: 29069438
The stem cell niche normally prevents aberrant stem cell behaviors that lead to cancer formation. Recent studies suggest that some cancers are derived from endogenous populations of adult stem cells that have somehow escaped from normal control by the niche. However, the molecular mechanisms by which the niche retains stem cells locally and tightly controls their divisions are poorly understood. This study demonstrates that the presence of heparan sulfate (HS), a class glygosaminoglycan chains, in the Drosophila germline stem cell niche prevents tumor formation in the testis. Loss of HS in the niche, called the hub, led to gross changes in the morphology of testes as well as the formation of both somatic and germline tumors. This loss of hub HS resulted in ectopic signaling events in the Jak/Stat pathway outside the niche. This ectopic Jak/Stat signaling disrupted normal somatic cell differentiation, leading to the formation of tumors. This finding indicates a novel non-autonomous role for niche HS in ensuring the integrity of the niche and preventing tumor formation.
Roubinet, C., Tsankova, A., Pham, T. T., Monnard, A., Caussinus, E., Affolter, M. and Cabernard, C. (2017). Spatio-temporally separated cortical flows and spindle geometry establish physical asymmetry in fly neural stem cells. Nat Commun 8(1): 1383. PubMed ID: 29123099
Asymmetric cell division, creating sibling cells with distinct developmental potentials, can be manifested in sibling cell size asymmetry. This form of physical asymmetry occurs in several metazoan cells, but the underlying mechanisms and function are incompletely understood. This study used Drosophila neural stem cells to elucidate the mechanisms involved in physical asymmetry establishment. Myosin relocalizes to the cleavage furrow via two distinct cortical Myosin flows: at anaphase onset, a polarity induced, basally directed Myosin flow clears Myosin from the apical cortex. Subsequently, mitotic spindle cues establish a Myosin gradient at the lateral neuroblast cortex, necessary to trigger an apically directed flow, removing Actomyosin from the basal cortex. On the basis of the data presented in this study, it is proposed that spatiotemporally controlled Myosin flows in conjunction with spindle positioning and spindle asymmetry are key determinants for correct cleavage furrow placement and cortical expansion, thereby establishing physical asymmetry.
Artoni, F., Kreipke, R., Palmeira, O., Dixon, C., Goldberg, Z. and Ruohola-Baker, H. (2017). Loss of foxo rescues stem cell aging in Drosophila germ line. Elife 6. PubMed ID: 28925355
Aging stem cells lose the capacity to properly respond to injury and regenerate their residing tissues. This study utilized the ability of Drosophila melanogaster germline stem cells (GSCs) to survive exposure to low doses of ionizing radiation (IR) as a model of adult stem cell injury and identified a regeneration defect in aging GSCs: while aging GSCs survive exposure to IR, they fail to reenter the cell cycle and regenerate the germline in a timely manner. Mechanistically, foxo and mTOR homologue Tor were identified as important regulators of GSC quiescence following exposure to ionizing radiation. foxo is required for entry in quiescence, while Tor is essential for cell cycle reentry. Importantly, it was further shown that the lack of regeneration in aging germ line stem cells after IR can be rescued by loss of foxo.
Sawyer, J. K., Cohen, E. and Fox, D. T. (2017). Inter-organ regulation of Drosophila intestinal stem cell proliferation by a hybrid organ boundary zone. Development 144(22):4091-4102. PubMed ID: 28947534
This study showed that a specific population of adult midgut organ boundary intestinal stem cells (OB-ISCs) is regulated by the neighboring hindgut, a developmentally distinct organ. This distinct OB-ISC control is due to proximity to a specialized transition zone between the endodermal midgut and ectodermal hindgut that shares molecular signatures of both organs, which has been termed the hybrid zone (HZ). During homeostasis, proximity to the HZ restrains OB-ISC proliferation. However, injury to the adult HZ/hindgut drives up-regulation of unpaired-3 cytokine, which signals through STAT protein to promote cell division only in OB-ISCs. If HZ disruption is severe, hyperplastic OB-ISCs expand across the inter-organ boundary. These data suggest that inter-organ signaling plays an important role in controlling OB-ISCs in homeostasis and injury repair, which is likely critical in prevention of disease.

Monday, January 22nd

Moschall, R., Strauss, D., Garcia-Bayert, M., Gebauer, F. and Medenbach, J. (2018). Drosophila Sister of Sex-lethal is a repressor of translation. RNA 24(2):149-158. PubMed ID: 29089381
The RNA-binding protein Sex Lethal (Sxl) is an important post-transcriptional regulator of gene expression in female Drosophila. To prevent the assembly of the dosage compensation complex in female flies, Sxl acts as a repressor of msl-2 mRNA translation. It employs two distinct and mutually reinforcing blocks to translation that operate on the 5' and 3' untranslated regions (UTRs) of msl-2 mRNA, respectively. While 5' UTR-mediated translational control involves an upstream open reading frame, 3' UTR-mediated regulation strictly requires the co-repressor protein Upstream of N-ras (Unr) which is recruited to the msl-2 mRNA by Sxl. This study has identified the protein Sister of Sex Lethal (Ssx) as a novel repressor of translation with Sxl-like activity. Sxl and Ssx have a comparable RNA-binding specificity and can associate with Uracil-rich RNA regulatory elements present in msl-2 mRNA. Moreover, both repress translation when bound to the 5' UTR of msl-2 However, Ssx is inactive in 3' UTR-mediated regulation as it cannot engage the co-repressor protein Unr. The difference in activity maps to the first RNA-recognition motif (RRM) of Ssx. Conversion of three amino acids within this domain into their Sxl counterpart results in a gain-of-function and repression via the 3' UTR, allowing detailed insights into the evolutionary origin of the two proteins and into the molecular requirements of an important translation regulatory pathway.
Rogers, A. K., Situ, K., Perkins, E. M. and Toth, K. F. (2017). Zucchini-dependent piRNA processing is triggered by recruitment to the cytoplasmic processing machinery. Genes Dev 31(18): 1858-1869. PubMed ID: 29021243
The piRNA pathway represses transposable elements in the gonads and thereby plays a vital role in protecting the integrity of germline genomes of animals. Mature piRNAs are processed from longer transcripts, piRNA precursors (pre-piRNAs). In Drosophila, processing of pre-piRNAs is initiated by piRNA-guided Slicer cleavage or the endonuclease Zucchini (Zuc). As Zuc does not have any sequence or structure preferences in vitro, it is not known how piRNA precursors are selected and channeled into the Zuc-dependent processing pathway. This study shows that a heterologous RNA that lacks complementary piRNAs is processed into piRNAs upon recruitment of several piRNA pathway factors. This processing requires Zuc and the helicase Armitage (Armi). Aubergine (Aub), Argonaute 3 (Ago3), and components of the nuclear RDC complex, which are required for normal piRNA biogenesis in germ cells, are dispensable. This approach allows discrimination of proteins involved in the transcription and export of piRNA precursors from components required for the cytoplasmic processing steps. piRNA processing correlates with localization of the substrate RNA to nuage, a distinct membraneless cytoplasmic compartment, which surrounds the nucleus of germ cells, suggesting that sequestration of RNA to this subcellular compartment is both necessary and sufficient for selecting piRNA biogenesis substrates.
Ota, R., Morita, S., Sato, M., Shigenobu, S., Hayashi, M. and Kobayashi, S. (2017). Transcripts immunoprecipitated with Sxl protein in primordial germ cells of Drosophila embryos. Dev Growth Differ 59(9):713-723. PubMed ID: 29124738
In Drosophila, Sex lethal (Sxl), an RNA binding protein, is required for induction of female sexual identity in both somatic and germline cells. Although the Sxl-dependent feminizing pathway in the soma was previously elucidated, the downstream targets for Sxl in the germline remained elusive. To identify these target genes, transcripts associated with Sxl in primordial germ cells (PGCs) of embryos were selected using RNA immunoprecipitation coupled to sequencing (RIP-seq) analysis. A total of 308 transcripts encoded by 282 genes were obtained. Seven of these genes, expressed at higher levels in PGCs as determined by microarray and in situ hybridization analyses, were subjected to RNAi-mediated functional analyses. Knockdown of Neos, Kap-alpha3, and CG32075 throughout germline development caused gonadal dysgenesis in a sex-dependent manner, and Su(var)2-10 knockdown caused gonadal dysgenesis in both sexes. Moreover, as with knockdown of Sxl, knockdown of Su(var)2-10 in PGCs gave rise to a tumorous phenotype of germline cells in ovaries. Because this phenotype indicates loss of female identity of germline cells, Su(var)2-10 is considered to be a strong candidate target of Sxl in PGCs. These results represent a first step toward elucidating the Sxl-dependent feminizing pathway in the germline.
Ilik, I. A., Maticzka, D., Georgiev, P., Gutierrez, N. M., Backofen, R. and Akhtar, A. (2017). A mutually exclusive stem-loop arrangement in roX2 RNA is essential for X-chromosome regulation in Drosophila. Genes Dev 31(19): 1973-1987. PubMed ID: 29066499
The X chromosome provides an ideal model system to study the contribution of RNA-protein interactions in epigenetic regulation. In male flies, roX long noncoding RNAs (lncRNAs) harbor several redundant domains to interact with the ubiquitin ligase male-specific lethal 2 (MSL2) and the RNA helicase Maleless (MLE) for X-chromosomal regulation. However, how these interactions provide the mechanics of spreading remains unknown. By using the uvCLAP (UV cross-linking and affinity purification) methodology, which provides unprecedented information about RNA secondary structures in vivo, the minimal functional unit of roX2 RNA was identified. By using wild-type and various MLE mutant derivatives, including a catalytically inactive MLE derivative, MLE(GET), this study showed that the minimal roX RNA contains two mutually exclusive stem-loops that exist in a peculiar structural arrangement: When one stem-loop is unwound by MLE, an alternate structure can form, likely trapping MLE in this perpetually structured region. This functional unit is necessary for dosage compensation, as mutations that disrupt this formation lead to male lethality. Thus, it is proposed that roX2 lncRNA contains an MLE-dependent affinity switch to enable reversible interactions of the MSL complex to allow dosage compensation of the X chromosome.
Liang, D., Tatomer, D. C., Luo, Z., Wu, H., Yang, L., Chen, L. L., Cherry, S. and Wilusz, J. E. (2017). The output of protein-coding genes shifts to circular RNAs when the pre-mRNA processing machinery is limiting. Mol Cell 68(5):940-954. PubMed ID: 29174924
Many eukaryotic genes generate linear mRNAs and circular RNAs, but it is largely unknown how the ratio of linear to circular RNA is controlled or modulated. Using RNAi screening in Drosophila cells, this study identified many core spliceosome and transcription termination factors that control the RNA outputs of reporter and endogenous genes. When spliceosome components were depleted or inhibited pharmacologically, the steady-state levels of circular RNAs increased while expression of their associated linear mRNAs concomitantly decreased. Upon inhibiting RNA polymerase II termination via depletion of the cleavage/polyadenylation machinery, circular RNA levels were similarly increased. This is because readthrough transcripts now extend into downstream genes and are subjected to backsplicing. In total, these results demonstrate that inhibition or slowing of canonical pre-mRNA processing events shifts the steady-state output of protein-coding genes toward circular RNAs. This is in part because nascent RNAs become directed into alternative pathways that lead to circular RNA production.
Rojas-Rios, P., Chartier, A., Pierson, S. and Simonelig, M. (2017). Aubergine and piRNAs promote germline stem cell self-renewal by repressing the proto-oncogene Cbl. EMBO J 36(21): 3194-3211. PubMed ID: 29030484
PIWI proteins play essential roles in germ cells and stem cell lineages. In Drosophila, Piwi is required in somatic niche cells and germline stem cells (GSCs) to support GSC self-renewal and differentiation. Whether and how other PIWI proteins are involved in GSC biology remains unknown. This study shows that Aubergine (Aub), another PIWI protein, is intrinsically required in GSCs for their self-renewal and differentiation. Aub needs to be loaded with piRNAs to control GSC self-renewal and acts through direct mRNA regulation. The Cbl proto-oncogene, a regulator of mammalian hematopoietic stem cells, was identified as a novel GSC differentiation factor. Aub stimulates GSC self-renewal by repressing Cbl mRNA translation and does so in part through recruitment of the CCR4-NOT complex. This study reveals the role of piRNAs and PIWI proteins in controlling stem cell homeostasis via translational repression and highlights piRNAs as major post-transcriptional regulators in key developmental decisions.

Friday, January 19th

Rohith, B. N. and Shyamala, B. V. (2017). Scalloped a member of the Hippo tumor suppressor pathway controls mushroom body size in Drosophila brain by non-canonical regulation of neuroblast proliferation.Dev Biol 432(2):203-214. PubMed ID: 29080790
Cell proliferation, growth and survival are three different basic processes which converge at determining a fundamental property -the size of an organism. Scalloped (Sd) is the first characterised transcriptional partner to Yorkie (Yki), the downstream effector of the Hippo pathway which is a highly potential and evolutionarily conserved regulator of organ size. The hypomorphic effect of sd on the development of Mushroom Bodies (MBs) in Drosophila brain was studied. sd non-function results in an increase in the size of MBs. sd regulation on MB size operates through multiple routes. Sd expressed in the differentiated MB neurons, imposes non-cell autonomous repression on the proliferation of MB precursor cells, and Sd expression in the MB neuroblasts (NB) cell autonomously represses mushroom body neuroblast (MBNB) proliferation. Further Sd in Kenyon cells (KCs) imparts a cell autonomous restriction on their growth. These findings are distinctive because, while the classical sd loss of function phenotypes in eye, wing and lymph gland are reported as loss of tissue or reduced organ size, the present study shows that, Sd inactivation in the developing MB, promotes precursor cell proliferation and results in an increase in the organ size.
Mauss, A. S., Busch, C. and Borst, A. (2017). Optogenetic neuronal silencing in Drosophila during visual processing. Sci Rep 7(1): 13823. PubMed ID: 29061981
Optogenetic channels and ion pumps have become indispensable tools in neuroscience to manipulate neuronal activity and thus to establish synaptic connectivity and behavioral causality. Inhibitory channels are particularly advantageous to explore signal processing in neural circuits since they permit the functional removal of selected neurons on a trial-by-trial basis. However, applying these tools to study the visual system poses a considerable challenge because the illumination required for their activation usually also stimulates photoreceptors substantially, precluding the simultaneous probing of visual responses. This study explored the utility of the recently discovered anion channelrhodopsins GtACR1 and GtACR2 for application in the visual system of Drosophila. First, their properties were characterized using a larval crawling assay. Further whole-cell recordings were obtained from cells expressing GtACR1, which mediated strong and light-sensitive photocurrents. Tangential cells of the lobula plate lend themselves well for this purpose since whole-cell patch-clamp recordings can be readily obtained from their large cell bodies. Finally, using physiological recordings and a behavioral readout, it was demonstrated that GtACR1 enables the fast and reversible silencing of genetically targeted neurons within circuits engaged in visual processing.
Meltzer, S., Bagley, J. A., Perez, G. L., O'Brien, C. E., DeVault, L., Guo, Y., Jan, L. Y. and Jan, Y. N. (2017). Phospholipid homeostasis regulates dendrite morphogenesis in Drosophila sensory neurons. Cell Rep 21(4): 859-866. PubMed ID: 29069593
Disruptions in lipid homeostasis have been observed in many neurodevelopmental disorders that are associated with dendrite morphogenesis defects. However, the molecular mechanisms of how lipid homeostasis affects dendrite morphogenesis are unclear. This study found that easily shocked (eas), which encodes a kinase with a critical role in phospholipid phosphatidylethanolamine (PE) synthesis, and two other enzymes in this synthesis pathway are required cell autonomously in sensory neurons for dendrite growth and stability. Furthermore, this study shows that the level of Sterol Regulatory Element-Binding Protein (SREBP) activity is important for dendrite development. SREBP activity increases in eas mutants, and decreasing the level of SREBP and its transcriptional targets in eas mutants largely suppresses the dendrite growth defects. Furthermore, reducing Ca(2+) influx in neurons of eas mutants ameliorates the dendrite morphogenesis defects. This study uncovers a role for EAS kinase and reveals the in vivo function of phospholipid homeostasis in dendrite morphogenesis.
Onodera, K., Baba, S., Murakami, A., Uemura, T. and Usui, T. (2017). Small conductance Ca(2+)-activated K(+) channels induce the firing pause periods during the activation of Drosophila nociceptive neurons. Elife 6. PubMed ID: 29035200
In Drosophila larvae, Class IV sensory neurons respond to noxious thermal stimuli and provoke heat avoidance behavior. Previous work showed that the activated neurons displayed characteristic fluctuations of firing rates, which consisted of repetitive high-frequency spike trains and subsequent pause periods, and it was proposed that the firing rate fluctuations enhanced the heat avoidance. This study further substantiate this idea by showing that the pause periods and the frequency of fluctuations are regulated by small conductance Ca(2+)-activated K(+) (SK) channels, and the SK knockdown larvae display faster heat avoidance than control larvae. The regulatory mechanism of the fluctuations in the Class IV neurons resembles that in mammalian Purkinje cells, which display complex spikes. Furthermore, these results suggest that such fluctuation coding in Class IV neurons is required to convert noxious thermal inputs into effective stereotyped behavior as well as general rate coding.
Qian, Y., Cao, Y., Deng, B., Yang, G., Li, J., Xu, R., Zhang, D., Huang, J. and Rao, Y. (2017). Sleep homeostasis regulated by 5HT2b receptor in a small subset of neurons in the dorsal fan-shaped body of Drosophila. Elife 6. PubMed ID: 28984573
Understanding of the molecular mechanisms underlying sleep homeostasis is limited. This paper describes a systematic approach to study neural signaling by the transmitter 5-hydroxytryptamine (5-HT) in Drosophila. Knockout and knockin lines were generated for Trh, the 5-HT synthesizing enzyme and all five 5-HT receptors, making it possible to determine their expression patterns and to investigate their functional roles. Loss of the Trh, 5HT1a or 5HT2b gene decreased sleep time whereas loss of the Trh or 5HT2b gene diminished sleep rebound after sleep deprivation. 5HT2b expression in a small subset of, probably a single pair of, neurons in the dorsal fan-shaped body (dFB) is functionally essential: elimination of the 5HT2b gene from these neurons led to loss of sleep homeostasis. Genetic ablation of 5HT2b neurons in the dFB decreased sleep and impaired sleep homeostasis. These results have shown that serotonergic signaling in specific neurons is required for the regulation of sleep homeostasis.
Reichardt, I., Bonnay, F., Steinmann, V., Loedige, I., Burkard, T. R., Meister, G. and Knoblich, J. A. (2017). The tumor suppressor Brat controls neuronal stem cell lineages by inhibiting Deadpan and Zelda. EMBO Rep 19(1):102-117. PubMed ID: 29191977
The TRIM-NHL protein Brain tumor (Brat) acts as a tumor suppressor in the Drosophila brain, but how it suppresses tumor formation is not completely understood. This study combined temperature-controlled brat RNAi with transcriptome analysis to identify the immediate Brat targets in Drosophila neuroblasts. Besides the known target Deadpan (Dpn), these experiments identified the transcription factor Zelda (Zld) as a critical target of Brat. These data show that Zld is expressed in neuroblasts and required to allow re-expression of Dpn in transit-amplifying intermediate neural progenitors. Upon neuroblast division, Brat is enriched in one daughter cell where its NHL domain directly binds to specific motifs in the 3'UTR of dpn and zld mRNA to mediate their degradation. In brat mutants, both Dpn and Zld continue to be expressed, but inhibition of either transcription factor prevents tumorigenesis. These genetic and biochemical data indicate that Dpn inhibition requires higher Brat levels than Zld inhibition and suggest a model where stepwise post-transcriptional inhibition of distinct factors ensures sequential generation of fates in a stem cell lineage.

