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


Wednesday, April 30th, 2015

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Head, L.M., et al. (2015). The influence of light on temperature preference in Drosophila. Curr Biol [Epub ahead of print]. PubMed ID: 25866391
Ambient light affects multiple physiological functions and behaviors, such as circadian rhythms, sleep-wake activities, and development, from flies to mammals. Mammals exhibit a higher body temperature when exposed to acute light compared to when they are exposed to the dark, but the underlying mechanisms are largely unknown. The body temperature of small ectotherms, such as Drosophila, relies on the temperature of their surrounding environment, and these animals exhibit a robust temperature preference behavior. This study demonstrates that Drosophila prefer a ~1 ° higher temperature when exposed to acute light rather than the dark. This acute light response, light-dependent temperature preference (LDTP), was observed regardless of the time of day, suggesting that LDTP was regulated separately from the circadian clock. However, screening of eye and circadian clock mutants suggested that the circadian clock neurons posterior dorsal neurons 1 (DN1ps) and Pigment-Dispersing Factor Receptor (PDFR) play a role in LDTP. To further investigate the role of DN1ps in LDTP, PDFR in DN1ps was knocked down, resulting in an abnormal LDTP. The phenotype of the pdfr mutant was rescued sufficiently by expressing PDFR in DN1ps, indicating that PDFR in DN1ps was responsible for LDTP. These results suggest that light positively influences temperature preference via the circadian clock neurons, DN1ps, which may result from the integration of light and temperature information. Given that both Drosophila and mammals respond to acute light by increasing their body temperature, the effect of acute light on temperature regulation may be conserved evolutionarily between flies and humans

Doll, C. A. and Broadie, K. (2015). Activity-dependent FMRP requirements in development of the neural circuitry of learning and memory. Development 142: 1346-1356. PubMed ID: 25804740
The activity-dependent refinement of neural circuit connectivity during critical periods of brain development is essential for optimized behavioral performance. It was hypothesized that this mechanism is defective in fragile X syndrome (FXS), the leading heritable cause of intellectual disability and autism spectrum disorders. This study used optogenetic tools in the Drosophila FXS disease model to test activity-dependent dendritogenesis in two extrinsic neurons of the mushroom body (MB) learning and memory brain center: (1) the input projection neuron (PN) innervating Kenyon cells (KCs) in the MB calyx microglomeruli and (2) the output MVP2 neuron innervated by KCs in the MB peduncle. Both input and output neuron classes exhibit distinctive activity-dependent critical period dendritic remodeling. MVP2 arbors expand in Drosophila mutants null for fragile X mental retardation 1 (dfmr1), as well as following channelrhodopsin-driven depolarization during critical period development, but are reduced by halorhodopsin-driven hyperpolarization. Optogenetic manipulation of PNs causes the opposite outcome - reduced dendritic arbors following channelrhodopsin depolarization and expanded arbors following halorhodopsin hyperpolarization during development. Importantly, activity-dependent dendritogenesis in both neuron classes absolutely requires dfmr1 during one developmental window. These results show that dfmr1 acts in a neuron type-specific activity-dependent manner for sculpting dendritic arbors during early-use, critical period development of learning and memory circuitry in the Drosophila brain.

Lee, Y.M. and Sun, Y.H. (2015). Maintenance of glia in the optic lamina is mediated by EGFR signaling by photoreceptors in adult Drosophila. PLoS Genet 11: e1005187. PubMed ID: 25909451
The late onset of neurodegeneration in humans indicates that the survival and function of cells in the nervous system must be maintained throughout adulthood. In the optic lamina of the adult Drosophila, the photoreceptor axons are surrounded by multiple types of glia. This study demonstrated that the adult photoreceptors actively contribute to glia maintenance in their target field within the optic lamina. This effect was dependent on the epidermal growth factor receptor (EGFR) ligands produced by the R1-6 photoreceptors and transported to the optic lamina to act on EGFR in the lamina glia. EGFR signaling was necessary and sufficient to act in a cell-autonomous manner in the lamina glia. These results suggest that EGFR signaling is required for the trafficking of the autophagosome/endosome to the lysosome. The loss of EGFR signaling resulted in cell degeneration most likely because of the accumulation of autophagosomes. These findings provide in vivo evidence for the role of adult neurons in the maintenance of glia and a novel role for EGFR signaling in the autophagic flux.

Frickenhaus, M., Wagner, M., Mallik, M., Catinozzi, M. and Storkebaum, E. (2015). Cell-type-specific gene inactivation reveals a key function for the Drosophila FUS homolog cabeza in neurons. Sci Rep 5: 9107. PubMed ID: 25772687
This study evaluated whether introducing FRT or LoxP sites in endogenous genes could allow for cell-type-specific gene inactivation in both dividing and postmitotic cells by GAL4-driven expression of FLP or Cre recombinase. For proof of principle, conditional alleles were generated for cabeza (caz), the Drosophila homolog of human FUS, a gene implicated in the neurodegenerative disorders amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Upon selective expression in neurons or muscle, both FLP and Cre mediated caz inactivation in all neurons or muscle cells, respectively. Neuron-selective caz inactivation resulted in failure of pharate adult flies to eclose from the pupal case, and adult escapers displayed motor performance defects and reduced life span. Due to Cre-toxicity, FLP/FRT is the preferred system for cell-type-specific gene inactivation, and this strategy outperforms RNAi-mediated knock-down. Furthermore, the GAL80 target system allowed for temporal control over gene inactivation, as induction of FLP expression from the adult stage onwards still inactivated caz in >99% of neurons. Remarkably, selective caz inactivation in adult neurons did not affect motor performance and life span, indicating that neuronal caz is required during development, but not for maintenance of adult neuronal function.

Wednesday, April 29th

Dubruille, R. and Loppin, B. (2015). Protection of #Drosophila chromosome ends with minimal #telomere capping. J Cell Sci [Epub ahead of print]. PubMed ID: 25908850
In Drosophila, telomere capping proteins have the remarkable capacity to recognize chromosome ends in a sequence-independent manner. This epigenetic protection is essential to prevent catastrophic ligations of chromosome extremities. Interestingly, capping proteins occupy a large telomere chromatin domain of several kilobases whose functional relevance to end protection is unknown. This study investigated the role of the large capping domain by manipulating HOAP capping protein expression in the male germ cells where telomere protection could be challenged without compromising viability. The exhaustion of HOAP resulted in a dramatic reduction of other capping proteins at telomeres, including K81, which is essential for male fertility. Strikingly however, although capping complexes were barely detected in HOAP-depleted male germ cells, telomere protection and male fertility were not dramatically affected. This study thus demonstrates that efficient protection of Drosophila telomeres can be achieved with surprisingly low amounts of capping complexes. It is proposed that these complexes prevent fusions by acting at the very extremity of chromosomes, reminiscent of the protection conferred by extremely short telomeric arrays in yeast or mammalian systems.

Guo, J. and Jin, D. (2015). A genetic screen in #Drosophila implicates Sex comb on midleg (Scm) in tissue overgrowth and mechanisms of Scm degradation by Wds. Mech Dev 136: 1-7. PubMed ID: 25772304
The sex comb on midleg (scm) gene encodes a transcriptional repressor and belongs to the Polycomb group (PcG) of genes, which regulates growth in Drosophila. Scm interacts with Polyhomeotic (a PcG protein) in vitro by recognizing its SPM domain. The homologous human protein, Sex comb on midleg-like 2 (Scml2), has been implicated in malignant brain tumors. Will die slowly (Wds) is another factor that regulates Drosophila development, and its homologous human protein, WD repeat domain 5(Wdr5), is part of the mixed lineage leukemia 1(MLL1) complex that promotes histone H3Lys4 methylation. Like Scml2, Wdr5 has been implicated in certain cancers; this protein plays an important role in leukemogenesis. This study finds that loss-of-function mutations in Scm result in non-autonomous tissue overgrowth in Drosophila, and determines that Scm is essential for ommatidium development and important for cell survival in Drosophila. Furthermore, this research suggests a relationship between Wds and Scm; Wds promotes Scm degradation through ubiquitination in vitro in Drosophila.

Rao, S. G., et al. (2015). Invadolysin acts genetically via the SAGA complex to modulate chromosome structure in #Drosophila. Nucleic Acids Res [Epub ahead of print]. PubMed ID: 25779050
Identification of components essential to chromosome structure and behaviour remains a vibrant area of study. Invadolysin has been shown to be essential in Drosophila, with roles in cell division and cell migration. Mitotic chromosomes are hypercondensed in length, but display an aberrant fuzzy appearance. It was additionally demonstrated that in human cells, invadolysin is localized on the surface of lipid droplets, organelles that store not only triglycerides and sterols but also free histones H2A, H2Av and H2B. Is there a link between the storage of histones in lipid droplets and the aberrantly structured chromosomes of invadolysin mutants? This study has identified a genetic interaction between invadolysin and Nonstop, the de-ubiquitinating protease component of the SAGA (Spt-Ada-Gcn5-acetyltransferase) chromatin-remodelling complex. invadolysin and nonstop mutants exhibit phenotypic similarities in terms of chromosome structure in both diploid and polyploid cells. Furthermore, IX-141/not1 transheterozygous animals accumulate mono-ubiquitinated histone H2B (ubH2B) and histone H3 tri-methylated at lysine 4 (H3K4me3). Whole mount immunostaining of IX-141/not1 transheterozygous salivary glands revealed that ubH2B accumulates surprisingly in the cytoplasm, rather than the nucleus. Over-expression of the Bre1 ubiquitin ligase phenocopies the effects of mutating either the invadolysin or nonstop genes. Intriguingly, nonstop and mutants of other SAGA subunits (gcn5, ada2b and sgf11) all suppress an invadolysin-induced rough eye phenotype. It is concluded that the abnormal chromosome phenotype of invadolysin mutants is likely the result of disrupting the histone modification cycle. It is thus proposed that Invadolysin plays a crucial role in chromosome organization via its interaction with the SAGA complex.

Eisert, R. J., Kennedy, S. A. and Waters, M. L. (2015). Investigation of the β-sheet interactions between #Drosophila HP1 chromodomain and Histone 3. Biochemistry [Epub ahead of print]. PubMed ID: 25790265
Methylated lysine 9 on the histone 3 (H3) tail recruits Heterochromatin protein 1 from Drosophila (dHP1) via its chromodomain and results in gene silencing. The dHP1 chromodomain binds H3 K9Me3 with an aromatic cage surrounding the trimethyllysine. The sequence selectivity of binding comes from insertion of the histone tail between two beta-strands of the chromodomain to form a three-stranded beta-sheet. This study investigated the sequence selectivity provided by the beta-sheet interactions and how those interactions compare to other model systems. Residue Thr6 of the histone tail forms cross-strand interactions with Ala25 and Asp62 of the chromodomain. Each of these three residues was substituted for amino acids known to have high beta-sheet propensities and/or to form favorable side chain-side chain (SC-SC) interactions in beta-sheets, including hydrophobic, H-bonding, and aromatic interactions. About 50% of the chromodomain mutants resulted in equal or tighter binding to the histone tail and about 25% of the histone tail mutants provided tighter binding compared to that of the native histone tail sequence. These studies provide novel insights into the sequence selectivity of the dHP1 chromodomain for the histone tail and relates the information gleaned from model systems and statistical studies to beta-sheet-mediated protein-protein interactions. Moreover, this work suggests that the development of designer histone-chromodomain pairs for chemical biology applications is feasible.

Tuesday, April 28th

Kovacs, L., et al., (2015). Role of the deubiquitylating enzyme DmUsp5 in coupling #ubiquitin equilibrium to development and #apoptosis in #Drosophila melanogaster. PLoS One 10: e0120875. PubMed ID: 25806519
Protein ubiquitylation is regulated through the balanced action of E3 ubiquitin ligases and deubiquitylating enzymes (DUB) which conjugate ubiquitins to, and remove them from target proteins, respectively. Genetic analysis has revealed that the deubiquitylating enzyme Ubiquitin specific protease 5 (DmUsp5) is required for maintenance of the ubiquitin equilibrium, cell survival and normal development in Drosophila. Loss of the DmUsp5 function leads to late larval lethality accompanied by the induction of apoptosis. Detailed analyses at a cellular level demonstrated that DmUsp5 mutants carry multiple abnormalities, including a drop in the free monoubiquitin level, the excessive accumulation of free polyubiquitins, polyubiquitylated proteins and subunits of the 26S proteasome. A shortage in free ubiquitins results in the induction of a ubiquitin stress response previously described only in the unicellular budding yeast. It is characterized by the induction of the proteasome-associated deubiquitylase DmUsp14 and sensitivity to cycloheximide. Removal of DmUsp5 also activates the pro-apoptotic machinery thereby resulting in widespread apoptosis, indicative of an anti-apoptotic role of DmUsp5. Collectively, the pleiotropic effects of a loss of DmUsp5 function can be explained in terms of the existence of a limited pool of free monoubiquitins which makes the ubiquitin-dependent processes mutually interdependent.

Querenet, M., Goubard, V., Chatelain, G., Davoust, N. and Mollereau, B. (2015). Spen is required for pigment cell survival during pupal development in #Drosophila. Dev Biol [Epub ahead of print]. PubMed ID: 25872184
Apoptosis is required during development to eliminate superfluous cells and sculpt tissues; spatial and timed control of apoptosis ensuring that the necessary number of cells is eliminated at a precise time in a given tissue. The elimination of supernumerary pigment or inter-ommatidial cells (IOCs) depends on cell-cell communication and is necessary for the formation of the honeycomb-like structure of the Drosophila eye. However, the mechanisms occurring during pupal development and controlling apoptosis of superfluous IOC in space and time remain unclear. This study found that split-ends (spen) is required for IOC survival at the time of removal of superfluous IOCs. Loss of spen function led to abnormal removal of IOCs by apoptosis. spen was required non-autonomously in cone cells for the survival of IOCs by positively regulating the Spitz/EGFR pathway. The study proposes that Spen is an important survival factor that ensures spatial control of the apoptotic wave that is necessary for the correct patterning and formation of the Drosophila eye.

Xu, T., Nicolson, S., Denton, D. and Kumar, S. (2015). Distinct requirements of #autophagy-related genes in programmed cell death in #Drosophila. Cell Death Differ [Epub ahead of print]. PubMed ID: 25882046
Although most programmed cell death (PCD) during animal development occurs by caspase-dependent apoptosis, autophagy-dependent cell death is also important in specific contexts. Previous studies established that PCD of the obsolete Drosophila larval midgut tissue is dependent on autophagy and can occur in the absence of the main components of the apoptotic pathway. As autophagy is primarily a survival mechanism in response to stress such as starvation, it is currently unclear if the regulation and mechanism of autophagy as a pro-death pathway is distinct to that as pro-survival. To establish the requirement of the components of the autophagy pathway during cell death, this study examined the effect of systematically knocking down components of the autophagy machinery on autophagy induction and timing of midgut PCD. The study found that there was a distinct requirement of the individual components of the autophagy pathway in a pro-death context. Furthermore, TORC1 was upstream of autophagy induction in the midgut indicating that while the machinery may be distinct the activation may occur similarly in PCD and during starvation-induced autophagy signalling. These data reveal that while autophagy initiation occurs similarly in different cellular contexts, there is a tissue/function-specific requirement for the components of the autophagic machinery.

Zimmermann, M., Kugler, S.J., Schulz, A. and Nagel, A.C. (2015). Loss of putzig activity results in #apoptosis during wing imaginal development in #Drosophila. PLoS One 10: e0124652. PubMed ID: 25894556
The Drosophila gene putzig (pzg) encodes a nuclear protein that is an integral component of the Trf2/Dref complex involved in the transcription of proliferation-related genes. Moreover, Pzg is found in a complex together with the nucleosome remodeling factor NURF, where it promotes Notch target gene activation. This study shows that downregulation of pzg activity in the developing wing imaginal discs induces an apoptotic response, accompanied by the induction of the pro-apoptotic gene reaper, repression of Drosophila inhibiΩ of apoptosis protein accumulation and the activation of the caspases Drice, Caspase3 and Dcp1. As a further consequence 'Apoptosis induced Proliferation' (AiP) and 'Apoptosis induced Apoptosis' (AiA) were triggered. As expected, the activity of the stress kinase Jun N-terminal kinase (JNK), proposed to mediate both processes, was ectopically induced in response to pzg loss. In addition, the expression of the mitogen wingless (wg) but not of decapentaplegic (dpp) was observed. Downregulation of Notch activated Dcp1 caspase and JNK signaling, however, neither induced ectopic wg nor dpp expression. In contrast, the consequences of Dref-RNAi were largely indistinguishable from pzg-RNAi with regard to apoptosis induction. Moreover, overexpression of Dref ameliorated the downregulation of pzg compatible with the notion that the two were required together to maintain cell and tissue homeostasis in Drosophila.

Monday, April 27th

Zhang, S., Chen, C., Wu, C., Yang, Y., Li, W. and Xue, L. (2015). The canonical #Wingless signaling modulates Basket-mediated cell death in #Drosophila. Cell Death Dis 6: e1713. PubMed ID: 25855961
Cell death is an essential regulatory mechanism for removing unneeded cells in animal development and tissue homeostasis. The c-Jun N-terminal kinase (JNK) pathway has pivotal roles in the regulation of cell death in response to various intrinsic and extrinsic stress signals. The canonical Wingless (Wg) signaling has been implicated in cell proliferation and cell fate decisions, whereas its role in cell death remains largely elusive. This study reports that activated Bsk (the Drosophila JNK homolog) induced cell death is mediated by the canonical Wg signaling. First, loss of Wg signaling abrogates Bsk-mediated caspase-independent cell death. Second, activation of Wg signaling promotes cell death in a caspase-independent manner. Third, activation of Bsk signaling results in upregulated transcription of wingless (wg) gene. Finally, Wg pathway participates in the physiological function of Bsk signaling in development. These findings not only reveal a previously undiscovered role of Wg signaling in Bsk-mediated cell death, but also provide a novel mechanism for the interplay between the two important signaling pathways in development.

Jambor, H., Surendranath, V., Kalinka, A. T., Mejstrik, P., Saalfeld, S. and Tomancak, P. (2015). Systematic imaging reveals features and changing localization of mRNAs in #Drosophila development. Elife 4 [Epub ahead of print]. PubMed ID: 25838129
mRNA localization is critical for eukaryotic cells and affects numerous transcripts, yet how cells regulate distribution of many mRNAs to their subcellular destinations is still unknown. This study combined transcriptomics and systematic imaging to determine the tissue-specific expression and subcellular distribution of 5862 mRNAs during Drosophila oogenesis. mRNA localization is widespread in the ovary and detectable in all of its cell types-the somatic epithelial, the nurse cells, and the oocyte. Genes defined by a common RNA localization share distinct gene features and differ in expression level, 3'UTR length and sequence conservation from unlocalized mRNAs. Comparison of mRNA localizations in different contexts revealed that localization of individual mRNAs changes over time in the oocyte and between ovarian and embryonic cell types. This genome scale image-based resource (Dresden Ovary Table, DOT) enables the transition from mechanistic dissection of singular mRNA localization events towards global understanding of how mRNAs transcribed in the nucleus distribute in cells.

Chittaranjan, S., et al. (2015). The #Drosophila TIPE family member Sigmar interacts with the Ste20-like kinase Misshapen and modulates #JNK-signaling, cytoskeletal remodeling and #autophagy. Biol Open [Epub ahead of print]. PubMed ID: 25836674
TNFAIP8 and other mammalian TIPE family proteins have attracted increased interest due to their associations with disease-related processes including oncogenic transformation, metastasis, and inflammation. The molecular and cellular functions of TIPE family proteins are still not well understood. This study reports the molecular and genetic characterization of the Drosophila TNFAIP8 homolog, CG4091/sigmar. Previous gene expression studies revealed dynamic expression of sigmar in larval salivary glands prior to histolysis. In sigmar loss-of-function mutants, the salivary glands are morphologically abnormal with defects in the tubulin network and decreased autophagic flux. Sigmar localizes subcellularly to microtubule-containing projections in Drosophila S2 cells, and co-immunoprecipitates with the Ste20-like kinase Misshapen, a regulator of the JNK pathway. Further, the Drosophila TNF ligand Eiger can induce sigmar expression, and sigmar loss-of-function leads to altered localization of pDJNK in salivary glands. Together, these findings link Sigmar to the JNK pathway, cytoskeletal remodeling and autophagy activity, and provide new insights into TIPE family member function.

