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


Sunday, July 31st, 2016

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Jodoin, J. N. and Martin, A. C. (2016). Abl suppresses cell extrusion and intercalation during epithelium folding. Mol Biol Cell [Epub ahead of print]. PubMed ID: 27440923
Tissue morphogenesis requires control over cell shape changes and rearrangements. In the Drosophila mesoderm, linked epithelial cells apically constrict, without cell extrusion or intercalation, to fold the epithelium into a tube that will then undergo a epithelial-to-mesenchymal transition (EMT). Apical constriction drives tissue folding or cell extrusion in different contexts, but the mechanisms that dictate the specific outcomes are poorly understood. Using live-imaging, this study found that Abelson (Abl) tyrosine kinase depletion during gastrulation causes apically constricting cells to undergo aberrant basal cell extrusion and cell intercalation. abl depletion disrupted apical-basal polarity and adherens junction organization in mesoderm cells, suggesting that extruding cells undergo premature EMT. The polarity loss was associated with abnormal basolateral contractile actomyosin and Enabled (Ena) accumulation. Depletion of the Abl effector Enabled (Ena) in abl depleted embryos suppressed the abl phenotype, consistent with cell extrusion resulting from misregulated abl. This work provides new insight as to how Abl loss and Ena misregulation promote cell extrusion and EMT.

Horn, T. and Panfilio, K. A. (2016). Novel functions for Dorsocross in epithelial morphogenesis in the beetle Tribolium castaneum. Development [Epub ahead of print]. PubMed ID: 27407103
Epithelial morphogenesis, the progressive restructuring of tissue sheets, is fundamental to embryogenesis. In insects not only embryonic tissues, but also extraembryonic (EE) epithelia play a crucial role in shaping the embryo. In Drosophila, the T-box transcription factor Dorsocross (Doc) is essential for EE tissue maintenance and therefore embryo survival. However, Drosophila possesses a single amnioserosa, whereas most insects have a distinct amnion and serosa. How does this derived situation compare with Doc function in the ancestral context of two EE epithelia? This study investigated the Doc orthologue in the flour beetle Tribolium castaneum, an excellent model for EE tissue complement and for functional, fluorescent live imaging approaches. Surprisingly, it was found that Tc-Doc controls all major events in Tribolium EE morphogenesis without affecting EE tissue specification or maintenance. These macroevolutionary changes in function between Tribolium and Drosophila are accompanied by regulatory network changes, where BMP signaling and possibly the transcription factor Hindsight are downstream mediators. It is proposed that Doc's ancestral function was to control morphogenesis and discuss how Tc-Doc could provide spatial precision for remodeling the amnion-serosa border.

Dubois, L., Frendo, J. L., Chanut-Delalande, H., Crozatier, M. and Vincent, A. (2016). Genetic dissection of the transcription factor code controlling serial specification of muscle identities in Drosophila. Elife 5 [Epub ahead of print]. PubMed ID: 27438571
Each Drosophila muscle is seeded by one Founder Cell issued from terminal division of a Progenitor Cell (PC). Muscle identity reflects the expression by each PC of a specific combination of identity Transcription Factors (iTFs). Sequential emergence of several PCs at the same position raised the question of how developmental time controlled muscle identity. This study identified roles of Anterior Open and ETS domain lacking in controlling PC birth time and Eyes absent, No Ocelli, and Sine oculis in specifying PC identity. The windows of transcription of these and other TFs in wild type and mutant embryos, revealed a cascade of regulation integrating time and space, feed-forward loops and use of alternative transcription start sites. These data provide a dynamic view of the transcriptional control of muscle identity in Drosophila and an extended framework for studying interactions between general myogenic factors and iTFs in evolutionary diversification of muscle shapes.
Hampoelz, B., Mackmull, M. T., Machado, P., Ronchi, P., Bui, K. H., Schieber, N., Santarella-Mellwig, R., Necakov, A., Andres-Pons, A., Philippe, J. M., Lecuit, T., Schwab, Y. and Beck, M. (2016). Pre-assembled nuclear pores insert into the nuclear envelope during early development. Cell [Epub ahead of print]. PubMed ID: 27397507
Nuclear pore complexes (NPCs) span the nuclear envelope (NE) and mediate nucleocytoplasmic transport. In metazoan oocytes and early embryos, NPCs reside not only within the NE, but also at some endoplasmic reticulum (ER) membrane sheets, termed annulate lamellae (AL). Although a role for AL as NPC storage pools has been discussed, it remains controversial whether and how they contribute to the NPC density at the NE. This study shows that AL insert into the NE as the ER feeds rapid nuclear expansion in Drosophila blastoderm embryos. NPCs within AL resemble pore scaffolds that mature only upon insertion into the NE. This paper delineates a topological model in which NE openings are critical for AL uptake that nevertheless occurs without compromising the permeability barrier of the NE. This unanticipated mode of pore insertion is developmentally regulated and operates prior to gastrulation.

Saturday, July 30th

Borg, R. M., Fenech Salerno, B., Vassallo, N., Bordonne, R. and Cauchi, R. J. (2016). Disruption of snRNP biogenesis factors Tgs1 and pICln induces phenotypes that mirror aspects of SMN-Gemins complex perturbation in Drosophila, providing new insights into spinal muscular atrophy. Neurobiol Dis 94: 245-258. PubMed ID: 27388936
The neuromuscular disorder, spinal muscular atrophy (SMA), results from insufficient levels of the survival motor neuron (SMN; see Drosophila Smn) protein. Together with Gemins 2-8 and Unrip, SMN forms the large macromolecular SMN-Gemins complex, which is known to be indispensable for chaperoning the assembly of spliceosomal small nuclear ribonucleoproteins (snRNPs). It remains unclear whether disruption of this function is responsible for the selective neuromuscular degeneration in SMA. This present study shows that loss of wing morphogenesis defect (wmd), the Drosophila Unrip orthologue, has a negative impact on the motor system. However, due to lack of a functional relationship between wmd/Unrip and Gemin3, it is likely that Unrip joined the SMN-Gemins complex only recently in evolution. Second, disruption of either Tgs1 or pICln, two cardinal players in snRNP biogenesis, results in viability and motor phenotypes that closely resemble those previously uncovered on loss of the constituent members of the SMN-Gemins complex. Interestingly, overexpression of both factors leads to motor dysfunction in Drosophila, a situation analogous to that of Gemin2. Toxicity is conserved in the yeast S. pombe where pICln overexpression induces a surplus of Sm proteins in the cytoplasm, indicating that a block in snRNP biogenesis is partly responsible for this phenotype. Importantly, this study shows a strong functional relationship and a physical interaction between Gemin3 and either Tgs1 or pICln. It is proposed that snRNP biogenesis is the pathway connecting the SMN-Gemins complex to a functional neuromuscular system, and its disturbance most likely leads to the motor dysfunction that is typical in SMA.

Bergkvist, L., Sandin, L., Kagedal, K. and Brorsson, A. C. (2016). AβPP processing results in greater toxicity per amount of Aβ1-42 than individually expressed and secreted Aβ1-42 in Drosophila melanogaster. Biol Open [Epub ahead of print]. PubMed ID: 27387531
The aggregation of the amyloid-β (Aβ; see Drosophila Appl) peptide into fibrillar deposits has long been considered the key neuropathological hallmark of Alzheimer's disease (AD). Aβ peptides are generated from proteolytic processing of the transmembrane Aβ precursor protein (AβPP) via sequential proteolysis through the β-secretase activity of β-site AβPP-cleaving enzyme (BACE1) and by the intramembranous enzyme γ-secretase. For over a decade, Drosophila melanogaster has been used as a model organism to study AD, and two different approaches have been developed to investigate the toxicity caused by AD-associated gene products in vivo. In one model, the Aβ peptide is directly over-expressed fused to a signal peptide, allowing secretion of the peptide into the extracellular space. In the other model, human AβPP is co-expressed with human BACE1, resulting in production of the Aβ peptide through the processing of AβPP by BACE1 and by endogenous fly γ-secretase. This study consisted of a parallel study of flies that expressed the Aβ1-42 peptide alone or that co-expressed AβPP and beta-secretase 1 (BACE1). Toxic effects (assessed by eye phenotype, longevity and locomotor assays) and levels of the Aβ1-42, Aβ1-40 and Aβ1-38 peptides were examined. The data reveal that the toxic effect per amount of detected Aβ1-42 peptide was higher in the flies co-expressing AβPP and BACE1 than in the Aβ1-42-expressing flies, and that the co-existence of Aβ1-42 and Aβ1-40 in the flies co-expressing AβPP and BACE1 could be of significant importance to the neurotoxic effect detected in these flies. Thus, the toxicity detected in these two fly models seems to have different modes of action and is highly dependent on how and where the peptide is generated rather than on the actual level of the Aβ1-42 peptide in the flies. This is important knowledge that needs to be taken into consideration when using Drosophila models to investigate disease mechanisms or therapeutic strategies in AD research.

Sung, H., Tandarich, L.C., Nguyen, K. and Hollenbeck, P.J. (2016). Compartmentalized regulation of Parkin-mediated mitochondrial quality control in the Drosophila nervous system in vivo. J Neurosci 36: 7375-7391. PubMed ID: 27413149
In neurons, the normal distribution and selective removal of mitochondria are considered essential for maintaining the functions of the large asymmetric cell and its diverse compartments. Parkin, a E3 ubiquitin ligase associated with familial Parkinson's disease, has been implicated in mitochondrial dynamics and removal in cells including neurons. However, it is not clear how Parkin functions in mitochondrial turnover in vivo, or whether Parkin-dependent events of the mitochondrial life cycle occur in all neuronal compartments. Using the live Drosophila nervous system, this study investigated the involvement of Parkin in mitochondrial dynamics, distribution, morphology, and removal. Contrary to expectations, it was found that Parkin-deficient animals do not accumulate senescent mitochondria in their motor axons or neuromuscular junctions; instead, they contain far fewer axonal mitochondria, and these displayed normal motility behavior, morphology, and metabolic state. However, the loss of Parkin does produce abnormal tubular and reticular mitochondria restricted to the motor cell bodies. In addition, in contrast to drug-treated, immortalized cells in vitro, mature motor neurons rarely display Parkin-dependent mitophagy. These data indicate that the cell body is the focus of Parkin-dependent mitochondrial quality control in neurons, and argue that a selection process allows only healthy mitochondria to pass from cell bodies to axons, perhaps to limit the impact of mitochondrial dysfunction.

Wiemerslage, L., Ismael, S. and Lee, D. (2016). Early alterations of mitochondrial morphology in dopaminergic neurons from Parkinson's disease-like pathology and time-dependent neuroprotection with D2 receptor activation. Mitochondrion [Epub ahead of print]. PubMed ID: 27423787
Neuroprotection, to prevent vulnerable cell populations from dying, is perhaps the main strategy for treating Parkinson's disease (PD). Yet in clinical practice, therapy is introduced after the disease is well established and many neurons have already disappeared, while experimentally, treatment is typically added at the same time that PD pathology is instigated. This study uses an already established Drosophila melanogaster model of PD to test for early markers of neurodegeneration and if those markers are reversible following neuroprotective treatment. Specifically, primary neuronal cultures were treated with the neurotoxin 1-methyl-4-phenylpyridinium (MPP+), and neuritic, dopaminergic mitochondria were tracked over time, observing a fragmenting change in their morphology before cell death. A neuroprotective treatment (quinpirole, a D2 receptor agonist) was added at different timepoints to determine if the changes in mitochondrial morphology are reversible. Neuroprotective treatment must be added concomitantly to prevent changes in mitochondrial morphology and subsequent cell death. This work further supports Drosophila's use as a model organism and mitochondria's use as a biomarker for neurodegenerative disease. But mainly, this work highlights an import factor for experiments in neuroprotection - time of treatment. These results highlight the problem that current neuroprotective treatments for PD may not be used the same way that they are tested experimentally.

Friday, July 29th

Bras-Pereira, C., Potier, D., Jacobs, J., Aerts, S., Casares, F. and Janody, F. (2016). dachshund potentiates Hedgehog signaling during Drosophila retinogenesis. PLoS Genet 12: e1006204. PubMed ID: 27442438
Proper organ patterning depends on a tight coordination between cell proliferation and differentiation. The patterning of Drosophila retina occurs both very fast and with high precision. This process is driven by the dynamic changes in signaling activity of the conserved Hedgehog (Hh) pathway, which coordinates cell fate determination, cell cycle and tissue morphogenesis. This study shows that during Drosophila retinogenesis, the retinal determination gene dachshund (dac) is not only a target of the Hh signaling pathway, but is also a modulator of its activity. Using developmental genetics techniques, dac was demonstrated to enhance Hh signaling by promoting the accumulation of the Gli transcription factor Cubitus interruptus (Ci) parallel to or downstream of fused. In the absence of dac, all Hh-mediated events associated to the morphogenetic furrow are delayed. One of the consequences is that, posterior to the furrow, dac- cells cannot activate a Roadkill-Cullin3 negative feedback loop that attenuates Hh signaling and which is necessary for retinal cells to continue normal differentiation. Therefore, dac is part of an essential positive feedback loop in the Hh pathway, guaranteeing the speed and the accuracy of Drosophila retinogenesis.

Huang, D., Li, X., Sun, L., Huang, P., Ying, H., Wang, H., Wu, J. and Song, H. (2016). Regulation of Hippo signalling by p38 signalling. J Mol Cell Biol [Epub ahead of print]. PubMed ID: 27402810
The Hippo signalling pathway has a crucial role in growth control during development, and its dysregulation contributes to tumorigenesis. Recent studies uncover multiple upstream regulatory inputs into Hippo signalling, which affects phosphorylation of the transcriptional coactivator Yorkie/YAP/TAZ by Warts/Lats. This study identifies the p38 MAPK pathway as a new upstream branch of the Hippo pathway. In Drosophila, overexpression of MAPKK gene licorne (lic), or MAPKKK gene Mekk1, promotes Yki activity and induces Hippo target gene expression. Loss-of-function studies show that lic regulates Hippo signalling in ovary follicle cells and in the wing disc. Epistasis analysis indicates that Mekk1 and lic affect Hippo signalling via p38b and wts It was further demonstrated that the Mekk1-Lic-p38b cascade inhibits Hippo signalling by promoting F-actin accumulation and Jelly belly phosphorylation. In addition, p38 signalling modulates actin filaments and Hippo signalling in parallel to small GTPases Ras, Rac1, and Rho1. Lastly, p38 signalling was shown to regulate Hippo signalling in mammalian cell lines. The Lic homolog MKK3 promotes nuclear localization of YAP via the actin cytoskeleton. Upregulation or downregulation of the p38 pathway regulates YAP-mediated transcription. This work thus reveals a conserved crosstalk between the p38 MAPK pathway and the Hippo pathway in growth regulation.

Yuan, Z., Praxenthaler, H., Tabaja, N., Torella, R., Preiss, A., Maier, D. and Kovall, R. A. (2016). Structure and function of the Su(H)-Hairless repressor complex, the major antagonist of Notch signaling in Drosophila melanogaster. PLoS Biol 14: e1002509. PubMed ID: 27404588
Notch is a conserved signaling pathway that specifies cell fates in metazoans. Receptor-ligand interactions induce changes in gene expression, which is regulated by the transcription factor CBF1/Su(H)/Lag-1 (CSL). CSL interacts with coregulators to repress and activate transcription from Notch target genes. While the molecular details of the activator complex are relatively well understood, the structure-function of CSL-mediated repressor complexes is poorly defined. In Drosophila, the antagonist Hairless directly binds Su(H) (the fly CSL ortholog) to repress transcription from Notch targets. This study determined the X-ray structure of the Su(H)-Hairless complex bound to DNA. Hairless binding produces a large conformational change in Su(H) by interacting with residues in the hydrophobic core of Su(H), illustrating the structural plasticity of CSL molecules to interact with different binding partners. Based on the structure, mutants in Hairless and Su(H) were designed that affect binding, but do not affect formation of the activator complex. These mutants were validated in vitro by isothermal titration calorimetry and yeast two- and three-hybrid assays. Moreover, these mutants allowed characterization the repressor function of Su(H) in vivo.

Yu, H., Takeuchi, H., Takeuchi, M., Liu, Q., Kantharia, J., Haltiwanger, R.S. and Li, H. (2016). Structural analysis of Notch-regulating Rumi reveals basis for pathogenic mutations. Nat Chem Biol [Epub ahead of print]. PubMed ID: 27428513
Rumi O-glucosylates the EGF repeats of a growing list of proteins essential in metazoan development, including Notch. Rumi is essential for Notch signaling, and Rumi dysregulation is linked to several human diseases. Despite Rumi's critical roles, it is unknown how Rumi glucosylates a serine of many but not all EGF repeats. This study reports crystal structures of Drosophila Rumi as binary and ternary complexes with a folded EGF repeat and/or donor substrates. These structures provide insights into the catalytic mechanism and show that Rumi recognizes structural signatures of the EGF motif, the U-shaped consensus sequence, C-X-S-X-(P/A)-C and a conserved hydrophobic region. It was found that five Rumi mutations identified in cancers and Dowling-Degos disease are clustered around the enzyme active site and adversely affect its activity. These data suggest that loss of Rumi activity may underlie these diseases, and the mechanistic insights may facilitate the development of modulators of Notch signaling.

Thursday, July 28th

Mavromatakis, Y.E. and Tomlinson, A. (2016). R7 photoreceptor specification in the developing Drosophila eye: The role of the transcription factor Deadpan. PLoS Genet 12: e1006159. PubMed ID: 27427987
Sequential cell fate decisions need to be made in a robust manner so there is no ambiguity in the state of the cell as it proceeds to the next stage. This study examines the decision made by the R7 precursor cell to become a photoreceptor. The transcription factor Tramtrack (Ttk) inhibits photoreceptor assignment, and previous studies have shown that the RTK-induced degradation of Ttk is critically required for R7 specification. This study found that the transcription factor Deadpan (Dpn) is also required; it is needed to silence ttk transcription, and only when Ttk protein degradation and transcriptional silencing occur together is the photoreceptor fate robustly achieved. Dpn expression needs to be tightly restricted to R7 precursors. Dpn and Ttk act as mutually repressive transcription factors, with Dpn acting to ensure that Ttk is effectively removed from R7, and Ttk acting to prevent Dpn expression in other cells. Furthermore, it was found that N activity is required to promote dpn transcription, and only in R7 precursors does the removal of Ttk coincide with high N activity, and only in this cell does Dpn expression result.

Yagi, R., Mabuchi, Y., Mizunami, M. and Tanaka, N. K. (2016). Convergence of multimodal sensory pathways to the mushroom body calyx in Drosophila melanogaster. Sci Rep 6: 29481. PubMed ID: 27404960
Detailed structural analyses of the mushroom body which plays critical roles in olfactory learning and memory revealed that it is directly connected with multiple primary sensory centers in Drosophila, for example the γ lobe neurons innervating the ventral accessory calyces respond to visual stimuli, the antennal lobe tracts neuron terminating in the lateral accessory calyces shows calcium responses to temperature shifts, and taste activity has been observed in the dorsal accessory calyces. Connectivity patterns between the mushroom body and primary sensory centers suggest that each mushroom body lobe processes information on different combinations of multiple sensory modalities. This finding provides a novel focus of research by Drosophila genetics for perception of the external world by integrating multisensory signals.

Perry, M., Kinoshita, M., Saldi, G., Huo, L., Arikawa, K. and Desplan, C. (2016). Molecular logic behind the three-way stochastic choices that expand butterfly colour vision. Nature 535: 280-284. PubMed ID: 27383790
Butterflies rely extensively on colour vision to adapt to the natural world. Most species express a broad range of colour-sensitive Rhodopsin proteins in three types of ommatidia (unit eyes), which are distributed stochastically across the retina. The retinas of Drosophila melanogaster use just two main types, in which fate is controlled by the binary stochastic decision to express the transcription factor Spineless in R7 photoreceptors. This study investigated how butterflies instead generate three stochastically distributed ommatidial types, resulting in a more diverse retinal mosaic that provides the basis for additional colour comparisons and an expanded range of colour vision. The Japanese yellow swallowtail (Papilio xuthus, Papilionidae) and the painted lady (Vanessa cardui, Nymphalidae) butterflies have a second R7-like photoreceptor in each ommatidium. Independent stochastic expression of Spineless in each R7-like cell results in expression of a blue-sensitive (SpinelessON) or an ultraviolet (UV)-sensitive (SpinelessOFF) Rhodopsin. In P. xuthus these choices of blue/blue, blue/UV or UV/UV sensitivity in the two R7 cells are coordinated with expression of additional Rhodopsin proteins in the remaining photoreceptors, and together define the three types of ommatidia. Knocking out spineless using CRISPR/Cas9 leads to the loss of the blue-sensitive fate in R7-like cells and transforms retinas into homogeneous fields of UV/UV-type ommatidia, with corresponding changes in other coordinated features of ommatidial type. Hence, the three possible outcomes of Spineless expression define the three ommatidial types in butterflies. This developmental strategy allowed the deployment of an additional red-sensitive Rhodopsin in P. xuthus, allowing for the evolution of expanded colour vision with a greater variety of receptors. This surprisingly simple mechanism that makes use of two binary stochastic decisions coupled with local coordination may prove to be a general means of generating an increased diversity of developmental outcomes.