Thursday, January 18th

Ray, A., Speese, S. D. and Logan, M. A. (2017). Glial Draper rescues Abeta toxicity in a Drosophila model of Alzheimer's Disease. J Neurosci 37(49):11881-11893. PubMed ID: 29109235
Pathological hallmarks of Alzheimer's disease (AD) include amyloid-beta (Abeta) plaques, neurofibrillary tangles, and reactive gliosis. Glial cells offer protection against AD by engulfing extracellular Abeta peptides, but the repertoire of molecules required for glial recognition and destruction of Abeta are still unclear. This study shows that the highly conserved glial engulfment receptor Draper/MEGF10 provides neuroprotection in an AD model of Drosophila (both sexes). Neuronal expression of human Abeta42arc in adult flies results in robust Abeta accumulation, neurodegeneration, locomotor dysfunction, and reduced lifespan. Notably, all of these phenotypes are more severe in draper mutant animals, while enhanced expression of glial Draper reverses Abeta accumulation, as well as behavioral phenotypes. Stat92E, c-Jun N-terminal Kinase (JNK)/AP-1 signaling, and expression of matrix metalloproteinase-1 (Mmp1) are activated downstream of Draper in glia in response to Abeta42arc exposure. Furthermore, Abeta42-induced upregulation of the phagolysosomal markers Atg8 and p62 was notably reduced in draper mutant flies. Based on these findings, it is proposed that glia clear neurotoxic Abeta peptides in the AD model Drosophila brain through a Draper/STAT92E/JNK cascade that may be coupled to protein degradation pathways such as autophagy or more traditional phagolysosomal destruction methods.
Llavero Hurtado, M., Fuller, H. R., Wong, A. M. S., Eaton, S. L., Gillingwater, T. H., Pennetta, G., Cooper, J. D. and Wishart, T. M. (2017). Proteomic mapping of differentially vulnerable pre-synaptic populations identifies regulators of neuronal stability in vivo. Sci Rep 7(1): 12412. PubMed ID: 28963550
Synapses are an early pathological target in many neurodegenerative diseases such as spinal muscular atrophy (SMA) and neuronal ceroid lipofuscinosis (NCLs). However, the reasons why synapses are particularly vulnerable to such a broad range of neurodegeneration inducing stimuli remains unknown. To identify molecular modulators of synaptic stability and degeneration, the Cln3-/- mouse model of a juvenile form of NCL. The molecular composition of anatomically-distinct, differentially-affected pre-synaptic populations were profiled and compared from the Cln3-/- mouse brain using proteomics followed by bioinformatic analyses. Identified protein candidates were then tested using a Drosophila CLN3 model to study their ability to modify the CLN3-neurodegenerative phenotype in vivo. Differential perturbations were identified in a range of molecular cascades correlating with synaptic vulnerability, including valine catabolism and rho signalling pathways. Genetic and pharmacological targeting of key 'hub' proteins in such pathways was sufficient to modulate phenotypic presentation in a Drosophila CLN3 model. It is proposed that such a workflow provides a target rich method for the identification of novel disease regulators which could be applicable to the study of other conditions where appropriate models exist.
Lo Piccolo, L., Jantrapirom, S., Nagai, Y. and Yamaguchi, M. (2017). FUS toxicity is rescued by the modulation of lncRNA hsromega expression in Drosophila melanogaster. Sci Rep 7(1): 15660. PubMed ID: 29142303
FUS is an aggregation-prone hnRNP involved in transcriptional and post-transcriptional regulation that aberrantly forms immunoreactive inclusion bodies in a range of neurological diseases classified as FUS-proteinopathies. Although FUS has been extensively examined, the underlying molecular mechanisms of these diseases have not yet been elucidated in detail. Previous work has shown that RNAi of the lncRNA hsromega altered the expression and sub-cellular localization of Drosophila FUS in the central nervous system of the fly. In order to obtain a clearer understanding of the role of hsromega in FUS toxicity, this study drove the expression of human FUS in Drosophila eyes with and without a hsromega RNAi background. hFUS was largely soluble and also able to form aggregates. As such, hFUS was toxic, inducing an aberrant eye morphology with the loss of pigmentation. The co-expression of hsromega double-stranded RNA reduced hFUS transcript levels and induced the formation of cytoplasmic non-toxic hFUS-LAMP1-insoluble inclusions. The combination of these events caused the titration of hFUS molar excess and a removal of hFUS aggregates to rescue toxicity. These results revealed the presence of a lncRNA-dependent pathway involved in the management of aggregation-prone hnRNPs, suggesting that properly formed FUS inclusions are not toxic to cells.
Perry, S., Han, Y., Das, A. and Dickman, D. (2017). Homeostatic plasticity can be induced and expressed to restore synaptic strength at neuromuscular junctions undergoing ALS-related degeneration. Hum Mol Genet 26(21): 4153-4167. PubMed ID: 28973139
Amyotrophic lateral sclerosis (ALS) is debilitating neurodegenerative disease characterized by motor neuron dysfunction and progressive weakening of the neuromuscular junction (NMJ). Hereditary ALS is strongly associated with variants in the human C9orf72 gene. This study characterized C9orf72 pathology at the Drosophila NMJ and utilized several approaches to restore synaptic strength in this model. First, a dramatic reduction was demonstrated in synaptic arborization and active zone number at NMJs following C9orf72 transgenic expression in motor neurons. Further, neurotransmission is similarly reduced at these synapses, consistent with severe degradation. However, despite these defects, C9orf72 synapses still retain the ability to express presynaptic homeostatic plasticity, a fundamental and adaptive form of NMJ plasticity in which perturbation to postsynaptic neurotransmitter receptors leads to a retrograde enhancement in presynaptic release. Next, it was shown that these endogenous but dormant homeostatic mechanisms can be harnessed to restore synaptic strength despite C9orf72 pathogenesis. Finally, activation of regenerative signaling is not neuroprotective in motor neurons undergoing C9orf72 toxicity. Together, these experiments define synaptic dysfunction at NMJs experiencing ALS-related degradation and demonstrate the potential to activate latent plasticity as a novel therapeutic strategy to restore synaptic strength.
Raj, A., Shah, P. and Agrawal, N. (2017). Sedentary behavior and altered metabolic activity by AgNPs ingestion in Drosophila melanogaster. Sci Rep 7(1): 15617. PubMed ID: 29142316
Among several nanoparticles, silver nanoparticles (AgNPs) are extensively used in a wide variety of consumer products due to its unique antimicrobial property. However, dosage effect of AgNPs on behavior and metabolic activity in an in vivo condition is not well studied. Therefore, to elucidate the impact of AgNPs on behavior and metabolism, systematic and detailed dosages study of AgNPs was performed by rearing Drosophila melanogaster on food without and with AgNPs. Dietary intake of AgNPs at early larval stage was shown to lead to behavioral abnormalities such as poor crawling and climbing ability of larvae and adults respectively. Interestingly, intake of higher dosage of AgNPs at larval stage significantly altered metabolic activity that includes lipid, carbohydrate and protein levels in adult flies. Further, detailed analysis revealed that AgNPs causes remarkable reduction in the number of lipid droplets (LDs) which are lipid storage organelles in Drosophila. An increased production of reactive oxygen species (ROS) was observed in AgNPs ingested larval tissues. These results strongly imply that higher dosage of AgNPs ingestion from early larval stage of Drosophila is inimical and thereby draws concern towards the usage of AgNPs in consumer goods.
Lee, G. J., Lim, J. J. and Hyun, S. (2017). Minocycline treatment increases resistance to oxidative stress and extends lifespan in Drosophila via FOXO. Oncotarget 8(50): 87878-87890. PubMed ID: 29152127
Minocycline is a semi-synthetic tetracycline derivative antibiotic that has received increasing attention for its non-antibiotic properties, mainly anti-inflammatory, tumor-suppressive, and neuroprotective effects. Drosophila is a widely used genetically tractable model organism for studying organismal aging by virtue of its short lifespan and ease of cultivation. This study examined the effects of minocycline on Drosophila lifespan and its associated traits. Minocycline-supplemented food significantly extended lifespan in both Canton S and w(1118) Drosophila strains. The drug-induced lifespan extension was not associated with reduced dietary intake or reduced female fecundity, but rather with increased resistance to an oxidative stressor (hydrogen peroxide). Notably, minocycline's effects on lifespan and resistance to oxidative stress were largely abrogated in Forkhead box O (FOXO) null mutant, and the drug treatment increased the activity of FOXO. These results may further understanding of minocycline's beneficial effects against several age-associated deteriorations observed in animal models.