Kim, W., Kim, H. D., Jung, Y., Kim, J. and Chung, J. (2015). #Drosophila LTV1 (Low Temperature Viability Protein 1) is required for #ribosome biogenesis and cell growth downstream of dMyc. J Biol Chem [Epub ahead of print]. PubMed ID: 25858587
During animal development, various signaling pathways converge to regulate cell growth. This study identified LTV1 as a novel cell growth regulator in Drosophila. LTV1 mutant larvae exhibit developmental delays and lethality at the second larval stage. Using biochemical studies, it was discovered that LTV1 interacts with ribosomal protein S3 (RpS3) and co-purifies with free 40S ribosome subunits. It was further demonstrated that LTV1 is crucial for ribosome biogenesis through 40S ribosome subunit synthesis and pre-ribosomal RNA processing, suggesting that LTV1 is required for cell growth by regulating protein synthesis. It was also demonstrated that Drosophila Myc (dMyc) directly regulates LTV1 transcription and requires LTV1 to stimulate ribosome biogenesis. Importantly, the loss of LTV1 blocks the cell growth and endoreplication induced by dMyc. Combined, these results suggest that LTV1 is a key downstream factor of dMyc-induced cell growth by properly maintaining ribosome biogenesis.

Sunday, April 26th

Schlichting, M., Grebler, R., Menegazzi, P. and Helfrich-Forster, C. (2015). Twilight dominates over moonlight in adjusting #Drosophila activity pattern. J Biol Rhythms 30: 117-128. PubMed ID: 25838418
Light is the most important zeitgeber for the synchronization of the Drosophila melanogaster circadian clock. In nature, there is twilight, and the nights are rarely completely dark, a fact that is usually disregarded in lab experiments. Recent studies showed contrary effects of simulated twilight and moonlight on fly locomotor activity, with twilight shifting morning and evening activity into the day and moonlight shifting it into the night. A currently unanswered question is, what may happen to locomotor activity when flies are exposed to more natural conditions in which both moonlight and twilight are simulated? The data demonstrate that flies are able to integrate twilight and moonlight. However, twilight seems to dominate over moonlight as both, morning and evening activity peaks, take place at dawn or at dusk, respectively, and not during the night. Furthermore, nocturnal activity decreases in the presence of twilight. The compound eyes are essential for this behavior, and by investigating different photoreceptor mutants, this study unraveled the importance of photoreceptor cells 7 and 8 for wild-type phases of the activity peaks. To adjust nocturnal activity levels to a wild-type manner, all photoreceptor cells work together in a complex way, with rhodopsin 6 having a prominent role.

Klichko, V. I., Chow, E. S., Kotwica-Rolinska, J., Orr, W. C., Giebultowicz, J. M. and Radyuk, S. N. (2015). #Aging alters circadian regulation of redox in #Drosophila. Front Genet 6: 83. PubMed ID: 25806044
Circadian coordination of metabolism, physiology, and neural functions contributes to healthy aging and disease prevention. Clock genes govern the daily rhythmic expression of target genes whose activities underlie such broad physiological parameters as maintenance of redox homeostasis. It has been reported that glutathione (GSH) biosynthesis is controlled by the circadian system via effects of the clock genes on expression of the catalytic (Gclc) and modulatory (Gclm) subunits comprising the glutamate cysteine ligase (GCL) holoenzyme. The objective of this study was to determine whether and how aging, which leads to weakened circadian oscillations, affects the daily profiles of redox-active biomolecules. Fly aging was found to be associated with altered profiles of Gclc and Gclm expression at both the mRNA and protein levels. Analysis of free aminothiols and GCL activity revealed that aging abolishes daily oscillations in GSH levels and alters the activity of glutathione biosynthetic pathways. Unlike GSH, its precursors and products of catabolism, methionine, cysteine and cysteinyl-glycine, were not rhythmic in young or old flies, while rhythms of the glutathione oxidation product, GSSG, were detectable. It is concluded that the temporal regulation of GSH biosynthesis is altered in the aging organism and that age-related loss of circadian modulation of pathways involved in glutathione production is likely to impair temporal redox homeostasis.

Garcia-Gonzalez, F. and Dowling, D. K. (2015). Transgenerational effects of sexual interactions and sexual conflict: non-sires boost the fecundity of #Drosophila females in the following generation. Biol Lett 11 [Epub ahead of print]. PubMed ID: 25788486
The consequences of sexual interactions extend beyond the simple production of offspring. These interactions typically entail direct effects on female fitness, but may also impact the life histories of later generations. Evaluating the cross-generational effects of sexual interactions provides insights into the dynamics of sexual selection and conflict. Such studies can elucidate whether offspring fitness optima diverge across sexes upon heightened levels of sexual interaction among parents. This study found that, in Drosophila melanogaster, components of reproductive success in females, but not males, were contingent on the nature of sexual interactions experienced by their mothers. In particular, maternal sexual interactions with non-sires enhanced female fecundity in the following generation. This highlights the importance of non-sire influences of sexual interactions on the expression of offspring life histories.

LeDue, E. E., Chen, Y. C., Jung, A. Y., Dahanukar, A. and Gordon, M. D. (2015). Pharyngeal sense organs drive robust sugar consumption in #Drosophila. Nat Commun 6: 6667. PubMed ID: 25807033
The fly pharyngeal sense organs lie at the transition between external and internal nutrient-sensing mechanisms. This study investigated the function of pharyngeal sweet gustatory receptor neurons, demonstrating that they express a subset of the nine previously identified sweet receptors and respond to stimulation with a panel of sweet compounds. pox-neuro (poxn) mutants lacking taste function in the legs and labial palps have intact pharyngeal sweet taste, which is both necessary and sufficient to drive preferred consumption of sweet compounds by prolonging ingestion. Moreover, flies putatively lacking all sweet taste show little preference for nutritive or non-nutritive sugars in a short-term feeding assay. Together, these data demonstrate that pharyngeal sense organs play an important role in directing sustained consumption of sweet compounds, and suggest that post-ingestive sugar sensing does not effectively drive food choice in a simple short-term feeding paradigm.

Saturday, April 25th

Kanke, M., Jambor, H., Reich, J., Marches, B., Gstir, R., Ryu, Y.H., Ephrussi, A. and Macdonald, P.M. (2015). oskarRNA plays multiple noncoding roles to support oogenesis and maintain integrity of the germline/soma distinction. RNA [Epub ahead of print]. PubMed ID: 25862242
The Drosophila oskar (osk) mRNA is unusual in that it has both coding and noncoding functions. As an mRNA, osk encodes a protein required for embryonic patterning and germ cell formation. Independent of that function, the absence of osk mRNA disrupts formation of the karyosome and blocks progression through oogenesis. This study shows that loss of osk mRNA also affects the distribution of regulatory proteins, relaxing their association with large RNPs within the germline, and allowing them to accumulate in the somatic follicle cells. This and other noncoding functions of the osk mRNA are mediated by multiple sequence elements with distinct roles. One role, provided by numerous binding sites in two distinct regions of the osk 3' UTR, was to sequester the translational regulator Bruno (Bru), which itself controlled translation of osk mRNA. This defined a novel regulatory circuit, with Bru restricting the activity of osk, and osk in turn restricting the activity of Bru. Other functional elements, which did not bind Bru and were positioned close to the 3' end of the RNA, acted in the oocyte and were essential. Despite the different roles played by the different types of elements contributing to RNA function, mutation of any led to accumulation of the germline regulatory factors in the follicle cells.

Simmons, M. J., Peterson, M. P., Thorp, M. W., Buschette, J. T., DiPrima, S. N., Harter, C. L. and Skolnick, M. J. (2015). piRNA-mediated #transposon regulation and the germ-line mutation rate in #Drosophila melanogaster males. Mutat Res 773: 16-21. PubMed ID: 25769182
Transposons, especially retrotransposons, are abundant in the genome of Drosophila melanogaster. These mobile elements are regulated by small RNAs that interact with the Piwi family of proteins-the piwi-interacting or piRNAs. The Piwi proteins are encoded by the genes argonaute3 (ago3), aubergine (aub), and piwi. Heterochromatin Protein 1 (HP1), a chromatin-organizing protein encoded by the Suppressor of variegation 205 [Su(var)205] gene, also plays a role in this regulation. To assess the mutational impact of weakening the system for transposon regulation, this study measured the frequency of recessive X-linked lethal mutations occurring in the germ lines of males from stocks that were heterozygous for mutant alleles of the ago3, aub, piwi, or Su(var)205 genes. These mutant alleles are expected to deplete the wild-type proteins encoded by these genes by as much as 50%. The mutant alleles of piwi and Su(var)205 significantly increased the X-linked lethal mutation frequency, whereas the mutant alleles of ago3 did not. An increased mutation frequency was also observed in males from one of two mutant aub stocks, but this increase may not have been due to the aub mutant. The increased mutation frequency caused by depleting Piwi or HP1 suggests that chromatin-organizing proteins play important roles in minimizing the germ-line mutation rate, possibly by stabilizing the structure of the heterochromatin in which many transposons are situated.

Zhang Y1, Pu J1, Li L., Wang, X. Y., Zhang, Y., Pu, J., Li, L., Shao, W., Zhan, S., Hao, J. and Xu, Y. Z. (2015). A conserved intronic U1 snRNP-binding sequence promotes trans-splicing in #Drosophila. Genes Dev 29: 760-771. PubMed ID: 25838544
Unlike typical cis-splicing, trans-splicing joins exons from two separate transcripts to produce chimeric mRNA and has been detected in most eukaryotes. Trans-splicing in trypanosomes and nematodes has been characterized as a spliced leader RNA-facilitated reaction; in contrast, its mechanism in higher eukaryotes remains unclear. This study investigate mod(mdg4), a classic trans-spliced gene in Drosophila, and report that two critical RNA sequences in the middle of the last 5' intron, TSA and TSB, promote trans-splicing of mod(mdg4). In TSA, a 13-nucleotide (nt) core motif is conserved across Drosophila species and is essential and sufficient for trans-splicing, which binds U1 small nuclear RNP (snRNP) through strong base-pairing with U1 snRNA (see U1A snRNP ). In TSB, a conserved secondary structure acts as an enhancer. Deletions of TSA and TSB using the CRISPR/Cas9 system result in developmental defects in flies. Although it is not clear how the 5' intron finds the 3' introns, compensatory changes in U1 snRNA rescue trans-splicing of TSA mutants, demonstrating that U1 recruitment is critical to promote trans-splicing in vivo. Furthermore, TSA core-like motifs are found in many other trans-spliced Drosophila genes, including lola. These findings represent a novel mechanism of trans-splicing, in which RNA motifs in the 5' intron are sufficient to bring separate transcripts into close proximity to promote trans-splicing.

Waldron, J. A., Jones, C. I., Towler, B. P., Pashler, A. L., Grima, D. P., Hebbes, S., Crossman, S. H., Zabolotskaya, M. V. and Newbury, S. F. (2015).. Xrn1/Pacman affects #apoptosis and regulates expression of hid and reaper. Biol Open [Epub ahead of print]. PubMed ID: 25836675
Programmed cell death, or apoptosis, is a highly conserved cellular process that is crucial for tissue homeostasis under normal development as well as environmental stress. Misregulation of apoptosis is linked to many developmental defects and diseases such as tumour formation, autoimmune diseases and neurological disorders. This paper shows a novel role for the exoribonuclease Pacman/Xrn1 in regulating apoptosis. Using Drosophila wing imaginal discs as a model system, a null mutation in pacman was demonstrated to result in small imaginal discs as well as lethality during pupation. Mutant wing discs show an increase in the number of cells undergoing apoptosis, especially in the wing pouch area. Compensatory proliferation also occurs in these mutant discs, but this is insufficient to compensate for the concurrent increase in apoptosis. The phenotypic effects of the pacman null mutation are rescued by a deletion that removes one copy of each of the pro-apoptotic genes reaper, hid and grim, demonstrating that pacman acts through this pathway. The null pacman mutation also results in a significant increase in the expression of the pro-apoptotic mRNAs, hid and reaper, with this increase mostly occurring at the post-transcriptional level, suggesting that Pacman normally targets these mRNAs for degradation. These results uncover a novel function for the conserved exoribonuclease Pacman and suggest that this exoribonuclease is important in the regulation of apoptosis in other organisms.

Friday, April 24th

Lu, Q. and Adler, P. N. (2015). The diaphanous gene of #Drosophila interacts antagonistically with multiple wing hairs and plays a key role in wing hair morphogenesis. PLoS One 10: e0115623. PubMed ID: 25730111
The Drosophila wing is covered by an array of distally pointing hairs that has served as a key model system for studying planar cell polarity (PCP). The adult cuticular hairs are formed in the pupae from cell extensions that contain extensive actin filaments and microtubules. The importance of the actin cytoskeleton for hair growth and morphogenesis is clear from the wide range of phenotypes seen in mutations in well-known actin regulators. Formin proteins promote the formation of long actin filaments of the sort thought to be important for hair growth. This study reports that the formin encoding diaphanous (dia) gene plays a key role in hair morphogenesis. Both loss of function mutations and the expression of a constitutively active Dia leads to cells forming both morphologically abnormal hairs and multiple hairs. The conserved frizzled (fz)/starry night (stan) PCP pathway functions to restrict hair initiation and activation of the cytoskeleton to the distal most part of wing cells. It also ensures the formation of a single hair per cell. These data suggest that the localized inhibition of Dia activity may be part of this mechanism. The expression of constitutively active Dia was found to greatly expand the region for activation of the cytoskeleton, and dia was found to function antagonistically with multiple wing hairs (mwh), the most downstream member of the fz/stan pathway. Further this study established that purified fragments of Dia and Mwh could be co-immunoprecipitated suggesting the genetic interaction could reflect a direct physical interaction.

Grode, K.D. and Rogers, S.L. (2015). The non-catalytic domains of #Drosophila #katanin regulate its abundance and microtubule-disassembly activity. PLoS One 10: e0123912. PubMed ID: 25886649
Microtubule severing is a biochemical reaction that generates an internal break in a microtubule and regulation of microtubule severing is critical for cellular processes such as ciliogenesis, morphogenesis, and meiosis and mitosis. Katanin is a conserved heterodimeric ATPase that severs and disassembles microtubules, but the molecular determinants for regulation of microtubule severing by katanin remain poorly defined. This study shows that the non-catalytic domains of Drosophila katanin regulate its abundance and activity in living cells. The microtubule-interacting and trafficking (MIT) domain and adjacent linker region of the Drosophila katanin catalytic subunit Kat60 cooperates to regulate microtubule severing in two distinct ways. First, the MIT domain and linker region of Kat60 decreases its abundance by enhancing its proteasome-dependent degradation. The Drosophila katanin regulatory subunit Kat80, which is required to stabilize Kat60 in cells, conversely reduces the proteasome-dependent degradation of Kat60. Second, the MIT domain and linker region of Kat60 augmented its microtubule-disassembly activity by enhancing its association with microtubules. On the basis of these data, it is proposed that the non-catalytic domains of Drosophila katanin serve as the principal sites of integration of regulatory inputs, thereby controlling its ability to sever and disassemble microtubules.

Groen, C. M., Jayo, A., Parsons, M. and Tootle, T. L. (2015). #Prostaglandins regulate nuclear localization of Fascin and its function in nucleolar architecture. Mol Biol Cell [Epub ahead of print]. PubMed ID: 25808493
Fascin, a highly conserved actin bundling protein, localizes and functions at new cellular sites in both Drosophila and multiple mammalian cell types. During Drosophila follicle development, in addition to being cytoplasmic, Fascin (Singed) is in the nuclei of the germline-derived nurse cells during stages 10B-12 (S10B-12) and at the nuclear periphery during stage 13 (S13). This localization is specific to Fascin, as other actin binding proteins, Villin and Profilin, do not exhibit the same subcellular distribution. Additionally, localization of Fascin to the nucleus and nuclear periphery is observed in multiple mammalian cell types. Thus, the regulation and function of Fascin at these new cellular locations is likely to be highly conserved. In Drosophila, loss of prostaglandin signaling (see Singed) causes a global reduction in nuclear Fascin, and a failure to relocalize to the nuclear periphery. Alterations in nuclear Fascin levels result in defects in nucleolar morphology in both Drosophila follicles and cultured mammalian cells, suggesting that nuclear Fascin plays an important role in nucleolar architecture. Given the numerous roles of Fascin in development and disease, including cancer, a novel finding that Fascin has functions within the nucleus sheds new light on the potential roles of Fascin in these contexts.

Davis, J. R., Luchici, A., Mosis, F., Thackery, J., Salazar, J. A., Mao, Y., Dunn, G. A., Betz, T., Miodownik, M. and Stramer, B. M. (2015). Inter-cellular forces in #Drosophila #hemocytes orchestrate contact inhibition of locomotion. Cell 161: 361-373. PubMed ID: 25799385
Contact inhibition of locomotion (CIL) is a multifaceted process that causes many cell types to repel each other upon collision. During development, this seemingly uncoordinated reaction is a critical driver of cellular dispersion within embryonic tissues. This study shows that Drosophila hemocytes require a precisely orchestrated CIL response for their developmental dispersal. Hemocyte collision and subsequent repulsion involves a stereotyped sequence of kinematic stages that are modulated by global changes in cytoskeletal dynamics. Tracking actin retrograde flow within hemocytes in vivo reveals synchronous reorganization of colliding actin networks through engagement of an inter-cellular adhesion. This inter-cellular actin-clutch leads to a subsequent build-up in lamellar tension, triggering the development of a transient stress fiber, which orchestrates cellular repulsion. These findings reveal that the physical coupling of the flowing actin networks during CIL acts as a mechanotransducer, allowing cells to haptically sense each other and coordinate their behaviors.

Thursday, April 23rd

Aldrich, J.C. and Maggert, K.A. (2015). #Transgenerational inheritance of #diet-induced #genome rearrangements in #Drosophila. PLoS Genet 11: e1005148. PubMed ID: 25885886
Ribosomal RNA gene (rDNA) copy number variation modulates heterochromatin formation and influences the expression of a large fraction of the Drosophila genome. This discovery, along with the link between rDNA, aging, and disease, highlights the importance of understanding how natural rDNA copy number variation arises. Pursuing the relationship between rDNA expression and stability, this study discovered that increased dietary yeast concentration, emulating periods of dietary excess during life, results in somatic rDNA instability and copy number reduction. Modulation of Insulin/TOR signaling produced similar results, indicating a role for known nutrient sensing signaling pathways in this process. Furthermore, adults fed elevated dietary yeast concentrations produced offspring with fewer rDNA copies demonstrating that these effects also occurred in the germline, and were transgenerationally heritable. This finding explains one source of natural rDNA copy number variation revealing a clear long-term consequence of diet.

Jeon, H.J. et al. (2015). Age-related change in gammaH2AX of #Drosophila #muscle: its significance as a marker for muscle damage and #longevity. Biogerontology [Epub ahead of print]. PubMed ID: 25860864
Muscle aging is closely related to unhealthy late-life and organismal aging. The state of differentiated cells has been shown to be critical to tissue homeostasis. Thus, understanding how fully differentiated muscle cells age is required for ensuring healthy aging. This study investigated age-related changes of γH2AX, an indicator of DNA strand breaks, in adult Drosophila muscle to document whether its changes are correlated with muscle degeneration and lifespan. The results demonstrated that γH2AX accumulation increased in adult Drosophila thoracic and leg muscles with age. Analyses of short-, normal-, and long-lived strains indicated that the age-related increase of γH2AX was closely associated with the extent of muscle degeneration, cleaved caspase-3 and poly-ubiquitin aggregates, and longevity. Further analysis of muscle-specific knockdown of heterochromatin protein 1a revealed that the excessive γH2AX accumulation in thoracic and leg muscles induced accelerated degeneration and decreased longevity. These data suggest a strong correlation between age-related muscle damage and lifespan in Drosophila. These findings indicate that γH2AX may be a reliable biomarker for assessing muscle aging in Drosophila.

Blake, M. R., Holbrook, S. D., Kotwica-Rolinska, J., Chow, E. S., Kretzschmar, D. and Giebultowicz, J. M. (2015). Manipulations of #amyloid precursor protein cleavage disrupt the #circadian clock in #aging #Drosophila. Neurobiol Dis 77: 117-126. PubMed ID: 25766673
Alzheimer's disease (AD) is a neurodegenerative disease characterized by severe cognitive deterioration. While causes of AD pathology are debated, a large body of evidence suggests that increased cleavage of Amyloid Precursor Protein (APP) producing the neurotoxic Amyloid-beta (Aβ) peptide plays a fundamental role in AD pathogenesis. One of the detrimental behavioral symptoms commonly associated with AD is the fragmentation of sleep-activity cycles with increased nighttime activity and daytime naps in humans. Sleep-activity cycles, as well as physiological and cellular rhythms, which may be important for neuronal homeostasis, are generated by a molecular system known as the circadian clock. Links between AD and the circadian system are increasingly evident but not well understood. This study examined whether genetic manipulations of APP-like (APPL) protein cleavage in Drosophila melanogaster affect rest-activity rhythms and core circadian clock function in this model organism. The increased β-cleavage of endogenous APPL by the β-secretase (dBACE) was shown to severely disrupts circadian behavior and leads to reduced expression of clock protein Per in central clock neurons of aging flies. The data suggest that behavioral rhythm disruption is not a product of APPL-derived Aβ production but rather may be caused by a mechanism common to both α and β-cleavage pathways. Specifically, it was shown that increased production of the endogenous Drosophila Amyloid Intracellular Domain (dAICD) caused disruption of circadian rest-activity rhythms, while flies overexpressing endogenous APPL maintained stronger circadian rhythms during aging. In summary, this study offers a novel entry point toward understanding the mechanism of circadian rhythm disruption in Alzheimer's disease.