Narbonne-Reveau, K., Lanet, E., Dillard, C., Foppolo, S., Chen, C. H., Parrinello, H., Rialle, S., Sokol, N. S. and Maurange, C. (2016). Neural stem cell-encoded temporal patterning delineates an early window of malignant susceptibility in Drosophila. Elife 5 [Epub ahead of print]. PubMed ID: 27296804
Pediatric neural tumors are often initiated during early development and can undergo very rapid transformation. However, the molecular basis of this early malignant susceptibility remains unknown. During Drosophila development, neural stem cells (NSCs) divide asymmetrically and generate intermediate progenitors that rapidly differentiate in neurons. Upon gene inactivation, these progeny can dedifferentiate and generate malignant tumors. This study found that intermediate progenitors, are prone to malignancy only when born during an early window of development, during early larval stages, while expressing the transcription factor Chinmo, and the mRNA-binding proteins Imp/IGF2BP and Lin-28. These genes compose an oncogenic module that is coopted upon dedifferentiation of early-born intermediate progenitors to drive unlimited tumor growth. In late larvae, temporal transcription factor progression in NSCs silences the module, thereby limiting mitotic potential and terminating the window of malignant susceptibility. Thus, this study identifies the gene regulatory network that confers malignant potential to neural tumors with early developmental origins.

Li, S. A., Cheng, L., Yu, Y. and Chen, Q. (2016). Structural basis of Dscam1 homodimerization: Insights into context constraint for protein recognition. Sci Adv 2: e1501118. PubMed ID: 27386517
The Drosophila neural receptor Dscam1 (Down syndrome cell adhesion molecule 1) plays an essential role in neuronal wiring and self-avoidance. Dscam1 potentially encodes 19,008 ectodomains through alternative RNA splicing and exhibits exquisite isoform-specific homophilic binding, which makes it an exceptional example for studying protein binding specificity. However, structural information on Dscam1 is limited, which hinders illumination of the mechanism of Dscam1 isoform-specific recognition. Whether different Dscam1 isoforms adopt the same dimerization mode remains a subject of debate. This study presents 12 Dscam1 crystal structures, provide direct evidence indicating that all isoforms adopt a conserved homodimer geometry in a modular fashion, identifies two mechanisms for the Ig2 binding domain to dispel electrostatic repulsion during dimerization, decodes Ig2 binding specificity by a central motif at its symmetry center, uncovers the role of glycosylation in Dscam1 homodimerization, and finds electrostatic potential complementarity to help define the binding region and the antiparallel binding mode. A concept is proposed that the context of a protein may set restrictions to regulate its binding specificity, which provides a better understanding of protein recognition.

Wednesday, July 27th

Waghmare, I. and Kango-Singh, M. (2016). Loss of cell adhesion increases tumorigenic potential of polarity deficient Scribble mutant cells. PLoS One 11: e0158081. PubMed ID: 27327956
Epithelial polarity genes are important for maintaining tissue architecture, and regulating growth. The Drosophila neoplastic tumor suppressor gene scribble (scrib) belongs to the basolateral polarity complex. Loss of scrib results in disruption of its growth regulatory functions, and downregulation or mislocalization of Scrib is correlated to tumor growth. Somatic scribble mutant cells (scrib-) surrounded by wild-type cells undergo apoptosis, which can be prevented by introduction of secondary mutations that provide a growth advantage. Using genetic tools in Drosophila, the phenotypic effects were analyzed of loss of scrib in different growth promoting backgrounds. This study investigated if a central mechanism that regulates cell adhesion governs the growth and invasive potential of scrib mutant cells. Increased proliferation, and survival abilities of scrib- cells in different genetic backgrounds affect their differentiation, and intercellular adhesion. Further, loss of scrib is sufficient to cause reduced cell survival, activation of the JNK pathway and a mild reduction of cell adhesion. These data show that for scrib cells to induce aggressive tumor growth characterized by loss of differentiation, cell adhesion, increased proliferation and invasion, cooperative interactions that derail signaling pathways play an essential role in the mechanisms leading to tumorigenesis. Thus, this study provides new insights on the effects of loss of scrib and the modification of these effects via cooperative interactions that enhance the overall tumorigenic potential of scrib deficient cells.

Wu, J., Wang, H., Guo, X. and Chen, J. (2016). Cofilin-mediated actin dynamics promotes actin bundle formation during Drosophila bristle development. Mol Biol Cell [Epub ahead of print]. PubMed ID: 27385345
The actin bundle is an array of linear actin filaments cross-linked by actin bundling proteins, but its assembly and dynamics are not as well understood as those of branched actin network. This study used the Drosophila bristle as a model system to study actin bundle formation. cofilin, a major actin disassembly factor of branched actin network, was found to promote the formation and positioning of actin bundles in the developing bristles. Loss-of-function of cofilin or AIP1, a cofactor of cofilin, each resulted in increased F-actin levels and severe defects in actin bundle organization, with the defects from cofilin deficiency being more severe. Further analyses revealed that cofilin likely regulates actin bundle formation and positioning by the following means. First, cofilin promotes a large G-actin pool both locally and globally, likely ensuring rapid actin polymerization for bundle initiation and growth. Second, cofilin limits the size of a nonbundled actin-myosin network to regulate positioning of actin bundles. Third, cofilin prevents incorrect assembly of branched and myosin-associated actin filament into bundles. Together, these results demonstrate that the interaction between the dynamic dendritic actin network and the assembling actin bundles is critical for actin bundle formation and needs to be closely regulated.

Cho, A., Kato, M., Whitwam, T., Kim, J.H. and Montell, D.J. (2016). An atypical tropomyosin in Drosophila with intermediate filament-like properties. Cell Rep [Epub ahead of print]. PubMed ID: 27396338
A longstanding mystery has been the absence of cytoplasmic intermediate filaments (IFs) from Drosophila despite their importance in other organisms. In the course of characterizing the in vivo expression and functions of Drosophila Tropomyosin (Tm) isoforms, this study discovered an essential but unusual product of the Tm1 locus, Tm1-I/C, which resembles an IF protein in some respects. Like IFs, Tm1-I/C spontaneously forms filaments in vitro that are intermediate in diameter between F-actin and microtubules. Like IFs but unlike canonical Tms, Tm1-I/C contains N- and C-terminal low-complexity domains flanking a central coiled coil. In vivo, Tm1-I/C forms cytoplasmic filaments that do not associate with F-actin or canonical Tms. Tm1-I/C is essential for collective border cell migration, in epithelial cells for proper cytoarchitecture, and in the germline for the formation of germ plasm. These results suggest that flies have evolved a distinctive type of cytoskeletal filament from Tm.

Nashchekin, D., Fernandes, A. R. and St Johnston, D. (2016). Patronin/Shot cortical foci assemble the noncentrosomal microtubule array that specifies the Drosophila anterior-posterior axis. Dev Cell 38: 61-72. PubMed ID: 27404359
Noncentrosomal microtubules play an important role in polarizing differentiated cells, but little is known about how these microtubules are organized. This study identified the spectraplakin, Short stop (Shot), as the cortical anchor for noncentrosomal microtubule organizing centers (ncMTOCs) in the Drosophila oocyte. Shot interacts with the cortex through its actin-binding domain and recruits the microtubule minus-end-binding protein, Patronin, to form cortical ncMTOCs. Shot/Patronin foci do not co-localize with gamma-tubulin, suggesting that they do not nucleate new microtubules. Instead, they capture and stabilize existing microtubule minus ends, which then template new microtubule growth. Shot/Patronin foci are excluded from the oocyte posterior by the Par-1 polarity kinase to generate the polarized microtubule network that localizes axis determinants. Both proteins also accumulate apically in epithelial cells, where they are required for the formation of apical-basal microtubule arrays. Thus, Shot/Patronin ncMTOCs may provide a general mechanism for organizing noncentrosomal microtubules in differentiated cells.

Tuesday, July 26th

La Fortezza, M., Schenk, M., Cosolo, A., Kolybaba, A., Grass, I. and Classen, A.K. (2016). JAK/STAT signalling mediates cell survival in response to tissue stress. Development [Epub ahead of print]. PubMed ID: 27385008
Tissue homeostasis relies on the ability of tissues to respond to stress. Tissue regeneration and tumour models in Drosophila have shown that JNK is a prominent stress-response pathway promoting injury-induced apoptosis and compensatory proliferation. A central question remaining unanswered is how both responses are balanced by activation of a single pathway. JAK/STAT signalling, a potential JNK target, is implicated in promoting compensatory proliferation. While JAK/STAT activation in imaginal discs was observed upon damage, it was also found that JAK/STAT and its downstream effector Zfh2 promote survival of JNK-signalling cells instead. The JNK component fos and the pro-apoptotic gene hid are regulated in a JAK/STAT-dependent manner. This molecular pathway restrains JNK-induced apoptosis and spatial propagation of JNK-signalling, thereby limiting the extent of tissue damage, as well as facilitating systemic and proliferative responses to injury. It was found that the pro-survival function of JAK/STAT also drives tumour growth under conditions of chronic stress. Altogether, these results define JAK/STAT function in tissue stress and illustrate how crosstalk between conserved signalling pathways establish an intricate equilibrium between proliferation, apoptosis and survival to restore tissue homeostasis.

Shimaji, K., Konishi, T., Yoshida, H., Kimura, H. and Yamaguchi, M. (2016). Genome-wide genetic screen identified the link between dG9a and epidermal growth factor receptor signaling pathway in vivo. Exp Cell Res [Epub ahead of print]. PubMed ID: 27343629
G9a is one of the histone H3 Lys 9 (H3K9) specific methyltransferases first identified in mammals. Drosophila G9a (dG9a) has been reported to induce H3K9 dimethylation in vivo, and the target genes of dG9a were identified during embryonic and larval stages. Although dG9a is important for a variety of developmental processes, the link between dG9a and signaling pathways are not addressed yet.By genome-wide genetic screen, taking advantage of the rough eye phenotype of flies that over-express dG9a in eye discs, this study identified 16 genes that enhanced the rough eye phenotype induced by dG9a over-expression. These 16 genes included Star, anterior open, bereft and F-box and leucine-rich repeat protein 6 which are components of Epidermal growth factor receptor (EGFR) signaling pathway. When dG9a over-expression was combined with mutation of Star, differentiation of R7 photoreceptors in eye imaginal discs as well as cone cells and pigment cells in pupal retinae was severely inhibited. Furthermore, the dG9a over-expression reduced the activated ERK signals in eye discs. These data demonstrate a strong genetic link between dG9a and the EGFR signaling pathway.

Top, D., Harms, E., Syed, S., Adams, E. L. and Saez, L. (2016). GSK-3 and CK2 kinases converge on Timeless to regulate the master clock. Cell Rep [Epub ahead of print]. PubMed ID: 27346344
The molecular clock relies on a delayed negative feedback loop of transcriptional regulation to generate oscillating gene expression. Although the principal components of the clock are present in all circadian neurons, different neuronal clusters have varying effects on rhythmic behavior, suggesting that the clocks they house are differently regulated. Combining biochemical and genetic techniques in Drosophila, this study identified a phosphorylation program native to the master pacemaker neurons that regulates the timing of nuclear accumulation of the Period/Timeless repressor complex. GSK-3/SGG binds and phosphorylates Period-bound Timeless, triggering a CK2-mediated phosphorylation cascade. Mutations that block the hierarchical phosphorylation of Timeless in vitro also delay nuclear accumulation in both tissue culture and in vivo and predictably change rhythmic behavior. This two-kinase phosphorylation cascade is anatomically restricted to the eight master pacemaker neurons, distinguishing the regulatory mechanism of the molecular clock within these neurons from the other clocks that cooperate to govern behavioral rhythmicity.

Squarr, A. J., Brinkmann, K., Chen, B., Steinbacher, T., Ebnet, K., Rosen, M. K. and Bogdan, S. (2016). Fat2 acts through the WAVE regulatory complex to drive collective cell migration during tissue rotation. J Cell Biol 212: 591-603. PubMed ID: 26903538
Directional cell movements during morphogenesis require the coordinated interplay between membrane receptors and the actin cytoskeleton. The WAVE regulatory complex (WRC; see Drosophila Arp2/3 component Actin-related protein 2/3 complex) is a conserved actin regulator. This study found that the atypical cadherin Fat2 recruits the WRC to basal membranes of tricellular contacts where a new type of planar-polarized whip-like actin protrusion is formed. Loss of either Fat2 function or its interaction with the WRC disrupts tricellular protrusions and results in the formation of nonpolarized filopodia. Evidence is provided for a molecular network in which the receptor tyrosine phosphatase Dlar interacts with the WRC to couple the extracellular matrix, the membrane, and the actin cytoskeleton during egg elongation. The data uncover a mechanism by which polarity information can be transduced from a membrane receptor to a key actin regulator to control collective follicle cell migration during egg elongation. 4D-live imaging of rotating MCF10A mammary acini further suggests an evolutionary conserved mechanism driving rotational motions in epithelial morphogenesis.

Monday, July 25th

Trost, M., Blattner, A. C., Leo, S. and Lehner, C. F. (2016). Drosophila dany is essential for transcriptional control and nuclear architecture in spermatocytes. Development 143: 2664-2676. PubMed ID: 27436041
The terminal differentiation of adult stem cell progeny depends on transcriptional control. A dramatic change in gene expression programs accompanies the transition from proliferating spermatogonia to postmitotic spermatocytes, which prepare for meiosis and subsequent spermiogenesis. More than a thousand spermatocyte-specific genes are transcriptionally activated in early Drosophila spermatocytes. This study describes the identification and initial characterization of dany (CG30401), a gene required in spermatocytes for the large-scale change in gene expression. Similar to tMAC (a testis-specific meiotic arrest complex (see Always early)) and tTAFs (see Cannonball), the known major activators of spermatocyte-specific genes, dany has a recent evolutionary origin, but it functions independently. Like dan and danr, its primordial relatives with functions in somatic tissues, dany encodes a nuclear Psq domain protein. Dany associates preferentially with euchromatic genome regions. In dany mutant spermatocytes, activation of spermatocyte-specific genes and silencing of non-spermatocyte-specific genes are severely compromised and the chromatin no longer associates intimately with the nuclear envelope. Therefore, as suggested recently for Dan/Danr, it is proposed that Dany is essential for the coordination of change in cell type-specific expression programs and large-scale spatial chromatin reorganization.

Boija, A. and Mannervik, M. (2016). Initiation of diverse epigenetic states during nuclear programming of the Drosophila body plan. Proc Natl Acad Sci U S A [Epub ahead of print]. PubMed ID: 27439862
Epigenetic patterns of histone modifications contribute to the maintenance of tissue-specific gene expression. This study shows that such modifications also accompany the specification of cell identities by the NF-kappaB transcription factor Dorsal in the precellular Drosophila embryo. Evidence is provided that the maternal pioneer factor, Zelda, is responsible for establishing poised RNA polymerase at Dorsal target genes before Dorsal-mediated zygotic activation. At the onset of cell specification, Dorsal recruits the CBP/p300 coactivator to the regulatory regions of defined target genes in the presumptive neuroectoderm, resulting in their histone acetylation and transcriptional activation. These genes are inactive in the mesoderm due to transcriptional quenching by the Snail repressor, which precludes recruitment of CBP and prevents histone acetylation. By contrast, inactivation of the same enhancers in the dorsal ectoderm is associated with Polycomb-repressed H3K27me3 chromatin. Thus, the Dorsal morphogen gradient produces three distinct histone signatures including two modes of transcriptional repression, active repression (hypoacetylation), and inactivity (H3K27me3). Whereas histone hypoacetylation is associated with a poised polymerase, H3K27me3 displaces polymerase from chromatin. These results link different modes of RNA polymerase regulation to separate epigenetic patterns and demonstrate that developmental determinants orchestrate differential chromatin states, providing new insights into the link between epigenetics and developmental patterning.

Hayashi, D., Tanabe, K., Katsube, H. and Inoue, Y. H. (2016). B-type nuclear lamin and the nuclear pore complex Nup107-160 influences maintenance of the spindle envelope required for cytokinesis in Drosophila male meiosis. Biol Open [Epub ahead of print]. PubMed ID: 27402967
In higher eukaryotes, nuclear envelope (NE) disassembly allows chromatin to condense and spindle microtubules to access kinetochores. The nuclear lamina, which strengthens the NE, is composed of a polymer meshwork made of A- and B-type lamins. This study found that the B-type lamin (Lam) is not fully disassembled and continues to localize along the spindle envelope structure during Drosophila male meiosis I, while the A-type lamin (LamC) is completely dispersed throughout the cytoplasm. Among the nuclear pore complex proteins, Nup107 co-localized with Lam during this meiotic division. Surprisingly, Lam depletion resulted in a higher frequency of cytokinesis failure in male meiosis. The similar meiotic phenotype was observed in Nup107-depleted cells. Abnormal localization of Lam was found in the Nup-depleted cells at premeiotic and meiotic stages. The central spindle microtubules became abnormal and recruitment of a contractile ring component to the cleavage sites was disrupted in Lam-depleted cells and Nup107-depleted cells. Therefore, it is speculated that both proteins are required for a reinforcement of the spindle envelope, which supports the formation of central spindle microtubules essential for cytokinesis in Drosophila male meiosis.

Mendiratta, S., Bhatia, S., Jain, S., Kaur, T. and Brahmachari, V. (2016). Interaction of the chromatin remodeling protein hINO80 with DNA. PLoS One 11: e0159370. PubMed ID: 27428271
The presence of a highly conserved DNA binding domain in INO80 subfamily predicted that INO80 directly interacts with DNA and this study demonstrates its DNA binding activity in vitro. The consensus motif recognized by the DBINO domain was identified by SELEX method and the specific interaction of INO80 with the consensus motif was demonstrated. INO80 significantly down regulates the reporter gene expression through its binding motif, and the repression is dependent on the presence of INO80 but not YY1 (see Drosophila Pho) in the cell. The interaction is lost if specific residues within the consensus motif are altered. A large number of potential target sites of INO80 were identified in the human genome through in silico analysis that can grouped into three classes; sites that contain the recognition sequence for INO80 and YY1, only YY1 and only INO80. The binding of INO80 to a representative set of sites in HEK cells and the correlated repressive histone modifications around the binding motif were demonstrated. In the light of the role of INO80 in homeotic gene regulation in Drosophila as an Enhancer of trithorax and polycomb protein (ETP) that can modify the effect of both repressive complexes like polycomb as well as the activating complex like trithorax, it remains to be seen if INO80 can act as a recruiter of chromatin modifying complexes.

Sunday, July 24th

Apitz, H. and Salecker, I. (2016). Retinal determination genes coordinate neuroepithelial specification and neurogenesis modes in the Drosophila optic lobe. Development 143: 2431-2442. PubMed ID: 27381228
Differences in neuroepithelial patterning and neurogenesis modes contribute to area-specific diversifications of neural circuits. In the Drosophila visual system, two neuroepithelia, the outer (OPC) and inner (IPC) proliferation centers, generate neuron subtypes for four ganglia in several ways. Whereas neuroepithelial cells in the medial OPC directly convert into neuroblasts, in an IPC subdomain they generate migratory progenitors by epithelial-mesenchymal transition that mature into neuroblasts in a second proliferative zone. The molecular mechanisms that regulate the identity of these neuroepithelia, including their neurogenesis modes, remain poorly understood. Analysis of Polycomblike revealed that loss of Polycomb group-mediated repression of the Hox gene Abdominal-B (Abd-B) causes the transformation of OPC to IPC neuroepithelial identity. This suggests that the neuroepithelial default state is IPC-like, whereas OPC identity is derived. Ectopic Abd-B blocks expression of the highly conserved retinal determination gene network members Eyes absent (Eya), Sine oculis (So) and Homothorax (Hth). These factors are essential for OPC specification and neurogenesis control. Finally, eya and so are also sufficient to confer OPC-like identity, and, in parallel with hth, the OPC-specific neurogenesis mode on the IPC. 

Panser, K., Tirian, L., Schulze, F., Villalba, S., Jefferis, G.S., Bühler, K. and Straw, A.D. (2016). Automatic segmentation of Drosophila neural compartments using GAL4 expression data reveals novel visual pathways. Curr Biol [Epub ahead of print]. PubMed ID: 27426516
Identifying distinct anatomical structures within the brain and developing genetic tools to target them are fundamental steps for understanding brain function. This study hypothesized that enhancer expression patterns can be used to automatically identify functional units such as neuropils and fiber tracts. Two recent, genome-scale Drosophila GAL4 libraries and associated confocal image datasets were used to segment large brain regions into smaller subvolumes. Obtained results (available at Straw Lab BrainCode) support this hypothesis because regions with well-known anatomy, namely the antennal lobes and central complex, were automatically segmented into familiar compartments. The basis for the structural assignment is clustering of voxels based on patterns of enhancer expression. These initial clusters are agglomerated to make hierarchical predictions of structure. The algorithm was applied to central brain regions receiving input from the optic lobes. Based on the automated segmentation and manual validation, promising driver lines for 11 previously identified and 14 novel types of visual projection neurons and their associated optic glomeruli could be identified and are available for sharing. The same strategy can be used in other brain regions and likely other species, including vertebrates.