Wednesday, January 17th

Murakami, A., Nagao, K., Juni, N., Hara, Y. and Umeda, M. (2017). An N-terminal di-proline motif is essential for fatty acid-dependent degradation of Delta9-desaturase in Drosophila. J Biol Chem [Epub ahead of print]. PubMed ID: 28972163
Delta9-fatty acid desaturase introduces a double bond at the Delta9 position of the acyl moiety of acyl-CoA and regulates the cellular levels of unsaturated fatty acids. However, it is unclear how Delta9-desaturase expression is regulated in response to changes in the levels of fatty acid desaturation. This study found that the degradation of DESAT1, the sole Delta9-desaturase in Drosophila cell line S2, was significantly enhanced when the amounts of unsaturated acyl chains of membrane phospholipids were increased by supplementation with unsaturated fatty acids, such as oleic and linoleic acids. In contrast, inhibition of DESAT1 activity remarkably suppressed its degradation. Of note, removal of DESAT1 N-terminal domain abolished the responsiveness of DESAT1 degradation to the level of fatty acid unsaturation. Further truncation and amino acid replacement analyses revealed that two sequential prolines, the second and third residues of DESAT1, were responsible for the unsaturated fatty acid-dependent degradation. Although degradation of mouse stearoyl-CoA desaturase 1 (SCD1) was unaffected by changes in fatty acid unsaturation, introduction of the N-terminal, sequential proline residues into SCD1 conferred responsiveness to unsaturated fatty acid-dependent degradation. Furthermore, this study also found that the Ca(2+)-dependent cysteine protease calpain is involved in the sequential proline-dependent degradation of DESAT1. In light of these findings, the sequential prolines at the second and third positions of DESAT1 were designated as a di-proline motif, which plays a crucial role in the regulation of Delta9-desaturase expression in response to changes in the level of cellular unsaturated fatty acids.
Meliambro, K., Wong, J. S., Ray, J., Calizo, R. C., Towne, S., Cole, B., El Salem, F., Gordon, R. E., Kaufman, L., He, J. C., Azeloglu, E. U. and Campbell, K. N. (2017). The Hippo pathway regulator KIBRA promotes podocyte injury by inhibiting YAP signaling and disrupting actin cytoskeletal dynamics. J Biol Chem. PubMed ID: 28982981
Evolutionary Homolog Study
(KIdney BRAin protein (KIBRA); see Drosophila Kibra), an upstream regulator of the Hippo signaling pathway encoded by the Wwc1 gene, shares the pro-injury properties of its putative binding partner dendrin and antagonizes the pro-survival signaling of the downstream Hippo pathway effector YAP (Yes-associated protein; see Drosophila Yorkie) in Drosophila and MCF10A cells. YAP has been identified an essential component of the glomerular filtration barrier that promotes podocyte survival by inhibiting dendrin pro-apoptotic function. This study tested the hypothesis that KIBRA promotes podocyte injury. Increased expression of KIBRA and phosphorylated YAP (P-YAP) protein in glomeruli of patients with biopsy-proven focal segmental glomerulosclerosis (FSGS). KIBRA/WWc1 overexpression in murine podocytes promoted LATS kinase phosphorylation, leading to subsequent YAP S127 phosphorylation, YAP cytoplasmic sequestration, and reduction in YAP target gene expression. Functionally, KIBRA overexpression induced significant morphological changes in podocytes including disruption of the actin cytoskeletal architecture and reduction of focal adhesion size and number, all of which were rescued by subsequent YAP overexpression. Conversely, constitutive KIBRA knockout mice displayed reduced P-YAP and increased YAP expression at baseline. These mice were protected from acute podocyte foot process effacement following protamine sulfate perfusion. KIBRA knockdown podocytes were also protected against protamine-induced injury. These findings suggest an important role for KIBRA in the pathogenesis of podocyte injury and the progression of proteinuric kidney disease.
Liu, T., Mahesh, G., Yu, W. and Hardin, P. E. (2017). CLOCK stabilizes CYCLE to initiate clock function in Drosophila. Proc Natl Acad Sci U S A 114(41): 10972-10977. PubMed ID: 28973907
The Drosophila circadian clock keeps time via transcriptional feedback loops. These feedback loops are initiated by CLOCK-CYCLE (CLK-CYC) heterodimers, which activate transcription of genes encoding the feedback repressors PERIOD and TIMELESS. Circadian clocks normally operate in approximately 150 brain pacemaker neurons and in many peripheral tissues in the head and body, but can also be induced by expressing CLK in nonclock cells. These ectopic clocks also require cyc, yet CYC expression is restricted to canonical clock cells despite evidence that cyc mRNA is widely expressed. This study shows that CLK binds to and stabilizes CYC in cell culture and in nonclock cells in vivo. Ectopic clocks also require the blue light photoreceptor CRYPTOCHROME (CRY), which is required for both light entrainment and clock function in peripheral tissues. These experiments define the genetic architecture required to initiate circadian clock function in Drosophila, reveal mechanisms governing circadian activator stability that are conserved in perhaps all eukaryotes, and suggest that Clk, cyc, and cry expression is sufficient to drive clock expression in naive cells.
Lin, X., Wang, F., Li, Y., Zhai, C., Wang, G., Zhang, X., Gao, Y., Yi, T., Sun, D. and Wu, S. (2017). The SCF ubiquitin ligase Slimb controls Nerfin-1 turnover in Drosophila. Biochem Biophys Res Commun. PubMed ID: 29154825
The C2H2 type zinc-finger transcription factor Nerfin-1 expresses dominantly in Drosophila nervous system and plays an important role in early axon guidance decisions and preventing neurons dedifferentiation. Recently, increasing reports indicated that INSM1 (homologue to nerfin-1 in mammals) is a useful marker for prognosis of neuroendocrine tumors. The dynamic expression of Nerfin-1 is regulated post-transcriptionally by multiple microRNAs; however, its post-translational regulation is still unclear. This study showed that the protein turnover of Nerfin-1 is regulated by Slimb, the substrate adaptor of SCF(Slimb) ubiquitin ligase complex. Mechanistically, Slimb associates with Nerfin-1 and promotes it ubiquitination and degradation in Drosophila S2R(+) cells. Furthermore, it was determined that the C-terminal half of Nerfin-1 (Nerfin-1(CT)) is required for its binding to Slimb. Genetic epistasis assays showed that Slimb misexpression antagonizes, while knock-down enhances the activity of Nerfin-1(CT) in Drosophila eyes. These data revealed a new link to understand the underlying mechanism for Nerfin-1 turnover in post-translational level, and provided useful insights in animal development and disease treatment by manipulating the activity of Slimb and Nerfin-1.
Maniati, K., Haralambous, K. J., Turin, L. and Skoulakis, E. M. C. (2017). Vibrational detection of odorant functional groups by Drosophila melanogaster. eNeuro 4(5). PubMed ID: 29094064
A remarkable feature of olfaction, and perhaps the hardest one to explain by shape-based molecular recognition, is the ability to detect the presence of functional groups in odorants, irrespective of molecular context. Previous work has shown that Drosophila trained to avoid deuterated odorants could respond to a molecule bearing a nitrile group, which shares the vibrational stretch frequency with the CD bond. This study reproduces and extends this finding by showing analogous olfactory responses of Drosophila to the chemically vastly different functional groups, thiols and boranes, that nevertheless possess a common vibration at 2600 cm(-1). Furthermore, it was shown that Drosophila do not respond to a cyanohydrin structure that renders nitrile groups invisible to IR spectroscopy. It is argued that the response of Drosophila to these odorants which parallels their perception in humans, supports the hypothesis that odor character is encoded in odorant molecular vibrations, not in the specific shape-based activation pattern of receptors.
Nagy, P., Szatmari, Z., Sandor, G. O., Lippai, M., Hegedus, K. and Juhasz, G. (2017). Drosophila Atg16 promotes enteroendocrine cell differentiation via regulation of intestinal Slit/Robo signaling. Development 144(21): 3990-4001. PubMed ID: 28982685
Genetic variations of Atg16l1, Slit2 and Rab19 predispose to the development of inflammatory bowel disease (IBD), but the relationship between these mutations is unclear. This study shows that in Drosophila guts lacking the WD40 domain of Atg16, pre-enteroendocrine (pre-EE) cells accumulate that fail to differentiate into properly functioning secretory EE cells. Mechanistically, loss of Atg16 or its binding partner Rab19 impairs Slit production, which normally inhibits EE cell generation by activating Robo signaling in stem cells. Importantly, loss of Atg16 or decreased Slit/Robo signaling triggers an intestinal inflammatory response. Surprisingly, analysis of Rab19 and domain-specific Atg16 mutants indicates that their stem cell niche regulatory function is independent of autophagy. These study reveals how mutations in these different genes may contribute to IBD.

Tuesday, January 16th

Kang, H., Jung, Y. L., McElroy, K. A., Zee, B. M., Wallace, H. A., Woolnough, J. L., Park, P. J. and Kuroda, M. I. (2017). Bivalent complexes of PRC1 with orthologs of BRD4 and MOZ/MORF target developmental genes in Drosophila. Genes Dev 31(19): 1988-2002. PubMed ID: 29070704
Regulatory decisions in Drosophila require Polycomb group (PcG) proteins to maintain the silent state and Trithorax group (TrxG) proteins to oppose silencing. Since PcG and TrxG are ubiquitous and lack apparent sequence specificity, a long-standing model is that targeting occurs via protein interactions; for instance, between repressors and PcG proteins. Instead, this study found that Pc-repressive complex 1 (PRC1) purifies with coactivators Fs(1)h [female sterile (1) homeotic] and Enok/Br140 during embryogenesis. Fs(1)h is a TrxG member and the ortholog of BRD4, a bromodomain protein that binds to acetylated histones and is a key transcriptional coactivator in mammals. Enok and Br140, another bromodomain protein, are orthologous to subunits of a mammalian MOZ/MORF acetyltransferase complex. This study confirmed PRC1-Br140 and PRC1-Fs(1)h interactions and identified their genomic binding sites. PRC1-Br140 bind developmental genes in fly embryos, with analogous co-occupancy of PRC1 and a Br140 ortholog, BRD1, at bivalent loci in human embryonic stem (ES) cells. It is proposed that identification of PRC1-Br140 "bivalent complexes" in fly embryos supports and extends the bivalency model posited in mammalian cells, in which the coexistence of H3K4me3 and H3K27me3 at developmental promoters represents a poised transcriptional state. It is further speculated that local competition between acetylation and deacetylation may play a critical role in the resolution of bivalent protein complexes during development.
Maksimov, D. A., Laktionov, P. P., Posukh, O. V., Belyakin, S. N. and Koryakov, D. E. (2017). Genome-wide analysis of SU(VAR)3-9 distribution in chromosomes of Drosophila melanogaster. Chromosoma [Epub ahead of print]. PubMed ID: 28975408
Histone modifications represent one of the key factors contributing to proper genome regulation. One of histone modifications involved in gene silencing is methylation of H3K9 residue. Present in the chromosomes across different eukaryotes, this epigenetic mark is controlled by SU(VAR)3-9 and its orthologs. Despite SU(VAR)3-9 was discovered over two decades ago, little is known about the details of its chromosomal distribution pattern. To fill in this gap, DamID-seq approach was used and high-resolution genome-wide profiles were obtained for SU(VAR)3-9 in two somatic (salivary glands and brain ganglia) and two germline (ovarian nurse cells and testes) tissues of Drosophila melanogaster. Analysis of tissue and developmental expression of SU(VAR)3-9-bound genes indicates that in the somatic tissues tested, as well as in the ovarian nurse cells, SU(VAR)3-9 tends to associate with transcriptionally silent genes. In contrast, in the testes, SU(VAR)3-9 shows preferential association with testis-specific genes, and its binding appears dynamic during spermatogenesis. In somatic cells, the mere presence/absence of SU(VAR)3-9 binding correlates with lower/higher expression. No such correlation is found in the male germline. Interestingly, transcription units in piRNA clusters (particularly flanks thereof) are frequently targeted by SU(VAR)3-9, and Su(var)3-9 mutation affects the expression of select piRNA species. This analyses suggest a context-dependent role of SU(VAR)3-9. In euchromatin, SU(VAR)3-9 may serve to fine-tune the expression of individual genes, whereas in heterochromatin, chromosome 4, and piRNA clusters, it may act more broadly over large chromatin domains.
Khuong, M. T., Fei, J., Cruz-Becerra, G. and Kadonaga, J. T. (2017). A simple and versatile system for the ATP-dependent assembly of chromatin. J Biol Chem 292(47): 19478-19490. PubMed ID: 28982979
Chromatin is the natural form of DNA in the eukaryotic nucleus and is the substrate for diverse biological phenomena. The functional analysis of these processes ideally would be carried out with nucleosomal templates that are assembled with customized core histones, DNA sequences, and chromosomal proteins. This study reports a simple, reliable, and versatile method for the ATP-dependent assembly of evenly spaced nucleosome arrays. This minimal chromatin assembly system comprises the Drosophila nucleoplasmin-like protein (dNLP) histone chaperone, the imitation switch (ISWI) ATP-driven motor protein, core histones, template DNA, and ATP. The dNLP and ISWI components were synthesized in bacteria, and each protein could be purified in a single step by affinity chromatography. The dNLP-ISWI system can be used with different DNA sequences, linear or circular DNA, bulk genomic DNA, recombinant or native Drosophila core histones, native human histones, the linker histone H1, the non-histone chromosomal protein HMGN2, and the core histone variants H3.3 and H2A.V. The dNLP-ISWI system should be accessible to a wide range of researchers and enable the assembly of customized chromatin with specifically desired DNA sequences, core histones, and other chromosomal proteins.
Melnikova, L., Kostyuchenko, M., Molodina, V., Parshikov, A., Georgiev, P. and Golovnin, A. (2017). Multiple interactions are involved in a highly specific association of the Mod(mdg4)-67.2 isoform with the Su(Hw) sites in Drosophila. Open Biol 7(10). PubMed ID: 29021216
The best-studied Drosophila insulator complex consists of two BTB-containing proteins, the Mod(mdg4)-67.2 isoform and CP190, which are recruited to the chromatin through interactions with the DNA-binding Su(Hw) protein. It was shown previously that Mod(mdg4)-67.2 is critical for the enhancer-blocking activity of the Su(Hw) insulators and it differs from more than 30 other Mod(mdg4) isoforms by the C-terminal domain required for a specific interaction with Su(Hw) only. The mechanism of the highly specific association between Mod(mdg4)-67.2 and Su(Hw) is not well understood. Therefore, a detailed analysis of domains involved in the interaction of Mod(mdg4)-67.2 with Su(Hw) and CP190 was performed. The N-terminal region of Su(Hw) interacts with the glutamine-rich domain common to all the Mod(mdg4) isoforms. The unique C-terminal part of Mod(mdg4)-67.2 contains the Su(Hw)-interacting domain and the FLYWCH domain that facilitates a specific association between Mod(mdg4)-67.2 and the CP190/Su(Hw) complex. Finally, interaction between the BTB domain of Mod(mdg4)-67.2 and the M domain of CP190 has been demonstrated. By using transgenic lines expressing different protein variants, this study has shown that all the newly identified interactions are to a greater or lesser extent redundant, which increases the reliability in the formation of the protein complexes.
Korandova, M., Krucek, T., Szakosova, K., Kodrik, D., Kuhnlein, R. P., Tomaskova, J. and Capkova Frydrychova, R. (2017). Chronic low-dose pro-oxidant treatment stimulates transcriptional activity of telomeric retroelements and increases telomere length in Drosophila. J Insect Physiol. PubMed ID: 29122549
It has been proposed that oxidative stress, elicited by high levels of reactive oxygen species, accelerates telomere shortening by erosion of telomeric DNA repeats. While most eukaryotes counteract telomere shortening by telomerase-driven addition of these repeats, telomeric loss in Drosophila is compensated by retrotransposition of the telomeric retroelements HeT-A, TART and TAHRE to chromosome ends. This study tested the effect of chronic exposure of flies to non-/sub-lethal doses of paraquat, which is a redox cycling compound widely used to induce oxidative stress in various experimental paradigms including telomere length analyses. Indeed, chronic paraquat exposure for five generations resulted in elevated transcriptional activity of both telomeric and non-telomeric transposable elements, and extended telomeric length in the tested fly lines. It is proposed that low oxidative stress leads to increased telomere length within Drosophila populations. For a mechanistic understanding of the observed phenomenon two scenarios are discusser: adaption, acting through a direct stimulation of telomere extension, or positive selection favoring individuals with longer telomeres within the population.
Yan, J., Chen, S. A., Local, A., Liu, T., Qiu, Y., Dorighi, K. M., Preissl, S., Rivera, C. M., Wang, C., Ye, Z., Ge, K., Hu, M., Wysocka, J. and Ren, B. (2018). Histone H3 lysine 4 monomethylation modulates long-range chromatin interactions at enhancers. Cell Res. PubMed ID: 29313530
Evolutionary Homolog Study
Long-range chromatin interactions between enhancers and promoters are essential for transcription of many developmentally controlled genes in mammals and other metazoans. Currently, the exact mechanisms that connect distal enhancers to their specific target promoters remain to be fully elucidated. This study shows that the enhancer-specific histone H3 lysine 4 monomethylation (H3K4me1) and the histone methyltransferases MLL3 and MLL4 (MLL3/4; see Drosophila Trithorax) play an active role in this process. This study demonstrates that in differentiating mouse embryonic stem cells, MLL3/4-dependent deposition of H3K4me1 at enhancers correlates with increased levels of chromatin interactions, whereas loss of this histone modification leads to reduced levels of chromatin interactions and defects in gene activation during differentiation. H3K4me1 facilitates recruitment of the Cohesin complex, a known regulator of chromatin organization, to chromatin in vitro and in vivo, providing a potential mechanism for MLL3/4 to promote chromatin interactions between enhancers and promoters. Taken together, these results support a role for MLL3/4-dependent H3K4me1 in orchestrating long-range chromatin interactions at enhancers in mammalian cells.