Oka, S., Hirai, J., Yasukawa, T., Nakahara, Y. and Inoue, Y. H. (2015). A correlation of reactive oxygen species accumulation by depletion of superoxide dismutases with age-dependent impairment in the nervous system and muscles of #Drosophila adults. Biogerontology [Epub ahead of print]. PubMed ID: 25801590
The theory that accumulation of reactive oxygen species (ROS) in internal organs is a major promoter of aging has been considered negatively. However, it is still controversial whether overexpression of superoxide dismutases (SODs), which remove ROS, extends the lifespan in Drosophila adults. This paper examined whether ROS accumulation by depletion of Cu/Zn-SOD (SOD1) or Mn-SOD (SOD2) influenced age-related impairment of the nervous system and muscles in Drosophila. The efficient depletion of Sod1 and Sod2 was confirmed through RNAi and ROS accumulation by monitoring of ROS-inducible gene expression. Both RNAi flies displayed accelerated impairment of locomotor activity with age and shortened lifespan. Similarly, adults with nervous system-specific depletion of Sod1 or Sod2 also showed reduced lifespan. An accelerated loss of dopaminergic neurons was found in the flies with suppressed SOD expression. A half-dose reduction of three pro-apoptotic genes resulted in a significant suppression of the neuronal loss, suggesting that apoptosis was involved in the neuronal loss caused by SOD silencing. In addition, depletion of Sod1 or Sod2 in musculature is also associated with enhancement of age-related locomotion impairment. In indirect flight muscles from SOD-depleted adults, abnormal protein aggregates containing poly-ubiquitin accumulated at an early adult stage and continued to increase as the flies aged. Most of these protein aggregates were observed between myofibril layers. Moreover, immuno-electron microscopy indicated that the aggregates were predominantly localized in damaged mitochondria. These findings suggest that muscular and neuronal ROS accumulation may have a significant effect on age-dependent impairment of the Drosophila adults.

Wednesday, April 22nd

Figueroa-Clarevega, A. and Bilder, D. (2015). Malignant Drosophila tumors interrupt insulin signaling to induce cachexia-like wasting. Dev Cell 33: 47-55. PubMed ID: 25850672
Tumors kill patients not only through well-characterized perturbations to their local environment but also through poorly understood pathophysiological interactions with distant tissues. This study uses a Drosophila tumor model to investigate the elusive mechanisms underlying such long-range interactions. Transplantation of tumors into adults induced robust wasting of adipose, muscle, and gonadal tissues that were distant from the tumor, phenotypes that resembled the cancer cachexia seen in human patients. Notably, malignant, but not benign, tumors induced peripheral wasting. The study identified the insulin growth factor binding protein (IGFBP) homolog ImpL2, an antagonist of insulin signaling, as a secreted factor mediating wasting. ImpL2 was sufficient to drive tissue loss, and insulin activity was reduced in peripheral tissues of tumor-bearing hosts. Importantly, knocking down ImpL2, specifically in the tumor, ameliorated wasting phenotypes. The study proposes that the tumor-secreted IGFBP creates insulin resistance in distant tissues, thus driving a systemic wasting response.

Kim, J. and Neufeld, T.P. (2015). Dietary sugar promotes systemic TOR activation in Drosophila through AKH-dependent selective secretion of Dilp3. Nat Commun 6: 6846. PubMed ID: 25882208
Secreted ligands of the insulin family promote cell growth and maintain sugar homeostasis. Insulin release is tightly regulated in response to dietary conditions, but how insulin-producing cells (IPCs) coordinate their responses to distinct nutrient signals is unclear. This study shows that regulation of insulin secretion in Drosophila larvae has been segregated into distinct branches- whereas amino acids promote the secretion of Drosophila insulin-like peptide 2 (Dilp2), circulating sugars promote the selective release of Dilp3. Dilp3 was uniquely required for the sugar-mediated activation of TOR signalling and suppression of autophagy in the larval fat body. Sugar levels were not sensed directly by the IPCs, but rather by the adipokinetic hormone (AKH)-producing cells of the corpora cardiaca, and the study demonstrated that AKH signalling was required in the IPCs for sugar-dependent Dilp3 release. Thus, IPCs integrate multiple cues to regulate the secretion of distinct insulin subtypes under varying nutrient conditions.

Vicidomini, R., Di Giovanni, A., Petrizzo, A., Iannucci, L. F., Benvenuto, G., Nagel, A. C., Preiss, A. and Furia, M. (2015). Loss of Drosophila pseudouridine synthase triggers apoptosis-induced proliferation and promotes cell-nonautonomous EMT. Cell Death Dis 6: e1705. PubMed ID: 25811802
Many developing tissues display regenerative capability that allows them to compensate cell loss and preserve tissue homeostasis. Because of their remarkable regenerative capability, Drosophila wing discs are extensively used for the study of regenerative phenomena. This study used the developing wing to investigate the role played in tissue homeostasis by the evolutionarily conserved eukaryotic H/ACA small nucleolar ribonucleoprotein pseudouridine synthase (Nucleolar protein at 60B or minifly). The study shows that localized depletion of this enzyme can act as an endogenous stimulus capable of triggering apoptosis-induced proliferation, and that context-dependent effects are elicited in different sub-populations of the silenced cells. In fact, some cells undergo apoptosis, whereas those surrounding the apoptotic foci, although identically depleted, overproliferate. This overproliferation correlates with ectopic induction of the Wg and JAK-STAT mitogenic pathways. Expression of a baculovirus p35 transgene, which blocks the complete execution of the death program and generates the so-called 'undead cells', amplifies the proliferative response. Pseudouridine synthase depletion also causes loss of apicobasal polarity, disruption of adherens cell junctions and ectopic induction of JNK (c-Jun N-terminal kinase) and Mmp1 (matrix metalloproteinase-1) activity, leading to a significant epithelial reorganization. Unexpectedly, cell-nonautonomous effects, such as epithelial mesenchymal transition in the contiguous unsilenced squamous epithelium, are also promoted. Collectively, these data point out that cell-cell communication and long-range signaling can take a relevant role in the response to pseudouridine synthase decline. These results can add new light on the still unexplained tumor predisposition that characterizes X-linked dyskeratosis, the human disease caused by reduced pseudouridine synthase activity.

Takada, S., Collins, E. R. and Kurahashi, K. (2015). The FHA domain determines Drosophila Chk2/Mnk localization to key mitotic structures and is essential for early embryonic DNA damage responses. Mol Biol Cell [Epub ahead of print]. PubMed ID: 25808488
DNA damage responses including mitotic centrosome inactivation, cell-cycle delay in mitosis, and nuclear dropping from embryo cortex maintain genome integrity in syncytial Drosophila embryos. A conserved signaling kinase Chk2 known as Mnk/Loki is essential for the responses. This study demonstrates that functional EGFP-Mnk expressed from a transgene localizes to the nucleus, centrosomes, the interkinetochore/centromere region, the midbody, and pseudocleavage furrows without DNA damage, and additionally forms numerous foci/aggregates on mitotic chromosomes upon DNA damage. EGFP-tagged Mnk deletion or point mutation variants and investigated domain functions of Mnk in vivo. A triple-mutation in the phosphopeptide-binding site of the fork-head associated (FHA) domain disrupted normal Mnk localization except to the nucleus. The mutation also disrupted Mnk foci formation on chromosomes upon DNA damage. FHA mutations and deletion of the SQ/TQ-cluster domain (SCD) abolished Mnk trans- and auto-phosphorylations indicative of kinase activation after DNA damage. A potent NLS was found at the C-terminus, which is required for normal Mnk function. It is proposed that the FHA domain in Mnk plays essential dual-functions in mediating embryonic DNA damage responses by means of its phosphopeptide binding ability: activating Mnk in the nucleus upon DNA damage and recruiting Mnk to multiple subcellular structures independently of DNA damage.

Tuesday, April 21st

Kirkhart, C. and Scott, K. (2015). #Gustatory #learning and processing in the #Drosophila mushroom bodies. J Neurosci 35: 5950-5958. PubMed ID: 25878268
The Drosophila mushroom bodies are critical association areas whose role in olfactory associative learning has been well characterized. Recent behavioral studies using a taste association paradigm revealed that gustatory conditioning also requires the mushroom bodies. This study examines the representations of tastes and the neural sites for taste associations in the mushroom bodies. Using molecular genetic approaches to target different neuronal populations, they found that the gamma lobes of the mushroom bodies and a subset of dopaminergic input neurons were required for taste associative learning. Monitoring responses to taste compounds in the mushroom body calyx with calcium imaging revealed sparse, taste-specific and organ-specific activation in the Kenyon cell dendrites of the main calyx and the dorsal accessory calyx. This work provides insight into gustatory representations in the mushroom bodies, revealing the essential role of gustatory inputs not only as rewards and punishments but also as adaptive cues.

Matsuno, M., Horiuchi, J., Yuasa, Y., Ofusa, K., Miyashita, T., Masuda, T., and Saitoe, M. (2015). Long-term #memory formation in #Drosophila requires training-dependent #glial transcription. J Neurosci 35: 5557-5565. PubMed ID: 25855172
Long-term memory (LTM) formation requires de novo gene expression in neurons, and subsequent structural and functional modification of synapses. However, the importance of gene expression in glia during this process has not been well studied. This report characterizes a cell adhesion molecule, Klingon (Klg), which is required for LTM formation in Drosophila. Klg localized to the juncture between neurons and glia, and expression in both cell types was required for LTM. Further, expression of a glial gene, repo, was reduced in klg mutants and knockdown lines. repo expression was required for LTM, and expression increased upon LTM induction. In addition, increasing repo expression in glia was sufficient to restore LTM in klg knockdown lines. These data indicate that neuronal activity enhances Klg-mediated neuron-glia interactions, causing an increase in glial expression of repo. Repo is a homeodomain transcription factor, suggesting that further downstream glial gene expression is also required for LTM.

Omoto, J. J., Yogi, P. and Hartenstein, V. (2015). Origin and development of neuropil #glia of the Drosophila larval and adult #brain: Two distinct glial populations derived from separate progenitors. Dev Biol [Epub ahead of print]. PubMed ID: 25779704
Glia comprise a conspicuous population of non-neuronal cells in vertebrate and invertebrate nervous systems. Drosophila serves as a favorable model to elucidate basic principles of glial biology in vivo. The Drosophila neuropil glia (NPG), subdivided into astrocyte-like (ALG) and ensheathing glia (EG), extend reticular processes which associate with synapses and sheath-like processes which surround neuropil compartments, respectively. This paper characterizes the development of NPG throughout fly brain development. Differentiated neuropil glia of the larval brain originate as a cluster of precursors derived from embryonic progenitors located in the basal brain. These precursors undergo a characteristic migration to spread over the neuropil surface while specifying/differentiating into primary ALG and EG. Embryonically-derived primary NPG are large cells which are few in number, and occupy relatively stereotyped positions around the larval neuropil surface. During metamorphosis, primary NPG undergo cell death. Neuropil glia of the adult (secondary NPG) are derived from type II lineages during the postembryonic phase of neurogliogenesis. These secondary NPG are much smaller in size but greater in number than primary NPG. Lineage tracing reveals that both NPG subtypes derive from intermediate neural progenitors of multipotent type II lineages. Taken together, this study reveals previously uncharacterized dynamics of NPG development and provides a framework for future studies utilizing Drosophila glia as a model.

Lovick, J. K. and Hartenstein, V. (2015). Hydroxyurea-mediated neuroblast ablation establishes birth dates of secondary lineages and addresses neuronal interactions in the developing #Drosophila #brain. Dev Biol. PubMed ID: 25773365
The Drosophila brain is comprised of neurons formed by approximately 100 lineages, each of which is derived from a stereotyped, asymmetrically dividing neuroblast. Lineages serve as structural and developmental units of Drosophila brain anatomy and reconstruction of lineage projection patterns represents a suitable map of Drosophila brain circuitry at the level of neuron populations ("macro-circuitry"). Two phases of neuroblast proliferation, the first in the embryo and the second during the larval phase (following a period of mitotic quiescence), produce primary and secondary lineages, respectively. Using temporally controlled pulses of hydroxyurea (HU) to ablate neuroblasts and their corresponding secondary lineages during the larval phase, this study analyzed the effect on development of primary and secondary lineages in the late larval and adult brain. The findings indicate that timing of neuroblast re-activation is highly stereotyped, allowing establishment of "birth dates" for all secondary lineages. Furthermore, the results demonstrate that, whereas the trajectory and projection pattern of primary and secondary lineages is established in a largely independent manner, the final branching pattern of secondary neurons is dependent upon the presence of appropriate neuronal targets. Taken together, these data provide new insights into the degree of neuronal plasticity during Drosophila brain development.

Monday, April 2039

Yuan, K., and O'Farrell, P.H. (2015). Cyclin B3 is a mitotic cyclin that promotes the metaphase-anaphase transition. Curr Biol [Epub ahead of print]. PubMed ID: 25754637
The timing mechanism for mitotic progression is still poorly understood. The spindle assembly checkpoint (SAC), whose reversal upon chromosome alignment is thought to time anaphase, is functional during the rapid mitotic cycles of the Drosophila embryo; but its genetic inactivation had no consequence on the timing of the early mitoses. Mitotic cyclins-Cyclin A, Cyclin B, and Cyclin B3-influence mitotic progression and are degraded in a stereotyped sequence. RNAi knockdown of Cyclins A and B resulted in a Cyclin B3-only mitosis in which anaphase initiated prior to chromosome alignment. Furthermore, in such a Cyclin B3-only mitosis, colchicine-induced SAC activation failed to block Cyclin B3 destruction, chromosome decondensation, or nuclear membrane re-assembly. Injection of Cyclin B proteins restored the ability of SAC to prevent Cyclin B3 destruction. Thus, SAC function depends on particular cyclin types. Changing Cyclin B3 levels showed that it accelerated progress to anaphase, even in the absence of SAC function. The impact of Cyclin B3 on anaphase initiation appeared to decline with developmental progress. These results show that different cyclin types affect anaphase timing differently in the early embryonic divisions. The early-destroyed cyclins-Cyclins A and B-restrained anaphase-promoting complex/cyclosome (APC/C) function, whereas the late-destroyed cyclin, Cyclin B3, stimulated function. It is proposed that the destruction schedule of cyclin types guides mitotic exit by affecting both Cdk1 and APC/C, whose activities change as each cyclin type is lost.

Defachelles, L., Raich, N., Terracol, R., Baudin, X., Williams, B., Goldberg, M. and Karess, R. E. (2015). RZZ and Mad1 dynamics in Drosophila mitosis. Chromosome Res [Epub ahead of print]. PubMed ID: 25772408
The presence or absence of Mad1 at kinetochores is a major determinant of spindle assembly checkpoint (SAC) activity, the surveillance mechanism that delays anaphase onset if one or more kinetochores remain unattached to spindle fibers. Among the factors regulating the levels of Mad1 at kinetochores is the Rod, Zw10, and Zwilch (RZZ) complex, which is required for Mad1 recruitment through a mechanism that remains unknown. The relative dynamics and interactions of Mad1 and RZZ at kinetochores have not been extensively investigated, although Mad1 has been reported to be stably recruited to unattached kinetochores. This study directly compare Mad1-green fluorescent protein (GFP) turnover dynamics on unattached Drosophila kinetochores with that of RZZ, tagged either with GFP-Rod or GFP-Zw10. Nearly 40 % of kinetochore-bound Mad1 has a significant dynamic component, turning over with a half-life of 12 s. RZZ in contrast is essentially stable on unattached kinetochores. In addition, this study reports that a fraction of RZZ and Mad1 can co-immunoprecipitate, indicating that the genetically determined recruitment hierarchy (in which Mad1 depends on RZZ) may reflect a physical association of the two complexes.

Gottardo, M., Callaini, G. and Riparbelli, M. G. (2015). Aurora A inhibition by MNL8054 promotes centriole elongation during Drosophila male meiosis. Cell Cycle [Epub ahead of print]. PubMed ID: 25785740
Aurora A kinase plays an important role in several aspects of cell division, including centrosome maturation and separation, a crucial step for the correct organization of the bipolar spindle. Although it has long been showed that this kinase accumulates at the centrosome throughout mitosis its precise contribution to centriole biogenesis and structure has until now not been reported. It is not surprising that so little is known, due to the small size of somatic centrioles, where only dramatic structural changes may be identified by careful electron microscopy analysis. Conversely, centrioles of Drosophila primary spermatocytes increase tenfold in length during the first prophase, thus making any change easily detectable. Therefore, this study examined the consequence of the pharmacological inhibition of Aurora A by MLN8054 on centriole biogenesis during early Drosophila gametogenesis. Depletion of this kinase was shown to result in longer centrioles, mainly during transition from prophase to prometaphase of the first meiosis. Abnormal ciliogenesis was found, characterized by irregularly growing axonemal doublets. These results represent the first documentation of a potential requirement of Aurora A in centriole integrity and elongation.

Sabino, D., Gogendeau, D., Gambarotto, D., Nano, M., Pennetier, C., Dingli, F., Arras, G., Loew, D. and Basto, R. (2015). Moesin is a major regulator of centrosome behavior in epithelial cells with extra centrosomes. Curr Biol 25: 879-889. PubMed ID: 25772448
Centrosome amplification has severe consequences during development and is thought to contribute to a variety of diseases such as cancer and microcephaly. However, the adverse effects of centrosome amplification in epithelia are still not known. This study investigated the consequences of centrosome amplification in the Drosophila wing disc epithelium. Epithelial cells were found to exhibit mechanisms of clustering but also inactivation of extra centrosomes. Importantly, these mechanisms are not fully efficient, and both aneuploidy and cell death can be detected. Epithelial cells with extra centrosomes generate tumors when transplanted into wild-type hosts and inhibition of cell death results in tissue over-growth and disorganization. Using SILAC-fly, this study found that Moesin, a FERM domain protein, is specifically upregulated in wing discs with extra centrosomes. Moesin localizes to the centrosomes and mitotic spindle during mitosis, and Moesin upregulation was shown to influence extra-centrosome behavior and robust bipolar spindle formation. This study provides a mechanistic explanation for the increased aneuploidy and transformation potential primed by centrosome amplification in epithelial tissues.

Sunday, April 19th

Rane, R. V., Rako, L., Kapun, M., Lee, S. F. and Hoffmann, A. A. (2015). Genomic evidence for role of inversion 3RP of #Drosophila melanogaster in facilitating #climate change #adaptation. Mol Ecol [Epub ahead of print]. PubMed ID: 25789416
Chromosomal inversion polymorphisms are common in animals and plants and recent models suggest that alternative arrangements spread by capturing different combinations of alleles acting additively or epistatically to favour local adaptation. It is also thought that inversions typically maintain favoured combinations for a long time by suppressing recombination between alternative chromosomal arrangements. This paper considers patterns of linkage disequilibrium and genetic divergence in an old inversion polymorphism in Drosophila melanogaster (In(3R)Payne) known to be associated with climate change adaptation and a recent invasion event into Australia. Karyotyped and sequenced whole chromosomes were extracted from two Australian populations so that changes in the arrangement of the alleles between geographically separated tropical and temperate areas could be compared. Chromosome-wide linkage disequilibrium (LD) analysis revealed strong LD within the region spanned by In(3R)Payne. This genomic region also showed strong differentiation between the tropical and the temperate populations but no differentiation between different karyotypes from the same population, after controlling for chromosomal arrangement. Patterns of differentiation across the chromosome arm and in gene ontologies were enhanced by the presence of the inversion. These data support the notion that inversions are strongly selected by bringing together combinations of genes but it is still not clear if such combinations act additively or epistatically. These data suggest that climatic adaptation through inversions can be dynamic, reflecting changes in the relative abundance of different forms of an inversion and ongoing evolution of allelic content within an inversion.