Yuan, L., Hu, S., Okray, Z., Ren, X., De Geest, N., Claeys, A., Yan, J., Bellefroid, E., Hassan, B.A. and Quan, X.J. (2016). The Drosophila Neurogenin, Tap, functionally interacts with the Wnt-PCP pathway to regulate neuronal extension and guidance. Development [Epub ahead of print]. PubMed ID: 27385016
The Neurogenin (Ngn) transcription factors control early neurogenesis and neurite outgrowth in mammalian cortex. In contrast to their proneural activity, their function in neurite growth is poorly understood. Drosophila has a single predicted Ngn homologue called Tap, whose function is completely unknown. This study shows that Tap is not a proneural protein in Drosophila but is required for proper axonal growth and guidance of neurons of the mushroom body (MB), a neuropile required for associative learning and memory. Genetic and expression analyses suggest that Tap inhibits excessive axonal growth by fine regulation of the levels of the Wnt signaling adaptor protein, Dishevelled.

Yakimova, A. O., Pugacheva, O. M., Golubkova, E. V. and Mamon, L. A. (2016). Cytoplasmic localization of SBR (Dm NXF1) protein and its zonal distribution in the ganglia of Drosophila melanogaster larvae. Invert Neurosci 16: 9. PubMed ID: 27389771
The Drosophila gene Dm nxf1 (nuclear export factor 1) previously known as small bristles (sbr) controls nuclear export of various mRNA transcripts. Dm NXF1 is present not only in nucleoplasm or at the nuclear rim but also in the cytoplasm. On the spatiotemporal level, anti-SBR antibodies labeled some neuroblasts and their lineages in the brains of Drosophila larvae. The number of Dm NXF1-rich lineages increased during larval development, but Dm NXF1 expression was not evident in all lineages. In all larval stages, Dm NXF1 concentrated in the midline cells of the ventral nerve cord, which reflects a specific status of those cells. In neurites, Dm NXF1 was present in the form of cytoplasmic granules, which is similar to the behavior of another RNA-binding protein, dFMR. Interestingly, though, the granule expression pattern of Dm NXF1 and dFMR did not always overlap, as some granules stained exclusively for one or the other protein. It suggests the existence of specific mRNA partners for Dm NXF1 in neurites.

Saturday, July 23rd

Okuda, K., Tong, M., Dempsey, B., Moore, K. J., Gazzinelli, R. T. and Silverman, N. (2016). Leishmania amazonensis engages CD36 to drive parasitophorous vacuole maturation. PLoS Pathog 12: e1005669. PubMed ID: 27280707
Leishmania amastigotes manipulate the activity of macrophages to favor their own success. However, very little is known about the role of innate recognition and signaling triggered by amastigotes in this host-parasite interaction. This work developed a new infection model in adult Drosophila to take advantage of its superior genetic resources to identify novel host factors limiting Leishmania amazonensis infection. The model is based on the capacity of macrophage-like cells, plasmatocytes, to phagocytose and control the proliferation of parasites injected into adult flies. Using this model, a collection of RNAi-expressing flies were screened for anti-Leishmania defense factors. Notably, three CD36-like scavenger receptors (croquemort, CG31741, and CG10345) were found that were important for defending against Leishmania infection. Mechanistic studies in mouse macrophages showed that CD36 accumulates specifically at sites where the parasite contacts the parasitophorous vacuole membrane. Furthermore, CD36-deficient macrophages were defective in the formation of the large parasitophorous vacuole typical of L. amazonensis infection, a phenotype caused by inefficient fusion with late endosomes and/or lysosomes. These data identify an unprecedented role for CD36 in the biogenesis of the parasitophorous vacuole and further highlight the utility of Drosophila as a model system for dissecting innate immune responses to infection.

Shiratsuchi, A., Nitta, M., Kuroda, A., Komiyama, C., Gawasawa, M., Shimamoto, N., Tuan, T. Q., Morita, T., Aiba, H. and Nakanishi, Y. (2016). Inhibition of phagocytic killing of Escherichia coli in Drosophila hemocytes by RNA chaperone Hfq. J Immunol [Epub ahead of print]. PubMed ID: 27357148
An RNA chaperone of Escherichia coli, called host factor required for phage Qbeta RNA replication (Hfq), forms a complex with small noncoding RNAs to facilitate their binding to target mRNA for the alteration of translation efficiency and stability. Although the role of Hfq in the virulence and drug resistance of bacteria has been suggested, how this RNA chaperone controls the infectious state remains unknown. The present study addressed this issue using Drosophila melanogaster as a host for bacterial infection. In an assay for abdominal infection using adult flies, an E. coli strain with mutation in hfq was eliminated earlier, whereas flies survived longer compared with infection with a parental strain. The same was true with flies deficient in humoral responses, but the mutant phenotypes were not observed when a fly line with impaired hemocyte phagocytosis was infected. The results from an assay for phagocytosis in vitro revealed that Hfq inhibits the killing of E. coli by Drosophila phagocytes after engulfment. Furthermore, Hfq seemed to exert this action partly through enhancing the expression of E. coli σ38, a stress-responsive sigma factor that was previously shown to be involved in the inhibition of phagocytic killing of E. coli, by a posttranscriptional mechanism. This study indicates that the RNA chaperone Hfq contributes to the persistent infection of E. coli by maintaining the expression of bacterial genes, including one coding for sigma38, that help bacteria evade host immunity.

Yang, W., Dierking, K., Rosenstiel, P. C. and Schulenburg, H. (2016). GATA transcription factor as a likely key regulator of the Caenorhabditis elegans innate immune response against gut pathogens .Zoology (Jena) [Epub ahead of print]. PubMed ID: 27372411
Invertebrate defence against pathogens exclusively relies on components of the innate immune system. Comprehensive information has been collected over the last years on the molecular components of invertebrate immunity and the involved signalling processes, especially for the main invertebrate model species, the fruitfly Drosophila melanogaster and the nematode Caenorhabditis elegans. Yet, the exact regulation of general and specific defences is still not well understood. In the current study, advantage was taken of a recently established database, WormExp, which combines all available gene expression studies for C. elegans, in order to explore commonalities and differences in the regulation of nematode immune defence against a large variety of pathogens versus food microbes. Significant overlaps were identified in the transcriptional response towards microbes, especially pathogenic bacteria. It was also found that the GATA motif is overrepresented in many microbe-induced gene sets and in targets of other previously identified regulators of worm immunity. Moreover, the activated targets of one of the known C. elegans GATA transcription factors, ELT-2 (see Drosophila Serpent), are significantly enriched in the gene sets, which are differentially regulated by gut-infecting pathogens. These findings strongly suggest that GATA transcription factors and particularly ELT-2 play a central role in regulating the C. elegans immune response against gut pathogens. More specific responses to distinct pathogens may be mediated by additional transcription factors, either acting alone or jointly with GATA transcription factors. Taken together, this analysis of the worm's transcriptional response to microbes provides a new perspective on the C. elegans immune system, which is proposed to be coordinated by GATA transcription factor ELT-2 in the gut.

Zemanova, M., Staskova, T. and Kodrik, D. (2016). Role of adipokinetic hormone and adenosine in the anti-stress response in Drosophila melanogaster. J Insect Physiol 91-92: 39-47. PubMed ID: 27374982
The role of adipokinetic hormone (AKH) and adenosine in the anti-stress response was studied in Drosophila melanogaster larvae and adults carrying a mutation in the Akh gene (Akh1), the adenosine receptor gene (AdoR1), or in both of these genes (Akh1 AdoR1 double mutant). Stress was induced by starvation or by the addition of an oxidative stressor paraquat (PQ) to food. Mortality tests revealed that the AAkh1 mutant was the most resistant to starvation, while the AdoR1 mutant was the most sensitive. Conversely, the Akh1 AdoR1 double mutant was more sensitive to PQ toxicity than either of the single mutants. Administration of PQ significantly increased the Drome-AKH level in w1118 and AdoR1 larvae; however, this was not accompanied by a simultaneous increase in Akh gene expression. In contrast, PQ significantly increased the expression of the glutathione S-transferase D1 (GstD1) gene. The presence of both a functional adenosine receptor and AKH seem to be important for the proper control of GstD1 gene expression under oxidative stress, however, the latter appears to play more dominant role. On the other hand, differences in glutathione S-transferase (GST) activity among the strains, and between untreated and PQ-treated groups were minimal. In addition, the glutathione level was significantly lower in all untreated AKH- or AdoR-deficient mutant flies as compared with the untreated control w1118 flies and further declined following treatment with PQ. All oxidative stress characteristics modified by mutations in Akh gene were restored or even improved by 'rescue' mutation in flies which ectopically express Akh. Thus, the results demonstrate the important roles of AKH and adenosine in the anti-stress response elicited by PQ in a Drosophila model, and provide the first evidence for the involvement of adenosine in the anti-oxidative stress response in insects.

Friday, July 22nd

Quijano, J.C., Wisotzkey, R.G., Tran, N.L., Huang, Y., Stinchfield, M.J., Haerry, T.E., Shimmi, O. and Newfeld, S.J. (2016). lolal is an evolutionarily new epigenetic regulator of dpp transcription during dorsal-ventral axis formation. Mol Biol Evol [Epub ahead of print]. PubMed ID: 27401231
Secreted ligands in the Dpp/BMP family drive dorsal-ventral (D/V) axis formation in all Bilaterian species. However, maternal factors regulating Dpp/BMP transcription in this process are largely unknown. This study identified the BTB domain protein longitudinals lacking-like (lolal) as a modifier of decapentaplegic (dpp) mutations. It was shown that Lolal is evolutionarily related to the Trithorax group of chromatin regulators and that lolal interacts genetically with the epigenetic factor Trithorax-like during Dpp D/V signaling. Maternally driven LolalHA is found in oocytes and translocates to zygotic nuclei prior to the point at which dpp transcription begins. lolal maternal and zygotic mutant embryos display significant reductions in dpp, pMad and zerknullt expression, but they are never absent. The data suggest that lolal is required to maintain dpp transcription during D/V patterning. Phylogenetic data reveals that lolal is an evolutionarily new gene present only in insects and crustaceans. The study concludes that Lolal is the first maternal protein with a role in dpp D/V transcriptional maintenance, that Lolal and the epigenetic protein Trithorax-like are essential for Dpp D/V signaling and that the architecture of the Dpp D/V pathway evolved in the arthropod lineage after the separation from vertebrates via the incorporation of new genes such as lolal

McCracken, A. and Locke, J. (2016). Mutations in ash1 and trx enhance P-element-dependent silencing in Drosophila melanogaster. Genome [Epub ahead of print]. PubMed ID: 27373142
In Drosophila melanogaster, the mini-w+ transgene in Pci (a transgene inserted proximally on chromosome 4 between Ribosomal protein S3A (RpS3A) and cubitus interruptus (ci), is normally expressed throughout the adult eye; however, when other P or KP elements are present, a variegated-eye phenotype results, indicating random w+ silencing during development called P-element-dependent silencing (PDS). Mutant Su(var)205 and Su(var)3-7 alleles act as haplo-suppressors/triplo-enhancers of this variegated phenotype, indicating that these heterochromatic modifiers act dose dependently in PDS. Previously, a spontaneous mutation of P{lacW}ciDplac called P{lacW}ciDplacE1 (E1) was recovered that variegated in the absence of P elements, presumably due to the insertion of an adjacent gypsy element. From a screen for genetic modifiers of E1 variegation, this study describes the isolation of five mutations in ash1 and three in trx that enhance the E1 variegated phenotype in a dose-dependent and cumulative manner. These mutant alleles enhance PDS at E1, and in E1/P{lacW}ciDplac, but suppress position effect variegation (PEV) at In(1)wm4. This opposite action is consistent with a model where ASH1 and TRX mark transcriptionally active chromatin domains. If ASH1 or TRX function is lost or reduced, heterochromatin can spread into these domains creating a sink that diverts heterochromatic proteins from other variegating locations, which then may express a suppressed phenotype.

Iwasaki, Y. W., Murano, K., Ishizu, H., Shibuya, A., Iyoda, Y., Siomi, M. C., Siomi, H. and Saito, K. (2016). Piwi modulates chromatin accessibility by regulating multiple factors including Histone H1 to repress transposons.Mol Cell [Epub ahead of print]. PubMed ID: 27425411
PIWI-interacting RNAs (piRNAs) mediate transcriptional and post-transcriptional silencing of transposable element (TE) in animal gonads. In Drosophila ovaries, Piwi-piRNA complexes (Piwi-piRISCs) repress TE transcription by modifying the chromatin state, such as by H3K9 trimethylation. This study demonstrates that Piwi physically interacts with linker histone H1. Depletion of Piwi decreases H1 density at a subset of TEs, leading to their derepression. Silencing at these loci separately requires H1 and H3K9me3 and Heterochromatin protein 1a (HP1a). Loss of H1 increases target loci chromatin accessibility without affecting H3K9me3 density at these loci, while loss of HP1a does not impact H1 density. Thus, Piwi-piRISCs require both H1 and HP1a to repress TEs, and the silencing is correlated with the chromatin state rather than H3K9me3 marks. These findings suggest that Piwi-piRISCs regulate the interaction of chromatin components with target loci to maintain silencing of TEs through the modulation of chromatin accessibility.

Du, J., Zhang, J., He, T., Li, Y., Su, Y., Tie, F., Liu, M., Harte, P. J. and Zhu, A. J. (2016). Stuxnet facilitates the degradation of Polycomb protein during development. Dev Cell 37: 507-519. PubMed ID: 27326929
Polycomb-group (PcG) proteins function to ensure correct deployment of developmental programs by epigenetically repressing target gene expression. Despite the importance, few studies have been focused on the regulation of PcG activity itself. This study reports a Drosophila gene, stuxnet (stx), that controls Pc protein stability. Heightened stx activity leads to homeotic transformation, reduced Pc activity, and de-repression of PcG targets. Conversely, stx mutants, which can be rescued by decreased Pc expression, display developmental defects resembling hyperactivation of Pc. Biochemical analyses provide a mechanistic basis for the interaction between stx and Pc; Stx facilitates Pc degradation in the proteasome, independent of ubiquitin modification. Furthermore, this mode of regulation is conserved in vertebrates. Mouse stx promotes degradation of Cbx4, an orthologous Pc protein, in vertebrate cells and induces homeotic transformation in Drosophila. These results highlight an evolutionarily conserved mechanism of regulated protein degradation on PcG homeostasis and epigenetic activity.

Thursday, July 21st

Nakazawa, M., Matsubara, H., Matsushita, Y., Watanabe, M., Vo, N., Yoshida, H., Yamaguchi, M. and Kataoka, T. (2016). The human Bcl-2 family member Bcl-rambo localizes to mitochondria and induces apoptosis and morphological aberrations in Drosophila. PLoS One 11: e0157823. PubMed ID: 27348811
Bcl-2 family proteins play a central role in regulating apoptosis. It has been previously reported that human Bcl-rambo, also termed BCL2L13, localizes to mitochondria and induces apoptosis when overexpressed in human embryonic kidney 293T cells. However, the physiological function of Bcl-rambo currently remains unclear. In the present study, human Bcl-rambo was ectopically expressed in Drosophila melanogaster. It was found to mainly localize to the mitochondria of Drosophila Schneider 2 (S2) cells. The overexpression of Bcl-rambo, but not Bcl-rambo lacking a C-terminal transmembrane domain, induces apoptosis in S2 cells. Moreover, the ectopic expression of Bcl-rambo by a GAL4-UAS system induces aberrant morphological changes characterized by atrophied wing, split thorax, and rough eye phenotypes. Bcl-rambo induces the activation of effector caspases in eye imaginal discs. The rough eye phenotype induced by Bcl-rambo is partly rescued by the co-expression of p35, Diap1, and Diap2. By using this Drosophila model, it was shown that human Bcl-rambo interacts genetically with Drosophila homologues of adenine nucleotide translocators and the autophagy-related 8 protein. These data demonstrate that human Bcl-rambo localizes to mitochondria and at least regulates an apoptosis signaling pathway in Drosophila.

Dinter, E., Saridaki, T., Nippold, M., Plum, S., Diederichs, L., Komnig, D., Fensky, L., May, C., Marcus, K., Voigt, A., Schulz, J. B. and Falkenburger, B. H. (2016). Rab7 induces clearance of α-synuclein aggregates. J Neurochem [Epub ahead of print]. PubMed ID: 27333324
Parkinson's disease can be caused by mutations in the α-synuclein gene and is characterized by aggregates of α-synuclein protein. Aggregates are degraded by the autophago-lysosomal pathway. Since Rab7 has been shown to regulate trafficking of late endosomes and autophagosomes, it was hypothesized that overexpressing Rab7 might be beneficial in Parkinson's disease. To test this hypothesis this study expressed the pathogenic A53T mutant of α-synuclein in HEK293 cells and Drosophila. In HEK293 cells, EGFP-Rab7 decorated vesicles contain α-synuclein. Rab7 overexpression reduced the percentage of cells with α-synuclein particles and the amount of α-synuclein protein. Clearance of α-synuclein is explained by the increased occurrence of acidified α-synuclein vesicles with Rab7 overexpression, presumably representing autolysosomes. In the fly model, Rab7 rescued the locomotor deficit induced by neuronal expression of A53T-α-synuclein. Rab7 might be involved in the biogenesis of protective, autophagosome-like organelles in dopaminergic neurons. Taken together, Rab7 increased the clearance of α-synuclein aggregates, reduced cell death, and rescued the phenotype in a fly model of Parkinson's disease. These findings indicate that Rab7 is rate-limiting for aggregate clearance, and that Rab7 activation may offer a therapeutic strategy for Parkinson's disease.

Celardo, I., Costa, A. C., Lehmann, S., Jones, C., Wood, N., Mencacci, N. E., Mallucci, G. R., Loh, S. H. and Martins, L. M. (2016). Mitofusin-mediated ER stress triggers neurodegeneration in pink1/parkin models of Parkinson's disease. Cell Death Dis 7: e2271. PubMed ID: 27336715
Mutations in PINK1 and PARKIN cause early-onset Parkinson's disease (PD), thought to be due to mitochondrial toxicity. This study shows that in Drosophila pink1 and parkin mutants, defective mitochondria also give rise to endoplasmic reticulum (ER) stress signalling, specifically to the activation of the protein kinase R-like endoplasmic reticulum kinase (PERK) branch of the unfolded protein response (UPR). Enhanced ER stress signalling in pink1 and parkin mutants is mediated by mitofusin bridges, which occur between defective mitochondria and the ER. Reducing mitofusin contacts with the ER is neuroprotective, through suppression of PERK signalling, while mitochondrial dysfunction remains unchanged. Further, both genetic inhibition of dPerk-dependent ER stress signalling and pharmacological inhibition using the PERK inhibitor GSK2606414 were neuroprotective in both pink1 and parkin mutants. It is concluded that activation of ER stress by defective mitochondria is neurotoxic in pink1 and parkin flies and that the reduction of this signalling is neuroprotective, independently of defective mitochondria.

Tio, M., Wen, R., Lim, Y.L., Wang, H., Ling, S.C., Zhao, Y. and Tan, E.K. (2016). FUS-linked essential tremor associated with motor dysfunction in Drosophila. Hum Genet [Epub ahead of print]. PubMed ID: 27395408
Essential tremor (ET) is one of the most common adult-onset neurological disorders which produce motor and non-motor symptoms. To date, there are no gold standard pathological hallmarks of ET, and despite a strong genetic contribution toward ET development, only a few pathogenic mutations have been identified. Recently, a pathogenic FUS-Q290X mutation has been reported in a large ET-affected family; however, the pathophysiologic mechanism underlying FUS-linked ET is unknown. This study generated transgenic Drosophila expressing hFUS-WT and hFUS-Q290X and targeted their expression in different tissues. It was found that the targeted expression of hFUS-Q290X in the dopaminergic and the serotonergic neurons does not cause obvious neuronal degeneration, but results in motor dysfunction which is accompanied by impairment in the GABAergic pathway. The involvement of the GABAergic pathway is supported by rescue of motor symptoms with gabapentin. Interestingly, gender specific downregulation of GABA-R and NMDA-R expression and reduction in serotonin level were also observed. Overexpression of hFUS-Q290X also causes an increase in longevity and this is accompanied by downregulation of the IIS/TOR signalling pathway. These in vivo studies of the hFUS-Q290X mutation in Drosophila link motor dysfunction to impairment in the GABAergic pathway.

Wednesday, July 20th

Rogers, E.M, Spracklen, A.J., Bilancia, C.G., Sumigray, K.D., Allred, S.C., Nowotarski, S.H., Schaefer, K.N., Ritchie, B.J. and Peifer, M. (2016). Abelson kinase acts as a robust, multifunctional scaffold in regulating embryonic morphogenesis. Mol Biol Cell [Epub ahead of print]. PubMed ID: 27385341
Abelson family kinases (Abl) are key regulators of cell behavior and the cytoskeleton during development and in leukemia. Abl's SH3, SH2, and tyrosine kinase domains are joined via a linker to an F-actin-binding domain (FABD). Research on Abl's roles in cell culture have led to several hypotheses for its mechanism of action: 1) Abl phosphorylates other proteins, modulating their activity. 2) Abl directly regulates the cytoskeleton via its cytoskeletal interaction domains, and/or 3) Abl is a scaffold for a signaling complex. The importance of these roles during normal development remains untested. This study tested these mechanistic hypotheses during Drosophila morphogenesis using a series of mutants to examine Abl's many cell biological roles. Strikingly, Abl lacking the FABD fully rescues morphogenesis, cell shape change, actin regulation, and viability, while kinase dead Abl, though reduced in function, retains substantial rescuing ability in some but not all Abl functions. The function of four conserved motifs in the linker region was also tested, revealing a key role for a conserved PXXP motif known to bind Crk and Abi. The study proposes that Abl acts as a robust multi-domain scaffold with different protein motifs and activities contributing differentially to diverse cellular behaviors.