Monday, January 15th

Li, Q., Li, Y., Wang, X., Qi, J., Jin, X., Tong, H., Zhou, Z., Zhang, Z. C. and Han, J. (2017). Fbxl4 serves as a clock output molecule that regulates sleep through promotion of rhythmic degradation of the GABAA receptor. Curr Biol [Epub ahead of print]. PubMed ID: 29174887
The timing of sleep is tightly governed by the circadian clock, which contains a negative transcriptional feedback loop and synchronizes the physiology and behavior of most animals to daily environmental oscillations. However, how the circadian clock determines the timing of sleep is largely unclear. In vertebrates and invertebrates, the status of sleep and wakefulness is modulated by the electrical activity of pacemaker neurons that are circadian regulated and suppressed by inhibitory GABAergic inputs. This study showed that Drosophila GABAA receptors undergo rhythmic degradation in arousal-promoting large ventral lateral neurons (lLNvs) and their expression level in lLNvs displays a daily oscillation. This study also demonstrated that the E3 ligase Fbxl4 promotes GABAA receptor ubiquitination and degradation and revealed that the transcription of fbxl4 in lLNvs is CLOCK dependent. Finally, it was demonstrated that Fbxl4 regulates the timing of sleep through rhythmically reducing GABA sensitivity to modulate the excitability of lLNvs. This study uncovered a critical molecular linkage between the circadian clock and the electrical activity of pacemaker neurons and demonstrated that CLOCK-dependent Fbxl4 expression rhythmically downregulates GABAA receptor level to increase the activity of pacemaker neurons and promote wakefulness.
Majot, A. T. and Bidwai, A. P. (2017). Analysis of transient hypermorphic activity of E(spl)D during R8 specification. PLoS One 12(10): e0186439. PubMed ID: 29036187
Drosophila atonal (ato) is required for the specification of founding R8 photoreceptors during retinal development. ato is regulated via dual eye-specific enhancers; ato-3' is subject to initial induction whereas 5'-ato facilitates Notch-mediated autoregulation. Notch is further utilized to induce bHLH repressors of the E(spl) locus to restrict Ato from its initial broad expression to individual cells. While Notch operates in two, distinct phases, it has remained unclear how the two phases maintain independence from one another. To probe this mechanism, evidence is provided that although E(spl) exert no influence on ato-3', E(spl) repress 5'-ato and deletion of the E(spl) locus elicits precocious 5'-ato activity. Thus, E(spl) imposes a delay to the timing in which Ato initiates autoregulation. Next, attempts were made to understand this finding in the context of E(spl)D, which encodes a dysregulated variant of E(spl)M8 that perturbs R8 patterning, though, as previously reported, only in conjunction with the mutant receptor Nspl. This study established a genetic interaction between E(spl)D and roughened eye (roe), a known modulator of Notch signaling in retinogenesis. This link further suggests a dosage-dependence between E(spl) and the proneural activators Ato and Sens, as indicated via interaction assays in which E(spl)D renders aberrant R8 patterning in conjunction with reduced proneural dosage. In total, the biphasicity of Notch signaling relies, to some degree, on the post-translational regulation of individual E(spl) members and, importantly, that post-translational regulation is likely necessary to modulate the level of E(spl) activity throughout the progression of Ato expression.
Kotoula, V., Moressis, A., Semelidou, O. and Skoulakis, E. M. C. (2017). Drk-mediated signaling to Rho kinase is required for anesthesia-resistant memory in Drosophila. Proc Natl Acad Sci U S A 114(41): 10984-10989. PubMed ID: 28973902
Anesthesia-resistant memory (ARM) was described decades ago, but the mechanisms that underlie this protein synthesis-independent form of consolidated memory in Drosophila remain poorly understood. Whether the several signaling molecules, receptors, and synaptic proteins currently implicated in ARM operate in one or more pathways and how they function in the process remain unclear. This study presents evidence that Drk, the Drosophila ortholog of the adaptor protein Grb2, is essential for ARM within adult mushroom body neurons. Significantly, Drk signals engage the Rho kinase Drok, implicating dynamic cytoskeletal changes in ARM, and this is supported by reduced F-actin in the mutants and after pharmacological inhibition of Drok. Interestingly, Drk-Drok signaling appears independent of the function of Radish (Rsh), a protein long implicated in ARM, suggesting that the process involves at least two distinct molecular pathways. Based on these results, it is proposed that signaling pathways involved in structural plasticity likely underlie this form of translation-independent memory.
Li, H., Horns, F., Wu, B., Xie, Q., Li, J., Li, T., Luginbuhl, D. J., Quake, S. R. and Luo, L. (2017). Classifying Drosophila olfactory projection neuron subtypes by single-cell RNA sequencing. Cell 171(5): 1206-1220 PubMed ID: 29149607
The definition of neuronal type and how it relates to the transcriptome are open questions. Drosophila olfactory projection neurons (PNs) are among the best-characterized neuronal types: different PN classes target dendrites to distinct olfactory glomeruli, while PNs of the same class exhibit indistinguishable anatomical and physiological properties. Using single-cell RNA sequencing, this study comprehensively characterized the transcriptomes of most PN classes and unequivocally mapped transcriptomes to specific olfactory function for six classes. Transcriptomes of closely related PN classes exhibit the largest differences during circuit assembly but become indistinguishable in adults, suggesting that neuronal subtype diversity peaks during development. Transcription factors and cell-surface molecules are the most differentially expressed genes between classes and are highly informative in encoding cell identity, enabling identification of a new lineage-specific transcription factor that instructs PN dendrite targeting. These findings establish that neuronal transcriptomic identity corresponds with anatomical and physiological identity defined by connectivity and function.
Marchetti, G. and Tavosanis, G. (2017). Steroid hormone ecdysone signaling specifies mushroom body neuron sequential fate via Chinmo. Curr Biol 27(19): 3017-3024.e3014. PubMed ID: 28966087
The functional variety in neuronal composition of an adult brain is established during development. Recent studies proposed that interactions between genetic intrinsic programs and external cues are necessary to generate proper neural diversity. However, the molecular mechanisms underlying this developmental process are still poorly understood. Three main subtypes of Drosophila mushroom body (MB) neurons are sequentially generated during development and provide a good example of developmental neural plasticity. The present data propose that the environmentally controlled steroid hormone ecdysone functions as a regulator of early-born MB neuron fate during larval-pupal transition. The BTB-zinc finger factor Chinmo acts upstream of ecdysone signaling to promote a neuronal fate switch. Indeed, Chinmo regulates the expression of the ecdysone receptor B1 isoform to mediate the production of gamma and alpha'beta' MB neurons. In addition, genetic evidence is provided for a regulatory negative feedback loop driving the alpha'beta' to alphabeta MB neuron transition in which ecdysone signaling in turn controls microRNA let-7 depression of Chinmo expression. Thus, these results uncover a novel interaction in the MB neural specification pathway for temporal control of neuronal identity by interplay between an extrinsic hormonal signal and an intrinsic transcription factor cascade.
Niens, J., Reh, F., Coban, B., Cichewicz, K., Eckardt, J., Liu, Y. T., Hirsh, J. and Riemensperger, T. D. (2017). Dopamine modulates serotonin innervation in the Drosophila brain. Front Syst Neurosci 11: 76. PubMed ID: 29085286
Parkinson's disease (PD) results from a progressive degeneration of the dopaminergic nigrostriatal system leading to a decline in movement control, with resting tremor, rigidity and postural instability. Several aspects of PD can be modeled in the fruit fly, Drosophila melanogaster, including alpha-synuclein-induced degeneration of dopaminergic neurons, or dopamine (DA) loss by genetic elimination of neural DA synthesis. Defective behaviors in this latter model can be ameliorated by feeding the DA precursor L-DOPA, analogous to the treatment paradigm for PD. Secondary complication from L-DOPA treatment in PD patients are associated with ectopic synthesis of DA in serotonin (5-HT)-releasing neurons, leading to DA/5-HT imbalance. This study examined the neuro-anatomical adaptations resulting from imbalanced DA/5-HT signaling in Drosophila mutants lacking neural DA. Similar to rodent models of PD, lack of DA leads to increased 5-HT levels and arborizations in specific brain regions. Conversely, increased DA levels by L-DOPA feeding leads to reduced connectivity of 5-HT neurons to their target neurons in the mushroom body (MB). The observed alterations of 5-HT neuron plasticity indicate that loss of DA signaling is not solely responsible for the behavioral disorders observed in Drosophila models of PD, but rather a combination of the latter with alterations of 5-HT circuitry.

Friday, January 12th

Liu, T., Li, L., Li, B. and Zhan, G. (2017). Phosphine inhibits transcription of the catalase gene through the DRE/DREF system in Drosophila melanogaster Sci Rep 7(1): 12913. PubMed ID: 29018235
Phosphine (PH3) is a toxin commonly used for pest control. Its toxicity is attributed primarily to its ability to induce oxidative damage. Previous work showed that phosphine could disrupt the cell antioxidant defence system by inhibiting expression of the catalase gene in Drosophila melanogaster (DmCAT). However, the exact mechanism of this inhibition remains unclear. This study implemented a luciferase reporter assay driven by the DmCAT promoter in D. melanogaster S2 cells and showed that this reporter could be inhibited by phosphine treatment. A minimal fragment of the promoter (-94 to 0 bp), which contained a DNA replication-related element (DRE) consensus motif (-78 to -85 bp), was sufficient for phosphine-mediated reporter inhibition, suggesting the involvement of the transcription factor DREF. Furthermore, phosphine treatment led to a reduction in DREF expression and consequent repression of DmCAT transcription. These results provide new insights on the molecular mechanism of phosphine-mediated catalase inhibition. Phosphine treatment leads to reduced levels of the transcription factor DREF, a positive regulator of the DmCAT gene, thereby resulting in the repression of DmCAT at transcriptional level.
Lockwood, B. L., Julick, C. R. and Montooth, K. L. (2017). Maternal loading of a small heat shock protein increases embryo thermal tolerance in Drosophila melanogaster. J Exp Biol 220(Pt 23): 4492-4501. PubMed ID: 29097593
Maternal investment is likely to have direct effects on offspring survival. In oviparous animals whose embryos are exposed to the external environment, maternal provisioning of molecular factors like mRNAs and proteins may help embryos cope with sudden changes in the environment. This study sought to modify the maternal mRNA contribution to offspring embryos and test for maternal effects on acute thermal tolerance in early embryos of Drosophila melanogaster, In vivo overexpression of a small heat shock protein gene (Hsp23) was driven in female ovaries, and the effects were measured of acute thermal stress on offspring embryonic survival and larval development. Overexpression of the Hsp23 gene in female ovaries produced offspring embryos with increased thermal tolerance. Brief heat stress in the early embryonic stage (0-1 h old) caused decreased larval performance later in life (5-10 days old), as indexed by pupation height. Maternal overexpression of Hsp23 protected embryos against this heat-induced defect in larval performance. These data demonstrate that transient products of single genes have large and lasting effects on whole-organism environmental tolerance. Further, the results suggest that maternal effects have a profound impact on offspring survival in the context of thermal variability.
Kang, P., Chang, K., Liu, Y., Bouska, M., Birnbaum, A., Karashchuk, G., Thakore, R., Zheng, W., Post, S., Brent, C. S., Li, S., Tatar, M. and Bai, H. (2017). Drosophila Kruppel homolog 1 represses lipolysis through interaction with dFOXO. Sci Rep 7(1): 16369. PubMed ID: 29180716
Transcriptional coordination is a vital process contributing to metabolic homeostasis. As one of the key nodes in the metabolic network, the forkhead transcription factor FOXO has been shown to interact with diverse transcription co-factors and integrate signals from multiple pathways to control metabolism, oxidative stress response, and cell cycle. Recently, insulin/FOXO signaling has been implicated in the regulation of insect development via the interaction with insect hormones, such as ecdysone and juvenile hormone. This study identified an interaction between Drosophila FOXO (dFOXO) and the zinc finger transcription factor Kruppel homolog 1 (Kr-h1), one of the key players in juvenile hormone signaling. Kr-h1 mutants show delayed larval development and altered lipid metabolism, in particular induced lipolysis upon starvation. Notably, Kr-h1 physically and genetically interacts with dFOXO in vitro and in vivo to regulate the transcriptional activation of insulin receptor (InR) and adipose lipase brummer (bmm). The transcriptional co-regulation by Kr-h1 and dFOXO may represent a broad mechanism by which Kruppel-like factors integrate with insulin signaling to maintain metabolic homeostasis and coordinate organism growth.
Hindle, S. J., Munji, R. N., Dolghih, E., Gaskins, G., Orng, S., Ishimoto, H., Soung, A., DeSalvo, M., Kitamoto, T., Keiser, M. J., Jacobson, M. P., Daneman, R. and Bainton, R. J. (2017).. Evolutionarily conserved roles for blood-brain barrier xenobiotic transporters in endogenous steroid partitioning and behavior. Cell Rep 21(5): 1304-1316. PubMed ID: 29091768
Central nervous system (CNS) chemical protection depends upon discrete control of small-molecule access by the blood-brain barrier (BBB). Curiously, some drugs cause CNS side-effects despite negligible transit past the BBB. To investigate this phenomenon, it was asked whether the highly BBB-enriched drug efflux transporter MDR1 has dual functions in controlling drug and endogenous molecule CNS homeostasis. If this is true, then brain-impermeable drugs could induce behavioral changes by affecting brain levels of endogenous molecules. Using computational, genetic, and pharmacologic approaches across diverse organisms, it was demonstrated that BBB-localized efflux transporters are critical for regulating brain levels of endogenous steroids and steroid-regulated behaviors (sleep in Drosophila and anxiety in mice). Furthermore, MDR1-interacting drugs were shown to be associated with anxiety-related behaviors in humans. A general mechanism is proposed for common behavioral side effects of prescription drugs: pharmacologically challenging BBB efflux transporters disrupts brain levels of endogenous substrates and implicates the BBB in behavioral regulation.
Housden, B. E., Li, Z., Kelley, C., Wang, Y., Hu, Y., Valvezan, A. J., Manning, B. D. and Perrimon, N. (2017). Improved detection of synthetic lethal interactions in Drosophila cells using variable dose analysis (VDA). Proc Natl Acad Sci U S A. PubMed ID: 29183982
Synthetic sick or synthetic lethal (SS/L) screens are a powerful way to identify candidate drug targets to specifically kill tumor cells, but this approach generally suffers from low consistency between screens. This study found that many SS/L interactions involve essential genes and are therefore detectable within a limited range of knockdown efficiency. Such interactions are often missed by overly efficient RNAi reagents. An assay was therefore developed that measures viability over a range of knockdown efficiency within a cell population. This method, called Variable Dose Analysis (VDA), is highly sensitive to viability phenotypes and reproducibly detects SS/L interactions. The VDA method was applied to search for SS/L interactions with TSC1 and TSC2, the two tumor suppressors underlying tuberous sclerosis complex (TSC), and generated a SS/L network for TSC. Using this network, four Food and Drug Administration-approved drugs were identified that selectively affect viability of TSC-deficient cells, representing promising candidates for repurposing to treat TSC-related tumors.
Liu, L., MacKenzie, K. R., Putluri, N., Maletic-Savatic, M. and Bellen, H. J. (2017). The glia-geuron lactate shuttle and elevated ROS promote lipid synthesis in neurons and lipid droplet accumulation in glia via APOE/D. Cell Metab 26(5): 719-737.e716. PubMed ID: 28965825
Elevated reactive oxygen species (ROS) induce the formation of lipids in neurons that are transferred to glia, where they form lipid droplets (LDs). This study shows that glial and neuronal monocarboxylate transporters (MCTs), fatty acid transport proteins (FATPs), and apolipoproteins are critical for glial LD formation. MCTs enable glia to secrete and neurons to absorb lactate, which is converted to pyruvate and acetyl-CoA in neurons. Lactate metabolites provide a substrate for synthesis of fatty acids, which are processed and transferred to glia by FATP and apolipoproteins. In the presence of high ROS, inhibiting lactate transfer or lowering FATP or apolipoprotein levels decreases glial LD accumulation in flies and in primary mouse glial-neuronal cultures. Human APOE can substitute for a fly glial apolipoprotein, and APOE4, an Alzheimer's disease susceptibility allele, is impaired in lipid transport and promotes neurodegeneration, providing insights into disease mechanisms.