Gerken, A. R., Eller, O. C., Hahn, D. A. and Morgan, T. J. (2015). Constraints, independence, and #evolution of thermal plasticity: Probing genetic architecture of long- and short-term #thermal acclimation. Proc Natl Acad Sci U S A 112: 4399-4404. PubMed ID: 25805817
Seasonal and daily thermal variation can limit species distributions because of physiological tolerances. Low temperatures are particularly challenging for ectotherms, which use both basal thermotolerance and acclimation, an adaptive plastic response, to mitigate thermal stress. Both basal thermotolerance and acclimation are thought to be important for local adaptation and persistence in the face of climate change. However, the evolutionary independence of basal and plastic tolerances remains unclear. Acclimation can occur over longer (seasonal) or shorter (hours to days) time scales, and the degree of mechanistic overlap is unresolved. Using a midlatitude population of Drosophila melanogaster, this study showed substantial heritable variation in both short- and long-term acclimation. Rapid cold hardening (short-term plasticity) and developmental acclimation (long-term plasticity) are positively correlated, suggesting shared mechanisms. However, there are independent components of these traits, because developmentally acclimated flies respond positively to short-term acclimation. A strong negative correlation between basal cold tolerance and developmental acclimation suggests that basal cold tolerance may constrain developmental acclimation, whereas a weaker negative correlation between basal cold tolerance and short-term acclimation suggests less constraint. Using genome-wide association mapping, The genetic architecture of rapid cold hardening and developmental acclimation responses shown nonoverlapping at the SNP and corresponding gene level. However, genes associated with each trait share functional similarities, including genes involved in apoptosis and autophagy, cytoskeletal and membrane structural components, and ion binding and transport. These results indicate substantial opportunity for short-term and long-term acclimation responses to evolve separately from each other and for short-term acclimation to evolve separately from basal thermotolerance.

Tanaka, K. M., Hopfen, C., Herbert, M. R., Schlotterer, C., Stern, D. L., Masly, J. P., McGregor, A. P. and Nunes, M. D. (2015). Genetic architecture and functional characterization of genes underlying the rapid diversification of male external genitalia between #Drosophila simulans and Drosophila mauritiana. Genetics [Epub ahead of print]. PubMed ID: 25783699
Male sexual characters are often among the first traits to diverge between closely related species and identifying the genetic basis of such changes can contribute to understanding of their evolutionary history. However, little is known about the genetic architecture or the specific genes underlying the evolution of male genitalia. The morphology of the claspers, posterior lobes and anal plates exhibit striking differences between Drosophila mauritiana and Drosophila simulans. Using QTL and introgression-based high-resolution mapping, several small regions were identified on chromosome arms 3L and 3R that contribute to differences in these traits. However, the loci underlying the evolution of clasper differences between these two species were found to be independent from those that contribute to posterior lobe and anal plate divergence. Furthermore, while most of the loci affect each trait in the same direction and act additively, evidence was found for epistasis between loci for clasper bristle number. In addition, an RNAi screen was conducted in D. melanogaster to investigate if positional and expression candidate genes, located on chromosome 3, are also involved in genital development. Six of these genes, including components of Wnt signaling and male-specific lethal 3 (msl3), were found to regulate the development of genital traits consistent with the effects of the introgressed regions where they are located and that thus represent promising candidate genes for the evolution these traits.

Ahmed-Braimah, Y. H. and Sweigart, A. L. (2015). A single gene causes an interspecific difference in pigmentation in Drosophila. Genetics [Epub ahead of print]. PubMed ID: 25769982
The genetic basis of species differences remains understudied. Studies in insects have contributed significantly to understanding of morphological evolution. Pigmentation traits in particular have received a great deal of attention and several genes in the insect pigmentation pathway have been implicated in inter- and intraspecific differences. Nonetheless, much remains unknown about many of the genes in this pathway and their potential role in understudied taxa. This study genetically analyze the puparium color difference between members of the virilis group of Drosophila. The puparium of Drosophila virilis is black, while those of D. americana, D. novamexicana, and D. lummei are brown. A series of backcross hybrid populations between D. americana and D. virilis were uded to map the genomic interval responsible for the difference between this species pair. First, it was shown that the pupal case color difference is caused by a single Mendelizing factor, which was ultimately mapped to an ~11kb region on chromosome 5. The mapped interval includes only the first exon and regulatory region(s) of the dopamine N-acetyltransferase gene (Dat). This gene encodes an enzyme that is known to play a part in the insect pigmentation pathway. Second, it was shown that this gene is highly expressed at the onset of pupation in light-brown taxa (D. americana and D. novamexicana) relative to D. virilis, but not in the dark-brown D. lummei. Finally, the role of Dat in adult pigmentation between D. americana (heavily melanized) and D. novamexicana (lightly melanized) was examined, and no discernible effect was found of this gene in adults. These results demonstrate that a single gene is entirely or almost entirely responsible for a morphological difference between species.

Saturday, April 18th

Sieber, M. H. and Spradling, A. C. (2015). Steroid signaling establishes a female metabolic state and regulates SREBP to control oocyte lipid accumulation. Curr Biol [Epub ahead of print]. PubMed ID: 25802149
Disruptions in energy homeostasis severely affect reproduction in many organisms and are linked to several reproductive disorders in humans. As a result, understanding the mechanisms that control nutrient accumulation in the oocyte will provide valuable insights into the links between metabolic disease and reproductive dysfunction. The steroid hormone ecdysone functions in Drosophila to control lipid metabolism and support oocyte production. First, local EcR-mediated signaling induces a stage-specific accumulation of lipids in stage-10 oocytes. EcR induces lipid accumulation by promoting the activation of the lipogenic transcription factor SREBP and by controlling the expression of the low-density lipoprotein (LDL) receptor homolog, LpR2. Second, global signaling via the ecdysone receptor, EcR, establishes a female metabolic state and promotes whole-body triglyceride and glycogen storage at high levels. EcR acts in the CNS to mediate these effects, in part by promoting higher levels of feeding in females. Thus, ecdysone functions at two levels to support reproduction: first by inducing lipid accumulation in the late stages of oocyte development and second by providing a signal that coordinates lipid metabolism in the germline with whole-animal lipid homeostasis. Ecdysone regulation allows females to assess the demands of oogenesis and alter their behavior and metabolic state to support the biosynthetic requirements of oocyte production.

Hamada-Kawaguchi, N., Nishida, Y. and Yamamoto, D. (2015). Btk29A-mediated tyrosine phosphorylation of Armadillo/β-Catenin promotes ring canal growth in Drosophila oogenesis. PLoS One 10: e0121484. PubMed ID: 25803041
Drosophila Btk29A is the ortholog of mammalian Btk, a Tec family nonreceptor tyrosine kinase whose deficit causes X-linked agammaglobulinemia in humans. The Btk29AficP mutation induces multiple abnormalities in oogenesis, including the growth arrest of ring canals, large intercellular bridges that allow the flow of cytoplasm carrying maternal products essential for embryonic development from the nurse cells to the oocyte during oogenesis. In this study, inactivation of Parcas, a negative regulator of Btk29A, was found to promote Btk29A accumulation on ring canals with a concomitant increase in the ring canal diameter, counteracting the Btk29AficP mutation. This mutation markedly reduced the accumulation of phosphotyrosine on ring canals and in the regions of cell-cell contact, where adhesion-supporting proteins such as DE-cadherin and β-catenin ortholog Armadillo (Arm) are located. Previous in vitro and in vivo analyses revealed that Btk29A directly phosphorylates Arm, leading to its release from DE-cadherin. In the present experiments, immunohistological analysis revealed that phosphorylation at tyrosine 150 (Y150) and Y667 of Arm was diminished in Btk29AficP mutant ring canals. Overexpression of an Arm mutant with unphosphorylatable Y150 inhibited ring canal growth. Thus Btk29A-induced Y150 phosphorylation is necessary for the normal growth of ring canals. It is suggested that the dissociation of tyrosine-phosphorylated Arm from DE-cadherin allows dynamic actin to reorganize, leading to ring canal expansion and cell shape changes during the course of oogenesis.

Reilly, E., Changela, N., Naryshkina, T., Deshpande, G. and Steward, R. (2015). Discs large 5, an essential gene in #Drosophila, regulates egg chamber organization. G3 (Bethesda) [Epub ahead of print]. PubMed ID: 25795662
Discs large 5 (Dlg5) is a member of the MAGUK family of proteins which typically serve as molecular scaffolds and mediate signaling complex formation and localization. In vertebrates, Dlg5 has been shown to be responsible for polarization of neural progenitors and to associate with Rab11 positive vesicles in epithelial cells. In Drosophila, however, the function of Dlg5 is not well documented. This study has identified dlg5 as an essential gene that shows embryonic lethality. dlg5 embryos displayed partial loss of primordial germ cells (PGCs) during gonad coalescence between stages 12-15 of embryogenesis. Loss of Dlg5 in germline and somatic stem cells in the ovary results in the depletion of both cell lineages. Reduced expression of Dlg5 in the follicle cells of the ovary leads to a number of distinct phenotypes, including defects in egg chamber budding, stalk cell overgrowth, and ectopic polar cell induction. Interestingly, loss of Dlg5 in follicle cells resulted in abnormal distribution of a critical component of cell adhesion, E-cadherin, shown to be essential for proper organization of egg chambers.

Ghosh, S and Lasko, P. (2015). Loss-of-function analysis reveals distinct requirements of the translation initiation factors eIF4E, eIF4E-3, eIF4G and eIF4G2 in Drosophila spermatogenesis. PLoS One 10: e0122519. PubMed ID: 25849588
Translational control is critical for spermatogenesis in Drosophila as many mRNAs synthesized in the spermatocytes are translated only much later during spermatid differentiation. Testes-specific translation initiation factors eIF4E-3 and eIF4G2 are essential specifically for male fertility. However, details of their roles during different stages of spermatogenesis are unknown, and the role of canonical translation initiation factors in spermatogenesis remains unexplored. This study addressed the functional role of eIF4E-1, eIF4E-3, eIF4G and eIF4G2 in testes development and formation of mature sperm. These four genes were systematically knocked down in different stages of germ cell development, and in the somatic cells. The results showed that eIF4E-1 function in early germ cells and the surrounding somatic cells is critical for spermatogenesis. Both eIF4E-1 and eIF4E-3 are required in spermatocytes for chromosome condensation and cytokinesis during the meiotic stages. Interestingly, eIF4G knockdown does not affect male fertility while eIF4G2 has distinct functions during spermatogenesis; it is required in early germ cells for proper meiotic divisions and spermatid elongation while its abrogation in spermatocytes causes meiotic arrest. Double knockdown of eIF4G and eIF4G2 showed that these proteins act redundantly during the early stages of spermatogenesis. Taken together, the analysis reveals spatio-temporal roles of the canonical and testes-specific translation initiation factors in coordinating developmental programs during spermatogenesis.

Friday, April 17th

Monier, B., Gettings, M., Gay, G., Mangeat, T., Schott, S., Guarner, A. and Suzanne, M. (2015). Apico-basal forces exerted by apoptotic cells drive epithelium folding. Nature 518: 245-248. PubMed ID: 25607361
Epithelium folding is a basic morphogenetic event that is essential in transforming simple two-dimensional epithelial sheets into three-dimensional structures in both vertebrates and invertebrates. Folding has been shown to rely on apical constriction. The resulting cell-shape changes depend either on adherens junction basal shift or on a redistribution of myosin II, which could be driven by mechanical signals. Yet the initial cellular mechanisms that trigger and coordinate cell remodelling remain largely unknown. This study unravels the active role of apoptotic cells in initiating morphogenesis, thus revealing a novel mechanism of epithelium folding. This study shows that, in the developing developing leg disk of Drosophila, apoptotic cells exert a transient pulling force upon the apical surface of the epithelium through a highly dynamic apico-basal myosin II cable. The apoptotic cells then induce a non-autonomous increase in tissue tension together with cortical myosin II apical stabilization in the surrounding tissue, eventually resulting in epithelium folding. Together these results, supported by a theoretical biophysical three-dimensional model, identify an apoptotic myosin-II-dependent signal as the initial signal leading to cell reorganization and tissue folding. This work further reveals that, far from being passively eliminated as generally assumed (for example, during digit individualization), apoptotic cells actively influence their surroundings and trigger tissue remodelling through regulation of tissue tension.

Tian, X., et al. (2015). A voltage-gated calcium channel regulates lysosomal fusion with endosomes and autophagosomes and is required for neuronal homeostasis. PLoS Biol 13: e1002103. PubMed ID: 25811491
Autophagy helps deliver sequestered intracellular cargo to lysosomes for proteolytic degradation and thereby maintains cellular homeostasis by preventing accumulation of toxic substances in cells. In a forward mosaic screen in Drosophila designed to identify genes required for neuronal function and maintenance, this study identified multiple cacophony (cac) mutant alleles. They exhibit an age-dependent accumulation of autophagic vacuoles (AVs) in photoreceptor terminals and eventually a degeneration of the terminals and surrounding glia. cac encodes an α1 subunit of a Drosophila voltage-gated calcium channel (VGCC) that is required for synaptic vesicle fusion with the plasma membrane and neurotransmitter release. This study shows that cac mutant photoreceptor terminals accumulate AV-lysosomal fusion intermediates, suggesting that Cac is necessary for the fusion of AVs with lysosomes, a poorly defined process. Loss of another subunit of the VGCC, α2δ or straightjacket (stj), causes phenotypes very similar to those caused by the loss of cac, indicating that the VGCC is required for AV-lysosomal fusion. The role of VGCC in AV-lysosomal fusion is evolutionarily conserved, as the loss of the mouse homologues, Cacna1a and Cacna2d2, also leads to autophagic defects in mice. Moreover, this study found that CACNA1A is localized to the lysosomes and that loss of lysosomal Cacna1a in cerebellar cultured neurons leads to a failure of lysosomes to fuse with endosomes and autophagosomes. Finally, it was shown that the lysosomal CACNA1A but not the plasma-membrane resident CACNA1A is required for lysosomal fusion. In summary, this study presents a model in which the VGCC plays a role in autophagy by regulating the fusion of AVs with lysosomes through its calcium channel activity and hence functions in maintaining neuronal homeostasis.

Park, S., Lee, Y., Pak, J.W., Kim, H., Choi, H., Kim, J.W., Roth, J. and Cho, J.W. (2015). O-GlcNAcylation modification is essential for the regulation of #autophagy in #Drosophila melanogaster. Cell Mol Life Sci [Epub ahead of print]. PubMed ID: 25840568
O-GlcNAcylation is a dynamic post-translational modification that takes place on ser/thr residues of nucleocytoplasmic proteins. O-GlcNAcylation regulates almost all cellular events as a nutrient sensor, a transcriptional and translational regulator, and a disease-related factor. Although the role of O-GlcNAcylation in insulin signaling and metabolism are well established, the relationship between O-GlcNAcylation and autophagy is largely unknown. This study manipulated O-GlcNAcylation in Drosophila and found that it regulates autophagy through Akt/dFOXO signaling. O-GlcNAcylation and the levels of O-GlcNAc transferase (OGT) were shown to increase during starvation. Furthermore, Atg proteins and autolysosomes were increased in OGT-reduced flies without fasting. Atg proteins and autophagosomes were reduced in OGT-overexpressing flies. These results suggest that not only autophagy gene expression but also autophagic structures are regulated by OGT through Akt and dFOXO. These data imply that O-GlcNAcylation is important in modulating autophagy as well as insulin signaling in Drosophila.

Zhang, S., Chen, C., Wu, C., Yang, Y., Li, W. and Xue, L. (2015). The canonical Wg signaling modulates Bsk-mediated cell death in #Drosophila. Cell Death Dis 6: e1713. PubMed ID: 25855961
Cell death is an essential regulatory mechanism for removing unneeded cells in animal development and tissue homeostasis. The c-Jun N-terminal kinase (JNK) pathway has pivotal roles in the regulation of cell death in response to various intrinsic and extrinsic stress signals. Canonical Wingless (Wg) signaling has been implicated in cell proliferation and cell fate decisions, whereas its role in cell death remains largely elusive. This study reports that activated Bsk (the Drosophila JNK homolog) induced cell death is mediated by the canonical Wg signaling. First, loss of Wg signaling abrogated Bsk-mediated caspase-independent cell death. Second, activation of Wg signaling promoted cell death in a caspase-independent manner. Third, activation of Bsk signaling results in upregulated transcription of wingless (wg) gene. Finally, Wg pathway participated in the physiological function of Bsk signaling in development. These findings not only reveal a previously undiscovered role of Wg signaling in Bsk-mediated cell death, but also provide a novel mechanism for the interplay between the two important signaling pathways in development.

Xiao, H., Wang, H., Silva, E. A., Thompson, J., Guillou, A., Yates, J. R., Jr., Buchon, N. and Franc, N. C. (2015). The Pallbearer E3 ligase promotes actin remodeling via RAC in efferocytosis by degrading the ribosomal protein S6. Dev Cell 32: 19-30. PubMed ID: 25533207
Clearance of apoptotic cells (efferocytosis) is achieved through phagocytosis by professional or amateur phagocytes. It is critical for tissue homeostasis and remodeling in all animals. Failure in this process can contribute to the development of inflammatory autoimmune or neurodegenerative diseases. Previous studies have shown that the PALL-SCF E3-ubiquitin ligase complex promotes apoptotic cell clearance, but it remained unclear how it did so. This study shows that the F-box protein Pallbearer (Pall) interacts with phosphorylated ribosomal protein S6 (RpS6) to promote its ubiquitylation and proteasomal degradation. This leads to Rac2 GTPase upregulation and activation and F-actin remodeling that promotes efferocytosis. This study further shows that the specific role of Pall in efferocytosis is driven by its apoptotic cell-induced nuclear export. Finding a role for RpS6 in the negative regulation of efferocytosis provides the opportunity to develop new strategies to regulate this process.

To, T. L., Piggott, B. J., Makhijani, K., Yu, D., Jan, Y. N. and Shu, X. (2015). Rationally designed fluorogenic protease reporter visualizes spatiotemporal dynamics of #apoptosis in vivo. Proc Natl Acad Sci U S A 112: 3338-3343. PubMed ID: 25733847
Fluorescence resonance energy transfer-based reporters have been widely used in imaging cell signaling; however, their in vivo application has been handicapped because of poor signal. Although fluorogenic reporters overcome this problem, no such reporter of proteases has been demonstrated for in vivo imaging. This paper reports the redesigning of an infrared fluorescent protein so that its chromophore incorporation is regulated by protease activity. Upon protease activation, the infrared fluorogenic protease reporter becomes fluorescent with no requirement of exogenous cofactor. To demonstrate biological applications, an infrared fluorogenic executioner-caspase reporter was designed that reveals spatiotemporal coordination between cell apoptosis and embryonic morphogenesis, as well as dynamics of apoptosis during tumorigenesis in Drosophila. The designed scaffold may be used to engineer reporters of other proteases with specific cleavage sequence.

Thursday, April 16th

Gill, S., Le, H. D., Melkani, G. C. and Panda, S. (2015). Time-restricted feeding attenuates age-related cardiac decline in Drosophila. Science 347: 1265-1269. PubMed ID: 25766238
Circadian clocks orchestrate periods of rest or activity and feeding or fasting over the course of a 24-hour day and maintain homeostasis. To assess whether a consolidated 24-hour cycle of feeding and fasting can sustain health, this study explored the effect of time-restricted feeding (TRF; food access limited to daytime 12 hours every day) on neural, peripheral, and cardiovascular physiology in Drosophila melanogaster. Improved sleep, prevention of body weight gain, and deceleration of cardiac aging under TRF, were detected even when caloric intake and activity were unchanged. This study used temporal gene expression profiling and validation through classical genetics to identify the TCP-1 ring complex (TRiC) chaperonin, the mitochondrial electron transport chain complexes, and the circadian clock as pathways mediating the benefits of TRF. In rodents, the daily cycle of feeding-fasting under TRF reinforces diurnal rhythms in multiple organs and prevents metabolic diseases when the animals are administered a high-fat diet. This study has shown that TRF protects against cardiac tissue aging in flies on either a normal or a fat-supplemented diet. This benefit appears to be mediated by the circadian clock, the TRiC chaperonin, and mitochondrial electron transport chain components.

Avila, F. W., Mattei, A. L. and Wolfner, M. F. (2015). Sex peptide receptor is required for the release of stored sperm by mated Drosophila melanogaster females. J Insect Physiol 76: 1-6. PubMed ID: 25783955
The storage of sperm in mated females is important for efficient reproduction. After sperm are transferred to females during mating, they need to reach and enter into the site(s) of storage, be maintained viably within storage, and ultimately be released from storage to fertilize eggs. Perturbation of these events can have drastic consequences on fertility. In Drosophila melanogaster, females store sperm for up to two weeks after a single mating. For sperm to be released normally from storage, Drosophila females need to receive the seminal fluid protein (SFP) sex peptide (SP) during mating. SP, which binds to sperm in storage, signals through the sex peptide receptor (SPR) to elicit two other effects on mated females: the persistence of egg laying and a reduction in sexual receptivity. However, it is not known whether SPR is also needed to mediate SP's effect on sperm release. By phenotypic analysis of flies deleted for SPR, and of flies knocked down for SPR, ubiquitously or in specific tissues, this study shows that SPR is required to mediate SP's effects on sperm release from storage. SPR expression in ppk+ neurons is needed for proper sperm release; these neurons include those that mediate SP's effect on receptivity and egg laying. However, this study found that SPR is also needed in the spermathecal secretory cells of the female reproductive tract for efficient sperm release. Thus, SPR expression is necessary in both the nervous system and in female reproductive tract cells to mediate the release of stored sperm.