Caviglia, S., Brankatschk, M., Fischer, E. J., Eaton, S. and Luschnig, S. (2016). Staccato/Unc-13-4 controls secretory lysosome-mediated lumen fusion during epithelial tube anastomosis. Nat Cell Biol 18: 727-739. PubMed ID: 27323327
A crucial yet ill-defined step during the development of tubular networks, such as the vasculature, is the formation of connections (anastomoses) between pre-existing lumenized tubes. By studying tracheal tube anastomosis in Drosophila melanogaster, this study uncovered a key role of secretory lysosome-related organelle (LRO) trafficking in lumen fusion. The conserved calcium-binding protein Unc-13-4/Staccato (Stac) and the GTPase Rab39 were identified as critical regulators of this process. Stac and Rab39 accumulate on dynamic vesicles, which form exclusively in fusion tip cells, move in a dynein-dependent manner, and contain late-endosomal, lysosomal, and SNARE components characteristic of LROs. The GTPase Arl3 is necessary and sufficient for Stac LRO formation and promotes Stac-dependent intracellular fusion of juxtaposed apical plasma membranes, thereby forming a transcellular lumen. Concomitantly, calcium is released locally from ER exit sites and apical membrane-associated calcium increases. It is proposed that calcium-dependent focused activation of LRO exocytosis restricts lumen fusion to appropriate domains within tip cells.

Czerniak, N.D., Dierkes, K., D'Angelo, A., Colombelli, J. and Solon, J. (2016). Patterned contractile forces promote epidermal spreading and regulate segment positioning during Drosophila head involution. Curr Biol [Epub ahead of print]. PubMed ID: 27397891
Epithelial spreading is a fundamental mode of tissue rearrangement occurring during animal development and wound closure. It has been associated either with the collective migration of cells or with actomyosin-generated forces acting at the leading edge (LE) and pulling the epithelial tissue. During the process of Drosophila head involution (HI), the epidermis spreads anteriorly to envelope the head tissues and fully cover the embryo. This results in epidermal segments of equal width that will give rise to the different organs of the fly. This study performed a quantitative analysis of tissue spreading during HI. Combining high-resolution live microscopy with laser microsurgery and genetic perturbations, it was shown that epidermal movement is in part, but not solely, driven by a contractile actomyosin cable at the LE. Additional driving forces are generated within each segment by a gradient of actomyosin-based circumferential tension. Interfering with Hedgehog (Hh) signaling can modulate this gradient, thus suggesting the involvement of polarity genes in the regulation of HI. In particular, it was shown that disruption of these contractile forces alters segment widths and leads to a mispositioning of segments. Within the framework of a physical description, it was confirmed that given the geometry of the embryo, a patterned profile of active circumferential tensions can indeed generate propelling forces and control final segment position. This study thus unravels a mechanism by which patterned tensile forces can regulate spreading and positioning of epithelial tissues.

Xie, J. and Hu, G. H. (2016). Hydrodynamic modeling of Bicoid morphogen gradient formation in Drosophila embryo. Biomech Model Mechanobiol. PubMed ID: 27193268
Bicoid is a maternal polarity determinant that mediates the anterior-posterior (AP) patterning in early Drosophila embryo. During oogenesis, its mRNA deposits at the anterior pole of the embryo and then translates to establish the Bicoid morphogen gradient soon after fertilization. Previous investigations indicated that the patterning is induced by the spatial gradient of Bicoid morphogen concentration, where the cytoplasmic convection plays a crucial role. This study examines the effect of advection transport on the formation of the Bicoid morphogen gradient using direct simulation of the cytoplasmic streaming described by Navier-Stokes equations, in which the cytoplasm behaves as an incompressible Newtonian fluid. To simulate the cytoplasmic streaming originated from membrane contractions, the flow is driven by slip velocities along the cortex and the anterior-posterior axis of the cell. Results show that the Bicoid concentration distribution this analysis obtained provides a quantitatively consistent picture with the experiment measurements, as well as the diffusive length scale. The competition among the diffusion, advection and degradation is analyzed when the cytoplasmic streaming is considered. It is found that the advection yields wavy phenomenon in the profiles of the Bicoid concentration at small diffusion coefficients, which might have important effects on the embryonic development. After the driven velocities is switched off, the interior flow evanesces gradually due to the viscous drag, the Bicoid degradation will overwhelm the advection effect.

Tuesday, July 19th

Zhang, Y., Guo, J., Guo, A. and Li, Y. (2016). Nicotine-induced acute hyperactivity is mediated by dopaminergic system in a sexually dimorphic manner. Neuroscience [Epub ahead of print]. PubMed ID: 27365175
Short-term exposure to nicotine induces positive effects in mice, monkeys and humans, including mild euphoria, hyperactivity, and enhanced cognition.Using a video recording system, this study found that acute nicotine administration induces locomotor hyperactivity in Drosophila. Suppressing dopaminergic neurons or down-regulating dopamine 1-like receptor (DopR) abolishes this acute nicotine response, but surprisingly, does so only in male flies. Using a GFP reconstitution across synaptic partners (GRASP) approach, dopaminergic neurons were shown to possess potential synaptic connections with acetylcholinergic neurons in wide regions of the brain. Furthermore, dopaminergic neurons are widely activated upon nicotine perfusion in both sexes, while the response curve differs significantly between the sexes. Moreover, knockdown of the β1 nicotine acetylcholine receptor (nAChR) in dopaminergic neurons abolishes the acute nicotine response only in male flies, while panneural knock-down occurs in both sexes. Taken together, these results reveal that in fruit flies, dopaminergic neurons mediate nicotine-induced acute locomotor hyperactivity in a sexually dimorphic manner, and Drosophila β1 nAChR subunit plays a crucial role in this nicotine response.

Costa, M., Manton, J. D., Ostrovsky, A. D., Prohaska, S. and Jefferis, G. S. (2016). NBLAST: Rapid, sensitive comparison of neuronal structure and construction of neuron family databases. Neuron [Epub ahead of print]. PubMed ID: 27373836
Neural circuit mapping is generating datasets of tens of thousands of labeled neurons. New computational tools are needed to search and organize these data. This paper presents NBLAST, a sensitive and rapid algorithm, for measuring pairwise neuronal similarity. NBLAST considers both position and local geometry, decomposing neurons into short segments; matched segments are scored using a probabilistic scoring matrix defined by statistics of matches and non-matches. NBLAST was validated on a published dataset of 16,129 single Drosophila neurons. NBLAST can distinguish neuronal types down to the finest level (single identified neurons) without a priori information. Cluster analysis of extensively studied neuronal classes identified new types and unreported topographical features. Fully automated clustering organized the validation dataset into 1,052 clusters, many of which map onto previously described neuronal types. NBLAST supports additional query types, including searching neurons against transgene expression patterns. Finally, NBLAST was shown to be effective with data from other invertebrates and zebrafish.

Bell, J. S. and Wilson, R. I. (2016). Behavior reveals selective summation and max pooling among olfactory processing channels. Neuron [Epub ahead of print]. PubMed ID: 27373835
The olfactory system is divided into processing channels (glomeruli), each receiving input from a different type of olfactory receptor neuron (ORN). This study investigated how glomeruli combine to control behavior in freely walking Drosophila. Optogenetically activating single ORN types were shown to typically produced attraction, although some ORN types produced repulsion. Attraction consisted largely of a behavioral program with the following rules: at fictive odor onset, flies walked upwind, and at fictive odor offset, they reversed. When certain pairs of attractive ORN types were co-activated, the level of the behavioral response resembled the sum of the component responses. However, other pairs of attractive ORN types produced a response resembling the larger component (max pooling). Although activation of different ORN combinations produced different levels of behavior, the rules of the behavioral program were consistent. The results illustrate a general method for inferring how groups of neurons work together to modulate behavioral programs.
Friederich, U., Billings, S. A., Hardie, R. C., Juusola, M. and Coca, D. (2016). Fly photoreceptors encode phase congruency. PLoS One 11: e0157993. PubMed ID: 27336733
More than five decades ago it was postulated that sensory neurons detect and selectively enhance behaviourally relevant features of natural signals. Although it is now known that sensory neurons are tuned to efficiently encode natural stimuli, until now it was not clear what statistical features of the stimuli they encode and how. This study reverse-engineered the neural code of Drosophila photoreceptors and shows for the first time that photoreceptors exploit nonlinear dynamics to selectively enhance and encode phase-related features of temporal stimuli, such as local phase congruency, which are invariant to changes in illumination and contrast. To mitigate for the inherent sensitivity to noise of the local phase congruency measure, the nonlinear coding mechanisms of the fly photoreceptors are tuned to suppress random phase signals, which explains why photoreceptor responses to naturalistic stimuli are significantly different from their responses to white noise stimuli.

Monday, July 18th

Chiang, Y. N., Tan, K. J., Chung, H., Lavrynenko, O., Shevchenko, A. and Yew, J. Y. (2016). Steroid hormone signaling is essential for pheromone production and oenocyte survival. PLoS Genet 12: e1006126. PubMed ID: 27333054
Many of the lipids found on the cuticles of insects function as pheromones and communicate information about age, sex, and reproductive status. To identify genes that control cuticular lipid production in Drosophila, a RNA interference screen was performed and Direct Analysis in Real Time and gas chromatography mass spectrometry were performed to quantify changes in the chemical profiles. Twelve putative genes were identified whereby transcriptional silencing led to significant differences in cuticular lipid production. Amongst them, a gene was identified that was named spidey (CG1444), which encodes a putative steroid dehydrogenase that has sex- and age-dependent effects on viability, pheromone production, and oenocyte survival. Transcriptional silencing or overexpression of spidey during embryonic development results in pupal lethality and significant changes in levels of the ecdysone metabolite 20-hydroxyecdysonic acid and 20-hydroxyecdysone. In contrast, inhibiting gene expression only during adulthood resulted in a striking loss of oenocyte cells and a concomitant reduction of cuticular hydrocarbons, desiccation resistance, and lifespan. Oenocyte loss and cuticular lipid levels were partially rescued by 20-hydroxyecdysone supplementation. Taken together, these results identify a novel regulator of pheromone synthesis and reveal that ecdysteroid signaling is essential for the maintenance of cuticular lipids and oenocytes throughout adulthood.

Zhang, T., Sheng, Z. and Du, W. (2016). Loss of histone deacetylase HDAC1 induces cell death in Drosophila epithelial cells through JNK and Hippo signaling. Mech Dev [Epub ahead of print]. PubMed ID: 27378074
Inactivation of HDAC1 and its homolog HDAC2 or addition of HDAC inhibitors in mammalian systems induces apoptosis, cell cycle arrest, and developmental defects. Although these phenotypes have been extensively characterized, the precise underlying mechanisms remain unclear, particularly in in vivo settings. This study shows that inactivation of Rpd3, the only HDAC1 and HDAC2 ortholog in Drosophila, induced apoptosis and clone elimination in the developing eye and wing imaginal discs. Depletion of Rpd3 by RNAi cell-autonomously increased JNK activities and decreased activities of Yki, the nuclear effecter of Hippo signaling pathway. In addition, inhibition of JNK activities largely rescued Rpd3 RNAi-induced apoptosis, but did not affect its inhibition of Yki activities. Conversely, increasing the Yki activities largely rescued Rpd3 RNAi-induced apoptosis, but did not affect its induction of JNK activities. Furthermore, inactivation of Mi-2, a core component of the Rpd3-containing NuRD complex strongly induced JNK activities; while inactivation of Sin3A, a key component of the Rpd3-containing Sin3 complex, significantly inhibited Yki activities. Taken together, these results reveal that inactivation of Rpd3 independently regulates JNK and Yki activities and that both Hippo and JNK signaling pathways contribute to Rpd3 RNAi-induced apoptosis.

Ou, Q., Zeng, J., Yamanaka, N., Brakken-Thal, C., O'Connor, M. B. and King-Jones, K. (2016). The insect prothoracic gland as a model for steroid hormone biosynthesis and regulation. Cell Rep 16: 247-262. PubMed ID: 27320926
Steroid hormones are ancient signaling molecules found in vertebrates and insects alike. Both taxa show intriguing parallels with respect to how steroids function and how their synthesis is regulated. As such, insects are excellent models for studying universal aspects of steroid physiology. This study presents a comprehensive genomic and genetic analysis of the principal steroid hormone-producing organs in two popular insect models, Drosophila and Bombyx. 173 genes were identified with previously unknown specific expression in steroid-producing cells, 15 of which had critical roles in development. The insect neuropeptide PTTH and its vertebrate counterpart ACTH both regulate steroid production, but molecular targets of these pathways remain poorly characterized. Identification of PTTH-dependent gene sets identified the nuclear receptor HR4 as a highly conserved target in both Drosophila and Bombyx. This study is a critical step toward understanding how steroid hormone production and release are regulated in all animal models (Ou, 2016).

Fu, C. L., Wang, X. F., Cheng, Q., Wang, D., Hirose, S. and Liu, Q. X. (2016). The T-box transcription factor Midline regulates wing development by repressing wingless and hedgehog in Drosophila. Sci Rep 6: 27981. PubMed ID: 27301278
Wingless (Wg) and Hedgehog (Hh) signaling pathways are key players in animal development. However, regulation of the expression of wg and hh are not well understood. This study shows that Midline (Mid), an evolutionarily conserved transcription factor, expresses in the wing disc of Drosophila and plays a vital role in wing development. Loss or knock down of mid in the wing disc induced hyper-expression of wingless (wg) and yielded cocked and non-flat wings. Over-expression of mid in the wing disc markedly repressed the expression of wg, DE-Cadherin (DE-Cad) and armadillo (arm), and resulted in a small and blistered wing. In addition, a reduction in the dose of mid enhanced phenotypes of a gain-of-function mutant of hedgehog (hh). Repression of hh was observed upon overexpression of mid in the wing disc. Taken together, it is proposed that Mid regulates wing development by repressing wg and hh in Drosophila.

Sunday, July 17th

Beatus, T. and Cohen, I. (2015). Wing-pitch modulation in maneuvering fruit flies is explained by an interplay between aerodynamics and a torsional spring. Phys Rev E Stat Nonlin Soft Matter Phys 92: 022712. PubMed ID: 26382437
While the wing kinematics of many flapping insects have been well characterized, understanding the underlying sensory, neural, and physiological mechanisms that determine these kinematics is still a challenge. Two main difficulties in understanding the physiological mechanisms arise from the complexity of the interaction between a flapping wing and its own unsteady flow, as well as the intricate mechanics of the insect wing hinge, which is among the most complicated joints in the animal kingdom. These difficulties call for the application of reduced-order approaches. This strategy was used to model the torques exerted by the wing hinge along the wing-pitch axis of maneuvering fruit flies as a damped torsional spring with elastic and damping coefficients as well as a rest angle. Furthermore, the air flows were modeled using simplified quasistatic aerodynamics. The findings suggest that flies take advantage of the passive coupling between aerodynamics and the damped torsional spring to indirectly control their wing-pitch kinematics by modulating the spring parameters. The damped torsional-spring model explains the changes measured in wing-pitch kinematics during roll correction maneuvers through modulation of the spring damping and elastic coefficients. These results, in conjunction with the previous literature, indicate that flies can accurately control their wing-pitch kinematics on a sub-wing-beat time scale by modulating all three effective spring parameters on longer time scales.

Honjo, K., Mauthner, S. E., Wang, Y., Skene, J. H. and Tracey, W. D. (2016). Nociceptor-enriched genes required for normal thermal nociception. Cell Rep [Epub ahead of print]. PubMed ID: 27346357
This study describe a targeted reverse genetic screen for thermal nociception genes in Drosophila larvae. Using laser capture microdissection and microarray analyses of nociceptive and non-nociceptive neurons, 275 nociceptor-enriched genes were identified. The function of the enriched genes were tested with nociceptor-specific RNAi and thermal nociception assays. Tissue-specific RNAi targeted against 14 genes caused insensitive thermal nociception while targeting of 22 genes caused hypersensitive thermal nociception. Previously uncategorized genes were named for heat resistance (i.e., boilerman (CG12681), fire dancer (CG8968), oven mitt (CG31976), trivet (CG12870), thawb (CG14608), and bunker gear (CG8297)) or heat sensitivity (firelighter (CG14946), black match (CG12524), eucalyptus (CG12269), primacord (CG15704), jet fuel (CG12858), detonator (CG14446), gasoline (CG6018), smoke alarm (CG13988), and jetboil (CG4398)). Insensitive nociception phenotypes were often associated with severely reduced branching of nociceptor neurites and hyperbranched dendrites were seen in two of the hypersensitive cases. Many genes that were identified are conserved in mammals.

Arena, P., Cali, M., Patane, L., Portera, A. and Strauss, R. (2016). A fly-inspired mushroom bodies model for sensory-motor control through sequence and subsequence learning. Int J Neural Syst: 1650035. PubMed ID: 27354193
Classification and sequence learning are relevant capabilities used by living beings to extract complex information from the environment for behavioral control. The insect world is full of examples where the presentation time of specific stimuli shapes the behavioral response. On the basis of previously developed neural models, inspired by Drosophila melanogaster, a new architecture for classification and sequence learning is here presented under the perspective of the Neural Reuse theory. Classification of relevant input stimuli is performed through resonant neurons, activated by the complex dynamics generated in a lattice of recurrent spiking neurons modeling the insect Mushroom Bodies neuropile. The network devoted to context formation is able to reconstruct the learned sequence and also to trace the subsequences present in the provided input. A sensitivity analysis to parameter variation and noise is reported. Experiments on a roving robot are reported to show the capabilities of the architecture used as a neural controller.

Badel, L., Ohta, K., Tsuchimoto, Y. and Kazama, H. (2016). Decoding of context-dependent olfactory behavior in Drosophila. Neuron [Epub ahead of print]. PubMed ID: 27321924
Odor information is encoded in the activity of a population of glomeruli in the primary olfactory center. However, how this information is decoded in the brain remains elusive. This question was addressed in Drosophila by combining neuronal imaging and tracking of innate behavioral responses. The behavior is accurately predicted by a model summing normalized glomerular responses, in which each glomerulus contributes a specific, small amount to odor preference. This model is further supported by targeted manipulations of glomerular input, which biased the behavior. Additionally, it was observed that relative odor preference changes and can even switch depending on the context, an effect correctly predicted by the normalization model. These results indicate that olfactory information is decoded from the pooled activity of a glomerular repertoire and demonstrate the ability of the olfactory system to adapt to the statistics of its environment.

Saturday, July 16th

Deignan, L., Pinheiro, M.T., Sutcliffe, C., Saunders, A., Wilcockson, S.G., Zeef, L.A., Donaldson, I.J. and Ashe, H.L. (2016). Regulation of the BMP signaling-responsive transcriptional network in the Drosophila embryo. PLoS Genet 12: e1006164. PubMed ID: 27379389
The BMP signaling pathway has a conserved role in dorsal-ventral axis patterning during embryonic development. In Drosophila, graded BMP signaling is transduced by the Mad transcription factor and opposed by the Brinker repressor. Using the Drosophila embryo as a model, this study combined RNA-seq with Mad and Brinker ChIP-seq to decipher the BMP-responsive transcriptional network underpinning differentiation of the dorsal ectoderm during dorsal-ventral axis patterning. Multiple new BMP target genes were identified, including positive and negative regulators of EGF signaling. Manipulation of EGF signaling levels by loss- and gain-of-function studies reveals that EGF signaling negatively regulates embryonic BMP-responsive transcription. Therefore, the BMP gene network has a self-regulating property in that it establishes a balance between its activity and that of the antagonistic EGF signaling pathway to facilitate correct patterning. In terms of BMP-dependent transcription, key roles for the Zelda and Zerknüllt transcription factors in establishing the resulting expression domain were identified, and widespread binding of insulator proteins to the Mad and Brinker-bound genomic regions was observed. Analysis of embryos lacking the BEAF-32 insulator protein shows reduced transcription of a peak BMP target gene and a reduction in the number of amnioserosa cells, the fate specified by peak BMP signaling. 