Thursday, January 11th

Couturier, L., Mazouni, K., Bernard, F., Besson, C., Reynaud, E. and Schweisguth, F. (2017). Regulation of cortical stability by RhoGEF3 in mitotic sensory organ precursor cells in Drosophila. Biol Open. PubMed ID: 29101098
In epithelia, mitotic cells round up and push against their neighbors to divide. Mitotic rounding results from increased assembly of F-actin and cortical recruitment of Myosin II, leading to increased cortical stability. Whether this process is developmentally regulated is not well known. This study examined the regulation of cortical stability in Sensory Organ precursor cells (SOPs) in the Drosophila pupal notum. SOPs differed in apical shape and actomyosin dynamics from their epidermal neighbors prior to division and appeared to have a more rigid cortex at mitosis. This study identified RhoGEF3 as an actin regulator expressed at higher levels in SOPs and showed that RhoGEF3 had in vitro GTPase Exchange Factor (GEF) activity for Cdc42. Additionally, RhoGEF3 genetically interacted with both Cdc42 and Rac1 when over-expressed in the fly eye. Using a null RhoGEF3 mutation generated by CRISPR-mediated homologous recombination, this study showed using live imaging that the RhoGEF3 gene, despite being dispensable for normal development, contributed to cortical stability in dividing SOPs. It is therefore suggested that cortical stability is developmentally regulated in dividing SOPs of the fly notum.
Kim, S., Nahm, M., Kim, N., Kwon, Y., Kim, J., Choi, S., Choi, E. Y., Shim, J., Lee, C. and Lee, S. (2017). Graf regulates hematopoiesis through GEEC endocytosis of EGFR. Development 144(22): 4159-4172. PubMed ID: 28993397
GTPase regulator associated with focal adhesion kinase 1 (GRAF1) is an essential component of the GPI-enriched endocytic compartment (GEEC) endocytosis pathway. Mutations in the human GRAF1 gene are associated with acute myeloid leukemia, but its normal role in myeloid cell development remains unclear. This study shows that Graf, the Drosophila ortholog of GRAF1, is expressed and specifically localizes to GEEC endocytic membranes in macrophage-like plasmatocytes. Loss of Graf impairs GEEC endocytosis, enhances EGFR signaling and induces a plasmatocyte overproliferation phenotype that requires the EGFR signaling cascade. Mechanistically, Graf-dependent GEEC endocytosis serves as a major route for EGFR internalization at high, but not low, doses of the predominant Drosophila EGFR ligand Spitz (Spi), and is indispensable for efficient EGFR degradation and signal attenuation. Finally, Graf interacts directly with EGFR in a receptor ubiquitylation-dependent manner, suggesting a mechanism by which Graf promotes GEEC endocytosis of EGFR at high Spi. Based on these findings, a model is proposed in which Graf functions to downregulate EGFR signaling by facilitating Spi-induced receptor internalization through GEEC endocytosis, thereby restraining plasmatocyte proliferation.
Kaur, P., Saunders, T. E. and Tolwinski, N. S. (2017). Coupling optogenetics and light-sheet microscopy, a method to study Wnt signaling during embryogenesis. Sci Rep 7(1): 16636. PubMed ID: 29192250
Optogenetics allows precise, fast and reversible intervention in biological processes. Light-sheet microscopy allows observation of the full course of Drosophila embryonic development from egg to larva. Bringing the two approaches together allows unparalleled precision into the temporal regulation of signaling pathways and cellular processes in vivo. To develop this method,the regulation of canonical Wnt signaling was investigated during anterior-posterior patterning of the Drosophila embryonic epidermis. Cryptochrome 2 (CRY2; see Drosophila Crptochrome) from Arabidopsis Thaliana was fused to mCherry fluorescent protein and Drosophila beta-catenin to form an easy to visualize optogenetic switch. Blue light illumination caused oligomerization of the fusion protein and inhibited downstream Wnt signaling in vitro and in vivo. Temporal inactivation of beta-catenin confirmed that Wnt signaling is required not only for Drosophila pattern formation, but also for maintenance later in development. It is anticipate that this method will be easily extendable to other developmental signaling pathways and many other experimental systems.
Jiang, K., Liu, Y., Zhang, J. and Jia, J. (2017). An intracellular activation of Smoothened independent of Hedgehog stimulation in Drosophila. J Cell Sci. PubMed ID: 29142103
Smoothened (Smo), a GPCR family protein, plays a critical role in the reception and transduction of Hedgehog (Hh) signal. Smo is phosphorylated and activated on the cell surface, however, it is unknown whether Smo can be intracellularly activated. This study demonstrates that inactivation of the ESCRT-III causes dramatic accumulation of Smo, and subsequent activation of Hh signaling. In contrast, inactivation of ESCRTs 0-II induces mild Smo accumulation. Evidence is porvided that Kurtz (Krz), the Drosophila beta-arrestin2, acts in parallel with the ESCRTs 0-II pathway to sort Smo to the multivesicular bodies and lysosome-mediated degradation. Additionally, upon inactivation of ESCRT-III, all active and inactive forms of Smo are accumulated. Endogenous Smo accumulated by ESCRT-III inactivation is highly activated, which is induced by phosphorylation but not sumoylation. Together, these findings demonstrate a model for intracellular activation of Smo, raising the possibility for tissue overgrowth caused by an excessive amount, rather than mutation of Smo.
Gibbs, E. B., Laremore, T. N., Usher, G. A., Portz, B., Cook, E. C. and Showalter, S. A. (2017). Substrate Specificity of the Kinase P-TEFb towards the RNA Polymerase II C-Terminal Domain. Biophys J 113(9): 1909-1911. PubMed ID: 28992937
The positive transcription elongation factor b (P-TEFb) promotes transcription elongation through phosphorylation of the RNA polymerase II C-terminal domain. This process is not well understood, partly due to difficulties in determining the specificity of P-TEFb toward the various heptad repeat motifs within the C-terminal domain. A simple assay using mass spectrometry was developed to identify the substrate specificity of the Drosophila melanogaster P-TEFb (DmP-TEFb) in vitro. This assay demonstrated that DmP-TEFb preferentially phosphorylates Ser5 and, surprisingly, that pre-phosphorylation or conserved amino acid variation at the 7-position in the heptad can alter DmP-TEFb specificity, leading to the creation of distinct double-phosphorylation marks.
Li, W., Li, W., Zou, L., Ji, S., Li, C., Liu, K., Zhang, G., Sun, Q., Xiao, F. and Chen, D. (2017). Membrane targeting of inhibitory Smads through palmitoylation controls TGF-beta/BMP signaling. Proc Natl Acad Sci U S A. PubMed ID: 29180412
TGF-beta/BMP (bone morphogenetic protein) signaling pathways play conserved roles in controlling embryonic development, tissue homeostasis, and stem cell regulation. Inhibitory Smads (I-Smads) have been shown to negatively regulate TGF-beta/BMP signaling by primarily targeting the type I receptors for ubiquitination and turnover. However, little is known about how I-Smads access the membrane to execute their functions. This study shows that Dad, the Drosophila I-Smad, associates with the cellular membrane via palmitoylation, thereby targeting the BMP type I receptor for ubiquitination. By performing systematic biochemistry assays, the specific cysteine (Cys556) essential for Dad palmitoylation and membrane association was characterized. Moreover, it was demonstrated that dHIP14, a Drosophila palmitoyl acyl-transferase, catalyzes Dad palmitoylation, thereby inhibiting efficient BMP signaling. Thus, these findings uncover a modification of the inhibitory Smads that controls TGF-beta/BMP signaling activity.

Wednesday, January 10th

Kadas, D., Klein, A., Krick, N., Worrell, J. W., Ryglewski, S. and Duch, C. (2017). Dendritic and axonal L-type calcium channels cooperate to enhance motoneuron firing output during Drosophila larval locomotion. J Neurosci 37(45): 10971-10982. PubMed ID: 28986465
Behaviorally adequate neuronal firing patterns are critically dependent on the specific types of ion channel expressed and on their subcellular localization. This study combines in situ electrophysiology with genetic and pharmacological intervention in larval Drosophila melanogaster of both sexes to address localization and function of L-type like calcium channels in motoneurons. Dmca1D (Cav1 homolog) L-type like calcium channels localize to both the somatodendritic and the axonal compartment of larval crawling motoneurons. In situ patch-clamp recordings in genetic mosaics reveal that Dmca1D channels increase burst duration and maximum intraburst firing frequencies during crawling-like motor patterns in semi-intact animals. Genetic and acute pharmacological manipulations suggest that prolonged burst durations are caused by dendritically localized Dmca1D channels, which activate upon cholinergic synaptic input and amplify EPSPs, thus indicating a conserved function of dendritic L-type channels from Drosophila to vertebrates. By contrast, maximum intraburst firing rates require axonal calcium influx through Dmca1D channels, likely to enhance sodium channel de-inactivation via a fast afterhyperpolarization through BK channel activation. Therefore, in unmyelinated Drosophila motoneurons different functions of axonal and dendritic L-type like calcium channels likely operate synergistically to maximize firing output during locomotion.
Kistenpfennig, C., Nakayama, M., Nihara, R., Tomioka, K., Helfrich-Forster, C. and Yoshii, T. (2017). A Tug-of-War between Cryptochrome and the visual system allows the adaptation of evening activity to long photoperiods in Drosophila melanogaster. J Biol Rhythms: 748730417738612. PubMed ID: 29179610
In many animals, the circadian clock plays a role in adapting to the coming season by measuring day length. The mechanism for measuring day length and its neuronal circuits remains elusive, however. Under laboratory conditions, the fruit fly, Drosophila melanogaster, displays 2 activity peaks: one in the morning and one in the evening. These peaks appear to be regulated by 2 separate circadian oscillators (the morning and evening oscillators) that reside in different subsets of pacemaker clock neurons in the brain. The morning and evening activity peaks can flexibly change their phases to adapt to different photoperiods by tracking dawn and dusk, respectively. This study found that cryptochrome (CRY) in the evening oscillators (the fifth small ventral lateral neuron [5th s-LNv] and the dorsal lateral neurons [LNds]) limits the ability of the evening peak to track dusk during long days. In contrast, light signaling from the external photoreceptors (compound eyes, ocelli, and Hofbauer-Buchner eyelets) increases the ability of the evening peak to track dusk. At the molecular level, CRY signaling dampens the amplitude of PAR-domain protein 1 (PDP1) oscillations in most clock neurons during long days, whereas signaling from the visual system increases these amplitudes. Thus, these results suggest that light inputs from the two major circadian photoreceptors, CRY and the visual system, have opposite effects on day length adaptation. Their tug-of-war appears to determine the precise phase adjustment of evening activity.
Jang, Y. H., Chae, H. S. and Kim, Y. J. (2017). Female-specific myoinhibitory peptide neurons regulate mating receptivity in Drosophila melanogaster. Nat Commun 8(1): 1630. PubMed ID: 29158481
Upon mating, fruit fly females become refractory to further mating for several days. An ejaculate protein called sex peptide (SP) acts on uterine neurons to trigger this behavioural change, but it is still unclear how the SP signal modifies the mating decision. This study describes two groups of female-specific local interneurons that are important for this process-the ventral abdominal lateral (vAL) and ventral abdominal medial (vAM) interneurons. Both vAL and vAM express myoinhibitory peptide (Mip)-GAL4. vAL is positive for Mip neuropeptides and the sex-determining transcriptional factor doublesex. Silencing the Mip neurons in females induces active rejection of male courtship attempts, whereas activation of the Mip neurons makes even mated females receptive to re-mating. vAL and vAM are located in the abdominal ganglion (AG) where they relay the SP signal to other AG neurons that project to the brain. Mip neuropeptides appear to promote mating receptivity both in virgins and mated females, although it is dispensable for normal mating in virgin females.
Klein, M., Krivov, S. V., Ferrer, A. J., Luo, L., Samuel, A. D. and Karplus, M. (2017). Exploratory search during directed navigation in C. elegans and Drosophila larva. Elife 6. PubMed ID: 29083306
Many organisms-from bacteria to nematodes to insect larvae-navigate their environments by biasing random movements. In these organisms, navigation in isotropic environments can be characterized as an essentially diffusive and undirected process. In stimulus gradients, movement decisions are biased to drive directed navigation toward favorable environments. How does directed navigation in a gradient modulate random exploration either parallel or orthogonal to the gradient? This study introduces methods originally used for analyzing protein folding trajectories to study the trajectories of the nematode Caenorhabditis elegans and the Drosophila larva in isotropic environments, as well as in thermal and chemical gradients. The statistics of random exploration in any direction are little affected by directed movement along a stimulus gradient. A key constraint on the behavioral strategies of these organisms appears to be the preservation of their capacity to continuously explore their environments in all directions even while moving toward favorable conditions.
Koppik, M. and Fricke, C. (2017). Gene expression changes in male accessory glands during ageing are accompanied by reproductive decline in Drosophila melanogaster. Mol Ecol. PubMed ID: 29055154
Senescence is accompanied by loss of reproductive functions. Reproductive ageing was studied in Drosophila melanogaster males and asked whether the expected decline in male reproductive success is due to diminished functionality of the male accessory gland (AG). The male AG produces the majority of seminal fluid proteins (SFPs) transferred to the female at mating. SFPs induce female postmating changes and are key to male reproductive success. Sge-dependent gene expression changes were measured for five representative SFP genes in males from four different age groups ranging from 1 to 6 weeks after eclosion. Simultaneously, male reproductive success was measured in postmating traits mediated by transfer of these five SFPs. A decreased was found in male SFP gene expression with advancing age and an accompanying decline in male postmating success. Hence, male reproductive senescence is associated with a decline in functionality of the male AG. While overall individual SFP genes decreased in expression, these results point towards the idea that the composition of an ejaculate might change with male age as the rate of change was variable for those five genes.
Lebreton, S., et al. (2017). A Drosophila female pheromone elicits species-specific long-range attraction via an olfactory channel with dual specificity for sex and food. Journal
Mate finding and recognition in animals evolves during niche adaptation and involves social signals and habitat cues. Drosophila melanogaster and related species are known to be attracted to fermenting fruit for feeding and egg-laying, which poses the question of whether species-specific fly odours contribute to long-range premating communication. This study has discovered an olfactory channel in D. melanogaster with a dual affinity to sex and food odorants. Female flies release a pheromone, (Z)-4-undecenal (Z4-11Al), that elicits flight attraction in both sexes. Its biosynthetic precursor is the cuticular hydrocarbon (Z,Z)-7,11-heptacosadiene (7,11-HD), which is known to afford reproductive isolation between the sibling species D. melanogaster and D. simulans during courtship. Twin olfactory receptors, Or69aB and Or69aA, are tuned to Z4-11Al and food odorants, respectively. They are co-expressed in the same olfactory sensory neurons, and feed into a neural circuit mediating species-specific, long-range communication; however, the close relative D. simulans, which shares food resources with D. melanogaster, does not respond to Z4-11Al. The Or69aA and Or69aB isoforms have adopted dual olfactory traits. The underlying gene yields a collaboration between natural and sexual selection, which has the potential to drive speciation.