May, C.M., Doroszuk, A. and Zwaan, B.J. (2015). The effect of developmental nutrition on life span and fecundity depends on the adult reproductive environment in Drosophila melanogaster. Ecol Evol 5: 1156-1168. PubMed ID: 25859322
This study used the fruit fly to determine whether life-history traits, particularly life span and fecundity, are affected by developmental nutrition, and whether this depends on the extent to which the adult environment allows females to realize their full reproductive potential. Flies were raised on three different developmental food levels containing increasing amounts of yeast and sugar: poor, control, and rich. Development on poor or rich larval food resulted in several life-history phenotypes indicative of suboptimal conditions, including increased developmental time, and, for poor food, decreased adult weight. However, development on poor larval food actually increased adult virgin life span. The study manipulated the reproductive potential of the adult environment by adding yeast or yeast and a male. This manipulation interacted with larval food to determine adult fecundity. Specifically, under two adult conditions, flies raised on poor larval food had higher reproduction at certain ages - when singly mated this occurred early in life and when continuously mated with yeast this occurred during midlife. Poor larval food was not necessarily detrimental to key adult life-history traits, but did exert an adult environment-dependent effect, especially by affecting virgin life span and altering adult patterns of reproductive investment. These findings are relevant because (1) they may explain differences between published studies on nutritional effects on life-history traits; (2) they indicate that optimal nutritional conditions are likely to be different for larvae and adults, potentially reflecting evolutionary history; and (3) they urge for the incorporation of developmental nutritional conditions into the central life-history concept of resource acquisition and allocation.

Neckameyer, W. S. and Nieto-Romero A. R. (2015). Response to stress in Drosophila is mediated by gender, age and stress paradigm. Stress 18: 1-13. PubMed ID: 25783197
All living organisms must maintain equilibrium in response to internal and external challenges within their environment. Changes in neural plasticity (alterations in neuronal populations, dendritic remodeling, and synaptic turnover) are critical components of the homeostatic response to stress, which has been strongly implicated in the onset of affective disorders. However, stress is differentially perceived depending on the type of stress and its context, as well as genetic background, age and sex; therefore, an individual's maintenance of neuronal homeostasis must differ depending upon these variables. This study established Drosophila as a model to analyze homeostatic responses to stress. Sexually immature and mature females and males from an isogenic wild-type strain raised under controlled environmental conditions were exposed to four reproducible and high-throughput translatable stressors to facilitate the analysis of a large number of animals for direct comparisons. These animals were assessed in an open-field arena, in a light-dark box, and in a forced swim test, as well as for sensitivity to the sedative effects of ethanol. These studies establish that immature and mature females and males represent behaviorally distinct populations under control conditions as well as after exposure to different stressors. Therefore, the neural substrates mediating the stress response must be differentially expressed depending upon the hormonal status of the brain. In addition, an adaptive response to a given stressor in one paradigm was not predictive for outcomes in other paradigms.

Wednesday, April 15th

Halstead, J. M., Lionnet, T., Wilbertz, J. H., Wippich, F., Ephrussi, A., Singer, R. H. and Chao, J. A. (2015). Translation. An RNA biosensor for imaging the first round of translation from single cells to living animals. Science 347: 1367-1671. PubMed ID: 25792328
Analysis of single molecules in living cells has provided quantitative insights into the kinetics of fundamental biological processes; however, the dynamics of messenger RNA (mRNA) translation have yet to be addressed. This study developed a fluorescence microscopy technique that reports on the first translation events of individual mRNA molecules. This allowed examination of the spatiotemporal regulation of translation during normal growth and stress and during Drosophila oocyte development. mRNAs were were shown not to be translated in the nucleus but are be translated within minutes after export, sequestration within P-bodies regulates translation, and oskar mRNA is not translated until it reaches the posterior pole of the oocyte. This methodology provides a framework for studying initiation of protein synthesis on single mRNAs in living cells.

Iwasaki, S., Sasaki, H.M., Sakaguchi, Y., Suzuki, T., Tadakuma, H. and Tomari, Y. (2015).Defining fundamental steps in the assembly of the #Drosophila #RNAi enzyme complex. Nature [Epub ahead of print]. PubMed ID: 25822791

Small RNAs such as small interfering RNAs (siRNAs) and microRNAs (miRNAs) silence the expression of their complementary target messenger RNAs via the formation of effector RNA-induced silencing complexes (RISCs), which contain Argonaute (Ago) family proteins at their core. Although loading of siRNA duplexes into Drosophila Ago2 requires the Dicer-2-R2D2 heterodimer and the Hsc70/Hsp90 (Hsp90 also known as Hsp83) chaperone machinery, the details of RISC assembly remain unclear. This study reconstitutes RISC assembly using only Ago2, Dicer-2, R2D2, Hsc70, Hsp90, Hop, Droj2 (an Hsp40 homologue) and p23. By following the assembly of single RISC molecules, this study found that, in the absence of the chaperone machinery, an siRNA bound to Dicer-2-R2D2 associated with Ago2 only transiently. The chaperone machinery extended the dwell time of the Dicer-2-R2D2-siRNA complex on Ago2, in a manner dependent on recognition of the 5'-phosphate on the siRNA guide strand. The study proposes that the chaperone machinery supports a productive state of Ago2, allowing it to load siRNA duplexes from Dicer-2-R2D2 and thereby assemble RISC.

Besnard-Guerin, C., Jacquier, C., Pidoux, J., Deddouche, S. and Antoniewsk, C. (2015). The cricket paralysis virus suppressor inhibits microRNA silencing mediated by the #Drosophila #Argonaute-2 protein. PLoS One 10: e0120205. PubMed ID: 25793377
Small RNAs are potent regulators of gene expression. They also act in defense pathways against invading nucleic acids such as transposable elements or viruses. To counteract these defenses, viruses have evolved viral suppressors of RNA silencing (VSRs). Plant viruses encoded VSRs interfere with siRNAs or miRNAs by targeting common mediators of these two pathways. In contrast, VSRs identified in insect viruses to date only interfere with the siRNA pathway whose effector Argonaute protein is Argonaute-2 (Ago-2). Although a majority of Drosophila miRNAs exerts their silencing activity through their loading into the #Argonaute-1 protein, recent studies highlighted that a fraction of miRNAs can be loaded into Ago-2, thus acting as siRNAs. In light of these recent findings, this study re-examined the role of insect VSRs on Ago-2-mediated miRNA silencing in Drosophila melanogaster. Using specific reporter systems in cultured Schneider-2 cells and transgenic flies, the Cricket Paralysis virus VSR CrPV1-A but not the Flock House virus B2 VSR was shown to abolish silencing by miRNAs loaded into the Ago-2 protein. Thus, these results provide the first evidence that insect VSR have the potential to directly interfere with the miRNA silencing pathway.

Haac, M. E., Anderson, M. A., Eggleston, H., Myles, K. M. and Adelman, Z. N. (2015). The hub protein #Loquacious connects the #microRNA and short interfering RNA pathways in #mosquitoes. Nucleic Acids Res [Epub ahead of print]. PubMed ID: 25765650
Aedes aegypti mosquitoes vector several arboviruses of global health significance, including dengue viruses and chikungunya virus. RNA interference (RNAi) plays an important role in antiviral immunity, gene regulation and protection from transposable elements. Double-stranded RNA binding proteins (dsRBPs) are important for efficient RNAi; in Drosophila functional specialization of the miRNA, endo-siRNA and exo-siRNA pathway is aided by the dsRBPs Loquacious (Loqs-PB, Loqs-PD) and R2D2, respectively. However, this functional specialization has not been investigated in other dipterans. No Loqs-PD was detected in Ae. aegypti; analysis of other dipteran genomes demonstrated that this isoform is not conserved outside of Drosophila. Overexpression experiments and small RNA sequencing following depletion of each dsRBP revealed that R2D2 and Loqs-PA cooperate non-redundantly in siRNA production, and that these proteins exhibit an inhibitory effect on miRNA levels. Conversely, Loqs-PB alone interacted with mosquito dicer-1 and was essential for full miRNA production. Mosquito Loqs interacts with both argonaute 1 and argonaute 2 in a manner independent of its interactions with dicer. It is concluded that the functional specialization of Loqs-PD in Drosophila is a recently derived trait, and that in other dipterans, including the medically important mosquitoes, Loqs-PA participates in both the miRNA and endo-siRNA based pathways.

Tuesday, April 14th

Won, J.H., Tsogtbartarr, O., Son, W., Singh, A., Choi, K.W. and Cho, K.O. (2015). Cell type-specific responses to Wingless, Hedgehog and Decapentaplegic are essential for patterning early eye-antenna disc in Drosophila. PLoS One 10: e0121999. PubMed ID: 25849899
The Drosophila eye-antenna imaginal disc (ead) is a flattened sac of two-layered epithelia, from which most head structures are derived. Secreted morphogens like Wingless (Wg), Hedgehog (Hh), and Decapentaplegic (Dpp) are important for early disc patterning, but the underlying mechanisms are still largely unknown. To understand how these morphogens function in the early larval discs, this study used wg-LacZ and dpp-Gal4 markers for the examination of wild-type and mutant eads. The ead immediately after hatching was crescent-shaped with the Bolwig's nerve at the ventral edge, suggesting that it consisted of dorsal domain. In a subsequent step, transcriptional induction of dpp in the cells along the Bolwig's nerve was followed by rapid growth of the ventral domain. Both Wg and Hh were shown to be required for the formation of the ventral domain. Wg was crucial for the growth of the entire disc, but Hh was shown to be essential for cell division only in the dorsal domain. In the ventral domain, Hh regulated dpp transcription. Based on these data, this study proposes that signaling among distinct groups of cells expressing Wg, Dpp, or Hh in the ead of the first-instar larvae are critical for coordinated disc growth and patterning.

Simon, E. and Guerrero, I. (2015). The transcription factor Optomotor-blind antagonizes Drosophila haltere growth by repressing Decapentaplegic and Hedgehog targets. PLoS One 10: e0121239. PubMed ID: 25793870
In Drosophila, decapentaplegic, which codes for a secreted signaling molecule, is activated by the Hedgehog signaling pathway at the anteroposterior compartment border of the two dorsal primordia; the wing and the haltere imaginal discs. In the wing disc, Decapentaplegic and Hedgehog signaling targets are implicated in cell proliferation and cell survival. However, most of their known targets in the wing disc are not expressed in the haltere disc due to their repression by the Hox gene Ultrabithorax. The T-box gene optomotor-blind escapes this repression in the haltere disc, and therefore is expressed in both the haltere and wing discs. Optomotor-blind is a major player during wing development and its function has been intensely investigated in this tissue, however, its role in haltere development has not been reported so far. This study shows that Optomotor-blind function in the haltere disc differs from that in the wing disc. Unlike its role in the wing, Optomotor-blind does not prevent apoptosis in the haltere but rather limits growth by repressing several Decapentaplegic and Hedgehog targets involved both in wing proliferation and in modulating the spread of morphogens similar to Ultrabithorax function but without disturbing Ultrabithorax expression.

Wen, D., Rivera-Perez, C., Abdou, M., Jia, Q., He, Q., Liu, X., Zyaan, O., Xu, J., Bendena, W. G., Tobe, S. S., Noriega, F. G., Palli, S. R., Wang, J. and Li, S. (2015). Methyl farnesoate plays a dual role in regulating Drosophila metamorphosis. PLoS Genet 11: e1005038. PubMed ID: 25774983
Corpus allatum (CA) ablation results in juvenile hormone (JH) deficiency and pupal lethality in Drosophila. The fly CA produces and releases three sesquiterpenoid hormones: JH III bisepoxide (JHB3), JH III, and methyl farnesoate (MF). In whole body extracts, MF is the most abundant sesquiterpenoid, followed by JHB3 and JH III. Knockout of juvenile hormone acid methyl transferase (jhamt) did not result in lethality; it decreased biosynthesis of JHB3, but MF biosynthesis was not affected. RNAi-mediated reduction of 3-hydroxy-3-methylglutaryl CoA reductase (Hmgcr) expression in the CA decreased biosynthesis and titers of the three sesquiterpenoids, resulting in partial lethality. Reducing Hmgcr expression in the CA of the jhamt mutant further decreased MF titer to a very low level, and caused complete lethality. JH III, JHB3, and MF function through Met and Gce, the two JH receptors, and induces expression of Kr-h1, a JH primary-response gene. As well, a portion of MF is converted to JHB3 in the hemolymph or peripheral tissues. Topical application of JHB3, JH III, or MF precluded lethality in JH-deficient animals, but not in the Met gce double mutant. Taken together, these experiments show that MF is produced by the larval CA and released into the hemolymph, from where it exerts its anti-metamorphic effects indirectly after conversion to JHB3, as well as acting as a hormone itself through the two JH receptors, Met and Gce.

Brunetti, T.M., Fremin, B.J. and Cripps, R.M. (2015). Identification of singles bar as a direct transcriptional target of Drosophila Myocyte enhancer factor-2 and a regulator of adult myoblast fusion. Dev Biol [Epub ahead of print]. PubMed ID: 25797154
In Drosophila, myoblast fusion is a conserved process in which founder cells (FCs) and fusion competent myoblasts (FCMs) fuse to form a syncytial muscle fiber. Mutants for the myogenic regulator Myocyte enhancer factor-2 (MEF2) show a failure of myoblast fusion, indicating that MEF2 regulates the fusion process. Indeed, chromatin immunoprecipitation studies show that several genes involved in myoblast fusion are bound by MEF2 during embryogenesis. Of these, the MARVEL domain gene singles bar (sing), is down-regulated in MEF2 knockdown pupae, and has five consensus MEF2 binding sites within a 9000-bp region. To determine if MEF2 is an essential and direct regulator of sing during pupal muscle development, a 315-bp myoblast enhancer of sing was identified. This enhancer was active during myoblast fusion, and mutation of two MEF2 sites significantly decreased enhancer activity. Lack of sing expression resulted in adult lethality and muscle loss, due to a failure of fusion during the pupal stage. Additionally, attempts were made to determine if sing was required in either FCs or FCMs to support fusion. Interestingly, knockdown of sing in either population did not significantly affect fusion, however, knockdown in both FCs and FCMs resulted in muscles with significantly reduced nuclei numbers, provisionally indicating that sing function is required in either cell type, but not both. Finally, MEF2 was found to regulated sing expression at the embryonic stage through the same 315-bp enhancer, indicating that sing is a MEF2 target at both critical stages of myoblast fusion. These studies define how MEF2 directly controls fusion at multiple stages of the life cycle, and provide further evidence that the mechanisms of fusion characterized in Drosophila embryos is also used in the formation of the more complex adult muscles.

Monday, April 13th

Besson, C., Bernard, F., Corson, F., Rouault, H., Reynaud, E., Keder, A., Mazouni, K. and Schweisguth, F. (2015). Planar cell polarity breaks the symmetry of PAR protein distribution prior to mitosis in Drosophila sensory organ precursor cells. Curr Biol [Epub ahead of print]. PubMed ID: 25843034
During development, cell-fate diversity can result from the unequal segregation of fate determinants at mitosis. Polarization of the mother cell is essential for asymmetric cell division (ACD). It often involves the formation of a cortical domain containing the PAR complex proteins Par3, Par6, and atypical protein kinase C (aPKC). In the fly notum, sensory organ precursor cells (SOPs) divide asymmetrically within the plane of the epithelium and along the body axis to generate two distinct cells. Fate asymmetry depends on the asymmetric localization of the PAR complex. In the absence of planar cell polarity (PCP), SOPs divide with a random planar orientation but still asymmetrically, showing that PCP is dispensable for PAR asymmetry at mitosis. To study when and how the PAR complex localizes asymmetrically, this study has used a quantitative imaging approach to measure the planar polarization of the proteins Bazooka (Baz, fly Par3), Par6, and aPKC in living pupae. By using imaging of functional GFP-tagged proteins with image processing and computational modeling, this study found that Baz, Par6, and aPKC become planar polarized prior to mitosis in a manner independent of the AuroraA kinase and that PCP was required for the planar polarization of Baz, Par6, and aPKC during interphase. This indicated that a "mitosis rescue" mechanism establishes asymmetry at mitosis in PCP mutants. This study therefore identifies PCP as the initial symmetry-breaking signal for the planar polarization of PAR proteins in asymmetrically dividing SOPs.

Shahab, J., Tiwari, M. D., Honemann-Capito, M., Krahn, M. P. and Wodarz, A. (2015). Bazooka/PAR3 is dispensable for polarity in Drosophila follicular epithelial cells. Biol Open [Epub ahead of print]. PubMed ID: 25770183
Apico-basal polarity is the defining characteristic of epithelial cells. In Drosophila, apical membrane identity is established and regulated through interactions between the highly conserved Par complex (Bazooka/Par3, atypical protein kinase C and Par6), and the Crumbs complex (Crumbs, Stardust and PATJ). It has been proposed that Bazooka operates at the top of a genetic hierarchy in the establishment and maintenance of apico-basal polarity. However, there is still ambiguity over the correct sequence of events and cross-talk with other pathways during this process. This study reassesses this issue by comparing the phenotypes of the commonly used baz4 and baz815-8 alleles with those of the so far uncharacterized bazXR11 and iEH747 null alleles in different Drosophila epithelia. While all these baz alleles display identical phenotypes during embryonic epithelial development, strong discrepancies were observed in the severity and penetrance of polarity defects in the follicular epithelium: polarity is mostly normal in bazEH747 and bazXR11 while baz4 and baz815-8 show loss of polarity, severe multilayering and loss of epithelial integrity throughout the clones. Further analysis reveals that the chromosomes carrying the baz4 and baz815-8 alleles may contain additional mutations that enhance the true baz loss-of-function phenotype in the follicular epithelium. This study clearly shows that Baz is dispensable for the regulation of polarity in the follicular epithelium, and that the requirement for key regulators of cell polarity is highly dependent on developmental context and cell type.

Grillo-Hill, B. K., Choi, C., Jimenez-Vidal, M. and Barber, D. L. (2015). Increased H efflux is sufficient to induce dysplasia and necessary for viability with oncogene expression. Elife 4. PubMed ID: 25793441
Intracellular pH (pHi) dynamics is increasingly recognized as an important regulator of a range of normal and pathological cell behaviors. Notably, increased pHi is now acknowledged as a conserved characteristic of cancers and in cell models is confirmed to increases proliferation and migration as well as limits apoptosis. However, the significance of increased pHi for cancer in vivo remains unresolved. Using Drosophila melanogaster, this study shows that increased pHi is sufficient to induce dysplasia in the absence of other transforming cues and potentiates growth and invasion with oncogenic Ras. Using a genetically encoded biosensor, increased pHi was also confirmed in situ. Moreover, in Drosophila models and clonal human mammary cells it was show that limiting H+ efflux with oncogenic Raf or Ras induces acidosis and synthetic lethality. Further, lethality in invasive, primary tumor cell lines with inhibiting H+ efflux was domonstrated. Synthetic lethality with reduced H+ efflux and activated oncogene expression could be exploited therapeutically to restrain cancer progression while limiting off-target effects.

Portela, M., Parsons, L. M., Grzeschik, N. A. and Richardson, H. E. (2015). Regulation of Notch signalling and endocytosis by the Lgl neoplastic tumor suppressor. Cell Cycle [Epub ahead of print]. PubMed ID: 25789785
The evolutionarily conserved neoplastic tumor suppressor protein, ≈Lethal (2) giant larvae (Lgl), plays roles in cell polarity and tissue growth via regulation of the Hippo pathway. In a recent study, it was shown that in the developing Drosophila eye epithelium, depletion of Lgl leads to increased ligand-dependent Notch signalling. lgl mutant tissue also exhibits an accumulation of early endosomes, recycling endosomes, early-multivesicular body markers and acidic vesicles. Elevated Notch signalling in lgl- tissue can be rescued by feeding larvae the vesicle de-acidifying drug chloroquine, revealing that Lgl attenuates Notch signalling by limiting vesicle acidification. Strikingly, chloroquine also rescued the lgl- overgrowth phenotype, suggesting that the Hippo pathway defects were also rescued. In this extraview, additional data is provided on the regulation of Notch signalling and endocytosis by Lgl, and possible mechanisms are discussed by which Lgl depletion contributes to signalling pathway defects and tumorigenesis

Sunday, April 12th

Ping, Y., Hahm, E. T., Waro, G., Song, Q., Vo-Ba, D. A., Licursi, A., Bao, H., Ganoe, L., Finch, K. and Tsunoda, S. (2015). Linking Aβ42-induced hyperexcitability to neurodegeneration, learning and motor deficits, and a shorter lifespan in an Alzheimer's model. PLoS Genet 11: e1005025. PubMed ID: 25774758
Alzheimer's disease (AD) is the most prevalent form of dementia in the elderly. β-amyloid (Aβ; see Drosophila Appl) accumulation in the brain is thought to be a primary event leading to eventual cognitive and motor dysfunction in AD. Aβ has been shown to promote neuronal hyperactivity, which is consistent with enhanced seizure activity in mouse models and AD patients. Little, however, is known about whether, and how, increased excitability contributes to downstream pathologies of AD. This study shows that overexpression of human Aβ42 in a Drosophila model indeed induces increased neuronal activity. The underlying mechanism involves the selective degradation of the A-type K+ channel, Kv4 (Shal). An age-dependent loss of Kv4 leads to an increased probability of action potential firing. Interestingly, loss of Kv4 alone results in learning and locomotion defects, as well as a shortened lifespan. To test whether the Aβ42-induced increase in neuronal excitability contributes to, or exacerbates, downstream pathologies, Kv4 was transgenically over-expressed to near wild-type levels in Aβ42-expressing animals. Restoration of Kv4 attenuated age-dependent learning and locomotor deficits, slowed the onset of neurodegeneration, and partially rescued premature death seen in Aβ42-expressing animals. It is concluded that Aβ42-induced hyperactivity plays a critical role in the age-dependent cognitive and motor decline of this Aβ42-Drosophila model, and possibly in AD.