Spong, K. E., Rodriguez, E. C. and Robertson, R. M. (2016). Spreading depolarization in the brain of Drosophila is induced by inhibition of the Na+/K+-ATPase and mitigated by a decrease in activity of protein kinase G. J Neurophysiol [Epub ahead of print] PubMed ID: 27358319
Spreading depolarization (SD) is characterized by a massive redistribution of ions accompanied by an arrest in electrical activity that slowly propagates through neural tissue. This study demonstrates the occurrence of SD in the brain of Drosophila providing a model system whereby cellular mechanisms can be dissected using molecular genetic approaches. Propagating waves of SD were reliably induced by disrupting the extracellular potassium concentration ([K+]o) either directly or by inhibition of the Na+/K+-ATPase with ouabain. The disturbance was monitored by recording the characteristic surges in [K+]o using K+-sensitive microelectrodes or by monitoring brain activity by measuring direct current (DC) potential. Using wild-type flies this study showed that young adults are more resistant to SD compared to older adults, evidenced by shorter bouts of SD activity and attenuated [K+]o disturbances. Furthermore, the susceptibility to SD was shown to differ between wild-type flies and w1118 mutants demonstrating that the ouabain-model is influenced by genetic strain. Lastly, flies with low levels of protein kinase G (PKG) had increased latencies to onset of both ouabain-induced SD and anoxic depolarization compared to flies with higher levels. These findings implicate the PKG pathway as a modulator of SD in the fly brain and given the conserved nature of the signalling pathway it could likely play a similar role during SD in the mammalian CNS.

Novak, Z. A., Wainman, A., Gartenmann, L. and Raff, J. W. (2016). Cdk1 phosphorylates Drosophila Sas-4 to recruit Polo to daughter centrioles and convert them to centrosomes. Dev Cell 37: 545-557. PubMed ID: 27326932
Centrosomes and cilia are organized by a centriole pair comprising an older mother and a younger daughter. Centriole numbers are tightly regulated, and daughter centrioles (which assemble in S phase) cannot themselves duplicate or organize centrosomes until they have passed through mitosis. It is unclear how this mitotic 'centriole conversion' is regulated, but it requires Plk1/Polo kinase. This study shows that in flies, Cdk1 phosphorylates the conserved centriole protein Sas-4 during mitosis. This creates a Polo-docking site that helps recruit Polo to daughter centrioles and is required for the subsequent recruitment of Asterless (Asl), a protein essential for centriole duplication and mitotic centrosome assembly. Point mutations in Sas-4 that prevent Cdk1 phosphorylation or Polo docking do not block centriole disengagement during mitosis, but block efficient centriole conversion and lead to embryonic lethality. These observations can explain why daughter centrioles have to pass through mitosis before they can duplicate and organize a centrosome.

Kelly, L. K., Wu, J., Yanfeng, W. A. and Mlodzik, M. (2016). Frizzled-induced Van Gogh phosphorylation by CK1epsilon promotes asymmetric localization of core PCP factors in Drosophila. Cell Rep. PubMed ID: 27346358
Epithelial tissues are polarized along two axes. In addition to apical-basal polarity, they are often polarized within the plane of the epithelium, so-called Planar Cell Polarity (PCP). PCP depends upon Wnt/Frizzled (Fz) signaling factors, including Fz itself and Van Gogh (Vang/Vangl). This study sought to understand how Vang interaction with other core PCP factors affects Vang function. Fz was found to induce Vang phosphorylation in a cell-autonomous manner. Vang phosphorylation occurs on conserved N-terminal serine/threonine residues, is mediated by CK1epsilon/Dco, and is critical for polarized membrane localization of Vang and other PCP proteins. This regulatory mechanism does not require Fz signaling through Dishevelled and thus represents a cell-autonomous upstream interaction between Fz and Vang. Furthermore, this signaling event appears to be related to Wnt5a-mediated Vangl2 phosphorylation during mouse limb patterning and may thus be a general mechanism underlying Wnt-regulated PCP establishment.

Friday, July 15th

Mottier-Pavie, V.I., Palacios, V., Eliazer, S., Scoggin, S. and Buszczak, M. (2016). The Wnt pathway limits BMP signaling outside of the germline stem cell niche in Drosophila ovaries. Dev Biol [Epub ahead of print]. PubMed ID: 27364467
The mechanisms that modulate and limit the signaling output of adult stem cell niches remain poorly understood. To gain further insights into how these microenvironments are regulated in vivo, this study performed a candidate gene screen designed to identify factors that restrict BMP signal production to the cap cells that comprise the germline stem cell (GSC) niche of Drosophila ovaries. It was found that disruption of Wnt4 and components of the canonical Wnt pathway results in a complex germ cell phenotype marked by an expansion of GSC-like cells, pre-cystoblasts and cystoblasts in young females. This phenotype correlates with an increase of decapentaplegic (dpp) mRNA levels within escort cells and varying levels of BMP responsiveness in the germline. Further genetic experiments showed that Wnt4, which exhibits graded expression in somatic cells of germaria, activates the Wnt pathway in posteriorly positioned escort cells. The activation of the Wnt pathway appears to be limited by the BMP pathway itself, as loss of Mad in escort cells results in the expansion of Wnt pathway activation. Wnt pathway activity changes within germaria during the course of aging, coincident with changes in dpp production. These data suggest that mutual antagonism between the BMP and Wnt pathways in somatic cells helps to regulate germ cell differentiation.

Yu, J., Liu, Y., Lan, X., Wu, H., Wen, Y., Zhou, Z., Hu, Z., Sha, J., Guo, X. and Tong, C. (2016). CHES-1-like, the ortholog of a non-obstructive azoospermia-associated gene, blocks germline stem cell differentiation by upregulating Dpp expression in Drosophila testis. Oncotarget [Epub ahead of print]. PubMed ID: 27281616
Azoospermia is a high risk factor for testicular germ cell tumors, whose underlying molecular mechanisms remain unknown. In a genome-wide association study to identify novel loci associated with human non-obstructive azoospermia (NOA), a single nucleotide polymorphism was uncovered in a human gene FOXN3. FOXN3 is an evolutionarily conserved gene. This study used Drosophila melanogaster as a model system to test whether CHES-1-like, the Drosophila FOXN3 ortholog, is required for male fertility. CHES-1-like knockout flies are viable and fertile, and show no defects in spermatogenesis. However, ectopic expression of CHES-1-like in germ cells significantly reduced male fertility. With CHES-1-like overexpression, spermatogonia fail to differentiate after four rounds of mitotic division, but continue to divide to form tumor like structures. In these testes, expression levels of differentiation factor, Bam, were reduced, but the expression region of Bam was expanded. Further reduced Bam expression in CHES-1-like expressing testes exhibited enhanced tumor-like structure formation. The expression of daughters against dpp (dad), a downstream gene of dpp signaling, was upregulated by CHES-1-like expression in testes. CHES-1-like directly binds to the dpp promoter. A model is proposed that CHES-1-like overexpression in germ cells activates dpp expression, inhibits spermatocyte differentiation, and finally leads to germ cell tumors.

Lee, H., Choi, H.W., Zhang, C., Park, Z.Y. and Kim, Y.J. (2016). A pair of oviduct-born Pickpocket neurons important for egg-laying in Drosophila melanogaster. Mol Cells [Epub ahead of print]. PubMed ID: 27378227
During copulation, male Drosophila transfers Sex Peptide (SP) to females where it acts on internal sensory neurons expressing pickpocket (ppk). These neurons induce a post-mating response (PMR) that includes elevated egglaying and refractoriness to re-mating. Exactly how ppk neurons regulate the different aspects of the PMR, however, remains unclear. This study identifies a small subset of the ppk neurons which require expression of a pre-mRNA splicing factor CG3542 for egg-laying, but not refractoriness to mating. Two CG3542-ppk expressing neurons were identified that innervate the upper oviduct and appear to be responsible for normal egg-laying. These results suggest specific subsets of the ppk neurons are responsible for each PMR component.

Williams, P. A., Krug, M. S., McMillan, E. A., Peake, J. D., Davis, T. L., Cocklin, S. and Strochlic, T. I. (2016). Phosphorylation of the RNA-binding protein Dazl by MAPKAP kinase 2 regulates spermatogenesis. Mol Biol Cell [Epub ahead of print]. PubMed ID: 27280388
The germ cell-specific RNA-binding protein Dazl (deleted in azoospermia-like, mammalian ortholog of Drosophila Boule) is shown to be phosphorylated by MK2 (MAPKAP kinase 2), a stress-induced protein kinase activated downstream of p38 MAPK. Phosphorylation of human Dazl by MK2 on an evolutionarily conserved serine residue inhibits its interaction with poly(A)-binding protein (PABP), resulting in reduced translation of Dazl-regulated target RNAs. Transgenic expression of wild-type human Dazl but not a phosphomimetic form in the Drosophila male germline can restore fertility to flies deficient in boule. These results suggest that signaling by the p38-MK2 pathway is a negative regulator of spermatogenesis via phosphorylation of Dazl.

Thursday, June 14th

Petit, M., Mongelli, V., Frangeul, L., Blanc, H., Jiggins, F. and Saleh, M.C. (2016). piRNA pathway is not required for antiviral defense in Drosophila melanogaster. Proc Natl Acad Sci U S A [Epub ahead of print]. PubMed ID: 27357659
Since its discovery, RNA interference has been identified as involved in many different cellular processes, and as a natural antiviral response in plants, nematodes, and insects. In insects, the small interfering RNA (siRNA) pathway is the major antiviral response. In recent years, the Piwi-interacting RNA (piRNA) pathway also has been implicated in antiviral defense in mosquitoes infected with arboviruses. Using Drosophila melanogaster and an array of viruses that infect the fruit fly acutely or persistently or are vertically transmitted through the germ line, this study investigated in detail the extent to which the piRNA pathway contributes to antiviral defense in adult flies. Following virus infection, the survival and viral titers of Piwi, Aubergine, Argonaute-3, and Zucchini mutant flies were similar to those of wild type flies. Using next-generation sequencing of small RNAs from wild type and siRNA mutant flies, it was shown that no viral-derived piRNAs are produced in fruit flies during different types of viral infection. This study provides the first evidence that the piRNA pathway does not play a major role in antiviral defense in adult Drosophila and demonstrates that viral-derived piRNA production depends on the biology of the host-virus combination rather than being part of a general antiviral process in insects.

Schmidts, I., Böttcher, R., Mirkovic-Hösle, M. and Förstemann, K. (2016). Homology directed repair is unaffected by the absence of siRNAs in Drosophila melanogaster. Nucleic Acids Res [Epub ahead of print]. PubMed ID: 27353331
Small interfering RNAs (siRNAs) defend the organism against harmful transcripts from exogenous (e.g. viral) or endogenous (e.g. transposons) sources. Recent publications describe the production of siRNAs induced by DNA double-strand breaks (DSB) in Neurospora crassa, Arabidopsis thaliana, Drosophila melanogaster and human cells, which suggests a conserved function. A current hypothesis is that break-induced small RNAs ensure efficient homologous recombination (HR). However, biogenesis of siRNAs is often intertwined with other small RNA species, such as microRNAs (miRNAs), which complicates interpretation of experimental results. In Drosophila, siRNAs are produced by Dcr-2 while miRNAs are processed by Dcr-1. Thus, it is possible to probe siRNA function without miRNA deregulation. This study examined DNA double-strand break repair after perturbation of siRNA biogenesis in cultured Drosophila cells as well as mutant flies. The assays comprised reporters for the single-strand annealing pathway, homologous recombination and sensitivity to the DSB-inducing drug camptothecin. Repair defects caused by the lack of siRNAs derived from the broken DNA locus were not detected. Since production of these siRNAs depends on local transcription, they may thus participate in RNA metabolism- an established function of siRNAs- rather than DNA repair.

Sysoev, V.O., Fischer, B., Frese, C.K., Gupta, I., Krijgsveld, J., Hentze, M.W., Castello, A. and Ephrussi, A. (2016). Global changes of the RNA-bound proteome during the maternal-to-zygotic transition in Drosophila. Nat Commun 7: 12128. PubMed ID: 27378189
The maternal-to-zygotic transition (MZT) is a process that occurs in animal embryos at the earliest developmental stages, during which maternally deposited mRNAs and other molecules are degraded and replaced by products of the zygotic genome. The zygotic genome is not activated immediately upon fertilization, and in the pre-MZT embryo post-transcriptional control by RNA-binding proteins (RBPs) orchestrates the first steps of development. To identify relevant Drosophila RBPs organism-wide, this study refined the RNA interactome capture method for comparative analysis of the pre- and post-MZT embryos. 523 proteins were determined to be high-confidence RBPs, half of which have not been previously reported to bind RNA. Comparison of the RNA interactomes of pre- and post-MZT embryos reveals high dynamicity of the RNA-bound proteome during early development, and suggests active regulation of RNA binding of some RBPs. This resource provides unprecedented insight into the system of RBPs that govern the earliest steps of Drosophila development.

Lazzaretti, D., Veith, K., Kramer, K., Basquin, C., Urlaub, H., Irion, U. and Bono, F. (2016). The bicoid mRNA localization factor Exuperantia is an RNA-binding pseudonuclease. Nat Struct Mol Biol [Epub ahead of print]. PubMed ID: 27376588
Anterior patterning in Drosophila is mediated by the localization of bicoid (bcd) mRNA at the anterior pole of the oocyte. Exuperantia (Exu) is a putative exonuclease (EXO) associated with bcd and required for its localization. This study presents the crystal structure of Exu, which reveals a dimeric assembly with each monomer consisting of a 3'-5' EXO-like domain and a sterile alpha motif (SAM)-like domain. The catalytic site is degenerate and inactive. Instead, the EXO-like domain mediates dimerization and RNA binding. It was shown that Exu binds RNA directly in vitro, that the SAM-like domain is required for RNA binding activity and that Exu binds a structured element present in the bcd 3' untranslated region with high affinity. Through structure-guided mutagenesis, it was shown that Exu dimerization is essential for bcd localization. These data demonstrate that Exu is a noncanonical RNA-binding protein with EXO-SAM-like domain architecture that interacts with its target RNA as a homodimer.

Wednesday, July 13th

Danielsen, E. T., Moeller, M. E., Yamanaka, N., Ou, Q., Laursen, J. M., Soenderholm, C., Zhuo, R., Phelps, B., Tang, K., Zeng, J., Kondo, S., Nielsen, C. H., Harvald, E. B., Faergeman, N. J., Haley, M. J., O'Connor, K. A., King-Jones, K., O'Connor, M. B. and Rewitz, K. F. (2016). A Drosophila genome-wide screen identifies regulators of steroid hormone production and developmental timing. Dev Cell 37: 558-570. PubMed ID: 27326933
Steroid hormones control important developmental processes and are linked to many diseases. To systematically identify genes and pathways required for steroid production,a Drosophila genome-wide in vivo RNAi screen was performed, and 1,906 genes were identified with potential roles in steroidogenesis and developmental timing. This screen was used as a resource to identify mechanisms regulating intracellular levels of cholesterol, a substrate for steroidogenesis. A conserved fatty acid elongase was identified that underlies a mechanism that adjusts cholesterol trafficking and steroidogenesis with nutrition and developmental programs. In addition, the existence of an autophagosomal cholesterol mobilization mechanism was identified and it was shown that activation of this system rescues Niemann-Pick type C1 deficiency that causes a disorder characterized by cholesterol accumulation. These cholesterol-trafficking mechanisms are regulated by TOR and feedback signaling that couples steroidogenesis with growth and ensures proper maturation timing. These results reveal genes regulating steroidogenesis during development that likely modulate disease mechanisms.

Feingold, D., Starc, T., O'Donnell, M. J., Nilson, L. and Dent, J. A. (2016). The orphan pentameric ligand-gated ion channel pHCl-2 is gated by pH and regulates fluid secretion in Drosophila Malpighian tubules. J Exp Biol [Epub ahead of print]. PubMed ID: 27358471
Pentameric ligand-gated ion channels (pLGICs) constitute a large protein superfamily in metazoa whose role as neurotransmitter receptors mediating rapid, ionotropic synaptic transmission has been extensively studied. Although the vast majority of pLGICs appear to be neurotransmitter receptors, the identification of pLGICs in non-neuronal tissues and homologous pLGIC-like proteins in prokaryotes points to biological functions, possibly ancestral, that are independent of neuronal signaling. This study reports the molecular and physiological characterization of a highly divergent, orphan pLGIC subunit, pHCl-2 (CG11340), in Drosophila melanogaster. pHCl-2 forms a channel that is insensitive to a wide array of neurotransmitters, but is instead gated by changes in extracellular pH. pHCl-2 is expressed in the Malpighian tubules, which are non-innervated renal-type secretory tissues. pHCl-2 is localized to the apical membrane of the epithelial principal cells of the tubules, and loss of pHCl-2 reduces urine production during diuresis. These data implicate pHCl-2 as an important source of chloride conductance required for proper urine production, highlighting a novel role for pLGICs in epithelial tissues regulating fluid secretion and osmotic homeostasis.

MacMillan, H.A., Knee, J.M., Dennis, A.B., Udaka, H., Marshall, K.E., Merritt, T.J. and Sinclair, B.J. (2016). Cold acclimation wholly reorganizes the Drosophila melanogaster transcriptome and metabolome. Sci Rep 6: 28999. PubMed ID: 27357258
Cold tolerance is a key determinant of insect distribution and abundance, and thermal acclimation can strongly influence organismal stress tolerance phenotypes, particularly in small ectotherms like Drosophila. However, there is limited understanding of the molecular and biochemical mechanisms that confer such impressive plasticity. This study used high-throughput mRNA sequencing (RNA-seq) and liquid chromatography - mass spectrometry (LC-MS) to compare the transcriptomes and metabolomes of D. melanogaster acclimated as adults to warm (rearing) (21.5°C) or cold conditions (6°C). Cold acclimation improves cold tolerance and leads to extensive biological reorganization: almost one third of the transcriptome and nearly half of the metabolome are differentially regulated. There is overlap in the metabolic pathways identified via transcriptomics and metabolomics, with proline and glutathione metabolism being the most strongly-supported metabolic pathways associated with increased cold tolerance. The study discusses several new targets in the study of insect cold tolerance (e.g. dopamine signaling and Na(+)-driven transport), but many previously identified candidate genes and pathways (e.g. heat shock proteins, Ca(2+) signaling, and ROS detoxification) are also identified in the present study, and these data are thus consistent with and extend the current understanding of the mechanisms of insect chilling tolerance.

Marcus, S. R. and Fiumera, A. C. (2016). Atrazine exposure affects longevity, development time and body size in Drosophila melanogaster. J Insect Physiol 91-92: 18-25. PubMed ID: 27317622
Atrazine is the one of the most widely used herbicides in the United States and non-target organisms may encounter it in the environment. Atrazine is known to affect male reproduction in both vertebrates and invertebrates but less is known about its effects on other fitness traits. This study assessed the effects of five different chronic exposure levels on a variety of fitness traits in Drosophila melanogaster. Male and female longevity, development time, proportion pupated, proportion emerged, body size, female mating rate, fertility and fecundity were measured. Atrazine exposure decreased the proportion pupated, the proportion emerged and adult survival. Development time was also affected by atrazine and exposed flies pupated and emerged earlier than controls. Although development time was accelerated, body size was actually larger in some of the exposures. Atrazine exposure had no effect on female mating rate and the effects on female fertility and fecundity were only observed in one of the two independent experimental blocks. Many of the traits showed non-monotonic dose response curves, where the intermediate concentrations showed the largest effects. Overall this study shows that atrazine influences a variety of life history traits in the model genetic system, D. melanogaster, and future studies should aim to identify the molecular mechanisms of toxicity.

Tuesday, July 12th

Meehan, T.L., Joudi, T.F., Timmons, A.K., Taylor, J.D., Habib, C.S., Peterson, J.S., Emmanuel, S., Franc, N.C. and McCall, K. (2016). Components of the engulfment machinery have distinct roles in corpse processing. PLoS One 11: e0158217. PubMed ID: 27347682
This study used the Drosophila ovarian follicle cells as a model for engulfment of apoptotic cells by epithelial cells. Engulfed material was shown to be processed using the canonical corpse processing pathway involving the small GTPases Rab5 and Rab7. The phagocytic receptor Draper is present on the phagocytic cup and on nascent, phosphatidylinositol 3-phosphate (PI(3)P)- and Rab7-positive phagosomes, whereas integrins are maintained on the cell surface during engulfment. Due to the difference in subcellular localization, the roles of Draper, integrins, and downstream signaling components in corpse processing were also investigated. It was found that some proteins are required for internalization only, while others have defects in corpse processing as well. This suggests that several of the core engulfment proteins are required for distinct steps of engulfment. By performing double mutant analysis, it was found that combined loss of draper and αPS3 still results in a small number of engulfed vesicles. Next, another known engulfment receptor, Crq, was investigated. It was found that loss of all three receptors does not inhibit engulfment any further, suggesting that Crq does not play a role in engulfment by the follicle cells. A more complete understanding of how the engulfment and corpse processing machinery interact may enable better understanding and treatment of diseases associated with defects in engulfment by epithelial cells. 

Bhogal, B., Plaza-Jennings, A. and Gavis, E. R. (2016). Nanos-mediated repression of hid protects larval sensory neurons after a global switch in sensitivity to apoptotic signals. Development 143: 2147-2159. PubMed ID: 27256879
Dendritic arbor morphology is a key determinant of neuronal function. Once established, dendrite branching patterns must be maintained as the animal develops to ensure receptive field coverage. The translational repressors Nanos (Nos) and Pumilio (Pum) are required to maintain dendrite growth and branching of Drosophila larval class IV dendritic arborization (da) neurons, but their specific regulatory role remains unknown. This study shows that Nos-Pum-mediated repression of the pro-apoptotic gene head involution defective (hid) is required to maintain a balance of dendritic growth and retraction in class IV da neurons and that upregulation of hid results in decreased branching because of an increase in caspase activity. The temporal requirement for nos correlates with an ecdysone-triggered switch in sensitivity to apoptotic stimuli that occurs during the mid-L3 transition. hid is required during pupariation for caspase-dependent pruning of class IV da neurons, and Nos and Pum delay pruning. Together, these results suggest that Nos and Pum provide a crucial neuroprotective regulatory layer to ensure that neurons behave appropriately in response to developmental cues.