Tuesday, January 9th

Dewar, A. D. M., Wystrach, A., Philippides, A. and Graham, P. (2017). Neural coding in the visual system of Drosophila melanogaster: How do small neural populations support visually guided behaviours?. PLoS Comput Biol 13(10): e1005735. PubMed ID: 29016606
All organisms wishing to survive and reproduce must be able to respond adaptively to a complex, changing world. Yet the computational power available is constrained by biology and evolution, favouring mechanisms that are parsimonious yet robust. This study investigated the information carried in small populations of visually responsive neurons in Drosophila melanogaster. These so-called 'ring neurons', projecting to the ellipsoid body of the central complex, are reported to be necessary for complex visual tasks such as pattern recognition and visual navigation. Recently the receptive fields of these neurons have been mapped, allowing an investigation of how well they can support such behaviours. For instance, in a simulation of classic pattern discrimination experiments, this study shows that the pattern of output from the ring neurons matches observed fly behaviour. However, performance of the neurons (as with flies) is not perfect and can be easily improved with the addition of extra neurons, suggesting the neurons' receptive fields are not optimised for recognising abstract shapes, a conclusion which casts doubt on cognitive explanations of fly behaviour in pattern recognition assays. Using artificial neural networks, it was then assessed how easy it is to decode more general information about stimulus shape from the ring neuron population codes. These neurons were shown to be well suited for encoding information about size, position and orientation, which are more relevant behavioural parameters for a fly than abstract pattern properties. This leads to a suggestion that in order to understand the properties of neural systems, one must consider how perceptual circuits put information at the service of behaviour.
Jacob, V., Scolari, F., Delatte, H., Gasperi, G., Jacquin-Joly, E., Malacrida, A. R. and Duyck, P. F. (2017). Current source density mapping of antennal sensory selectivity reveals conserved olfactory systems between tephritids and Drosophila. Sci Rep 7(1): 15304. PubMed ID: 29127313
Ecological specialization of insects involves the functional and morphological reshaping of olfactory systems. Little is known about the degree to which insect sensitivity to odorant compounds is conserved between genera, tribes, or families. This study compared the olfactory systems of six tephritid fruit fly species spanning two tribes and the distantly related Drosophila melanogaster at molecular, functional, and morphological levels. Olfaction in these flies is mediated by a set of olfactory receptors (ORs) expressed in different functional classes of neurons located in distinct antennal regions. A phylogenetic analysis was performed that revealed both family-specific OR genes and putative orthologous OR genes between tephritids and Drosophila. With respect to function, a current source density (CSD) analysis was used to map activity across antennae. Functional maps mirrored the intrinsic structure of antennae observed with scanning electron microscopy. Together, the results revealed partial conservation of the olfactory systems between tephritids and Drosophila. It was also demonstrated that the mapping of olfactory responses is necessary to decipher antennal sensory selectivity to olfactory compounds. CSD analysis can be easily applied to map antennae of other species and therefore enables the rapid deriving of olfactory maps and the reconstructing of the target organisms' history of evolution.
Gerhard, S., Andrade, I., Fetter, R. D., Cardona, A. and Schneider-Mizell, C. M. (2017). Conserved neural circuit structure across Drosophila larval development revealed by comparative connectomics. Elife 6. PubMed ID: 29058674
During postembryonic development, the nervous system must adapt to a growing body. How changes in neuronal structure and connectivity contribute to the maintenance of appropriate circuit function remains unclear. Previous work measured the cellular neuroanatomy underlying synaptic connectivity in Drosophila. This study examined how neuronal morphology and connectivity change between first instar and third instar larval stages using serial section electron microscopy. Nociceptive circuits were reconstructed in a larva of each stage, and consistent topographically arranged connectivity was found between identified neurons. Five-fold increases in each size, number of terminal dendritic branches, and total number of synaptic inputs were accompanied by cell type-specific connectivity changes that preserved the fraction of total synaptic input associated with each pre-synaptic partner. It is proposed that precise patterns of structural growth act to conserve the computational function of a circuit, for example determining the location of a dangerous stimulus.
Himmelreich, S., Masuho, I., Berry, J. A., MacMullen, C., Skamangas, N. K., Martemyanov, K. A. and Davis, R. L. (2017). Dopamine receptor DAMB signals via Gq to mediate forgetting in Drosophila. Cell Rep 21(8): 2074-2081. PubMed ID: 29166600
Prior studies have shown that aversive olfactory memory is acquired by dopamine acting on a specific receptor, dDA1, expressed by mushroom body neurons. Active forgetting is mediated by dopamine acting on another receptor, Damb, expressed by the same neurons. Surprisingly, prior studies have shown that both receptors stimulate cyclic AMP (cAMP) accumulation, presenting an enigma of how mushroom body neurons distinguish between acquisition and forgetting signals. This study surveyed the spectrum of G protein coupling of dDA1 and Damb, and it was confirmed that both receptors can couple to Gs to stimulate cAMP synthesis. However, the Damb receptor uniquely activates Gq to mobilize Ca(2+) signaling with greater efficiency and dopamine sensitivity. The knockdown of Galphaq with RNAi in the mushroom bodies inhibits forgetting but has no effect on acquisition. These findings identify a Damb/Gq-signaling pathway that stimulates forgetting and resolves the opposing effects of dopamine on acquisition and forgetting.
Hakeda-Suzuki, S., Takechi, H., Kawamura, H. and Suzuki, T. (2017). Two receptor tyrosine phosphatases dictate the depth of axonal stabilizing layer in the visual system. Elife 6. PubMed ID: 29116043
Formation of a functional neuronal network requires not only precise target recognition, but also stabilization of axonal contacts within their appropriate synaptic layers. Little is known about the molecular mechanisms underlying the stabilization of axonal connections after reaching their specifically targeted layers. This study shows that two receptor protein tyrosine phosphatases (RPTPs), LAR and Ptp69D, act redundantly in photoreceptor afferents to stabilize axonal connections to the specific layers of the Drosophila visual system. Surprisingly, by combining loss-of-function and genetic rescue experiments, this study found that the depth of the final layer of stable termination relied primarily on the cumulative amount of LAR and Ptp69D cytoplasmic activity, while specific features of their ectodomains contribute to the choice between two synaptic layers, M3 and M6, in the medulla. These data demonstrate how the combination of overlapping downstream but diversified upstream properties of two RPTPs can shape layer-specific wiring.
Guo, F., Chen, X. and Rosbash, M. (2017). Temporal calcium profiling of specific circadian neurons in freely moving flies. Proc Natl Acad Sci U S A 114(41): E8780-e8787. PubMed ID: 28973886
There are no general methods for reliably assessing the firing properties or even calcium profiles of specific neurons in freely moving flies. To this end, this study adapted a GFP-based calcium reporter to luciferase that was expressed in small subsets of circadian neurons. This Tric-LUC reporter allowed a direct comparison of luciferase activity with locomotor activity, which was assayed in the same flies with video recording. The LUC profile from activity-promoting E cells paralleled evening locomotor activity, and the LUC profile from sleep-promoting glutamatergic DN1s (gDN1s) paralleled daytime sleep. Similar profiles were generated by novel reporters recently identified based on transcription factor activation. As E cell and gDN1 activity is necessary and sufficient for normal evening locomotor activity and daytime sleep profiles, respectively, it is suggest that their luciferase profiles reflect their neuronal calcium and in some cases firing profiles in wake-behaving flies.

Monday, January 8th

Davis, T. L., Hoi, C. S. L. and Rebay, I. (2017). Mutations that impair Eyes absent tyrosine phosphatase activity in vitro reduce robustness of retinal determination gene network output in Drosophila. PLoS One 12(11): e0187546. PubMed ID: 29108015
A limited collection of signaling networks and transcriptional effectors directs the full spectrum of cellular behaviors that comprise development. One mechanism to diversify regulatory potential is to combine multiple biochemical activities into the same protein. Exemplifying this principle of modularity, Eyes absent (Eya), originally identified as a transcriptional co-activator within the retinal determination gene network (RDGN), also harbors tyrosine and threonine phosphatase activities. Although mounting evidence argues for the importance of Eya's phosphatase activities to mammalian biology, genetic rescue experiments in Drosophila have shown that the tyrosine phosphatase function is dispensable for normal development. This study repeats these rescue experiments in genetically sensitized backgrounds in which the dose of one or more RDGN factor was reduced. Heterozygosity for sine oculis or dachshund, both core RDGN members, compromised the ability of phosphatase-dead eya, but not of the control wild type eya transgene, to rescue the retinal defects and reduced viability associated with eya loss. It is speculated that Eya's tyrosine phosphatase activity, although non-essential, confers robustness to RDGN output.
Enya, S., et al. (2017). Dual roles of glutathione in ecdysone biosynthesis and antioxidant function during the larval development in Drosophila. Genetics [Epub ahead of print]. PubMed ID: 29021278
Ecdysteroid biosynthesis is achieved by a series of specialized enzymes encoded in the Halloween genes. Recently, noppera-bo (nobo), encoding a glutathione S-transferase (GST), has been identified. GSTs conjugate substrates with the reduced form of glutathione (GSH), a bioactive tripeptide composed of glutamate, cysteine, and glycine. It was hypothesized that GSH itself is required for ecdysteroid biosynthesis. This study reports phenotypic analysis of a mutant in the gamma-glutamylcysteine synthetase catalytic subunit (Gclc) gene in Drosophila. Gclc encodes the conserved catalytic component of the enzyme conjugating glutamate and cysteine in the GSH biosynthesis pathway. Loss of Gclc function leads to drastic GSH deficiency in the larval body fluid. Gclc-mutant animals show larval-arrest phenotype. Ecdysteroid titer in Gclc mutant larvae decreases, and the larval-arrest phenotype is rescued by oral administration of 20E or cholesterol. Moreover, Gclc mutant animals exhibit abnormal lipid deposition in the prothoracic gland, a steroidogenic organ during larval development. All of these phenotypes are reminiscent of nobo loss-of-function animals. On the other hand, Gclc mutant larvae also exhibit a significant reduction in antioxidant capacity. Ecdysteroid biosynthesis defect in Gclc mutant animals is not associated with loss of antioxidant function. It is hypothesized that a primary role of GSH in early D. melanogaster larval development is ecdysteroid biosynthesis, independent from antioxidant role of GSH.
Hsia, E. Y. C., Zhang, Y., Tran, H. S., Lim, A., Chou, Y. H., Lan, G., Beachy, P. A. and Zheng, X. (2017). Hedgehog mediated degradation of Ihog adhesion proteins modulates cell segregation in Drosophila wing imaginal discs. Nat Commun 8(1): 1275. PubMed ID: 29097673
The Drosophila Hedgehog receptor functions to regulate the essential downstream pathway component, Smoothened, and to limit the range of signaling by sequestering Hedgehog protein signal within imaginal disc epithelium. Hedgehog receptor function requires both Patched and Ihog activity, the latter interchangeably encoded by interference hedgehog (ihog) or brother of ihog (boi). This study shows that Patched and Ihog activity are mutually required for receptor endocytosis and degradation, triggered by Hedgehog protein binding, and causing reduced levels of Ihog/Boi proteins in a stripe of cells at the anterior/posterior compartment boundary of the wing imaginal disc. This Ihog spatial discontinuity may contribute to classically defined cell segregation and lineage restriction at the anterior/posterior wing disc compartment boundary, as suggested by observations that Ihog activity mediates aggregation of otherwise non-adherent cultured cells and that loss of Ihog activity disrupts wing disc cell segregation, even with downstream genetic rescue of Hedgehog signal response.
Chaturvedi, D., Reichert, H., Gunage, R. D. and VijayRaghavan, K. (2017). Identification and functional characterization of muscle satellite cells in Drosophila. Elife 6. PubMed ID: 29072161
Work on genetic model systems such as Drosophila and mouse has shown that the fundamental mechanisms of myogenesis are remarkably similar in vertebrates and invertebrates. Strikingly, however, satellite cells, the adult muscle stem cells that are essential for the regeneration of damaged muscles in vertebrates, have not been reported in invertebrates. This study shows that lineal descendants of muscle stem cells are present in adult muscle of Drosophila as small, unfused cells observed at the surface and in close proximity to the mature muscle fibers. Normally quiescent, following muscle fiber injury, these cells express Zfh1 and engage in Notch-Delta-dependent proliferative activity and generate lineal descendant populations, which fuse with the injured muscle fiber. In view of strikingly similar morphological and functional features, these novel cells are considered to be the Drosophila equivalent of vertebrate muscle satellite cells.
Dye, N. A., Popovic, M., Spannl, S., Etournay, R., Kainmuller, D., Ghosh, S., Myers, E. W., Julicher, F. and Eaton, S. (2017). Cell dynamics underlying oriented growth of the Drosophila wing imaginal disc. Development 144(23): 4406-4421. PubMed ID: 29038308
Quantitative analysis of the dynamic cellular mechanisms shaping the Drosophila wing during its larval growth phase has been limited, impeding the ability to understand how morphogen patterns regulate tissue shape. Such analysis requires explants to be imaged under conditions that maintain both growth and patterning, as well as methods to quantify how much cellular behaviors change tissue shape. This study demonstrates a key requirement for the steroid hormone 20-hydroxyecdysone (20E) in the maintenance of numerous patterning systems in vivo and in explant culture. Low concentrations of 20E support prolonged proliferation in explanted wing discs in the absence of insulin, incidentally providing novel insight into the hormonal regulation of imaginal growth. 20E-containing media was used to observe growth directly and to apply recently developed methods for quantitatively decomposing tissue shape changes into cellular contributions. Whereas cell divisions drive tissue expansion along one axis, their contribution to expansion along the orthogonal axis is cancelled by cell rearrangements and cell shape changes. This finding raises the possibility that anisotropic mechanical constraints contribute to growth orientation in the wing disc.
Gupta, R. P., Bajpai, A. and Sinha, P. (2017). Selector genes display tumor cooperation and inhibition in Drosophila epithelium in a developmental context-dependent manner. Biol Open 6(11): 1581-1591. PubMed ID: 29141951
During animal development, selector genes determine identities of body segments and those of individual organs. Selector genes are also misexpressed in cancers, although their contributions to tumor progression per se remain poorly understood. Using a model of cooperative tumorigenesis, this study shows that gain of selector genes results in tumor cooperation, but in only select developmental domains of the wing, haltere and eye-antennal imaginal discs of Drosophila larva. Thus, the field selector, Eyeless (Ey), and the segment selector, Ultrabithorax (Ubx), readily cooperate to bring about neoplastic transformation of cells displaying somatic loss of the tumor suppressor, Lgl, but in only those developmental domains that express the homeo-box protein, Homothorax (Hth), and/or the Zinc-finger protein, Teashirt (Tsh). In non-Hth/Tsh-expressing domains of these imaginal discs, however, gain of Ey in lgl- somatic clones induces neoplastic transformation in the distal wing disc and haltere, but not in the eye imaginal disc. Likewise, gain of Ubx in lgl- somatic clones induces transformation in the eye imaginal disc but not in its endogenous domain, namely, the haltere imaginal disc. These results reveal that selector genes could behave as tumor drivers or inhibitors depending on the tissue contexts of their gains.

Friday, January 5th

Moghaddam, M. B., Gross, T., Becker, A., Vilcinskas, A. and Rahnamaeian, M. (2017). The selective antifungal activity of Drosophila melanogaster metchnikowin reflects the species-dependent inhibition of succinate-coenzyme Q reductase. Sci Rep 7(1): 8192. PubMed ID: 28811531
Insect-derived antifungal peptides have a significant economic potential, particularly for the engineering of pathogen-resistant crops. However, the nonspecific antifungal activity of such peptides could result in detrimental effects against beneficial fungi, whose interactions with plants promote growth or increase resistance against biotic and abiotic stress. The antifungal peptide metchnikowin (Mtk) from Drosophila melanogaster acts selectively against pathogenic Ascomycota, including Fusarium graminearum, without affecting Basidiomycota such as the beneficial symbiont Piriformospora indica. This study investigated the mechanism responsible for the selective antifungal activity of Mtk by using the peptide to probe a yeast two-hybrid library of F. graminearum cDNAs. Mtk was found to specifically target the iron-sulfur subunit (SdhB) of succinate-coenzyme Q reductase (SQR). A functional assay based on the succinate dehydrogenase (SDH) activity of mitochondrial complex II clearly demonstrated that Mtk inhibited the SDH activity of F. graminearum mitochondrial SQR by up to 52%, but that the equivalent enzyme in P. indica was unaffected. A phylogenetic analysis of the SdhB family revealed a significant divergence between the Ascomycota and Basidiomycota. SQR is one of the key targets of antifungal agents and it is therefore proposed that Mtk as an environmentally sustainable and more selective alternative to chemical fungicides.
Dostalova, A., Rommelaere, S., Poidevin, M. and Lemaitre, B. (2017). Thioester-containing proteins regulate the Toll pathway and play a role in Drosophila defence against microbial pathogens and parasitoid wasps. BMC Biol 15(1): 79. PubMed ID: 28874153
Members of the thioester-containing protein (TEP) family contribute to host defence in both insects and mammals. However, their role in the immune response of Drosophila is elusive. This study addresses the role of TEPs in Drosophila immunity by generating a mutant fly line, referred to as TEPq Delta , lacking the four immune-inducible TEPs, TEP1, 2, 3 and 4. Survival analyses with TEPq Delta flies reveal the importance of these proteins in defence against entomopathogenic fungi, Gram-positive bacteria and parasitoid wasps. These results confirm that TEPs are required for efficient phagocytosis of bacteria, notably for the two Gram-positive species tested, Staphylococcus aureus and Enterococcus faecalis. Furthermore, TEPq Delta flies have reduced Toll pathway activation upon microbial infection, resulting in lower expression of antimicrobial peptide genes. Epistatic analyses suggest that TEPs function upstream or independently of the serine protease ModSP at an initial stage of Toll pathway activation. Collectively, this study brings new insights into the role of TEPs in insect immunity. It reveals that TEPs participate in both humoral and cellular arms of immune response in Drosophila. In particular, it shows the importance of TEPs in defence against Gram-positive bacteria and entomopathogenic fungi, notably by promoting Toll pathway activation.
Kubiak, M. and Tinsley, M. C. (2017). Sex-specific routes to immune senescence in Drosophila melanogaster. Sci Rep 7(1): 10417. PubMed ID: 28874758
Animal immune systems change dramatically during the ageing process, often accompanied by major increases in pathogen susceptibility. However, the extent to which senescent elevations in infection mortality are causally driven by deteriorations in canonical systemic immune processes is unclear. This study examined Drosophila melanogaster and compared the relative contributions of impaired systemic immune defences and deteriorating barrier defences to increased pathogen susceptibility in aged flies. To assess senescent changes in systemic immune response efficacy, one and four-week old flies with the entomopathogenic fungus Beauveria bassiana and subsequent mortality was studied; whereas to include the role of barrier defences flies were injected by dusting the cuticle with fungal spores. The processes underlying pathogen defence senescence differ between males and females. Both sexes became more susceptible to infection as they aged. However, it is concluded that for males, this was principally due to deterioration in barrier defences, whereas for females systemic immune defence senescence was mainly responsible. The potential roles of sex-specific selection on the immune system and behavioural variation between males and females in driving these different senescent trends is discussed.
Houtz, P., Bonfini, A., Liu, X., Revah, J., Guillou, A., Poidevin, M., Hens, K., Huang, H. Y., Deplancke, B., Tsai, Y. C. and Buchon, N. (2017). Hippo, TGF-beta, and Src-MAPK pathways regulate transcription of the upd3 cytokine in Drosophila enterocytes upon bacterial infection. PLoS Genet 13(11): e1007091. PubMed ID: 29108021
Cytokine signaling is responsible for coordinating conserved epithelial regeneration and immune responses in the digestive tract. In the Drosophila midgut, Upd3 is a major cytokine, which is induced in enterocytes (EC) and enteroblasts (EB) upon oral infection, and initiates intestinal stem cell (ISC) dependent tissue repair. To date, the genetic network directing upd3 transcription remains largely uncharacterized. This study has identified the key infection-responsive enhancers of the upd3 gene and shows that distinct enhancers respond to various stresses. Furthermore, through functional genetic screening, bioinformatic analyses and yeast one-hybrid screening, it was determined that the transcription factors Scalloped (Sd), Mothers against dpp (Mad), and D-Fos are principal regulators of upd3 expression. This study demonstrates that upd3 transcription in the gut is regulated by the activation of multiple pathways, including the Hippo, TGF-beta/Dpp, and Src, as well as p38-dependent MAPK pathways. Thus, these essential pathways, which are known to control ISC proliferation cell-autonomously, are also activated in ECs to promote tissue turnover the regulation of upd3 transcription.
Duneau, D., Ferdy, J. B., Revah, J., Kondolf, H., Ortiz, G. A., Lazzaro, B. P. and Buchon, N. (2017). Stochastic variation in the initial phase of bacterial infection predicts the probability of survival in D. melanogaster. Elife 6. PubMed ID: 29022878
A central problem in infection biology is understanding why two individuals exposed to identical infections have different outcomes. This study has developed an experimental model where genetically identical, co-housed Drosophila given identical systemic infections experience different outcomes, with some individuals succumbing to acute infection while others control the pathogen as an asymptomatic persistent infection. Differences in bacterial burden at the time of death did not explain the two outcomes of infection. Inter-individual variation in survival stems from variation in within-host bacterial growth, which is determined by the immune response. A model was developed that captures bacterial growth dynamics and identifies key factors that predict the infection outcome: the rate of bacterial proliferation and the time required for the host to establish an effective immunological control. The results provide a framework for studying the individual host-pathogen parameters governing the progression of infection and lead ultimately to life or death.
Hori, A., Kurata, S. and Kuraishi, T. (2017). Unexpected role of the IMD pathway in Drosophila gut defense against Staphylococcus aureus. Biochem Biophys Res Commun. PubMed ID: 29108998
This study used Drosophila as a model animal to investigate the molecular mechanisms of innate immunity. To combat orally transmitted pathogenic Gram-negative bacteria, the Drosophila gut is armed with the peritrophic matrix, which is a physical barrier composed of chitin and glycoproteins: the Duox system that produces reactive oxygen species (ROS), which in turn sterilize infected microbes, and the IMD pathway that regulates the expression of antimicrobial peptides (AMPs), which in turn control ROS-resistant pathogens. However, little is known about the defense mechanisms against Gram-positive bacteria in the fly gut. This study shows that the peritrophic matrix protects Drosophila against Gram-positive bacteria S. aureus. The few roles are described of ROS in response to the infection, and the IMD pathway was shown to be required for the clearance of ingested microbes, possibly independently from AMP expression. These findings provide a new aspect of the gut defense system of Drosophila, and helps to elucidate the processes of gut-microbe symbiosis and pathogenesis.