Martinez-Morentin, L., Martinez, L., Piloto, S., Yang, H., Schon, E. A., Garesse, R., Bodmer, R., Ocorr, K., Cervera, M. and Arredondo, J. J. (2015). Cardiac deficiency of single cytochrome oxidase assembly factor scox induces p53 dependent apoptosis in a Drosophila cardiomyopathy model. Hum Mol Genet [Epub ahead of print]. PubMed ID: 25792727
The heart is a muscle with high energy demands. Hence, most patients with mitochondrial disease produced by defects in the Oxidative Phosphorylation System (OXPHOS) are susceptible to cardiac involvement. The presentation of mitochondrial cardiomyopathy includes hypertrophic, dilated, and left ventricular (LV) noncompaction, but the molecular mechanisms involved in cardiac impairment are unknown. One of the most frequent OXPHOS defect in humans frequently associated with cardiomyopathy is cytochrome c oxidase (COX) deficiency caused by mutations in COX assembly factors like Sco1 and Sco2. To investigate the molecular mechanisms that underlie the cardiomyopathy associated with Sco deficiency, Scox, the single Drosophila Sco orthologue, was specifically knocked down. Cardiac-specific knockdown of Scox reduces fly lifespan, and it severely compromises heart function and structure, producing dilated cardiomyopathy. Cardiomyocytes with low levels of Scox have a significant reduction in COX activity and they undergo a metabolic switch from OXPHOS to glycolysis, mimicking the clinical features found in patients harbouring Sco mutations. The major cardiac defects observed are produced by a significant increase in apoptosis, which is p53-dependent. Genetic and molecular evidence strongly suggests that p53 is directly involved in the development of the cardiomyopathy induced by Scox deficiency. Remarkably, apoptosis is enhanced in the muscle and liver of Sco2 knock-out mice, clearly suggesting that cell death is a key feature of the COX deficiencies produced by mutations in Sco genes in humans.

Brunke, S., Quintin, J., Kasper, L., Jacobsen, I. D., Richter, M. E., Hiller, E., Schwarzmuller, T., d'Enfert, C., Kuchler, K., Rupp, S., Hube, B. and Ferrandon, D. (2015). Of mice, flies - and men? Comparing fungal infection models for large-scale screening efforts. Dis Model Mech [Epub ahead of print]. PubMed ID: 25786415
The use of non-mammalian models for studying infectious diseases is still controversial, as it is often unclear how well findings from these systems predict virulence potential in humans or other mammals. This study compared the commonly used models, fruit fly and mouse (representing invertebrate and mammalian hosts) for their similarities and degree of correlation upon infection with a library of mutants of an important fungal pathogen, the yeast Candida glabrata. Using two indices, for fly survival time and for mouse fungal burden in specific organs, a good agreement was shown between the models. A suitable predictive model is provided for estimating the virulence potential of mutants in the mouse from fly survival data. As examples, cell wall integrity mutants were attenuated in flies and mutants of a MAP kinase pathway defective in both fly virulence and relative fitness in mice. In addition, mutants with strongly reduced in vitro growth were generally, but not always, reduced in fly virulence. Overall, this study demonstrates that surveying Drosophila survival after infection is a suitable model to predict outcome of murine infections, especially for severely attenuated mutants. Pre-screening of mutants in an invertebrate Drosophila model can thus give a good estimate of the probability for finding a strain with reduced microbial burden in the mouse host.

Begum, R., Calaza, K., Kam, J. H., Salt, T. E., Hogg, C. and Jeffery, G. (2015). Near-infrared light increases ATP, extends lifespan and improves mobility in aged Drosophila melanogaster. Biol Lett 11 [Epub ahead of print]. PubMed ID: 25788488
Ageing is an irreversible cellular decline partly driven by failing mitochondrial integrity. Mitochondria accumulate DNA mutations and reduce ATP production necessary for cellular metabolism. This is associated with inflammation. Near-infrared exposure increases retinal ATP in old mice via cytochrome c oxidase absorption and reduces inflammation. This study exposed fruitflies daily to 670 nm red light radiation, revealing elevated ATP and reduced inflammation with age. Critically, there was a significant increase in average lifespan: 100-175% more flies survived into old age following 670 nm exposure and these had significantly improved mobility. This may be a simple route to extending lifespan and improving function in old age.

Saturday, April 11th

York-Andersen, A. H., Parton, R. M., Bi, C. J., Bromley, C. L., Davis, I. and Weil, T. T. (2015). A single and rapid calcium wave at egg activation in Drosophila. Biol Open [Epub ahead of print]. PubMed ID: 25750438
Activation is an essential process that accompanies fertilisation in all animals and heralds major cellular changes, most notably, resumption of the cell cycle. While activation involves wave-like oscillations in intracellular Ca2+ concentration in mammals, ascidians and polychaete worms and a single Ca2+ peak in fish and frogs, in insects, such as Drosophila, to date, it has not been shown what changes in intracellular Ca2+ levels occur. This study utilised ratiometric imaging of Ca2+ indicator dyes and genetically encoded Ca2+ indicator proteins to identify and characterise a single, rapid, transient wave of Ca2+ in the Drosophila egg at activation. Using genetic tools, physical manipulation and pharmacological treatments this study demonstrates that the propagation of the Ca2+ wave requires an intact actin cytoskeleton and an increase in intracellular Ca2+ can be uncoupled from egg swelling, but not from progression of the cell cycle. It was further shown that mechanical pressure alone is not sufficient to initiate a Ca2+ wave. It was also found that processing bodies, sites of mRNA decay and translational regulation, become dispersed following the Ca2+ transient. Based on this data the following model for egg activation in Drosophila is proposed: exposure to lateral oviduct fluid initiates an increase in intracellular Ca2+ at the egg posterior via osmotic swelling, possibly through mechano-sensitive Ca2+ channels; a single Ca2+ wave then propagates in an actin dependent manner; this Ca2+ wave co-ordinates key developmental events including resumption of the cell cycle and initiation of translation of mRNAs such as bicoid.

Ordan, E., Brankatschk, M., Dickson, B., Schnorrer, F. and Volk, T. (2015). Slit cleavage is essential for producing an active, stable, non-diffusible short-range signal that guides muscle migration. Development [Epub ahead of print]. PubMed ID: 25813540
During organogenesis, secreted signaling proteins direct cell migration towards their target tissue. In Drosophila embryos, developing muscles are guided by signals produced by tendons to promote the proper attachment of muscles to tendons, essential for proper locomotion. Previously, the repulsive protein Slit, secreted by tendon cells, has been proposed to be an attractant for muscle migration. However, this study demonstrates that through tight control of its distribution, Slit repulsion is used for both directing and arresting muscle migration. Slit cleavage was shown to restrict the distribution of Slit to tendon cells, allowing it to function as a short-range repellent that directs muscle migration and patterning, and promotes their halt upon reaching the target site. Mechanistically, Slit processing was shown to produce a rapidly degraded C-terminal fragment and an active, stable N-terminal polypeptide that was tethered to the tendon cell membrane, which further protects it from degradation. Consistently, the requirement for Slit processing could be bypassed by providing an uncleavable, membrane-bound form of Slit that is stable and is retained on expressing tendon cells. Moreover, muscle elongation appears to be extremely sensitive to Slit levels, as replacing the entire full-length Slit with the stable Slit-N-polypeptide resulted in excessive repulsion, which lead to a defective muscle pattern. These findings reveal a novel cleavage-dependent regulatory mechanism controlling Slit spatial distribution, which may operate in other Slit-dependent processes

Liu, N. and Lasko, P. (2015). Analysis of RNA interference lines identifies new functions of maternally-expressed genes involved in embryonic patterning in Drosophila melanogaster. G3 (Bethesda) [Epub ahead of print]. PubMed ID: 25834215
Embryonic patterning in Drosophila melanogaster is initially established through the activity of a number of maternally expressed genes that are expressed during oogenesis. mRNAs from some of these genes accumulate in the posterior pole plasm of the oocyte and early embryo, and localize further into RNA islands, transient ring-like structures that form around the nuclei of future primordial germ cells (pole cells) at stage 3 of embryogenesis. As mRNAs from several genes with known functions in anterior-posterior patterning and/or germ cell specification accumulate in RNA islands, it was hypothesized that some other mRNAs that localize in this manner might also function in these developmental processes. This study investigated the developmental functions of 51 genes whose mRNAs accumulate in RNA islands by abrogating their activity in the female germline using RNA interference. This analysis revealed requirements for ttk, pbl, Hip14, eIF5, eIF4G, and CG9977 for progression through early oogenesis. Dorsal appendage defects were observed in a proportion of eggs produced by females expressing double-stranded RNA targeting Mkrn1 or jvl, implicating these two genes in dorsal-ventral patterning. In addition, posterior patterning defects and a reduction in pole cell number were seen in the progeny of Mkrn1 females. As the mammalian orthologue of Mkrn1 acts as an E3 ubiquitin ligase, these results suggest an additional link between protein ubiquitination and pole plasm activity.

Holloway, D.M. and Spirov, A.V. (2015). Mid-embryo patterning and precision in Drosophila segmentation: Krüppel dual regulation of hunchback. PLoS One 10: e0118450. PubMed ID: 25793381
In early development, genes are expressed in spatial patterns which later define cellular identities and tissue locations. The mechanisms of such pattern formation have been studied extensively in early Drosophila embryos. The gap gene hunchback (hb) is one of the earliest genes to be expressed in anterior-posterior (AP) body segmentation. As a transcriptional regulator for a number of downstream genes, the spatial precision of hb expression can have significant effects in the development of the body plan. To investigate the factors contributing to hb precision, this study used fine spatial and temporal resolution data to develop a quantitative model for the regulation of hb expression in the mid-embryo. In particular, modelling hb pattern refinement in mid nuclear cleavage cycle 14 (NC14) revealed some of the regulatory contributions of simultaneously-expressed gap genes. Matching the model to recent data from wild-type (WT) embryos and mutants of the gap gene Krüppel (Kr) indicated that a mid-embryo Hb concentration peak important in thoracic development (at parasegment 4, PS4) is regulated in a dual manner by Kr, with low Kr concentration activating hb and high Kr concentration repressing hb. The processes of gene expression (transcription, translation, transport) are intrinsically random. Stochastic simulations were used to characterize the noise generated in hb expression. Kr regulation could limit the positional variability of the Hb mid-embryo border. This has been recently corroborated in experimental comparisons of WT and Kr- mutant embryos. Further, Kr regulation could decrease uncertainty in mid-embryo hb expression (i.e. contribute to a smooth Hb boundary) and decrease between-copy transcriptional variability within nuclei. Since many tissue boundaries are first established by interactions between neighbouring gene expression domains, these properties of Hb-Kr dynamics to diminish the effects of intrinsic expression noise may represent a general mechanism contributing to robustness in early development.

Friday, April 10th

Navarro, J.A., Botella, J.A., Metzendorf, C., Lind, M.I. and Schneuwly, S. (2015).. Mitoferrin modulates iron toxicity in a Drosophila model of Friedreich´s ataxia. Free Radic Biol Med [Epub ahead of print]. PubMed ID: 25841783
Friedreich's ataxia is the most important recessive ataxia in the Caucasian population. Loss of frataxin expression affects the production of iron-sulphur clusters and, therefore, mitochondrial energy production. One of the pathological consequences is an increase of iron transport into the mitochondrial compartment leading to a toxic accumulation of reactive iron. However, the mechanism underlying this inappropriate mitochondrial iron accumulation is still unknown. Control and frataxin-deficient flies were fed with an iron-diet in order to mimic an iron overload and used to assess various cellular as well as mitochondrial functions. This study shows that frataxin-deficient flies are hypersensitive towards dietary iron and developed an iron-dependent decay of mitochondrial functions. In the fly model exhibiting only partial frataxin loss, there was an inability to activate ferritin translation and the enhancement of mitochondrial iron uptake via mitoferrin upregulation were likely the key molecular events behind the iron-induced phenotype. Both defects are observed during the normal process of aging, confirming their importance in the progression of the pathology. In an effort to further assess the importance of these mechanisms, genetic interaction studies were carried out. Mitoferrin downregulation improved many of the frataxin-deficient conditions, including nervous system degeneration, whereas mitoferrin overexpression exacerbated most of them. Taken together, this study demonstrates the crucial role of mitoferrin dysfunction in the etiology of Friedreich´s ataxia and provides evidence that impairment of mitochondrial iron transport could be an effective treatment of the disease.

Cuesto, G., Jordan-Alvarez, S., Enriquez-Barreto, L., Ferrus, A., Morales, M. and Acebes, A. (2015). GSK3β inhibition promotes synaptogenesis in Drosophila and mammalian neurons. PLoS One 10: e0118475. PubMed ID: 25764078
The PI3K-dependent activation of AKT results in the inhibition of GSK3β in most signaling pathways. These kinases regulate multiple neuronal processes including the control of synapse number as shown for Drosophila and rodents. Alzheimer disease's patients exhibit high levels of circulating GSK3β and, consequently, pharmacological strategies based on GSK3beta antagonists have been designed. The approach, however, has yielded inconclusive results so far. This paper describes a comparative study in Drosophila and rats addressing the role of GSK3β in synaptogenesis. In flies, the genetic inhibition of the shaggy-encoded GSK3β increases the number of synapses, while its upregulation leads to synapse loss. Likewise, in three weeks cultured rat hippocampal neurons, the pharmacological inhibition of GSK3β increases synapse density and Synapsin (see Drosophila Synapsin) expression. However, experiments on younger cultures (12 days) yielded an opposite effect, a reduction of synapse density. This unexpected finding seems to unveil an age- and dosage-dependent differential response of mammalian neurons to the stimulation/inhibition of GSK3β, a feature that must be considered in the context of human adult neurogenesis and pharmacological treatments for Alzheimer's disease based on GSK3β antagonists.

Chen, N., Guo, A. and Li, Y. (2015). Aging accelerates memory extinction and impairs memory restoration in Drosophila. Biochem Biophys Res Commun [Epub ahead of print]. PubMed ID: 25842205
Age-related memory impairment (AMI) is a phenomenon observed from invertebrates to human. Memory extinction is proposed to be an active inhibitory modification of memory, however, whether extinction is affected in aging animals remains to be elucidated. Employing a modified paradigm for studying memory extinction in fruit flies, this study found that only the stable, but not the labile memory component was suppressed by extinction, thus effectively resulting in higher memory loss in aging flies. Strikingly, young flies are able to fully restore the stable memory component 3 h post extinction, while aging flies failed to do so. In conclusion, these findings reveal that both accelerated extinction and impaired restoration contribute to memory impairment in aging animals.

Begum, R., Calaza, K., Kam, J.H., Salt, T.E., Hogg, C. and Jeffery G. (2015). Near-infrared light increases ATP, extends lifespan and improves mobility in aged Drosophila melanogaster. Biol Lett 11 [Epub ahead of print]. PubMed ID: 25788488
Ageing is an irreversible cellular decline partly driven by failing mitochondrial integrity. Mitochondria accumulate DNA mutations and reduce ATP production necessary for cellular metabolism. This is associated with inflammation. Near-infrared exposure increases retinal ATP in old mice via cytochrome c oxidase absorption and reduces inflammation. In this study, fruit flies exposed daily to 670 nm radiation, revealed elevated ATP and reduced inflammation with age. Critically, there was a significant increase in average lifespan: 100-175% more flies survived into old age following 670 nm exposure and these had significantly improved mobility. This may be a simple route to extending lifespan and improving function in old age.

Thursday, April 9th

Birkholz, O., Rickert, C., Nowak, J., Coban, I.C. and Technau, G.M. (2015). Bridging the gap between postembryonic cell lineages and identified embryonic neuroblasts in the ventral nerve cord of Drosophila melanogaster. Biol Open [Epub ahead of print]. PubMed ID: 25819843
The clarification of complete cell lineages, which are produced by specific stem cells, is fundamental for understanding mechanisms, controlling the generation of cell diversity and patterning in an emerging tissue. In the developing Central Nervous System (CNS) of Drosophila, neural stem cells (neuroblasts) exhibit two periods of proliferation: During embryogenesis they produce primary lineages, which form the larval CNS. After a phase of mitotic quiescence, a subpopulation of them resumes proliferation in the larva to give rise to secondary lineages that build up the CNS of the adult fly. Within the ventral nerve cord (VNC) detailed descriptions exist for both primary and secondary lineages. However, while primary lineages have been linked to identified neuroblasts, the assignment of secondary lineages has so far been hampered by technical limitations. Therefore, primary and secondary neural lineages co-existed as isolated model systems. This study provides the missing link between the two systems for all lineages in the thoracic and abdominal neuromeres. Using the Flybow technique, embryonic neuroblasts were identified by their characteristic and unique lineages in the living embryo and their further development was traced into the late larval stage. This comprehensive analysis provides the first complete view of which embryonic neuroblasts are postembryonically reactivated along the anterior/posterior-axis of the VNC, and reveals the relationship between projection patterns of primary and secondary sublineages.

Liu, W. W., Mazor, O. and Wilson, R. I. (2015). Thermosensory processing in the Drosophila brain. Nature 519: 353-357. PubMed ID: 25739502
In Drosophila, just as in vertebrates, changes in external temperature are encoded by bidirectional opponent thermoreceptor cells: some cells are excited by warming and inhibited by cooling, whereas others are excited by cooling and inhibited by warming. The central circuits that process these signals are not understood. In Drosophila, a specific brain region receives input from thermoreceptor cells. This study shows that distinct genetically identified projection neurons (PNs) in this brain region are excited by cooling, warming, or both. The PNs excited by cooling receive mainly feed-forward excitation from cool thermoreceptors. In contrast, the PNs excited by warming ('warm-PNs') receive both excitation from warm thermoreceptors and crossover inhibition from cool thermoreceptors through inhibitory interneurons. Notably, this crossover inhibition elicits warming-evoked excitation, because warming suppresses tonic activity in cool thermoreceptors. This in turn disinhibits warm-PNs and sums with feed-forward excitation evoked by warming. Crossover inhibition could cancel non-thermal activity (noise) that is positively correlated among warm and cool thermoreceptor cells, while reinforcing thermal activity which is anti-correlated. These results show how central circuits can combine signals from bidirectional opponent neurons to construct sensitive and robust neural codes.

Frank, D. D., Jouandet, G. C., Kearney, P. J., Macpherson, L. J. and Gallio, M. (2015). Temperature representation in the Drosophila brain. Nature 519: 358-361. PubMed ID: 25739506
In Drosophila, rapid temperature changes are detected at the periphery by dedicated receptors forming a simple sensory map for hot and cold in the brain. However, flies show a host of complex innate and learned responses to temperature, indicating that they are able to extract a range of information from this simple input. This study defines the anatomical and physiological repertoire for temperature representation in the Drosophila brain. First, a photolabelling strategy was used to trace the connections that relay peripheral thermosensory information to higher brain centres, and show that they largely converge onto three target regions: the mushroom body, the lateral horn (both of which are well known centres for sensory processing) and the posterior lateral protocerebrum, a region that this study defines as a major site of thermosensory representation. Next, using in vivo calcium imaging, the thermosensory projection neurons selectively activated by hot or cold stimuli were described. Fast-adapting neurons display transient ON and OFF responses and track rapid temperature shifts remarkably well, while slow-adapting cell responses better reflect the magnitude of simple thermal changes. Unexpectedly, a population of broadly tuned cells were found that respond to both heating and cooling, and they were shown to be required for normal behavioural avoidance of both hot and cold in a simple two-choice temperature preference assay. Taken together, these results uncover a coordinated ensemble of neural responses to temperature in the Drosophila brain, demonstrate that a broadly tuned thermal line contributes to rapid avoidance behaviour, and illustrate how stimulus quality, temporal structure, and intensity can be extracted from a simple glomerular map at a single synaptic station.