Sarkissian, T., Arya, R., Gyonjyan, S., Taylor, B. and White, K. (2016). Cell death regulates muscle fiber number. Dev Biol [Epub ahead of print]. PubMed ID: 27131625
Cell death can have both cell autonomous and non-autonomous roles in normal development. Previous studies have shown that the central cell death regulators grim and reaper are required for the developmentally important elimination of stem cells and neurons in the developing central nervous system (CNS). This study shows that cell death in the nervous system is also required for normal muscle development. In the absence of grim and reaper, there is an increase in the number of fibers in the ventral abdominal muscles in the Drosophila adult. This phenotype can be partially recapitulated by inhibition of cell death specifically in the CNS, indicating a non-autonomous role for neuronal death in limiting muscle fiber number. It was also shown that FGFs produced in the cell death defective nervous system are required for the increase in muscle fiber number. Cell death in the muscle lineage during pupal stages also plays a role in specifying fiber number. Altogetger, data suggests that FGFs from the CNS act as a survival signal for muscle FCs. Thus, proper muscle fiber specification requires cell death in both the nervous system and in the developing muscle itself.

Hou, L., Liu, K., Li, Y., Ma, S., Ji, X. and Liu, L. (2016). Necrotic pyknosis is a morphologically and biochemically distinct event from apoptotic pyknosis. J Cell Sci [Epub ahead of print]. PubMed ID: 27358477
Classification of apoptosis and necrosis by morphological difference has been widely used for decades. However, this method has been seriously doubt in recent years, mainly due to lack of functional and biochemical evidence to interpret the morphology changes. To address these questions, this study devised genetic manipulations in Drosophila to study pyknosis, a process of nuclear shrinkage and chromatin condensation occurred in apoptosis and necrosis. By following the progression of necrotic pyknosis, a transient state was surprisingly observed of chromatin detachment from the nuclear envelope (NE), followed with the NE completely collapsed onto chromatin. This phenomenon lead to the discovery that phosphorylation of barrier-to-autointegration factor (BAF) mediates this initial separation of NE from chromatin. Functionally, inhibition of BAF phosphorylation suppressed the necrosis in both Drosophila and human cells, suggesting necrotic pyknosis is conserved in the propagation of necrosis. In contrast, apoptotic pyknosis did not show a detached state of chromatin from NE and inhibition of BAF phosphorylation had no effect on apoptotic pyknosis and apoptosis. This research provides the first genetic evidence supporting morphological classification of apoptosis and necrosis by pyknosis.

Monday, July 11th

Prasad, N., Tarikere, S., Khanale, D., Habib, F. and Shashidhara, L. S. (2016). A comparative genomic analysis of targets of Hox protein Ultrabithorax amongst distant insect species. Sci Rep 6: 27885. PubMed ID: 27296678
In the fruitfly Drosophila melanogaster, the differential development of wing and haltere is dependent on the function of the Hox protein Ultrabithorax (Ubx). This study compare Ubx-mediated regulation of wing patterning genes between the honeybee, Apis mellifera, the silkmoth, Bombyx mori and Drosophila. Orthologues of Ubx are expressed in the third thoracic segment of Apis and Bombyx, although they make functional hindwings. When over-expressed in transgenic Drosophila, Ubx derived from Apis or Bombyx could suppress wing development, suggesting evolutionary changes at the level of co-factors and/or targets of Ubx. To gain further insights into such events, direct targets of Ubx from Apis and Bombyx were identified by ChIP-seq and compared with those of Drosophila. While majority of the putative targets of Ubx are species-specific, a considerable number of wing-patterning genes are retained, over the past 300 millions years, as targets in all the three species. Interestingly, many of these are differentially expressed only between wing and haltere in Drosophila but not between forewing and hindwing in Apis or Bombyx. Detailed bioinformatics and experimental validation of enhancer sequences suggest that, perhaps along with other factors, changes in the cis-regulatory sequences of earlier targets contribute to diversity in Ubx function.

Kravchuk, O., Kim, M., Klepikov, P., Parshikov, A., Georgiev, P. and Savitsky, M. (2016). Transvection in Drosophila: trans-interaction between yellow enhancers and promoter is strongly suppressed by a cis-promoter only in certain genomic regions. Chromosoma [Epub ahead of print]. PubMed ID: 27300555
Transvection is a phenomenon of interallelic communication whereby enhancers of one allele can activate a promoter located on the homologous chromosome. It has been shown for many independent genes that enhancers preferentially act on the cis-linked promoter, but deletion of this promoter allows the enhancers to act in trans. This study tested whether this cis-preference in the enhancer-promoter interaction could be reconstituted outside of the natural position of a gene. The yellow gene was chosen as a model system. Transgenic flies were generated that carried the yellow gene modified by the inclusion of the strategically placed recognition sites for the Cre and Flp recombinases. To facilitate transvection, an endogenous Su(Hw) insulator (1A2) or gypsy insulator was placed behind the yellow gene. Independent action of the recombinases produced a pair of derivative alleles, one containing the promoter-driven yellow gene, and the other, the enhancers and promoter that failed to produce a functional yellow protein. As a result, strong transvection was observed in many genomic regions, suggesting that a complete cis-preference of the enhancer-promoter interactions is mainly restricted to genes in their natural loci.

Fukaya, T., Lim, B. and Levine, M. (2016). Enhancer control of transcriptional bursting. Cell [Epub ahead of print]. PubMed ID: 27293191
Transcription is episodic, consisting of a series of discontinuous bursts. Using live-imaging methods and quantitative analysis, transcriptional bursting was examined in living Drosophila embryos. Different developmental enhancers positioned downstream of synthetic reporter genes produce transcriptional bursts with similar amplitudes and duration but generate very different bursting frequencies, with strong enhancers producing more bursts than weak enhancers. Insertion of an insulator reduces the number of bursts and the corresponding level of gene expression, suggesting that enhancer regulation of bursting frequency is a key parameter of gene control in development. It was also shown that linked reporter genes exhibit coordinated bursting profiles when regulated by a shared enhancer, challenging conventional models of enhancer-promoter looping.

Zhang, Z., Boskovic, Z., Hussain, M. M., Hu, W., Inouye, C., Kim, H. J., Abole, A. K., Doud, M. K., Lewis, T. A., Koehler, A. N., Schreiber, S. L. and Tjian, R. (2015). Chemical perturbation of an intrinsically disordered region of TFIID distinguishes two modes of transcription initiation. Elife 4 [Epub ahead of print]. PubMed ID: 26314865
Intrinsically disordered proteins/regions (IDPs/IDRs) are proteins or peptide segments that fail to form stable 3-dimensional structures in the absence of partner proteins. They are abundant in eukaryotic proteomes and are often associated with human diseases, but their biological functions have been elusive to study. This study reports the identification of a tin(IV) oxochloride-derived cluster that binds an evolutionarily conserved IDR within the metazoan TFIID transcription complex. Binding arrests an isomerization of promoter-bound TFIID that is required for the engagement of Pol II during the first (de novo) round of transcription initiation. However, the specific chemical probe does not affect reinitiation, which requires the re-entry of Pol II, thus, mechanistically distinguishing these two modes of transcription initiation. This work also suggests a new avenue for targeting the elusive IDRs by harnessing certain features of metal-based complexes for mechanistic studies, and for the development of novel pharmaceutical interventions.

Sunday, July 10th

Lienard, M. A., Araripe, L. O. and Hartl, D. L. (2016). Neighboring genes for DNA-binding proteins rescue male sterility in Drosophila hybrids. Proc Natl Acad Sci U S A [Epub ahead of print]. PubMed ID: 27357670
Crosses between closely related animal species often result in male hybrids that are sterile, and the molecular and functional basis of genetic factors for hybrid male sterility is of great interest. This study reports a molecular and functional analysis of HMS1, a region of 9.2 kb in chromosome 3 of Drosophila mauritiana, which results in virtually complete hybrid male sterility when homozygous in the genetic background of sibling species Drosophila simulans. The HMS1 region contains two strong candidate genes for the genetic incompatibility, agt and Taf1 Both encode unrelated DNA-binding proteins, agt for an alkyl-cysteine-S-alkyltransferase and Taf1 for a subunit of transcription factor TFIID that serves as a multifunctional transcriptional regulator. The contribution of each gene to hybrid male sterility was assessed by means of germ-line transformation, with constructs containing complete agt and Taf1 genomic sequences as well as various chimeric constructs. Both agt and Taf1 contribute about equally to HMS1 hybrid male sterility. Transgenes containing either locus rescue sterility in about one-half of the males, and among fertile males the number of offspring is in the normal range. This finding suggests compensatory proliferation of the rescued, nondysfunctional germ cells. Results with chimeric transgenes imply that the hybrid incompatibilities result from interactions among nucleotide differences residing along both agt and Taf1. The results challenge a number of preliminary generalizations about the molecular and functional basis of hybrid male sterility, and strongly reinforce the role of DNA-binding proteins as a class of genes contributing to the maintenance of postzygotic reproductive isolation.

Suryamohan, K., Hanson, C., Andrews, E., Sinha, S., Scheel, M. D. and Halfon, M. S. (2016). Redeployment of a conserved gene regulatory network during Aedes aegypti development. Dev Biol [Epub ahead of print]. PubMed ID: 27341759
Changes in gene regulatory networks (GRNs) underlie the evolution of morphological novelty and developmental system drift. The fruitfly Drosophila melanogaster and the dengue and Zika vector mosquito Aedes aegypti have substantially similar nervous system morphology. Nevertheless, they show significant divergence in a set of genes co-expressed in the midline of the Drosophila central nervous system, including the master regulator single minded and downstream genes including short gastrulation, Star, and NetrinA. In contrast to Drosophila, this study found that midline expression of these genes is either absent or severely diminished in A. aegypti. Instead, they are co-expressed in the lateral nervous system. This suggests that in A. aegypti this "midline GRN" has been redeployed to a new location while lost from its previous site of activity. In order to characterize the relevant GRNs, the SCRMshaw method was employed to identify transcriptional cis-regulatory modules in both species. Analysis of these regulatory sequences in transgenic Drosophila suggests that the altered gene expression observed in A. aegypti is the result of trans-dependent redeployment of the GRN, potentially stemming from cis-mediated changes in the expression of sim and other as-yet unidentified regulators. The results illustrate a novel 'repeal, replace, and redeploy' mode of evolution in which a conserved GRN acquires a different function at a new site while its original function is co-opted by a different GRN. This represents a striking example of developmental system drift in which the dramatic shift in gene expression does not result in gross morphological changes, but in more subtle differences in development and function of the late embryonic nervous system.

Klok, C. J., Kaiser, A., Socha, J. J., Lee, W. K. and Harrison, J. F. (2016). Multigenerational effects of rearing atmospheric oxygen level on the tracheal dimensions and diffusing capacities of pupal and adult Drosophila melanogaster. Adv Exp Med Biol 903: 285-300. PubMed ID: 27343104
Insects are small relative to vertebrates, and were larger in the Paleozoic when atmospheric oxygen levels were higher. The safety margin for oxygen delivery does not increase in larger insects, due to an increased mass-specific investment in the tracheal system and a greater use of convection in larger insects. Prior studies have shown that the dimensions and number of tracheal system branches varies inversely with rearing oxygen in embryonic and larval insects. This study tested whether rearing in 10, 21, or 40 kPa atmospheric oxygen atmospheres for 5-7 generations affected the tracheal dimensions and diffusing capacities of pupal and adult Drosophila. Abdominal tracheae and pupal snorkel tracheae showed weak responses to oxygen, while leg tracheae showed strong, but imperfect compensatory changes. The diffusing capacity of leg tracheae appears closely matched to predicted oxygen transport needs by diffusion, perhaps explaining the consistent and significant responses of these tracheae to rearing oxygen. The reduced investment in tracheal structure in insects reared in higher oxygen levels may be important for conserving tissue oxygen partial pressure and may provide an important mechanism for insects to invest only the space and materials necessary into respiratory structure.

Corbett-Detig, R. (2016). Selection on inversion breakpoints favors proximity to pairing sensitive sites in Drosophila melanogaster. Genetics [Epub ahead of print]. PubMed ID: 27343234
Chromosomal inversions are widespread among taxa, and have been implicated in a number of biological processes including adaptation, sex chromosome evolution, and segregation distortion. Consistent with selection favoring linkage between loci, it is well established that length is a selected trait of inversions. However, the factors that affect the distribution of inversion breakpoints remain poorly understood. 'Sensitive sites' have been mapped on all euchromatic chromosome arms in Drosophila melanogaster, and may be a source of natural selection on inversion breakpoint positions. Briefly, sensitive sites are genomic regions wherein proximal structural rearrangements result in large reductions in local recombination rates in heterozygotes. This study shows that breakpoints of common inversions are significantly more likely to lie within a cytological band containing a sensitive site than are breakpoints of rare inversions. Furthermore, common inversions for which neither breakpoint intersects a sensitive site are significantly longer than rare inversions, but common inversions whose breakpoints intersect a sensitive site show no evidence for increased length. These results are interpreted to mean that selection favors inversions whose breakpoints disrupt synteny near to sensitive sites, possibly because these inversions suppress recombination in large genomic regions. This is the first evidence consistent with positive selection acting on inversion breakpoint positions.

Saturday, July 9th

Benton, M. A., Pechmann, M., Frey, N., Stappert, D., Conrads, K. H., Chen, Y. T., Stamataki, E., Pavlopoulos, A. and Roth, S. (2016). Toll genes have an ancestral role in axis elongation. Curr Biol [Epub ahead of print]. PubMed ID: 27212406
One of the key morphogenetic processes used during development is the controlled intercalation of cells between their neighbors. This process has been co-opted into a range of developmental events, and it also underlies an event that occurs in each major group of bilaterians: elongation of the embryo along the anterior-posterior axis. In Drosophila, a novel component of this process was recently discovered by Pare (2014), who showed that three Toll genes, Toll-2 (18 Wheeler), Toll-6 and Toll-8 (Tollo), function together to drive cell intercalation during germband extension. This finding raises the question of whether this role of Toll genes is an evolutionary novelty of flies or a general mechanism of embryonic morphogenesis. This study shows that the Toll gene function in axis elongation is, in fact, widely conserved among arthropods. First, two Toll genes were functionally demonstrated to be required for cell intercalation in the beetle Tribolium castaneum. These genes belong to a previously undescribed Toll subfamily, and members of this subfamily exhibit striped expression (as seen in Tribolium and previously reported in Drosophila) in embryos of six other arthropod species spanning the entire phylum. Last, it was shown that two of these Toll genes are required for normal morphogenesis during anterior-posterior embryo elongation in the spider Parasteatoda tepidariorum, a member of the most basally branching arthropod lineage. From these findings, it is hypothesized that Toll genes had a morphogenetic function in embryo elongation in the last common ancestor of all arthropods, which existed over 550 million years ago.

Battlay, P., Schmidt, J. M., Fournier-Level, A. and Robin, C. (2016). Genomic and transcriptomic associations identify a new insecticide resistance phenotype for the selective sweep at the Cyp6g1 locus of Drosophila melanogaster. G3 (Bethesda) [Epub ahead of print]. PubMed ID: 27317781
Scans of the Drosophila melanogaster genome have identified organophosphate resistance loci among those with the most pronounced signature of positive selection. In this study the molecular basis of resistance to the organophosphate insecticide azinphos-methyl was investigated using the Drosophila Genetic Reference Panel and genome-wide association. Recently released full transcriptome data was used to extend the utility of the Drosophila Genetic Reference Panel resource beyond traditional genomewide association studies to allow systems genetics analyses of phenotypes. Both genomic and transcriptomic associations independently identified Cyp6g1, a gene involved in resistance to DDT and neonicotinoid insecticides, as the top candidate for azinphos-methyl resistance. This was verified by transgenically overexpressing Cyp6g1 using natural regulatory elements from a resistant allele, resulting in a 6.5-fold increase in resistance. Four novel candidate genes associated with azinphos-methyl resistance were found, all of which are involved in either regulation of fat storage or nervous system development. In Cyp6g1, a demonstrable resistance locus was found, a verification that transcriptome data can be used to identify variants associated with insecticide resistance, and an overlap was found between peaks of a genome-wide association study and a genome-wide selective sweep analysis.

Palmer, W. H. and Obbard, D. J. (2016). Variation and evolution in the glutamine-rich repeat region of Drosophila Argonaute-2. G3 (Bethesda) [Epub ahead of print]. PubMed ID: 27317784
RNA interference pathways mediate biological processes through Argonaute-family proteins, which bind small RNAs as guides to silence complementary target nucleic acids. In insects and crustaceans Argonaute-2 silences viral nucleic acids, and therefore acts as a primary effector of innate antiviral immunity. Although the function of the major Argonaute-2 domains, which are conserved across most Argonaute-family proteins, are known, many invertebrate Argonaute-2 homologs contain a glutamine-rich repeat (GRR) region of unknown function at the N-terminus. This study combined long-read amplicon sequencing of Drosophila Genetic Reference Panel (DGRP) lines with publicly available sequence data from many insect species to show that this region evolves extremely rapidly and is hypervariable within species. Distinct GRR haplotype groups war defined in D. melanogaster, and it is suggested that one of these haplotype groups has recently risen to high frequency in a North American population. Finally, published data from genome-wide association studies of viral resistance in D. melanogaster was used to test whether GRR haplotypes are associated with survival after virus challenge. A marginally significant association with survival after challenge with Drosophila C Virus in the DGRP was found, but this finding could not be replicated using lines from the Drosophila Synthetic Population Resource panel.

Arguello, J. R., Cardoso-Moreira, M., Grenier, J. K., Gottipati, S., Clark, A. G. and Benton, R. (2016). Extensive local adaptation within the chemosensory system following Drosophila melanogaster's global expansion. Nat Commun 7: ncomms11855. PubMed ID: 27292132
How organisms adapt to new environments is of fundamental biological interest, but poorly understood at the genetic level. Chemosensory systems provide attractive models to address this problem, because they lie between external environmental signals and internal physiological responses. To investigate how selection has shaped the well-characterized chemosensory system of Drosophila melanogaster, this study analysed genome-wide data from five diverse populations. By couching population genomic analyses of chemosensory protein families, including odorant receptors, gustatory receptors, and odorant-binding proteins, within parallel analyses of other large families, it was demonstrated that chemosensory proteins are not outliers for adaptive divergence between species. However, chemosensory families often display the strongest genome-wide signals of recent selection within D. melanogaster. Recent adaptation has operated almost exclusively on standing variation, and patterns of adaptive mutations predict diverse effects on protein function. Finally, evidence is provided that chemosensory proteins have experienced relaxed constraint, and it is argued that this has been important for their rapid adaptation over short timescales.

Friday, July 8th

Spuhler, I. A., Conley, G. M., Scheffold, F. and Sprecher, S. G. (2016). Super resolution imaging of genetically labeled synapses in Drosophila brain tissue. Front Cell Neurosci 10: 142. PubMed ID: 27303270
Understanding synaptic connectivity and plasticity within brain circuits and their relationship to learning and behavior is a fundamental quest in neuroscience. Visualizing the fine details of synapses using optical microscopy remains however a major technical challenge. Super resolution microscopy opens the possibility to reveal molecular features of synapses beyond the diffraction limit. With direct stochastic optical reconstruction microscopy, dSTORM, this study imaged synaptic proteins, in the brain tissue of Drosophila. Super resolution imaging of brain tissue harbors difficulties due to light scattering and the density of signals. In order to reduce out of focus signal, advantage was taken of the genetic tools available in the Drosophila, and synaptic proteins, including Bruchpilot, Synaptotagmin, Drep-2 and cholinergic receptor nicotinic Acetylcholine Receptor α7, expressed in only a small number of neurons, were fluorescently tagged. These neurons form synapses within the calyx of the mushroom body, a distinct brain region involved in associative memory formation. The results show that super resolution microscopy, in combination with genetically labeled synaptic proteins, is a powerful tool to investigate synapses in a quantitative fashion providing an entry point for studies on synaptic plasticity during learning and memory formation.