Thursday, January 4th

Heyam, A., Coupland, C. E., Degut, C., Haley, R. A., Baxter, N. J., Jakob, L., Aguiar, P. M., Meister, G., Williamson, M. P., Lagos, D. and Plevin, M. J. (2017). Conserved asymmetry underpins homodimerization of Dicer-associated double-stranded RNA-binding proteins. Nucleic Acids Res. PubMed ID: 29045748
Double-stranded RNA-binding domains (dsRBDs) are commonly found in modular proteins that interact with RNA. Two varieties of dsRBD exist: canonical Type A dsRBDs interact with dsRNA, while non-canonical Type B dsRBDs lack RNA-binding residues and instead interact with other proteins. In higher eukaryotes, the microRNA biogenesis enzyme Dicer forms a 1:1 association with a dsRNA-binding protein (dsRBP). Human Dicer associates with HIV TAR RNA-binding protein (TRBP) or protein activator of PKR (PACT), while Drosophila Dicer-1 associates with Loquacious (Loqs). In each case, the interaction involves a region of the protein that contains a Type B dsRBD. All three dsRBPs are reported to homodimerize, with the Dicer-binding region implicated in self-association. This study reports that these dsRBD homodimers display structural asymmetry and that this unusual self-association mechanism is conserved from flies to humans. The core dsRBD is sufficient for homodimerization, and mutation of a conserved leucine residue abolishes self-association. Differences in the self-association properties of Loqs, TRBP and PACT are attributed to divergence of the composition of the homodimerization interface. Modifications that make TRBP more like PACT enhance self-association. These data are examined in the context of miRNA biogenesis and the protein/protein interaction properties of Type B dsRBDs.
Castillejo-Lopez, C., Cai, X., Fahmy, K. and Baumgartner, S. (2018). Drosophila exoribonuclease nibbler is a tumor suppressor, acts within the RNA(i) machinery and is not enriched in the nuage during early oogenesis. Hereditas 155: 12. PubMed ID: 28974923
micro RNAs (miRNAs) are important regulators of many biological pathways. A plethora of steps are required to form, from a precursor, the mature miRNA that eventually acts on its target RNA to repress its expression or to inhibit translation. Recently, Drosophila nibbler (nbr) has been shown to be an important player in the maturation process of miRNA and piRNA. Nbr is an exoribonuclease which helps to shape the 3' end of miRNAs by trimming the 3' overhang to a final length. In contrast to previous reports on the localization of Nbr, this study reports that 1) Nbr is expressed only during a short time of oogenesis and appears ubiquitously localized within oocytes, and that 2) Nbr was is not enriched in the nuage where it was shown to be involved in piwi-mediated mechanisms. To date, there is little information available on the function of nbr for cellular and developmental processes. Due to the fact that nbr mutants are viable with minor deleterious effects, this study used the GAL4/UAS over-expression system to define novel functions of nbr. Hitherto unknown functions of nbr are revealed; 1) as a tumor suppressor and 2) as a suppressor of RNA(i). Finally, it was confirmed that nbr is a suppressor of transposon activity. These data suggest that nbr exerts much more widespread functions than previously reported from trimming 3' ends of miRNAs only.
Bienkowski, R. S., Banerjee, A., Rounds, J. C., Rha, J., Omotade, O. F., Gross, C., Morris, K. J., Leung, S. W., Pak, C., Jones, S. K., Santoro, M. R., Warren, S. T., Zheng, J. Q., Bassell, G. J., Corbett, A. H. and Moberg, K. H. (2017). The conserved, disease-associated RNA binding protein dNab2 interacts with the Fragile X Protein ortholog in Drosophila neurons. Cell Rep 20(6): 1372-1384. PubMed ID: 28793261
The Drosophila dNab2 protein is an ortholog of human ZC3H14, a poly(A) RNA binding protein required for intellectual function. dNab2 supports memory and axon projection, but its molecular role in neurons is undefined. This study presents a network of interactions that links dNab2 to cytoplasmic control of neuronal mRNAs in conjunction with the fragile X protein ortholog dFMRP. dNab2 and dfmr1 interact genetically in control of neurodevelopment and olfactory memory, and their encoded proteins co-localize in puncta within neuronal processes. dNab2 regulates CaMKII, but not futsch, implying a selective role in control of dFMRP-bound transcripts. Reciprocally, dFMRP and vertebrate FMRP restrict mRNA poly(A) tail length, similar to dNab2/ZC3H14. Parallel studies of murine hippocampal neurons indicate that ZC3H14 is also a cytoplasmic regulator of neuronal mRNAs. Altogether, these findings suggest that dNab2 represses expression of a subset of dFMRP-target mRNAs, which could underlie brain-specific defects in patients lacking ZC3H14.
Dufourt, J., Bontonou, G., Chartier, A., Jahan, C., Meunier, A. C., Pierson, S., Harrison, P. F., Papin, C., Beilharz, T. H. and Simonelig, M. (2017). piRNAs and Aubergine cooperate with Wispy poly(A) polymerase to stabilize mRNAs in the germ plasm. Nat Commun 8(1): 1305. PubMed ID: 29101389
Piwi-interacting RNAs (piRNAs) and PIWI proteins play a crucial role in germ cells by repressing transposable elements and regulating gene expression. In Drosophila, maternal piRNAs are loaded into the embryo mostly bound to the PIWI protein Aubergine (Aub). Aub targets maternal mRNAs through incomplete base-pairing with piRNAs and can induce their destabilization in the somatic part of the embryo. Paradoxically, these Aub-dependent unstable mRNAs encode germ cell determinants that are selectively stabilized in the germ plasm. This study shows that piRNAs and Aub actively protect germ cell mRNAs in the germ plasm. Aub directly interacts with the germline-specific poly(A) polymerase Wispy, thus leading to mRNA polyadenylation and stabilization in the germ plasm. These results reveal a role for piRNAs in mRNA stabilization and identify Aub as an interactor of Wispy for mRNA polyadenylation. They further highlight the role of Aub and piRNAs in embryonic patterning through two opposite functions.
Gerstberger, S., Meyer, C., Benjamin-Hong, S., Rodriguez, J., Briskin, D., Bognanni, C., Bogardus, K., Steller, H. and Tuschl, T. (2017). The conserved RNA exonuclease Rexo5 is required for 3' end maturation of 28S rRNA, 5S rRNA, and snoRNAs. Cell Rep 21(3): 758-772. PubMed ID: 29045842
Non-coding RNA biogenesis in higher eukaryotes has not been fully characterized. Drosophila melanogaster Rexo5 (CG8368) protein is a metazoan-specific member of the DEDDh 3'-5' single-stranded RNA exonucleases on the basis genetic, biochemical, and RNA-sequencing tests. Rexo5 is required for small nucleolar RNA (snoRNA) and rRNA biogenesis and is essential in D. melanogaster. Loss-of-function mutants accumulate improperly 3' end-trimmed 28S rRNA, 5S rRNA, and snoRNA precursors in vivo. Rexo5 is ubiquitously expressed at low levels in somatic metazoan cells but extremely elevated in male and female germ cells. Loss of Rexo5 leads to increased nucleolar size, genomic instability, defective ribosome subunit export, and larval death. Loss of germline expression compromises gonadal growth and meiotic entry during germline development.
Gendron, C. M. and Pletcher, S. D. (2017). MicroRNAs mir-184 and let-7 alter Drosophila metabolism and longevity. Aging Cell 16(6): 1434-1438. PubMed ID: 28963741
MicroRNAs (miRNAs) are small RNA molecules that regulate gene expression associated with many complex biological processes. By comparing miRNA expression between long-lived cohorts of Drosophila melanogaster that were fed a low-nutrient diet with normal-lived control animals fed a high-nutrient diet, this study identified miR-184, let-7, miR-125, and miR-100 as candidate miRNAs involved in modulating aging. Ubiquitous, adult-specific overexpression of these individual miRNAs led to significant changes in fat metabolism and/or lifespan. Most impressively, adult-specific overexpression of let-7 in female nervous tissue increased median fly lifespan by ~22%. Evidence is provided that this lifespan extension is not due to alterations in nutrient intake or to decreased insulin signaling.