Wang, L., Du, Y., Nomura, Y. and Dong, K. (2015). Distinct modulating effects of TipE-homologs 2-4 on Drosophila sodium channel splice variants. Insect Biochem Mol Biol [Epub ahead of print]. PubMed ID: 25744892
The Drosophila melanogaster TipE protein is thought to be an insect sodium channel auxiliary subunit functionally analogous to the β subunits of mammalian sodium channels. Besides TipE, four TipE-homologous proteins (TEH1-TEH4) have been identified. It has been reported that TipE and TEH1 have both common and distinct effects on the gating properties of splice variants of the Drosophila sodium channel, DmNav (Para)D. However, limited information is available on the effects of TEH2, TEH3 and TEH4 on the function of DmNav channel variants. This study found that TEH2 increased the amplitude of peak current, but did not alter the gating properties of three examined DmNav splice variants expressed in Xenopus oocytes. In contrast, TEH4 had no effect on peak current, yet altered the gating properties of all three channel variants. Furthermore, TEH4 enhanced persistent current and slowed sodium current decay. The effects of TEH3 on DmNav variants are similar to those of TEH4, but the data were collected from a small portion of oocytes because co-expression of TEH3 with DmNav variants generated a large leak current in the majority of oocytes examined. In addition, TEH3 and TEH4 enhanced the expression of endogenous currents in oocytes. Taken together, these results reveal distinct roles of TEH proteins in modulating the function of sodium channels and suggest that TEH proteins might provide an important layer of regulation of membrane excitability in vivo. These results also raise an intriguing possibility of TEH3/TEH4 as auxiliary subunits of other voltage-gated ion channels besides sodium channels.

Wednesday, April 8th

Kim, Y. J., Igiesuorobo, O., Ramos, C. I., Bao, H., Zhang, B. and Serpe, M. (2015). Prodomain removal enables Neto to stabilize glutamate receptors at the Drosophila neuromuscular junction. PLoS Genet 11: e1004988. PubMed ID: 25723514
Stabilization of neurotransmitter receptors at postsynaptic specializations is a key step in the assembly of functional synapses. Drosophila Neto (Neuropillin and Tolloid-like protein) is an essential auxiliary subunit of ionotropic glutamate receptor (iGluR) complexes required for the iGluRs clustering at the neuromuscular junction (NMJ). This study shows that optimal levels of Neto are crucial for stabilization of iGluRs at synaptic sites and proper NMJ development. Genetic manipulations of Neto levels shifted iGluRs distribution to extrajunctional locations. Perturbations in Neto levels also produced small NMJs with reduced synaptic transmission, but only Neto-depleted NMJs showed diminished postsynaptic components. Drosophila Neto contains an inhibitory prodomain that is processed by Furin1-mediated limited proteolysis. neto null mutants rescued with a Neto variant that cannot be processed have severely impaired NMJs and reduced iGluRs synaptic clusters. Unprocessed Neto retains the ability to engage iGluRs in vivo and to form complexes with normal synaptic transmission. However, Neto prodomain must be removed to enable iGluRs synaptic stabilization and proper postsynaptic differentiation.

Lee, C. H., Blackband, S. J. and Fernandez-Funez, P. (2015). Visualization of synaptic domains in the Drosophila brain by magnetic resonance microscopy at 10 micron isotropic resolution. Sci Rep 5: 8920. PubMed ID: 25753480
Understanding the complex architecture, connectivity, and pathology of the human brain is a major application of magnetic resonance imaging (MRI). However, the cellular basis of MR signal is still poorly understood. The advent of MR microscopy (MRM) enables imaging biological samples at cellular resolution, helping to interpret the nature of MR signal at the cellular level. In this regard, the small Drosophila brain can reveal key aspects of MR signal through the visualization of complex, intact neuronal structures in their native spatial arrangement. Applying state-of-the-art MR technology, this study imaged fixed Drosophila heads at 10 mμm isotropic resolution by two endogenously contrasted MRM sequences. The improved MRM sensitivity described in this study delivered the highest 3D resolution of an intact animal head reported so far. 3D fast low angle shot (FLASH) revealed strong signal in most internal tissues, particularly in the brain cortex, which contains the cell bodies of neurons and glia. Remarkably, 3D diffusion weighted imaging (DWI) delivered unprecedented contrast within the modular brain neuropil, revealing hyperintense signal in synapse-rich microdomains. Thus, the complex Drosophila brain revealed unknown features of FLASH and DWI with potential applications in characterizing the structure and pathology of the mammalian brain.

Zhao, G., Wu, Y., Du, L., Li, W., Xiong, Y., Yao, A., Wang, Q. and Zhang, Y. Q. (2015). Drosophila S6 kinase like inhibits neuromuscular junction growth by downregulating the BMP receptor Thickveins. PLoS Genet 11: e1004984. PubMed ID: 25748449
Synaptic connections must be precisely controlled to ensure proper neural circuit formation. In Drosophila melanogaster, bone morphogenetic protein (BMP) promotes growth of the neuromuscular junction (NMJ) by binding and activating the BMP ligand receptors Wishful thinking (Wit) and Thickveins (Tkv) expressed in motor neurons. This paper reports that an evolutionally conserved, previously uncharacterized member of the S6 kinase (S6K) family S6K like (S6KL; Protein kinase-like 17E) acts as a negative regulator of BMP signaling. S6KL null mutants are viable and fertile but exhibit more satellite boutons, fewer and larger synaptic vesicles, larger spontaneous miniature excitatory junctional potential (mEJP) amplitudes, and reduced synaptic endocytosis at the NMJ terminals. Reducing the gene dose by half of tkv in S6KL mutant background reversed the NMJ overgrowth phenotype. The NMJ phenotypes of S6KL mutants were accompanied by an elevated level of Tkv protein and phosphorylated Mad, an effector of the BMP signaling pathway, in the nervous system. In addition, Tkv physically interacted with S6KL in cultured S2 cells. Furthermore, knockdown of S6KL enhanced Tkv expression, while S6KL overexpression downregulated Tkv in cultured S2 cells. This latter effect was blocked by the proteasome inhibitor MG132. These results together demonstrate for the first time that S6KL regulates synaptic development and function by facilitating proteasomal degradation of the BMP receptor Tkv.

Ueda, A. and Wu, C. F. (2015). The role of cAMP in synaptic homeostasis in response to environmental temperature challenges and hyperexcitability mutations. Front Cell Neurosci 9: 10. PubMed ID: 25698925
Homeostasis is the ability of physiological systems to regain functional balance following environment or experimental insults and synaptic homeostasis has been demonstrated in various species following genetic or pharmacological disruptions. Among environmental challenges, homeostatic responses to temperature extremes are critical to animal survival under natural conditions. Axon terminal arborization in Drosophila larval neuromuscular junctions (NMJs) has been shown to be enhanced at elevated temperatures; however, the amplitude of excitatory junctional potentials (EJPs) remains unaltered despite the increase in synaptic bouton numbers. This study determines the cellular basis of this homeostatic adjustment in larvae reared at high temperature (HT, 29 ° C). Synaptic current focally recorded from individual synaptic boutons was unaffected by rearing temperature (<15 ° C to >30 ° C). However, HT rearing decreased the quantal size (amplitude of spontaneous miniature EJPs, or mEJPs), which compensates for the increased number of synaptic releasing sites to retain a normal EJP size. The quantal size decrease is accounted for by a decrease in input resistance of the postsynaptic muscle fiber, indicating an increase in membrane area that matches the synaptic growth at HT. Interestingly, a mutation in rutabaga (rut) encoding adenylyl cyclase (AC) exhibited no obvious changes in quantal size or input resistance of postsynaptic muscle cells after HT rearing, suggesting an important role for rut AC in temperature-induced synaptic homeostasis in Drosophila. This study extends previous finding of rut-dependent synaptic homeostasis in hyperexcitable mutants, e.g., slowpoke (slo). In slo larvae, the lack of BK channel function is partially ameliorated by upregulation of presynaptic Shaker (Sh) IA current to limit excessive transmitter release in addition to postsynaptic glutamate receptor recomposition that reduces the quantal size.

Tuesday, April 7th

Ge, W., Deng, Q., Guo, (2015). Regulation of pattern formation and gene amplification during Drosophila oogenesis by the miR-318 microRNA. Genetics [Epub ahead of print]. PubMed ID: 25786856
Pattern formation during epithelial development requires the coordination of multiple signaling pathways. This study investigates the functions of an ovary-enriched miRNA, miR-318, in epithelial development during Drosophila oogenesis. miR-318 maternal loss-of-function mutants are female sterile and lay eggs with abnormal morphology. Removal of miR-318 disrupts the dorsal-anterior follicle cell patterning, resulting in abnormal dorsal appendages. miR-318 mutant females also produced thin and fragile eggshells, due to impaired chorion gene amplification. The study provided evidence that the ecdysone signaling pathway activates expression of miR-318 and that miR-318 cooperates with Tramtrack69 (Ttk69) to control the switch from endocycling to chorion gene amplification during differentiation of the follicular epithelium. The multiple functions of miR-318 in oogenesis illustrate the importance of miRNAs in maintaining cell fate and promoting the developmental transition in the female follicular epithelium.

Zaballos, M. A., Cantero, W. and Azpiazu, N. (2015). The TALE transcription factor Homothorax functions to assemble heterochromatin during Drosophila embryogenesis. PLoS One 10: e0120662. PubMed ID: 25794008
Previously studies have identified Homothorax (Hth) as an important factor for the correct assembly of the pericentromeric heterochromatin during the first fast syncytial divisions of the Drosophila embryo. These studies have been extended to later stages of embryonic development. It was shown that hth mutants exhibit a drastic overall reduction in the tri-methylation of H3 in Lys9, with no reduction of the previous di-methylation. One phenotypic outcome of such a reduction is a genome instability visualized by the many DNA breaks observed in the mutant nuclei. Moreover, loss of Hth leads to the opening of closed heterochromatic regions, including the rDNA genomic region. These data show that the satellite repeats get transcribed in wild type embryos and that this transcription depends on the presence of Hth, which binds to them as well as to the rDNA region. This work indicates that there is an important role of transcription of non-coding RNAs for constitutive heterochromatin assembly in the Drosophila embryo, and suggests that Hth plays an important role in this process.

Feltzin, V. L., Khaladkar, M., Abe, M., Parisi, M., Hendriks, G. J., Kim, J. and Bonini, N. M. (2015). The exonuclease Nibbler regulates age-associated traits and modulates piRNA length in Drosophila. Aging Cell [Epub ahead of print]. PubMed ID: 25754031
Nibbler (Nbr) is a 3'-to-5' exonuclease that trims the 3'end of microRNAs (miRNAs) to generate different length patterns of miRNAs in Drosophila. Despite its effect on miRNAs, its biological significance and whether Nbr affects other classes of small RNAs such as piRNAs and endo-siRNAs is not know. This study characterized the in vivo function of nbr by defining the Nbr protein expression pattern and loss-of-function effects. Nbr protein is enriched in the ovary and head. Analysis of nbr null animals reveals adult-stage defects that progress with age, including held-up wings, decreased locomotion, and brain vacuoles, indicative of accelerated age-associated processes upon nbr loss. Importantly, these effects depend on catalytic residues in the Nbr exonuclease domain, indicating that the catalytic activity is responsible for these effects. Given the impact of nbr on miRNAs, the effect of nbr on piRNA and endo-siRNA lengths was examined by deep-sequence analysis of libraries from ovaries. As with miRNAs, nbr mutation leads to longer length piRNAs - an effect that was dependent on the catalytic residues of the exonuclease domain. These analyses indicate a role of nbr on age-associated processes and to modulate length of multiple classes of small RNAs including miRNAs and piRNAs in Drosophila.

Kim, G., Pai, C. I., Sato, K., Person, M. D., Nakamura, A. and Macdonald, P. M. (2015). Region-specific activation of oskar mRNA translation by inhibition of Bruno-mediated repression. PLoS Genet 11: e1004992. PubMed ID: 25723530
A complex program of translational repression, mRNA localization, and translational activation ensures that Oskar (Osk) protein accumulates only at the posterior pole of the Drosophila oocyte. Inappropriate expression of Osk disrupts embryonic axial patterning, and is lethal. A key factor in translational repression is Bruno (Bru), which binds to regulatory elements in the osk mRNA 3' UTR. After posterior localization of osk mRNA, repression by Bru must be alleviated. This study describes an in vivo assay system to monitor the spatial pattern of Bru-dependent repression, separate from the full complexity of osk regulation. This assay reveals a form of translational activation-region-specific activation-which acts regionally in the oocyte, is not mechanistically coupled to mRNA localization, and functions by inhibiting repression by Bru. It was also shown that Bru dimerizes, and mutations were identified that disrupt this interaction to test its role in vivo. Loss of dimerization does not disrupt repression, as might have been expected from an existing model for the mechanism of repression. However, loss of dimerization does impair regional activation of translation, suggesting that dimerization may constrain, not promote, repression. This work provides new insight into the question of how localized mRNAs become translationally active, showing that repression of osk mRNA is locally inactivated by a mechanism acting independent of mRNA localization.

Monday, April 6th

Moreno, E., Fernandez-Marrero, Y., Meyer, P. and Rhiner, C. (2015). Brain regeneration in Drosophila involves comparison of neuronal fitness. Curr Biol [Epub ahead of print]. PubMed ID: 25754635
Darwinian-like cell selection has been studied during development and cancer. Cell selection is often mediated by direct intercellular comparison of cell fitness, using "fitness fingerprints". In Drosophila, cells compare their fitness via several isoforms of the transmembrane protein Flower. This paper reports a study of the role of intercellular fitness comparisons during regeneration. Regeneration-competent organisms are traditionally injured by amputation, whereas in clinically relevant injuries such as local ischemia or traumatic injury, damaged tissue remains within the organ. It was reasoned that "Darwinian" interactions between old and newly formed tissues may be important in the elimination of damaged cells. A model of adult brain regeneration in Drosophila was used in which mechanical puncture activates regenerative neurogenesis based on damage-responsive stem cells. It was found that apoptosis after brain injury occurs in damage-exposed tissue located adjacent to zones of de novo neurogenesis. Injury-affected neurons start to express isoforms of the Flower cell fitness indicator protein not found on intact neurons.This change in the neuronal fitness fingerprint is required to recognize and eliminate such neurons. Moreover, apoptosis is inhibited if all neurons express "low-fitness" markers, showing that the availability of new and healthy cells drives tissue replacement. In summary, this study found that elimination of impaired tissue during brain regeneration requires comparison of neuronal fitness and that tissue replacement after brain damage is coordinated by injury-modulated fitness fingerprints. Intercellular fitness comparisons between old and newly formed tissues could be a general mechanism of regenerative tissue replacement.

Roberts, L., Leise, T. L., Noguchi, T., Galschiodt, A. M., Houl, J. H., Welsh, D. K. and Holmes, T. C. (2015). Light evokes rapid circadian network oscillator desynchrony followed by gradual phase retuning of synchrony. Curr Biol [Epub ahead of print]. PubMed ID: 25754644
Circadian neural circuits generate near 24-hr physiological rhythms that can be entrained by light to coordinate animal physiology with daily solar cycles. To examine how a circadian circuit reorganizes its activity in response to light, period (per) clock gene cycling was imaged for up to 6 days at single-neuron resolution in whole-brain explant cultures prepared from per-luciferase transgenic flies. Cultures subjected to a phase-advancing light pulse (LP) were comapred to cultures maintained in darkness (DD). In DD, individual neuronal oscillators in all circadian subgroups are initially well synchronized but then show monotonic decrease in oscillator rhythm amplitude and synchrony with time. The small ventral lateral neurons (s-LNvs) and dorsal lateral neurons (LNds) exhibit this decrease at a slower relative rate. In contrast, the LP evokes a rapid loss of oscillator synchrony between and within most circadian neuronal subgroups, followed by gradual phase retuning of whole-circuit oscillator synchrony. The LNds maintain high rhythmic amplitude and synchrony following the LP along with the most rapid coherent phase advance. Immunocytochemical analysis of Per shows that these dynamics in DD and LP are recapitulated in vivo. Anatomically distinct circadian neuronal subgroups vary in their response to the LP, showing differences in the degree and kinetics of their loss, recovery and/or strengthening of synchrony, and rhythmicity. Transient desynchrony appears to be an integral feature of light response of the Drosophila multicellular circadian clock. Individual oscillators in different neuronal subgroups of the circadian circuit show distinct kinetic signatures of light response and phase retuning.

Brent Chen, Q., Das, S., Visic, P., Buford, K. D., Zong, Y., Buti, W., Odom, K. R., Lee, H. and Leal, S. M. (2015). The Drosophila T-box transcription factor Midline functions within Insulin/Akt and c-Jun-N terminal kinase stress-reactive signaling pathways to regulate interommatial bristle bormation and cell survival. Mech Dev [Epub ahead of print]. PubMed ID: 25748605
It was recently reported that the T-box transcription factor midline (mid) functions within the Notch-Delta signaling pathway to specify sensory organ precursor (SOP) cell fates in early-staged pupal eye imaginal discs and to suppress apoptosis. From genetic and allelic modifier screens, it is now reported that mid interacts with genes downstream of the insulin receptor(InR)/Akt, c-Jun-N-terminal kinase (JNK) and Notch signaling pathways to regulate interommatidial bristle (IOB) formation and cell survival. One of the most significant mid-interacting genes identified from the modifier screen is dFOXO, a transcription factor exhibiting a nucleocytoplasmic subcellular distribution pattern. In common with dFOXO, Mid exhibits a nucleocytoplasmic distribution pattern within WT third-instar larval tissue homogenates. Because dFOXO is a stress-responsive factor, the effects of either oxidative or metabolic stress responses were assayed on modifying the mid mutant phenotype which is characterized by a 50% loss of IOBs within the adult compound eye. While metabolic starvation stress does not affect the mid mutant phenotype, either 1 mM paraquat or 20% coconut oil, oxidative stress inducers, partially suppresses the mid mutant phenotype resulting in a significant recovery of IOBs. Another significant mid-interacting gene was groucho (gro). Mid and Gro are predicted to act as corepressors of the enhancer-of-split gene complex downstream of Notch. Immunolabeling WT and dFOXO null third-instar larval eye-antennal imaginal discs with anti-Mid and anti-Engrailed (En) antibodies indicates that dFOXO is required to activate Mid and En expression within photoreceptor neurons of the eye disc. Taken together, these studies show that Mid and dFOXO serve as critical effectors of cell fate specification and survival within integrated Notch, InR/dAkt, and JNK signaling pathways during third-instar larval and pupal eye imaginal disc development.

Somers, J., Nguyen, J., Lumb, C., Batterham, P. and Perry, T. (2015). In vivo functional analysis of the Drosophila melanogaster nicotinic acetylcholine receptor Dα6 using the insecticide spinosad. Insect Biochem Mol Biol [Epub ahead of print]. PubMed ID: 25747007
The vinegar fly, Drosophila melanogaster, has been used to identify and manipulate insecticide resistance genes. The advancement of genome engineering technology and the increasing availability of pest genome sequences has increased the predictive and diagnostic capacity of the Drosophila model. The Drosophila model can be extended to investigate the basic biology of the interaction between insecticides and the proteins they target. Recently an in vivo system was developed that permits the expression and study of key insecticide targets, the nicotinic acetylcholine receptors (nAChRs; see nAChRα5, nAChRα6 and nAChRα7), in controlled genetic backgrounds. This system was used to study the interaction between the insecticide spinosad and a nAChR subunit, Dalpha6. Reciprocal chimeric subunits were created from Dα6 and Dα7, a subunit that does not respond to spinosad. Using the in vivo system, the Dα6/Dα7 chimeric subunits were tested for their capacity to respond to spinosad. Only the subunits containing the C-terminal region of Dα6 were able to respond to spinosad, thus confirming the importance this region for spinosad binding. A new incompletely dominant, spinosad resistance mechanism that may evolve in pest species is also examined. First generated using chemical mutagenesis, the Dα6P146S mutation was recreated using the Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas9 system, the first use of this technology to introduce a resistant mutation into a controlled genetic background. Both alleles present with the same incompletely dominant, spinosad resistance phenotype, proving the P146S replacement to be the causal mutation. The proximity of the P146S mutation to the conserved Cys-loop indicates that it may impair the gating of the receptor. The results of this study enhance the understanding of nAChR structure:function relationships.