Weiss, K. R. and Littleton, J. T. (2016). Characterization of axonal transport defects in Drosophila huntingtin mutants. J Neurogenet: 1-30. PubMed ID: 27309588
Polyglutamine expansion within Huntingtin (Htt) causes the fatal neurodegenerative disorder Huntington's Disease (HD). Although Htt is ubiquitously expressed and conserved from Drosophila to humans, its normal biological function is still being elucidated. This study characterized a role for the Drosophila Htt homolog (dHtt) in fast axonal transport (FAT). Generation and expression of transgenic dHtt-mRFP and human Htt-mRFP fusion proteins in Drosophila revealed co-localization with mitochondria and synaptic vesicles undergoing FAT. However, Htt was not ubiquitously associated with the transport machinery, as it was excluded from dense-core vesicles and APLIP1 containing vesicles. Quantification of cargo movement in dHtt deficient axons revealed that mitochondria and synaptic vesicles show a decrease in the distance and duration of transport, and an increase in the number of pauses. In addition, the ratio of retrograde to anterograde flux was increased in mutant animals. The data suggest dHtt likely acts locally at cargo interaction sites to regulate processivity. An increase in dynein heavy chain expression was also observed in dHtt mutants, suggesting the altered flux observed for all cargo may represent secondary transport changes occurring independent of dHtt's primary function. Expression of dHtt in a milton (HAP1) mutant background revealed that the protein does not require mitochondria or HAP1 to localize along axons, suggesting Htt has an independent mechanism for coupling to motors to regulate their processivity during axonal transport.

Shirangi, T. R., Wong, A. M., Truman, J. W. and Stern, D. L. (2016). Doublesex regulates the connectivity of a neural circuit controlling Drosophila male courtship song. Dev Cell 37: 533-544. PubMed ID: 27326931
It is unclear how regulatory genes establish neural circuits that compose sex-specific behaviors. The Drosophila melanogaster male courtship song provides a powerful model to study this problem. Courting males vibrate a wing to sing bouts of pulses and hums, called pulse and sine song, respectively. This study reports the discovery of male-specific thoracic interneurons-the TN1A neurons-that are required specifically for sine song. The TN1A neurons can drive the activity of a sex-non-specific wing motoneuron, hg1, which is also required for sine song. The male-specific connection between the TN1A neurons and the hg1 motoneuron is regulated by the sexual differentiation gene doublesex. doublesex was shown to be required in the TN1A neurons during development to increase the density of the TN1A arbors that interact with dendrites of the hg1 motoneuron. These findings demonstrate how a sexual differentiation gene can build a sex-specific circuit motif by modulating neuronal arborization.

Gomez-Diaz, C., Bargeton, B., Abuin, L., Bukar, N., Reina, J. H., Bartoi, T., Graf, M., Ong, H., Ulbrich, M. H., Masson, J. F. and Benton, R. (2016). A CD36 ectodomain mediates insect pheromone detection via a putative tunnelling mechanism. Nat Commun 7: 11866. PubMed ID: 27302750
CD36 transmembrane proteins have diverse roles in lipid uptake, cell adhesion and pathogen sensing. Despite numerous in vitro studies, how they act in native cellular contexts is poorly understood. A Drosophila CD36 homologue, antennal protein Sensory neuron membrane protein 1 (SNMP1), was previously shown to facilitate detection of lipid-derived pheromones by their cognate receptors in olfactory cilia. This study investigated how SNMP1 functions in vivo. Structure-activity dissection demonstrates that SNMP1's ectodomain is essential, but intracellular and transmembrane domains dispensable, for cilia localization and pheromone-evoked responses. SNMP1 can be substituted by mammalian CD36, whose ectodomain can interact with insect pheromones. Homology modelling, using the mammalian LIMP-2 structure as template, reveals a putative tunnel in the SNMP1 ectodomain that is sufficiently large to accommodate pheromone molecules. Amino-acid substitutions predicted to block this tunnel diminish pheromone sensitivity. A model is proposed in which SNMP1 funnels hydrophobic pheromones from the extracellular fluid to integral membrane receptors.

Thursday, July 7th

Chouhan, A.K., Guo, C., Hsieh, Y.C., Ye, H., Senturk, M., Zuo, Z., Li, Y., Chatterjee, S., Botas, J., Jackson, G.R., Bellen, H.J. and Shulman, J.M. (2016). Uncoupling neuronal death and dysfunction in Drosophila models of neurodegenerative disease. Acta Neuropathol Commun 4: 62. PubMed ID: 27338814
Common neurodegenerative proteinopathies, such as Alzheimer's disease (AD) and Parkinson's disease (PD), are characterized by the misfolding and aggregation of toxic protein species, including the amyloid β (Aβ) peptide, microtubule-associated protein Tau (Tau), and alpha-synuclein (αSyn) protein. These factors also show toxicity in Drosophila. Using standardized conditions and medium-throughput assays, this study expressed human Tau, Aβ or αSyn selectively in neurons of the adult Drosophila retina and monitored age-dependent changes in both structure and function, based on tissue histology and recordings of the electroretinogram (ERG), respectively. Each protein was found to cause a unique profile of neurodegenerative pathology. Strikingly, expression of Tau leads to progressive loss of ERG responses whereas retinal architecture and neuronal numbers are largely preserved. By contrast, Aβ induces modest, age-dependent neuronal loss without degrading the retinal ERG. αSyn expression is characterized by marked retinal vacuolar change, progress photoreceptor cell death, and delayed-onset but modest ERG changes. Surprisingly, Tau and αSyn each cause prominent but distinct synaptotoxic profiles, including disorganization or enlargement of photoreceptor terminals, respectively. These findings suggest that Drosophila may be useful for revealing determinants of neuronal dysfunction that precede cell loss, including synaptic changes, in the adult nervous system. 

Carazo, P., Green, J., Sepil, I., Pizzari, T. and Wigby, S. (2016). Inbreeding removes sex differences in lifespan in a population of Drosophila melanogaster. Biol Lett 12. PubMed ID: 27354712
Sex differences in ageing rates and lifespan are common in nature, and an enduring puzzle for evolutionary biology. One possibility is that sex-specific mortality rates may result from recessive deleterious alleles in 'unguarded' heterogametic X or Z sex chromosomes (the unguarded X hypothesis). Empirical evidence for this is, however, limited. This study tests a fundamental prediction of the unguarded X hypothesis in Drosophila melanogaster, namely that inbreeding shortens lifespan more in females (the homogametic sex in Drosophila) than in males. To test for additional sex-specific social effects, the lifespan of males and females kept in isolation was studied, in related same-sex groups, and in unrelated same-sex groups. As expected, outbred females outlive outbred males and inbreeding shortens lifespan. However, inbreeding-mediated reductions in lifespan are stronger for females, such that lifespan is similar in inbred females and males. It was also shown that the social environment, independent of inbreeding, affects male, but not female lifespan. In conjunction with recent studies, these data suggest that asymmetric inheritance mechanisms may play an important role in the evolution of sex-specific lifespan and that social effects must be considered explicitly when studying these fundamental patterns. 

Yang, Y., He, S., Wang, Q., Li, F., Kwak, M. J., Chen, S., O'Connell, D., Zhang, T., Pirooz, S. D., Jeon, Y., Chimge, N. O., Frenkel, B., Choi, Y., Aldrovandi, G. M., Oh, B. H., Yuan, Z. and Liang, C. (2016). Autophagic UVRAG promotes UV-induced photolesion repair by activation of the CRL4(DDB2) E3 ligase. Mol Cell 62: 507-519. PubMed ID: 27203177
UV-induced DNA damage, a major risk factor for skin cancers, is primarily repaired by nucleotide excision repair (NER). UV radiation resistance-associated gene (UVRAG) is a tumor suppressor involved in autophagy. It was initially isolated as a cDNA partially complementing UV sensitivity in xeroderma pigmentosum (XP), but this was not explored further. This study shows that UVRAG plays an integral role in UV-induced DNA damage repair. It localizes to photolesions and associates with DDB1 to promote the assembly and activity of the DDB2-DDB1-Cul4A-Roc1 (CRL4(DDB2)) ubiquitin ligase complex, leading to efficient XPC recruitment and global genomic NER. UVRAG depletion decreased substrate handover to XPC and conferred UV-damage hypersensitivity. The importance of UVRAG for UV-damage tolerance was confirmed using a Drosophila model. Furthermore, increased UV-signature mutations in melanoma correlate with reduced expression of UVRAG. These results identify UVRAG as a regulator of CRL4(DDB2)-mediated NER and suggest that its expression levels may influence melanoma predisposition.

Chen, K., Lin, G., Haelterman, N.A., Ho, T.S., Li, T., Li, Z., Duraine, L., Graham, B.H., Jaiswal, M., Yamamoto, S., Rasband, M.N. and Bellen, H.J. (2016). Loss of Frataxin induces iron toxicity, sphingolipid synthesis, and Pdk1/Mef2 activation, leading to neurodegeneration. Elife [Epub ahead of print]. PubMed ID: 27343351
Mutations in Frataxin (FXN) cause Friedreich's ataxia (FRDA), a recessive neurodegenerative disorder. Previous studies have proposed that loss of FXN causes mitochondrial dysfunction, which triggers elevated reactive oxygen species (ROS) and leads to the demise of neurons. This study describes a ROS independent mechanism that contributes to neurodegeneration in fly FXN mutants. Loss of frataxin homolog (fh) in Drosophila was shown to leads to iron toxicity, which in turn induces sphingolipid synthesis and ectopically activates 3-phosphoinositide dependent protein kinase-1 (Pdk1) and myocyte enhancer factor-2 (Mef2). Dampening iron toxicity, inhibiting sphingolipid synthesis by Myriocin, or reducing Pdk1 or Mef2 levels, all effectively suppress neurodegeneration in fh mutants. Moreover, increasing dihydrosphingosine activates Mef2 activity through PDK1 in mammalian neuronal cell line suggesting that the mechanisms are evolutionarily conserved. These results indicate that an iron/sphingolipid/PDk1/Mef2 pathway may play a role in FRDA.

Wednesday, July 6th

Chance, R. K. and Bashaw, G. J. (2015). Slit-dependent endocytic trafficking of the Robo receptor is required for Son of Sevenless recruitment and midline axon repulsion. PLoS Genet 11: e1005402. PubMed ID: 26335920
Understanding how axon guidance receptors are activated by their extracellular ligands to regulate growth cone motility is critical to learning how proper wiring is established during development. Roundabout (Robo) is one such guidance receptor that mediates repulsion from its ligand Slit in both invertebrates and vertebrates. This study shows that endocytic trafficking of the Robo receptor in response to Slit-binding is necessary for its repulsive signaling output. Dose-dependent genetic interactions and in vitro Robo activation assays support a role for Clathrin-dependent endocytosis, and entry into both the early and late endosomes as positive regulators of Slit-Robo signaling. Two conserved motifs were identified in Robo's cytoplasmic domain that are required for its Clathrin-dependent endocytosis and activation in vitro; gain of function and genetic rescue experiments provide strong evidence that these trafficking events are required for Robo repulsive guidance activity in vivo. These data support a model in which Robo's ligand-dependent internalization from the cell surface to the late endosome is essential for receptor activation and proper repulsive guidance at the midline by allowing recruitment of the downstream effector Son of Sevenless in a spatially constrained endocytic trafficking compartment.

Lefebvre, F. A., Benoit Bouvrette, L. P., Perras, L., Blanchet-Cohen, A., Garnier, D., Rak, J. and Lecuyer, E. (2016). Comparative transcriptomic analysis of human and Drosophila extracellular vesicles. Sci Rep 6: 27680. PubMed ID: 27282340
Extracellular vesicles (EVs) are membrane-enclosed nanoparticles containing specific repertoires of genetic material. In mammals, EVs can mediate the horizontal transfer of various cargos and signaling molecules, notably miRNA and mRNA species. Whether this form of intercellular communication prevails in other metazoans remains unclear. This study reports the first parallel comparative morphologic and transcriptomic characterization of EVs from Drosophila and human cellular models. Electronic microscopy revealed that human and Drosophila cells release similar EVs with diameters ranging from 30 to 200 nm, which contain complex populations of transcripts. RNA-seq identified abundant ribosomal RNAs, related pseudogenes and retrotransposons in human and Drosophila EVs. Vault RNAs and Y RNAs abounded in human samples, whereas small nucleolar RNAs involved in pseudouridylation were most prevalent in Drosophila EVs. Numerous mRNAs were identified, largely consisting of exonic sequences displaying full-length read coverage and enriched for translation and electronic transport chain functions. By analogy with human systems, these sizeable similarities suggest that EVs could potentially enable RNA-mediated intercellular communication in Drosophila.

Lorincz, P., Lakatos, Z., Varga, A., Maruzs, T., Simon-Vecsei, Z., Darula, Z., Benko, P., Csordas, G., Lippai, M., Ando, I., Hegedus, K., Medzihradszky, K. F., Takats, S. and Juhasz, G. (2016). MiniCORVET is a Vps8-containing early endosomal tether in Drosophila. Elife 5 [Epub ahead of print]. PubMed ID: 27253064
Yeast studies identified two heterohexameric tethering complexes, which consist of 4 shared (Vps11, Vps16, Vps18 and Vps33) and 2 specific subunits: Vps3 and Vps8 (CORVET) versus Vps39 and Vps41 (HOPS). CORVET is an early and HOPS is a late endosomal tether. The function of HOPS is well known in animal cells, while CORVET is poorly characterized. This study shows that Drosophila Vps8 is highly expressed in hemocytes and nephrocytes, and localizes to early endosomes despite the lack of a clear Vps3 homolog. Vps8 forms a complex and acts together with Vps16A, Deep Orange/Vps18 and Carnation/Vps33A, and loss of any of these proteins leads to fragmentation of endosomes. Surprisingly, Vps11 deletion causes enlargement of endosomes, similar to loss of the HOPS-specific subunits Vps39 and Light/Vps41. This study thus identifies a 4 subunit-containing miniCORVET complex as an unconventional early endosomal tether in Drosophila.

Vonhoff, F. and Keshishian, H. (2016). Cyclic nucleotide signaling is required during synaptic refinement at the Drosophila neuromuscular junction. Dev Neurobiol [Epub ahead of print]. PubMed ID: 27281494
The removal of miswired synapses is a fundamental prerequisite for normal circuit development, leading to clinical problems when aberrant. However, the underlying activity-dependent molecular mechanisms involved in synaptic pruning remain incompletely resolved. This study examined the dynamic properties of intracellular calcium oscillations and tested a role for cAMP signaling during synaptic refinement in intact Drosophila embryos using optogenetic tools. In vivo evidence at the single gene level is provided that the calcium-dependent adenylyl cyclase rutabaga, the phosphodiesterase dunce, the kinase PKA, and Protein Phosphatase 1 (PP1) all operate within a functional signaling pathway to modulate Sema2a-dependent chemorepulsion. Presynaptic cAMP levels are required to be dynamically maintained at an optimal level to suppress connectivity defects. It is also proposed that PP1 may serve as a molecular link between cAMP signaling and CaMKII in the pathway underlying refinement. These results introduce an in vivo model where presynaptic cAMP levels, downstream of electrical activity and calcium influx, act via PKA and PP1 to modulate the neuron's response to chemorepulsion involved in the withdrawal of off-target synaptic contacts.

Tuesday, July 5th

Gerstner, J. R., Lenz, O., Vanderheyden, W. M., Chan, M. T., Pfeiffenberger, C. and Pack, A. I. (2016). Amyloid-β induces sleep fragmentation that is rescued by fatty acid binding proteins in Drosophila. J Neurosci Res [Epub ahead of print]. PubMed ID: 27320125
Disruption of sleep/wake activity in Alzheimer's disease (AD) patients significantly affects their quality of life and that of their caretakers and is a major contributing factor for institutionalization. Levels of amyloid-β (Aβ; see Drosophila Appl) have been shown to be regulated by neuronal activity and to correlate with the sleep/wake cycle. Whether consolidated sleep can be disrupted by Aβ alone is not well understood. It was hypothesized that Aβ42 can increase wakefulness and disrupt consolidated sleep. This study shows that flies expressing the human Aβ42 transgene in neurons have significantly reduced consolidated sleep compared with control flies. Fatty acid binding proteins (Fabp) are small hydrophobic ligand carriers that have been clinically implicated in AD. Aβ42 flies that carry a transgene of either the Drosophila Fabp or the mammalian brain-type Fabp show a significant increase in nighttime sleep and long consolidated sleep bouts, rescuing the Aβ42-induced sleep disruption. These studies suggest that alterations in Fabp levels and/or activity may be associated with sleep disturbances in AD. Future work to determine the molecular mechanisms that contribute to Fabp-mediated rescue of Aβ42-induced sleep loss will be important for the development of therapeutics in the treatment of AD.
Parkhitko, A.A., Binari, R., Zhang, N., Asara, J.M., Demontis, F. and Perrimon, N. (2016). Tissue-specific down-regulation of S-adenosyl-homocysteine via suppression of dAhcyL1/dAhcyL2 extends health span and life span in Drosophila. Genes Dev [Epub ahead of print]. PubMed ID: 27313316
Aging is a risk factor for many human pathologies and is characterized by extensive metabolic changes. Using targeted high-throughput metabolite profiling in Drosophila melanogaster at different ages, this study demonstrates that methionine metabolism changes strikingly during aging. Methionine generates the methyl donor S-adenosyl-methionine (SAM), which is converted via methylation to S-adenosyl-homocysteine (SAH), which accumulates during aging. A targeted RNAi screen against methionine pathway components reveals significant life span extension in response to down-regulation of two noncanonical Drosophila homologs of the SAH hydrolase Ahcy (S-adenosyl-L-homocysteine hydrolase [SAHH]), CG9977/dAhcyL1 and Ahcy89E/CG8956/dAhcyL2, which act as dominant-negative regulators of canonical AHCY. Importantly, tissue-specific down-regulation of dAhcyL1/L2 in the brain and intestine extends health and life span. Furthermore, metabolomic analysis of dAhcyL1-deficient flies reveals its effect on age-dependent metabolic reprogramming and H3K4 methylation. Altogether, reprogramming of methionine metabolism in young flies and suppression of age-dependent SAH accumulation lead to increased life span. These studies highlight the role of noncanonical Ahcy enzymes as determinants of healthy aging and longevity.

Dong, T., He, J., Wang, S., Wang, L., Cheng, Y. and Zhong, Y. (2016). Inability to activate Rac1-dependent forgetting contributes to behavioral inflexibility in mutants of multiple autism-risk genes. Proc Natl Acad Sci U S A [Epub ahead of print]. PubMed ID: 27335463
The etiology of autism is so complicated because it involves the effects of variants of several hundred risk genes along with the contribution of environmental factors. Therefore, it has been challenging to identify the causal paths that lead to the core autistic symptoms such as social deficit, repetitive behaviors, and behavioral inflexibility. As an alternative approach, extensive efforts have been devoted to identifying the convergence of the targets and functions of the autism-risk genes to facilitate mapping out causal paths. This study used a reversal-learning task to measure behavioral flexibility in Drosophila and determined the effects of loss-of-function mutations in multiple autism-risk gene homologs in flies. Mutations of five autism-risk genes with diversified molecular functions all lead to a similar phenotype of behavioral inflexibility indicated by impaired reversal-learning. These reversal-learning defects result from the inability to forget or rather, specifically, to activate Rac1 (Ras-related C3 botulinum toxin substrate 1)-dependent forgetting. Thus, behavior-evoked activation of Rac1-dependent forgetting has a converging function for autism-risk genes.

Li, T., Giagtzoglou, N., Eberl, D., Nagarkar-Jaiswal, S., Cai, T., Godt, D., Groves, A.K. and Bellen, H.J. (2016). The E3 ligase Ubr3 regulates Usher syndrome and MYH9 disorder proteins in the auditory organs of Drosophila and mammals. Elife [Epub ahead of print]. PubMed ID: 27331610
Myosins play essential roles in the development and function of auditory organs and multiple myosin genes are associated with hereditary forms of deafness. Using a forward genetic screen in Drosophila, this study identified an E3 ligase, Ubr3, as an essential gene for auditory organ development. Ubr3 negatively regulates the mono-ubiquitination of non-muscle Myosin II, a protein associated with hearing loss in humans. The mono-ubiquitination of Myosin II promotes its physical interaction with Myosin VIIa, a protein responsible for Usher syndrome type IB. It was shown that ubr3 mutants phenocopy pathogenic variants of Myosin II and that Ubr3 interacts genetically and physically with three Usher syndrome proteins. The interactions between Myosin VIIa and Myosin IIa are conserved in the mammalian cochlea and in human retinal pigment epithelium cells. These observations reveal a novel mechanism that regulates protein complexes affected in two forms of syndromic deafness and suggests a molecular function for Myosin IIa in auditory organs.

Monday, July 4th

Vienne, J., Spann, R., Guo, F. and Rosbash, M. (2016). Age-related reduction of recovery sleep and arousal threshold in Drosophila. Sleep [Epub ahead of print]. PubMed ID: 27306274
Physiological studies show that aging affects both sleep quality and quantity in humans, and sleep complaints increase with age. Along with knowledge about the negative effects of poor sleep on health, understanding the enigmatic relationship between sleep and aging is important. Because human sleep is similar to Drosophila (fruit fly) sleep in many ways, this study addressed the effects of aging on sleep in this model organism. Baseline sleep was recorded in five different Drosophila genotypes raised at either 21 ° C or 25 ° C. The amount of sleep recovered was then investigated after a nighttime of sleep deprivation (12 h) and after chronic sleep deprivation (3 h every night for multiple nights). Finally, the effects of aging on arousal, namely, sensitivity to neuronal and mechanical stimuli, were studied. Fly sleep was shown to be affected by age in a manner similar to that of humans and other mammals. Not only do older flies of several genotypes have more fragmented sleep and reduced total sleep time compared to young flies, but older flies also fail to recover as much sleep after sleep deprivation. This suggests either lower sleep homeostasis and/or a failure to properly recover sleep. Older flies also show a decreased arousal threshold, i.e., an increased response to neuronal and mechanical wake-promoting stimuli. The reduced threshold may either reflect or cause the reduced recovery sleep of older flies compared to young flies after sleep deprivation. It is concluded that further studies are certainly needed, but it is suggested that the lower homeostatic sleep drive of older flies causes their decreased arousal threshold.