Wednesday, January 3rd

Hebbar, S., Khandelwal, A., R, J., Hindle, S. J., Chiang, Y. N., Yew, J. Y., Sweeney, S. T. and Schwudke, D. (2017). Lipid metabolic perturbation is an early-onset phenotype in adult spin mutants: a Drosophila model for lysosomal storage disorders. Mol Biol Cell. PubMed ID: 29046397
Intracellular accumulation of lipids and swollen dysfunctional lysosomes are linked to several neurodegenerative diseases including lysosomal storage disorders (LSD). Detailed characterization of lipid metabolic changes in relation to the onset and progression of neurodegeneration is currently missing. This study systematically analyzed lipid perturbations in spinster (spin) mutants, a Drosophila model of LSD-like neurodegeneration. The results highlight an imbalance in brain ceramide and sphingosine in the early stages of neurodegeneration, preceding the accumulation of endomembranous structures, manifestation of altered behaviour, and buildup of lipofuscin. Manipulating levels of ceramidase, altering these lipids in spin mutants led to the conclusion that ceramide homeostasis is the driving force in disease progression and is integral to spin function in the adult nervous system. Twenty-nine novel physical interaction partners of Spin were identified, and focus was placed on the lipid carrier protein, Lipophorin (Lpp). A subset of Lpp and Spin co-localize in the brain and within organs specialized for lipid metabolism (fat bodies and oenocytes). Reduced Lpp protein was observed in spin mutant tissues. Finally, increased levels of lipid metabolites produced by oenocytes in spin mutants allude to a functional interaction between Spin and Lpp, underscoring the systemic nature of lipid perturbation in LSD.
Dean, D., Weinstein, H., Amin, S., Karno, B., McAvoy, E., Hoy, R., Recknagel, A., Jarvis, C. and Deitcher, D. (2017). Extending julius seizure, a bang-sensitive gene, as a model for studying epileptogenesis: Cold shock, and a new insertional mutation. Fly (Austin): [Epub ahead of print] PubMed ID: 29125376
The bang-sensitive (BS) mutants of Drosophila are an important model for studying epilepsy. A novel BS locus, julius seizure (jus), encoding a protein containing two transmembrane domains and an extracellular cysteine-rich loop has recently been identified. It was also determined that jussda iso7.8, a previously identified BS mutation, is an allele of jus by recombination, deficiency mapping, complementation testing, and genetic rescue. RNAi knockdown revealed that jus expression is important in cholinergic neurons and that the critical stage of jus expression is the mid-pupa. Finally, a functional, GFP-tagged genomic construct of jus was found to be expressed mostly in axons of the neck connectives and of the thoracic abdominal ganglia. This study shows that a MiMiC GFP-tagged Jus is localized to the same brain regions as the GFP-tagged genomic construct, but its expression is mostly confined to cell bodies and it causes bang-sensitivity. The MiMiC GFP-tag lies in the extracellular loop while the genomic construct is tagged at the C-terminus. This suggests that the alternate position of the GFP tag may disrupt Jus protein function by altering its subcellular localization and/or stability. A small subset of jus-expressing neurons are responsible for the BS phenotype. Extending the utility of the BS seizure model, it was shown that jus mutants exhibit cold-sensitive paralysis and are partially sensitive to strobe-induced seizures.
Del Signore, S. J., Biber, S. A., Lehmann, K. S., Heimler, S. R., Rosenfeld, B. H., Eskin, T. L., Sweeney, S. T. and Rodal, A. A. (2017). dOCRL maintains immune cell quiescence by regulating endosomal traffic. PLoS Genet 13(10): e1007052. PubMed ID: 29028801
Lowe Syndrome is a developmental disorder characterized by eye, kidney, and neurological pathologies, and is caused by mutations in the phosphatidylinositol-5-phosphatase OCRL. OCRL plays diverse roles in endocytic and endolysosomal trafficking, cytokinesis, and ciliogenesis, but it is unclear which of these cellular functions underlie specific patient symptoms. This study shows that mutation of Drosophila OCRL causes cell-autonomous activation of hemocytes, which are macrophage-like cells of the innate immune system. Among many cell biological defects that identified in docrl mutant hemocytes, the cause of innate immune cell activation was pinpointed to reduced Rab11-dependent recycling traffic and concomitantly increased Rab7-dependent late endosome traffic. Loss of docrl amplifies multiple immune-relevant signals, including Toll, Jun kinase, and STAT, and leads to Rab11-sensitive mis-sorting and excessive secretion of the Toll ligand Spatzle. Thus, docrl regulation of endosomal traffic maintains hemocytes in a poised, but quiescent state, suggesting mechanisms by which endosomal misregulation of signaling may contribute to symptoms of Lowe syndrome.
Hall, H., Medina, P., Cooper, D. A., Escobedo, S. E., Rounds, J., Brennan, K. J., Vincent, C., Miura, P., Doerge, R. and Weake, V. M. (2017). Transcriptome profiling of aging Drosophila photoreceptors reveals gene expression trends that correlate with visual senescence. BMC Genomics 18(1): 894. PubMed ID: 29162050
Aging is associated with functional decline of neurons and increased incidence of both neurodegenerative and ocular disease. This study sought to identify changes in differentially regulated genes in photoreceptors that contribute to visual senescence. To identify gene expression changes that could lead to visual senescence, the aging transcriptome was characterized of Drosophila sensory neurons highly enriched for photoreceptors. The nuclear transcriptome of genetically-labeled photoreceptors over a 40 day time course was characterized, and increased expression of genes involved in stress and DNA damage response and decreased expression of genes required for neuronal function were found. Long, highly expressed and heavily spliced genes are more likely to be downregulated with age, indicating that other mechanisms could contribute to expression changes at these genes. Lastly, circular RNAs (circRNAs) were found to strongly increase during aging in photoreceptors. Overall, this study identified changes in gene expression in aging Drosophila photoreceptors that could account for visual senescence. Further, it was shown that genomic features predict these age-related changes, suggesting potential mechanisms that could be targeted to slow the rate of age-associated visual decline.
Dissel, S., Klose, M., Donlea, J., Cao, L., English, D., Winsky-Sommerer, R., van Swinderen, B. and Shaw, P. J. (2017). Enhanced sleep reverses memory deficits and underlying pathology in Drosophila models of Alzheimer's disease. Neurobiol Sleep Circadian Rhythms 2: 15-26. PubMed ID: 29094110
To test the hypothesis that sleep can reverse cognitive impairment during Alzheimer's disease, sleep was enhanced in flies either co-expressing human amyloid precursor protein and Beta-secretase (APP:BACE), or in flies expressing human tau. The ubiquitous expression of APP:BACE or human tau disrupted sleep. The sleep deficits could be reversed and sleep could be enhanced when flies were administered the GABA-A agonist 4,5,6,7-tetrahydroisoxazolo-[5,4-c]pyridine-3-ol (THIP). Expressing APP:BACE disrupted both Short-term memory (STM) and Long-term memory (LTM) as assessed using Aversive Phototaxic Suppression (APS) and courtship conditioning. Flies expressing APP:BACE also showed reduced levels of the synaptic protein Discs large (DLG). Enhancing sleep in memory-impaired APP:BACE flies fully restored both STM and LTM and restored DLG levels. Sleep also restored STM to flies expressing human tau. Using live-brain imaging of individual clock neurons expressing both tau and the cAMP sensor Epac1-camps, tau was found to disrupt cAMP signaling. Importantly, enhancing sleep in flies expressing human tau restored proper cAMP signaling. Thus, this study demonstrated that sleep can be used as a therapeutic to reverse deficits that accrue during the expression of toxic peptides associated with Alzheimer's disease.
Gray, K. M., Kaifer, K. A., Baillat, D., Wen, Y., Bonacci, T. R., Ebert, A. D., Raimer, A. C., Spring, A. M., Have, S. T., Glascock, J. J., Gupta, K., Van Duyne, G. D., Emanuele, M. J., Lamond, A. I., Wagner, E. J., Lorson, C. L. and Matera, A. G. (2017). Self-oligomerization regulates stability of Survival Motor Neuron (SMN) protein isoforms by sequestering an SCF(Slmb) degron. Mol Biol Cell. PubMed ID: 29167380
Spinal muscular atrophy (SMA) is caused by homozygous mutations in human SMN1. Expression of a duplicate gene (SMN2) primarily results in skipping of exon 7 and production of an unstable protein isoform, SMNDelta7. Although SMN2 exon skipping is the principal contributor to SMA severity, mechanisms governing stability of SMN isoforms are poorly understood. This study used a Drosophila model system and label-free proteomics to identify the SCF(Slmb) ubiquitin E3 ligase complex as a novel SMN binding partner. SCF(Slmb) interacts with a phospho-degron embedded within the human and fruitfly SMN YG-box oligomerization domains. Substitution of a conserved serine (S270A) interferes with SCF(Slmb) binding and stabilizes SMNDelta7. SMA-causing missense mutations that block multimerization of full-length SMN are also stabilized in the degron mutant background. Overexpression of SMNDelta7(S270A), but not wild-type SMNDelta7, provides a protective effect in SMA model mice and human motor neuron cell culture systems. These findings support a model wherein the degron is exposed when SMN is monomeric, and sequestered when SMN forms higher-order multimers.

Tuesday, January 2nd

Chouhan, N. S., Mohan, K. and Ghose, A. (2017). cAMP signaling mediates behavioral flexibility and consolidation of social status in Drosophila aggression. J Exp Biol 220(Pt 23): 4502-4514. PubMed ID: 28993465
Social rituals, such as male-male aggression in Drosophila, are often stereotyped and the component behavioral patterns modular. The likelihood of transition from one behavioral pattern to another is malleable by experience and confers flexibility to the behavioral repertoire. Experience-dependent modification of innate aggressive behavior in flies alters fighting strategies during fights and establishes dominant-subordinate relationships. Dominance hierarchies resulting from agonistic encounters are consolidated to longer-lasting, social-status-dependent behavioral modifications, resulting in a robust loser effect. This study shows that cAMP dynamics regulated by the calcium-calmodulin-dependent adenylyl cyclase, Rut, and the cAMP phosphodiesterase, Dnc, but not the Amn gene product, in specific neuronal groups of the mushroom body and central complex, mediate behavioral plasticity necessary to establish dominant-subordinate relationships. rut and dnc mutant flies were unable to alter fighting strategies and establish dominance relationships during agonistic interactions. This real-time flexibility during a fight was independent of changes in aggression levels. Longer-term consolidation of social status in the form of a loser effect, however, required additional Amn-dependent inputs to cAMP signaling and involved a circuit-level association between the alpha/beta and gamma neurons of the mushroom body. These findings implicate cAMP signaling in mediating the plasticity of behavioral patterns in aggressive behavior and in the generation of a temporally stable memory trace that manifests as a loser effect.
Christ, P., Hill, S. R., Schachtner, J., Hauser, F. and Ignell, R. (2017). Functional characterization of the dual allatostatin-A receptors in mosquitoes. Peptides 99: 44-55. PubMed ID: 29103918
The neuropeptide allatostatin-A (AstA) and its cognate receptors (AstARs) are involved in the modulation of feeding behavior, which in hematophagous insects includes the regulation of the disease vector-related behaviors, host seeking and blood feeding. In mosquitoes and other dipterans, there are two copies of AstAR, contrasting with the single copy found in other insects. This study identified and cloned the dual AstAR system of two important disease vectors Aedes aegypti and Culex quinquefasciatus, and compared them with those previously described, including those in Anopheles coluzzii and Drosophila melanogaster. Phylogenetic analysis of the AstARs revealed that the mosquito AstAR1s has retained a similar amino acid sequence as the AstARs from non-dipteran insect species. Intron analysis revealed that the number of introns accumulated in the AstAR2s is similar to that in other insects, and that introns are conserved within the receptor types, but that only the final two introns are conserved across AstAR1s and 2s. The dual AstARs were characterized in An. coluzzii, Ae. aegypti and Cx. quinquefasciatus by stably expressing the receptors in a Chinese hamster oocyte cell line (CHO) also stably expressing a promiscuous G-protein (G16), and challenged them with the endogenous isoforms of AstA from the three mosquito species. In the culicine mosquitoes, Ae. aegypti and Cx. quinquefasciatus, the AstARs demonstrated differential sensitivity to AstA, with the AstAR2s displaying a higher sensitivity than the AstAR1s, suggesting a divergence of functional roles for these AstARs. In contrast, both An. coluzzii AstARs demonstrated a similar sensitivity to the AstA ligands. These findings are discussed in the light of AstA acting as a regulator of blood feeding in mosquitoes. A better understanding of the regulation of host seeking and blood feeding in vector mosquitoes will lead to the rational development of novel approaches for vector control.
Depetris-Chauvin, A., Galagovsky, D., Chevalier, C., Maniere, G. and Grosjean, Y. (2017). Olfactory detection of a bacterial short-chain fatty acid acts as an orexigenic signal in Drosophila melanogaster larvae. Sci Rep 7(1): 14230. PubMed ID: 29079812
Microorganisms inhabiting fermenting fruit produce chemicals that elicit strong behavioral responses in flies. Depending on their ecological niche, individuals confer a positive or a negative valence to a chemical and, accordingly, they trigger either attractive or repulsive behaviors. This study examined the case of bacterial short-chain fatty acids (SCFA) that trigger opposite behaviors in adult and larvae of Drosophila melanogaster. SCFA-attractive responses depend on two larval exclusive chemoreceptors, Or30a and Or94b. Of those SCFA, propionic acid improves larval survival in suboptimal rearing conditions and supports growth. Olfactory detection of propionic acid specifically is sufficient to trigger feeding behaviors, and this effect requires the correct activity of Or30a(+) and Or94b(+) olfactory sensory neurons. Additionally, the case was studied of the invasive pest Drosophila suzukii that lives on undamaged ripe fruit with less SCFA production. Contrary to D. melanogaster, D. suzukii larvae show reduced attraction towards propionic acid, which does not trigger feeding behavior in this invasive species. These results demonstrate the relevance of propionic acid as an orexigenic signal in D. melanogaster larvae. Moreover, this study underlines that the changes on ecological niche are accompanied with alterations of olfactory preferences and vital olfactory driven behaviors.
Gunawardhana, K. L. and Hardin, P. E. (2017). VRILLE controls PDF neuropeptide accumulation and arborization rhythms in small ventrolateral neurons to drive rhythmic behavior in Drosophila. Curr Biol 27(22): 3442-3453.e3444. PubMed ID: 29103936
In Drosophila, the circadian clock is comprised of transcriptional feedback loops that control rhythmic gene expression responsible for daily rhythms in physiology, metabolism, and behavior. The core feedback loop, which employs CLOCK-CYCLE (CLK-CYC) activators and PERIOD-TIMELESS (PER-TIM) repressors to drive rhythmic transcription peaking at dusk, is required for circadian timekeeping and overt behavioral rhythms. CLK-CYC also activates an interlocked feedback loop, which uses the PAR DOMAIN PROTEIN 1epsilon (PDP1epsilon) activator and the VRILLE (VRI) repressor to drive rhythmic transcription peaking at dawn. Although Pdp1epsilon mutants disrupt activity rhythms without eliminating clock function, whether vri is required for clock function and/or output is not known. Using a conditionally inactivatable transgene to rescue vri developmental lethality, this study shows that clock function persists after vri inactivation but that activity rhythms are abolished. The inactivation of vri disrupts multiple output pathways thought to be important for activity rhythms, including PDF accumulation and arborization rhythms in the small ventrolateral neuron (sLNv) dorsal projection. These results demonstrate that vri acts as a key regulator of clock output and suggest that the primary function of the interlocked feedback loop in Drosophila is to drive rhythmic transcription required for overt rhythms.
Davis, S. M., Thomas, A. L., Liu, L., Campbell, I. M. and Dierick, H. A. (2017). Isolation of aggressive behavior mutants in Drosophila using a screen for wing damage. Genetics [Epub ahead of print]. PubMed ID: 29109180
Aggression is a complex social behavior that is widespread in nature. To date only a limited number of genes that affect aggression have been identified, in large part because the complexity of the phenotype makes screening difficult and time consuming regardless of the species that is studied. Aggressive group-housed Drosophila melanogaster males inflict damage on each other's wings; wing damage negatively affects their ability to fly and mate. Using this wing-damage phenotype, males from ~1,400 chemically mutagenized strains were screened and ~40 mutant strains were found with substantial wing damage. Five of these mutants also had increased aggressive behavior. To identify the causal mutation in one of the top aggressive strains, whole genome sequencing and genomic duplications rescue strategies were used. A novel mutation was identified in the voltage-gated potassium channel Shaker (Sh) and a nearby previously identified Sh mutation was also shown to exhibit increased aggression. This simple screen can be used to dissect the molecular mechanisms underlying aggression.
Herrero, A., Duhart, J. M. and Ceriani, M. F. (2017). Neuronal and glial clocks underlying structural remodeling of pacemaker neurons in Drosophila. Front Physiol 8: 918. PubMed ID: 29184510
Ventral Lateral Neurons (LNvs), which are essential in the control of rest-activity cycles in Drosophila, undergo circadian remodeling of their axonal projections. This structural plasticity gives rise to changes in the degree of connectivity, which could provide a means of transmitting time of day information. Several neuronal types undergoing circadian remodeling hint to a differential effect of clock genes; while per mutants exhibited poorly developed axonal terminals giving rise to low complexity arbors, tim mutants displayed a characteristic hyper branching phenotype, suggesting these genes could be playing additional roles to those ascribed to core clock function. To shed light onto this possibility clock gene levels were altered through RNAi- mediated downregulation and expression of a dominant negative form exclusively in the adult LNvs. These experiments confirmed that the LNv clock is necessary to drive the remodeling process. The contribution of glia to the structural plasticity of the small LNvs was explored through acute disruption of their internal clock. Interestingly, impaired glial clocks also abolished circadian structural remodeling, without affecting other clock-controlled outputs. Taken together these data shows that both neuronal and glial clocks are recruited to define the architecture of the LNv projections along the day, thus enabling a precise reconfiguration of the circadian network.
Fedotov, S. A., Bragina, J. V., Besedina, N. G., Danilenkova, L. V., Kamysheva, E. A. and Kamyshev, N. G. (2017). Gene CG15630 (fipi) is involved in regulation of the interpulse interval in Drosophila courtship song. J Neurogenet: 1-12. PubMed ID: 29191114
To study the central pattern generators functioning, genes have been identified whose neurospecific knockdowns led to deviations in the courtship song of Drosophila melanogaster males. Reduced expression of the gene CG15630 caused a decrease in the interpulse interval. To investigate the role of CG15630, which is called fipi (factor of interpulse interval), in the courtship song production, at first, this study has characterized fipi transcripts and protein (FIPI) in the mutant flies carrying P insertion and deletions in this gene and in flies with its RNAi knockdown. FIPI is homologous to the mammalian NCAM2 protein, an important factor of neuronal development in the olfactory system. This study revealed that local fipi knockdown in the antennal olfactory sensory neurons (OR67d and IR84a), which are responsible for reception of chemosignals modulating courtship behavior, alters the interpulse interval in the opposite directions. Thus, a proper fipi expression seems to be necessary for perception of sexual chemosignals, and the effect of fipi knockdown on IPI value depends on the type of chemoreceptor neurons affected.
Hasegawa, T., Tomita, J., Hashimoto, R., Ueno, T., Kume, S. and Kume, K. (2017). Sweetness induces sleep through gustatory signalling independent of nutritional value in a starved fruit fly. Sci Rep 7(1): 14355. PubMed ID: 29084998
Starvation reduces sleep in various animal species including humans and fruit flies. Immediate hunger and the following insufficient nutritional status resulting from starvation may affect sleep and arousal differently. In order to clarify the mechanism underlying the relationship between diet and sleep, the sleep behaviour of Drosophila melanogaster was analyzed that were either starved or fed with different types of sugars. Starved flies showed longer activity bouts, short sleep bouts and a decreased arousal threshold. Non-nutritive sweeteners such as sucralose and arabinose, which are sweet but not nutritive, induced sleep in starved flies, but sleep bout length and the arousal threshold was short and decreased, respectively. On the other hand, sorbitol, which is not sweet but nutritive, did not induce sleep, but slightly increased the lowered arousal threshold. Activation of sweetness receptor expressing neurons induced sleep in starved flies. These results suggest that sweetness alone is sufficient to induce sleep in starved flies and that the nutritional status affects sleep homeostasis by decreasing the arousal threshold, which resulted in short sleep bouts in Drosophila.
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