Lihoreau, M., Poissonnier, L. A., Isabel, G. and Dussutour, A. (2016). Drosophila females trade off good nutrition with high quality oviposition sites when choosing foods. Exp Biol [Epub ahead of print]. PubMed ID: 27284071
Animals, from insects to human, select foods to regulate their acquisition of key nutrients in amounts and balances maximising fitness. In species where the nutrition of juveniles depends on parents, adults must make challenging foraging decisions that simultaneously address their own nutrient needs as well as those of the progeny. This study examined how fruit flies Drosophila melanogaster, a species where individuals eat and lay eggs in decaying fruits, integrate feeding decisions (individual nutrition) and oviposition decisions (offspring nutrition) when foraging. Using cafeteria assays with artificial diets varying in concentrations and ratios of protein to carbohydrates, Drosophila females were shown to exhibit complex foraging patterns, alternating between laying eggs on high carbohydrate foods and feeding on foods with different nutrient contents depending on their own nutritional state. Although larvae showed faster development on high protein foods, both survival and learning performances were higher on balanced foods. It is suggested that the apparent mismatch between the oviposition preference of females for high carbohydrate foods and the high performances of larvae on balanced foods reflects a natural situation where high carbohydrate ripened fruits gradually enrich in proteinaceous yeast as they start rotting, thereby yielding optimal nutrition for the developing larvae. The findings that animals with rudimentary parental care uncouple feeding and egg-laying decisions in order to balance their own diet and provide a nutritionally optimal environment to their progeny reveals unsuspected levels of complexity in the nutritional ecology of parent-offspring interactions.

Koseki, N., Mori, S., Suzuki, S., Tonooka, Y., Kosugi, S., Miyakawa, H. and Morimoto, T. (2016). Individual differences in sensory responses influence decision making by Drosophila melanogaster larvae on exposure to contradictory cues. J Neurogenet: 1-39. PubMed ID: 27309770
Animals make decisions on behavioral choice by evaluating internal and external signals. Individuals often make decisions in different ways, but the underlying neural mechanisms are not well understood. This study describes a system for observing the behavior of individual Drosophila melanogaster larvae simultaneously presented with contradictory signals, in this case attractive (yeast paste) and aversive (NaCl) signals. Olfaction was used to detect the yeast paste, whereas the ENaC/Pickpocket channel was important for NaCl detection. Wild-type (Canton-S) larvae fall into two decision making groups: one group decided to approach the yeast paste by overcoming the aversive signal, whereas the other group decided to forgo the yeast paste because of the aversive signal. These findings indicate that different endogenous sensitivities to NaCl contribute to make differences between two groups and that diverse decision making steps occur in individual animals.

Gomez-Marin, A., Oron, E., Gakamsky, A., Dan, V., Benjamini, Y. and Golani, I. (2016). Generative rules of Drosophila locomotor behavior as a candidate homology across phyla. Sci Rep 6: 27555. PubMed ID: 27271799
This study used immobility as an origin and reference for the measurement of fly locomotor a behavior; speed, walking direction and trunk orientation as the degrees of freedom shaping this behavior; and cocaine as the parameter inducing progressive transitions in and out of immobility. The generative rules that shape Drosophila locomotor behavior were characterized and quantified. Transitions into immobility unfold via sequential enhancement and then elimination of translation, curvature and finally rotation. Transitions out of immobility unfold by progressive addition of these degrees of freedom in the opposite order. The same generative rules have been found in vertebrate locomotor behavior in several contexts (pharmacological manipulations, ontogeny, social interactions) involving transitions in-and-out of immobility. Recent claims for deep homology between arthropod central complex and vertebrate basal ganglia provide an opportunity to examine whether the rules reported in this study also share common descent. The approach prompts the discovery of behavioral homologies, contributing to the elusive problem of behavioral evolution.

Sunday, July 3rd

Garcia, E. L., Wen, Y., Praveen, K. and Matera, A. G. (2016). Transcriptomic comparison of Drosophila snRNP biogenesis mutants reveals mutant-specific changes in pre-mRNA processing: implications for spinal muscular atrophy. Transcriptomic comparison of Drosophila snRNP biogenesis mutants reveals mutant-specific changes in pre-mRNA processing: implications for spinal muscular atrophy. RNA [Epub ahead of print]. PubMed ID: 27268418
Survival motor neuron (SMN) functions in the assembly of spliceosomal small nuclear ribonucleoproteins (snRNPs) that catalyze pre-mRNA splicing. This study used disruptions in Smn and two additional snRNP biogenesis genes, Phax and Ars2, to classify RNA processing differences as snRNP-dependent or gene-specific in Drosophila Phax and Smn mutants exhibited comparable reductions in snRNAs, and comparison of their transcriptomes uncovered shared sets of RNA processing changes. In contrast, Ars2 mutants displayed only small decreases in snRNA levels, and RNA processing changes in these mutants were generally distinct from those identified in Phax and Smn animals. Instead, RNA processing changes in Ars2 mutants support the known interaction of Ars2 protein with the cap-binding complex, as splicing changes showed a clear bias toward the first intron. Bypassing disruptions in snRNP biogenesis, direct knockdown of spliceosomal proteins caused similar changes in the splicing of snRNP-dependent events. However, these snRNP-dependent events were largely unaltered in three Smn mutants expressing missense mutations that were originally identified in human spinal muscular atrophy (SMA) patients. Hence, findings here clarify the contributions of Phax, Smn, and Ars2 to snRNP biogenesis in Drosophila, and loss-of-function mutants for these proteins reveal differences that help disentangle cause and effect in SMA model flies.

Wissel, S., Kieser, A., Yasugi, T., Duchek, P., Roitinger, E., Gokcezade, J. F., Steinmann, V., Gaul, U., Mechtler, K., Forstemann, K., Knoblich, J. A. and Neumuller, R. A. (2016). A combination of CRISPR/Cas9 and standardized RNAi as a versatile platform for the characterization of gene function. G3 (Bethesda) [Epub ahead of print]. PubMed ID: 27280787
Traditional loss-of-function studies in Drosophila suffer from a number of shortcomings including off-target effects in the case of RNA interference (RNAi) or the stochastic nature of mosaic clonal analysis. This study describes minimal in vivo GFP interference (miGFPi) as a versatile strategy to characterize gene function and to conduct highly stringent, cell-type specific loss-of-function experiments in Drosophila. miGFPi combines CRISPR/Cas9 mediated tagging of genes at their endogenous locus with an immunotag and an exogenous 21 nucleotide RNAi effector sequence with the use of a single reagent, highly validated RNAi line targeting this sequence. The utility and time effectiveness of this method was demonstrated by characterizing the function of the Polymerase I (Pol I) associated transcription factor Tif-1a and the previously uncharacterized gene MESR4 in the Drosophila female germline stem cell lineage. In addition, miGFPi was shown to serve as a powerful technique to functionally characterize individual isoforms of a gene. This aspect of miGFPi was exemplified by studying isoform specific loss-of-function phenotypes of the longitudinals lacking (lola) gene in neural stem cells. Altogether, the miGFPi strategy constitutes a generalized loss-of-function approach that is amenable to study the function of all genes in the genome in a stringent and highly time effective manner.

Chen, Y. A., Stuwe, E., Luo, Y., Ninova, M., Le Thomas, A., Rozhavskaya, E., Li, S., Vempati, S., Laver, J. D., Patel, D. J., Smibert, C. A., Lipshitz, H. D., Fejes Toth, K. and Aravin, A. A. (2016). Cutoff suppresses RNA Polymerase II termination to ensure expression of piRNA precursors. Mol Cell [Epub ahead of print]. PubMed ID: 27292797
Small non-coding RNAs called piRNAs serve as guides for an adaptable immune system that represses transposable elements in germ cells of Metazoa. In Drosophila the RDC complex, composed of Rhino, Deadlock and Cutoff (Cuff) bind chromatin of dual-strand piRNA clusters, special genomic regions, which encode piRNA precursors. The RDC complex is required for transcription of piRNA precursors, though the mechanism by which it licenses transcription remained unknown. This study shows that Cuff prevents premature termination of RNA polymerase II. Cuff prevents cleavage of nascent RNA at poly(A) sites by interfering with recruitment of the cleavage and polyadenylation specificity factor (CPSF) complex. Cuff also protects processed transcripts from degradation by the exonuclease Rat1. This work reveals a conceptually different mechanism of transcriptional enhancement. In contrast to other factors that regulate termination by binding to specific signals on nascent RNA, the RDC complex inhibits termination in a chromatin-dependent and sequence-independent manner.

Tripathi, B. K., Surabhi, S., Bhaskar, P. K., Mukherjee, A. and Mutsuddi, M. (2016). The RNA binding KH domain of Spoonbill depletes pathogenic non-coding spinocerebellar ataxia 8 transcripts and suppresses neurodegeneration in Drosophila. Biochim Biophys Acta 1862: 1732-1741. PubMed ID: 27302466
Spinocerebellar ataxia 8 (SCA8) pathogenesis is a resultant of gain-of-function machinery that primarily results at the RNA level. It has been reported that expanded non-coding CTG trinucleotide repeat in the ATXN8OS transcripts leads to SCA8 coupled neurodegeneration. Targeted depletion of pathogenic SCA8 transcripts is a viable therapeutic approach. This report focused on the suppression of toxic RNA gain-of-function associated with SCA8. Suppression of SCA8 associated neurodegeneration by KH RNA binding domain of Spoonbill is reported. KH domain suppresses pathogenic SCA8 associated phenotype in adult flies. Ectopic expression of KH domain leads to massive reduction in the number and size of SCA8 RNA foci. Spoonbill interacts with toxic SCA8 transcripts via its KH domain and promotes its depletion. To date, no attempts have been made for therapeutic intervention of SCA8 pathogenesis. Further characterization of Spoonbill KH domain may aid in designing peptide based therapeutics for SCA8 associated neurodegeneration.

Saturday, July 2nd

Jin, M., Eblimit, A., Pulikkathara, M., Corr, S., Chen, R. and Mardon, G. (2016). Conditional knockout of retinal determination genes in differentiating cells in Drosophila. FEBS J [Epub ahead of print]. PubMed ID: 27257739
Conditional gene knockout in post-mitotic cells is a valuable technique which allows the study of gene function with spatiotemporal control. Surprisingly, in contrast to its long-term and extensive use in mouse studies, this technology is lacking in Drosophila. This study used a novel method for generating complete loss of eyes absent (eya) or sine oculis (so) function in post-mitotic cells posterior to the morphogenetic furrow (MF). Specifically, genomic rescue constructs with flippase recognition target (FRT) sequences flanking essential exons are used to generate conditional null alleles. By removing gene function in differentiating cells, it was shown that eya and so are dispensable for larval photoreceptor differentiation, but are required for differentiation during pupal development. Both eya and so are necessary for photoreceptor survival and the apoptosis caused by loss of eya or so function is likely a secondary consequence of inappropriate differentiation. Their requirement for cone cell development was confirmed and a novel role in interommatidial bristle (IOB) formation was revealed. In addition, so is required for normal eye disc morphology. This is the first report of a knockout method to study eya and so function in post-mitotic cells. This technology will open the door to a large array of new functional studies in virtually any tissue and at any stage of development or in adults.

Misra, M., Edmund, H., Ennis, D., Schlueter, M. A., Marot, J. E., Tambasco, J., Barlow, I., Sigurbjornsdottir, S., Mathew, R., Valles, A. M., Davis, I., Leptin, M. and Gavis, E. R. (2016). A genome-wide screen for dendritically localized RNAs identifies genes required for dendrite morphogenesis. G3 (Bethesda) [Epub ahead of print]. PubMed ID: 27260999
Localizing messenger RNAs at specific subcellular sites is a conserved mechanism for targeting the synthesis of cytoplasmic proteins to distinct subcellular domains, thereby generating asymmetric protein distributions necessary for cellular and developmental polarity. However, the full range of transcripts that are asymmetrically distributed in specialized cell types and the significance of their localization, especially in the nervous system, are not known. This study used the EP-MS2 method, which combines EP transposon insertion with the MS2/MCP in vivo fluorescent labeling system to screen for novel localized transcripts in polarized cells, focusing on the highly branched Drosophila class IV dendritic arborization neurons. Of a total of 541 lines screened, 55 EP-MS2 insertions were identified producing transcripts that were enriched in neuronal processes, particularly in dendrites. The 47 genes identified by these insertions encode molecularly diverse proteins and are enriched for genes that function in neuronal development and physiology. RNAi-mediated knockdown confirmed roles for many of the candidate genes in dendrite morphogenesis. It is proposed that the transport of mRNAs encoded by these genes into the dendrites allows their expression to be regulated on a local scale during the dynamic developmental processes of dendrite outgrowth, branching, and/or remodeling.

Yang, H. H., St-Pierre, F., Sun, X., Ding, X., Lin, M. Z. and Clandinin, T. R. (2016). Subcellular imaging of voltage and calcium signals reveals neural processing in vivo. Cell [Epub ahead of print]. PubMed ID: 27264607
A mechanistic understanding of neural computation requires determining how information is processed as it passes through neurons and across synapses. However, it has been challenging to measure membrane potential changes in axons and dendrites in vivo. This study used in vivo, two-photon imaging of novel genetically encoded voltage indicators, as well as calcium imaging, to measure sensory stimulus-evoked signals in the Drosophila visual system with subcellular resolution. Across synapses, major transformations were found in the kinetics, amplitude, and sign of voltage responses to light. Distinct relationships were described between voltage and calcium signals in different neuronal compartments, a substrate for local computation. Finally, it was demonstrated that ON and OFF selectivity, a key feature of visual processing across species, emerges through the transformation of membrane potential into intracellular calcium concentration. By imaging voltage and calcium signals to map information flow with subcellular resolution, this study illuminates where and how critical computations arise.

Soldano, A., Alpizar, Y. A., Boonen, B., Franco, L., Lopez-Requena, A., Liu, G., Mora, N., Yaksi, E., Voets, T., Vennekens, R., Hassan, B. A. and Talavera, K. (2016). Gustatory-mediated avoidance of bacterial lipopolysaccharides via TRPA1 activation in Drosophila. Elife 5. PubMed ID: 27296646
Detecting pathogens and mounting immune responses upon infection is crucial for animal health. However, these responses come at a high metabolic price, and avoiding pathogens before infection may be advantageous. The bacterial endotoxins lipopolysaccharides (LPS) are important immune system infection cues, but it remains unknown whether animals possess sensory mechanisms to detect them prior to infection. This study shows that fruit flies display strong aversive responses to LPS and that gustatory neurons expressing Gr66a bitter receptors mediate avoidance of LPS in feeding and egg laying assays. The expression of the chemosensory cation channel dTRPA1 in these cells was found to be necessary and sufficient for LPS avoidance. Furthermore, LPS stimulates Drosophila neurons in a TRPA1-dependent manner and activates exogenous dTRPA1 channels in human cells. These findings demonstrate that flies detect bacterial endotoxins via a gustatory pathway through TRPA1 activation as conserved molecular mechanism.

Friday, July 1st

Kavi, H., Emelyanov, A. V., Fyodorov, D. V. and Skoultchi, A. I. (2016). Independent biological and biochemical functions for individual structural domains of Drosophila linker histone H1. J Biol Chem [Epub ahead of print]. PubMed ID: 27226620
Linker histone H1 is among the most abundant components of chromatin. H1 has profound effects on chromosome architecture. H1 also helps to tether DNA- and histone-modifying enzymes to chromatin. Metazoan linker histones have a conserved tri-partite structure comprising N-terminal, globular and long, unstructured C-terminal domains. Truncated Drosophila H1 polypeptides in vitro and H1 mutant transgenes in vivo were used to interrogate the roles of these domains in multiple biochemical and biological activities of H1. The globular domain and the proximal part of the C-terminal domain were shown to be essential for H1 deposition into chromosomes and for the stability of H1-chromatin binding. The two domains are also essential for fly viability and the establishment of a normal polytene chromosome structure. Additionally, through interaction with the heterochromatin-specific HMT Su(var)3-9, the H1 C-terminal domain makes important contributions to formation and H3K9 methylation of heterochromatin, as well as silencing of transposons in heterochromatin. Surprisingly, the N-terminal domain does not appear to be required for any of these functions. However, it is involved in formation of a single chromocenter in polytene chromosomes. In summary, this study has discovered that linker histone H1, similar to core histones, exerts its multiple biological functions through independent, biochemically separable activities of its individual structural domains.

Khoroshko, V. A., Levitsky, V. G., Zykova, T. Y., Antonenko, O. V., Belyaeva, E. S. and Zhimulev, I. F. (2016). Chromatin heterogeneity and distribution of regulatory elements in the late-replicating intercalary heterochromatin domains of Drosophila melanogaster chromosomes. PLoS One 11: e0157147. PubMed ID: 27300486
Late-replicating domains (intercalary heterochromatin) in the Drosophila genome are quite large and encompass clusters of functionally unrelated tissue-specific genes. They correspond to the topologically associating domains and conserved microsynteny blocks. This study has uncovered surprising heterogeneity of chromatin composition in these regions. Intercalary heterochromatin regions were found to host chromatin fragments with a particular epigenetic profile. Aquamarine chromatin fragments (spanning 0.67% of late-replicating regions) are heterogeneous in terms of their decompactization. These fragments are enriched with enhancer sequences and binding sites for insulator proteins. They likely mark the chromatin state that is related to the binding of cis-regulatory proteins. Malachite chromatin fragments (11% of late-replicating regions) appear to function as universal transitional regions between two contrasting chromatin states. Significant enrichment of insulator proteins CP190, SU(HW), and MOD2.2 was observed in malachite chromatin. Neither aquamarine nor malachite chromatin types appear to correlate with the positions of highly conserved non-coding elements (HCNE) that are typically replete in intercalary heterochromatin. Malachite chromatin found on the flanks of intercalary heterochromatin regions tends to replicate earlier than the malachite chromatin embedded in intercalary heterochromatin. The peculiar organization and features of replication in large late-replicating regions are discussed as possible factors shaping the evolutionary stability of intercalary heterochromatin.

Perez-Perri, J.I., Dengler, V.L., Audetat, K.A., Pandey, A., Bonner, E.A., Urh, M., Mendez, J., Daniels, D.L., Wappner, P., Galbraith, M.D. and Espinosa, J.M. (2016). The TIP60 complex is a conserved coactivator of HIF1A. Cell Rep [Epub ahead of print]. PubMed ID: 27320910
Hypoxia-inducible factors (HIFs) are critical regulators of the cellular response to hypoxia. Despite their established roles in normal physiology and numerous pathologies, the molecular mechanisms by which they control gene expression remain poorly understood. This study reports a conserved role for the TIP60 complex as a HIF1 transcriptional cofactor in Drosophila and human cells. TIP60 (KAT5) is required for HIF1-dependent gene expression in fly cells and embryos and colorectal cancer cells. HIF1A interacts with and recruits TIP60 to chromatin. TIP60 is dispensable for HIF1A association with its target genes but is required for HIF1A-dependent chromatin modification and RNA polymerase II activation in hypoxia. In human cells, global analysis of HIF1A-dependent gene activity reveals that most HIF1A targets require either TIP60, the CDK8-Mediator complex, or both as coactivators for full expression in hypoxia. Thus, HIF1A employs functionally diverse cofactors to regulate different subsets of genes within its transcriptional program.

Klebanow, L. R., Peshel, E. C., Schuster, A. T., De, K., Sarvepalli, K., Lemieux, M. E., Lenoir, J. J., Moore, A. W., McDonald, J. A. and Longworth, M. S. (2016). Drosophila Condensin II subunit, Chromosome Associated Protein-D3, regulates cell fate determination through non-cell autonomous signaling. Development [Epub ahead of print]. PubMed ID: 27317808
The pattern of the Drosophila melanogaster adult wing is heavily influenced by the expression of proteins that dictate cell fate decisions between intervein and vein during development. dSRF (Drosophila Serum Response Factor) expression in specific regions of the larval wing disc promotes intervein cell fate while EGFR (Epidermal Growth Factor Receptor) activity promotes vein cell fate. This study reports that the chromatin organizing protein, dCAP-D3, acts to dampen dSRF levels at the anterior/posterior (A/P) boundary in the larval wing disc, promoting differentiation of cells into the anterior crossvein (ACV). dCAP-D3 represses Knot expression in cells immediately adjacent to the A/P boundary, thus blocking Knot-mediated repression of EGFR activity and preventing cell death. Maintenance of EGFR activity in these cells depresses dSRF levels in the neighboring ACV progenitor cells, allowing them to differentiate into vein cells. These findings uncover a novel transcriptional regulatory network influencing Drosophila wing vein development, and are the first to identify a Condensin II subunit as an important regulator of EGFR activity and cell fate determination in vivo.
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