Sunday, April 5th

Bretscher, A. J., Honti, V., Binggeli, O., Burri, O., Poidevin, M., Kurucz, E., Zsamboki, J., Ando, I. and Lemaitre, B. (2015). The Nimrod transmembrane receptor Eater is required for hemocyte attachment to the sessile compartment in Drosophila melanogaster. Biol Open [Epub ahead of print]. PubMed ID: 25681394
Eater is an EGF-like repeat transmembrane receptor of the Nimrod family and is expressed in Drosophila hemocytes. Eater was initially identified for its role in phagocytosis of both Gram-positive and Gram-negative bacteria. This study describes the deletion of eater and shows that it appears to be required for efficient phagocytosis of Gram-positive but not Gram-negative bacteria. However, the most striking phenotype of eater deficient larvae is the near absence of sessile hemocytes, both plasmatocyte and crystal cell types. The eater deletion is the first loss of function mutation identified that causes absence of the sessile hemocyte state. This study shows that Eater is required cell-autonomously in plasmatocytes for sessility. However, the presence of crystal cells in the sessile compartment requires Eater in plasmatocytes. eater deficient hemocytes exhibit a cell adhesion defect. Collectively, these data uncover a new requirement of Eater in enabling hemocyte attachment at the sessile compartment and points to a possible role of Nimrod family members in hemocyte adhesion.

Lee, K. A., Kim, B., Bhin, J., Kim do, H., You, H., Kim, E. K., Kim, S. H., Ryu, J. H., Hwang, D. and Lee, W. J. (2015). Bacterial uracil modulates Drosophila DUOX-dependent gut Immunity via Hedgehog-induced signaling endosomes. Cell Host Microbe 17: 191-204. PubMed ID: 25639794
Genetic studies in Drosophila have demonstrated that generation of microbicidal reactive oxygen species (ROS) through the NADPH dual oxidase (DUOX) is a first line of defense in the gut epithelia. Bacterial uracil acts as DUOX-activating ligand through poorly understood mechanisms. This study shows that the Hedgehog (Hh) signaling pathway modulates uracil-induced DUOX activation. Uracil-induced Hh signaling is required for intestinal expression of the calcium-dependent cell adhesion molecule Cadherin 99C (Cad99C) and subsequent Cad99C-dependent formation of endosomes. These endosomes play essential roles in uracil-induced ROS production by acting as signaling platforms for PLCβ/PKC/Ca(2+)-dependent DUOX activation. Animals with impaired Hh signaling exhibit abolished Cad99C-dependent endosome formation and reduced DUOX activity, resulting in high mortality during enteric infection. Importantly, endosome formation, DUOX activation, and normal host survival are restored by genetic reintroduction of Cad99C into enterocytes, demonstrating the important role for Hh signaling in host resistance to enteric infection.

Markus, R., Lerner, Z., Honti, V., Csordas, G., Zsamboki, J., Cinege, G., Parducz, A., Lukacsovich, T., Kurucz, E. and Ando, I. (2015). Multinucleated giant hemocytes are effector cells in cell-mediated immune responses of Drosophila. J Innate Immun [Epub ahead of print]. PubMed ID: 25659341
This study identified and characterized a so far unrecognized cell type, dubbed the multinucleated giant hemocyte (MGH), in the ananassae subgroup of Drosophilidae. The functional and ultrastructural characteristics of this novel blood cell type are described as well as its characterization with a set of discriminative immunological markers. MGHs are encapsulating cells that isolate and kill the parasite without melanization. They share some properties with but differ considerably from lamellocytes, the encapsulating cells of Drosophila melanogaster, the broadly used model organism in studies of innate immunity. MGHs are nonproliferative effector cells that are derived from phagocytic cells of the sessile tissue and the circulation, but do not exhibit phagocytic activity. In contrast to lamellocytes, MGHs are gigantic cells with filamentous projections and contain many nuclei, which are the result of the fusion of several cells. Although the structure of lamellocytes and MGHs differ remarkably, their function in the elimination of parasites is similar, which is potentially the result of the convergent evolution of interactions between hosts and parasites in different geographic regions. MGHs are highly motile and share several features with mammalian multinucleated giant cells, a syncytium of macrophages formed during granulomatous inflammation.

Chrostek, E. and Teixeira, L. (2015). Mutualism breakdown by amplification of wolbachia genes. PLoS Biol 13: e1002065. PubMed ID: 25668031
Most insect species are associated with vertically transmitted endosymbionts. Because of the mode of transmission, the fitness of these symbionts is dependent on the fitness of the hosts. Therefore, these endosymbionts need to control their proliferation in order to minimize their cost for the host. The genetic bases and mechanisms of this regulation remain largely undetermined. The maternally inherited bacteria of the genus Wolbachia are the most common endosymbionts of insects, providing some of them with fitness benefits. In Drosophila melanogaster, Wolbachia wMelPop is a unique virulent variant that proliferates massively in the hosts and shortens their lifespan. The genetic bases of wMelPop virulence are unknown, and their identification would allow a better understanding of how Wolbachia levels are regulated. This study shows that amplification of a region containing eight Wolbachia genes, called Octomom, is responsible for wMelPop virulence. Using Drosophila lines selected for carrying Wolbachia with different Octomom copy numbers, it was demonstrated that the number of Octomom copies determines Wolbachia titers and the strength of the lethal phenotype. Octomom amplification is unstable, and reversion of copy number to one reverts all the phenotypes. These results provide a link between genotype and phenotype in Wolbachia and identify a genomic region regulating Wolbachia proliferation. These results show that transition from a mutualist to a pathogen may occur because of a single genomic change in the endosymbiont. This implies that there must be constant selection on endosymbionts to control their densities.

Saturday, April 4th

Kao, J. Y., Zubair, A., Salomon, M. P., Nuzhdin, S. V. and Campo, D. (2015). Population genomic analysis uncovers African and European admixture in Drosophila melanogaster populations from the southeastern United States and Caribbean Islands. Mol Ecol. PubMed ID: 25735402
Drosophila melanogaster is postulated to have colonized North America in the past several hundred years in two waves. Flies from Europe colonized the east coast United States while flies from Africa inhabited the Caribbean, which if true, make the southeast US and Caribbean Islands a secondary contact zone for African and European D. melanogaster. This scenario has been proposed based on phenotypes and limited genetic data. In this study, individual whole genomes of flies from populations in the southeast US and Caribbean Islands were sequenced and these populations were examined in conjunction with population sequences from the west coast US, Africa, and Europe. West coast US populations were found to be closely related to the European population, likely reflecting a rapid westward expansion upon first settlements into North America. Genomic evidence of African and European admixture was found in southeast US and Caribbean populations, with a clinal pattern of decreasing proportions of African ancestry with higher latitude. This genomic analysis of D. melanogaster populations from the southeast US and Caribbean Islands provides more evidence for the Caribbean Islands as the source of previously reported novel African alleles found in other east coast US populations. It was also found the border between the southeast US and the Caribbean island to be the admixture hot zone where distinctly African-like Caribbean flies become genomically more similar to European-like southeast US flies. These findings have important implications for previous studies examining the generation of east coast US clines via selection.

Fabian, D. K., Lack, J. B., Mathur, V., Schlotterer, C., Schmidt, P. S., Pool, J. E. and Flatt, T. (2015). Spatially varying selection shapes life history clines among populations of Drosophila melanogaster from sub-Saharan Africa. J Evol Biol [Epub ahead of print]. PubMed ID: 25704153
Clines in life history traits, presumably driven by spatially varying selection, are widespread. Major latitudinal clines have been observed, for example, in Drosophila melanogaster, an ancestrally tropical insect from Africa that has colonized temperate habitats on multiple continents. Yet, how geographic factors other than latitude, such as altitude or longitude, affect life history in this species remains poorly understood. Moreover, most previous work has been performed on derived European, American and Australian populations, but whether life history also varies predictably with geography in the ancestral Afro-tropical range has not been investigated systematically. This study has examined life history variation among populations of D. melanogaster from sub-Saharan Africa. Viability and reproductive diapause were found to be invariant with geography, but body size increases with altitude, latitude and longitude. Early fecundity covaries positively with altitude and latitude, whereas lifespan showed the opposite trend. Examination of genetic variance-covariance matrices revealed geographic differentiation also in trade-off structure. Methods for attributing morphological variation to drift or selection compare variation in neutral markers between populations (FST) to the analogous measure of quantitative genetic variation underlying phenotypes between populations (QST). QST-FST analysis showed that life history differentiation among populations is likely shaped by selection. Together, these results suggest that geographic and/or climatic factors drive adaptive phenotypic differentiation among ancestral African populations and confirm the widely held notion that latitude and altitude represent parallel gradients.

Sun, W., Valero, M. C., Seong, K. M., Steele, L. D., Huang, I. T., Lee, C. H., Clark, J. M., Qiu, X. and Pittendrigh, B. R. (2015). A glycine insertion in the Estrogen-Related Receptor (ERR) is associated with enhanced expression of three cytochrome P450 genes in transgenic Drosophila melanogaster. PLoS One 10: e0118779. PubMed ID: 25761142
Insecticide-resistant Drosophila melanogaster strains represent a resource for the discovery of the underlying molecular mechanisms of cytochrome P450 constitutive over-expression, even if some of these P450s are not directly involved in the resistance phenotype. For example, in select 4,4'-dichlorodiphenyltrichloroethane (DDT) resistant strains the glucocorticoid receptor-like (GR-like) potential transcription factor binding motifs (TFBMs) have previously been shown to be associated with constitutively differentially-expressed cytochrome P450s, Cyp12d1, Cyp6g2 and Cyp9c1. However, insects are not known to have glucocorticoids. The only ortholog to the mammalian glucocorticoid receptor (GR) in D. melanogaster is an estrogen-related receptor (ERR) gene, which has two predicted alternative splice isoforms (ERRa and ERRb). Sequencing of ERRa and ERRb in select DDT susceptible and resistant D. melanogaster strains has revealed a glycine (G) codon insertion which was only observed in the ligand binding domain of ERR from the resistant strains tested (ERR-G). Transgenic flies, expressing the ERRa-G allele, constitutively over-expressed Cyp12d1, Cyp6g2 and Cyp9c1. Only Cyp12d1 and Cyp6g2 were over-expressed in the ERRb-G transgenic flies. Phylogenetic studies show that the G-insertion appeared to be located in a less conserved domain in ERR and this insertion is found in multiple species across the Sophophora subgenera.

Nadimpalli, S., Persikov, A. V. and Singh, M. (2015). Pervasive variation of transcription factor orthologs contributes to regulatory network evolution. PLoS Genet 11: e1005011. PubMed ID: 25748510
Differences in transcriptional regulatory networks underlie much of the phenotypic variation observed across organisms. Changes to cis-regulatory elements are widely believed to be the predominant means by which regulatory networks evolve, yet examples of regulatory network divergence due to transcription factor (TF) variation have also been observed. To systematically ascertain the extent to which TFs contribute to regulatory divergence, this study analyzed the evolution of the largest class of metazoan TFs, Cys2-His2 zinc finger (C2H2-ZF) TFs, across 12 Drosophila species spanning ~45 million years of evolution. Remarkably, a significant fraction of all C2H2-ZF 1-to-1 orthologs in flies were found to exhibit variations that can affect their DNA-binding specificities. In addition to loss and recruitment of C2H2-ZF domains, diverging DNA-contacting residues were found in ~44% of domains shared between D. melanogaster and the other fly species. These diverging DNA-contacting residues, found in ~70% of the D. melanogaster C2H2-ZF genes in this analysis and corresponding to ~26% of all annotated D. melanogaster TFs, show evidence of functional constraint: they tend to be conserved across phylogenetic clades and evolve slower than other diverging residues. These same variations were rarely found as polymorphisms within a population of D. melanogaster flies, indicating their rapid fixation. The predicted specificities of these dynamic domains gradually change across phylogenetic distances, suggesting stepwise evolutionary trajectories for TF divergence. Further, whereas proteins with conserved C2H2-ZF domains are enriched in developmental functions, those with varying domains exhibit no functional enrichments. This work suggests that a subset of highly dynamic and largely unstudied TFs are a likely source of regulatory variation in Drosophila and other metazoans.

Friday, April 3rd

Wang, C., Zhang, W., Yin, M.X., Hu, L., Li, P., Xu, J., Huang, H., Wang, S., Lu, Y., Wu, W., Ho, M.S., Li, L., Zhao, Y. and Zhang, L. (2015). Suppressor of Deltex mediates Pez degradation and modulates Drosophila midgut homeostasis. Nat Commun 6: 6607. PubMed ID: 25814387
Pez functions as an upstream negative regulator of Yorkie (Yki) to regulate intestinal stem cell (ISC) proliferation and is essential for the activity of the Hippo pathway specifically in the Drosophila midgut epithelium. This study reports that Suppressor of Deltex (Su(dx)) acts as a negative regulator of Pez. Su(dx) was shown to target Pez for degradation both in vitro and in vivo. Overexpression of Su(dx) induced proliferation in the fly midgut epithelium, which could be rescued by overexpressed Pez. The study also demonstrated that the interaction between Su(dx) and Pez, bridged by WW domains and PY/PPxY motifs, is required for Su(dx)-mediated Pez degradation. Furthermore, Kibra, a binding partner of Pez, was shown to stabilize Pez via WW-PY/PPxY interaction. Moreover, PTPN14, a Pez mammalian homolog, is degraded by overexpressed Su(dx) or Su(dx) homologue WWP1 in mammalian cells. These results reveal a previously unrecognized mechanism of Pez degradation in maintaining the homeostasis of Drosophila midgut.

Okada, H., Schittenhelm, R. B., Straessle, A. and Hafen, E. (2015). Multi-functional regulation of 4E-BP gene expression by the Ccr4-Not complex. PLoS One 10: e0113902. PubMed ID: 25793896
The mechanistic target of rapamycin (mTOR) signaling pathway is highly conserved from yeast to humans. It senses various environmental cues to regulate cellular growth and homeostasis. Deregulation of the pathway has been implicated in many pathological conditions including cancer. Phosphorylation cascades through the pathway have been extensively studied but not much is known about the regulation of gene expression of the pathway components. This study reports that the mRNA level of eukaryotic translation initiation factor (eIF) subunit 4E-binding protein (4E-BP) gene, one of the key mTOR signaling components, is regulated by the highly conserved Ccr4-Not complex. RNAi knockdown of Not1, a putative scaffold protein of this protein complex, increases the mRNA level of 4E-BP in Drosophila Kc cells. Examination of the gene expression mechanism using reporter swap constructs reveals that Not1 depletion increases reporter mRNAs with the 3'UTR of 4E-BP gene, but decreases the ones with the 4E-BP promoter region, suggesting that Ccr4-Not complex regulates both degradation and transcription of 4E-BP mRNA. These results indicate that the Ccr4-Not complex controls expression of a single gene at multiple levels and adjusts the magnitude of the total effect. Thus, this study study reveals a novel regulatory mechanism of a key component of the mTOR signaling pathway at the level of gene expression.

Morawa, K.S., Schneider, M. and Klein, T. (2015). Lgd regulates the activity of the BMP/Dpp signalling pathway during Drosophila oogenesis. Development 142: 1325-1335. PubMed ID: 25804739
The tumour suppressor gene lethal (2) giant discs (lgd) is involved in endosomal trafficking of transmembrane proteins in Drosophila. Loss of function results in the ligand-independent activation of the Notch pathway in all imaginal disc cells and follicle cells. Analysis of lgd loss of function has largely been restricted to imaginal discs and suggests that no other signalling pathway is affected. The devotion of Lgd to the Notch pathway was puzzling given that lgd loss of function also affects trafficking of components of other signalling pathways, such as the Dpp pathway. Moreover, Lgd physically interacts with Shrub, a fundamental component of the ESCRT trafficking machinery, whose loss of function results in the activation of several signalling pathways. This study shows that during oogenesis lgd loss of function causes ectopic activation of the Drosophila BMP signalling pathway. This activation occurs in somatic follicle cells as well as in germline cells. The activation in germline cells causes an extra round of division, producing egg chambers with 32 instead of 16 cells. Moreover, more germline stem cells are formed. The lgd mutant cells are defective in endosomal trafficking, causing an accumulation of the type I Dpp receptor Thickveins in maturing endosomes, which probably causes activation of the pathway. Taken together, these results show that lgd loss of function causes various effects among tissues and can lead to the activation of signalling pathways other than Notch. They further show that there is a role for the endosomal pathway during oogenesis.

Nguyen, H. Q., Nye, J., Buster, D. W., Klebba, J. E., Rogers, G. C. and Bosco, G. (2015). Drosophila Casein Kinase I alpha regulates homolog pairing and genome organization by modulating Condensin II subunit Cap-H2 levels. PLoS Genet 11: e1005014. PubMed ID: 25723539
The spatial organization of chromosomes within interphase nuclei is important for gene expression and epigenetic inheritance. Although the extent of physical interaction between chromosomes and their degree of compaction varies during development and between different cell-types, it is unclear how regulation of chromosome interactions and compaction relate to spatial organization of genomes. Drosophila is an excellent model system for studying chromosomal interactions including homolog pairing. Recent work has shown that condensin II governs both interphase chromosome compaction and homolog pairing and condensin II activity is controlled by the turnover of its regulatory subunit Cap-H2. Specifically, Cap-H2 is a target of the SCFSlimb E3 ubiquitin-ligase which down-regulates Cap-H2 in order to maintain homologous chromosome pairing, chromosome length and proper nuclear organization. This study identifies Casein Kinase I α (CK1α) as an additional negative-regulator of Cap-H2. CK1α-depletion stabilizes Cap-H2 protein and results in an accumulation of Cap-H2 on chromosomes. Similar to Slimb mutation, CK1α depletion in cultured cells, larval salivary gland, and nurse cells results in several condensin II-dependent phenotypes including dispersal of centromeres, interphase chromosome compaction, and chromosome unpairing. Moreover, CK1alpha loss-of-function mutations dominantly suppress condensin II mutant phenotypes in vivo. Thus, CK1alpha facilitates Cap-H2 destruction and modulates nuclear organization by attenuating chromatin localized Cap-H2 protein.

Thursday, April 2nd

Jensen, K., McClure, C., Priest, N.K. and Hunt, J. (2015). Sex-specific effects of protein and carbohydrate intake on reproduction but not lifespan in Drosophila melanogaster. Aging Cell [Epub ahead of print]. PubMed ID: 25808180
Modest dietary restriction extends lifespan (LS) in a diverse range of taxa and typically has a larger effect in females than males. Traditionally, this has been attributed to a stronger trade-off between LS and reproduction in females than in males that is mediated by the intake of calories. Recent studies, however, suggest that it is the intake of specific nutrients that extends LS and mediates this trade-off. This study used the geometric framework (GF) to examine the sex-specific effects of protein (P) and carbohydrate (C) intake on LS and reproduction in Drosophila melanogaster. This study found that LS was maximized at a high intake of C and a low intake of P in both sexes, whereas nutrient intake had divergent effects on reproduction. Male offspring production rate and LS were maximized at the same intake of nutrients, whereas female egg production rate was maximized at a high intake of diets with a P:C ratio of 1:2. This resulted in larger differences in nutrient-dependent optima for LS and reproduction in females than in males, as well as an optimal intake of nutrients for lifetime reproduction that differed between the sexes. Under dietary choice, the sexes followed similar feeding trajectories regulated around a P:C ratio of 1:4. Consequently, neither sex reached their nutritional optimum for lifetime reproduction, suggesting intralocus sexual conflict over nutrient optimization. This study shows clear sex differences in the nutritional requirements of reproduction in D. melanogaster and joins the growing list of studies challenging the role of caloric restriction in extending LS.

Lee, K. P. (2015). Dietary protein:carbohydrate balance is a critical modulator of lifespan and reproduction in Drosophila melanogaster: A test using a chemically defined diet. J Insect Physiol 75: 12-19. PubMed ID: 25728576
Macronutrient balance is an important determinant of fitness in many animals, including insects. Previous studies have shown that altering the concentrations of yeast and sugar in the semi-synthetic media has a profound impact on lifespan in Drosophila melanogaster, suggesting that dietary protein:carbohydrate (P:C) balance is the main driver of lifespan and ageing processes. However, since yeast is rich in multiple nutrients other than proteins, this lifespan-determining role of dietary P:C balance needs to be further substantiated through trials using a chemically-defined, synthetic diet. The present study investigated the effects of dietary P:C balance on lifespan and fecundity in female D. melanogaster flies fed on one of eight isocaloric synthetic diets differing in P:C ratio (0:1, 1:16, 1:8, 1:4, 1:2, 1:1, 2:1 or 4:1). Lifespan and dietary P:C ratio were related in a convex manner, with lifespan increasing to a peak at the two intermediate P:C ratios (1:2 and 1:4) and falling at the imbalanced ratios (0:1 and 4:1). Ingesting nutritionally imbalanced diets not only caused an earlier onset of senescence but also accelerated the age-dependent increase in mortality. Egg production was suppressed when flies were fed on a protein-deficient food (0:1), but increased with increasing dietary P:C ratio. Long-lived flies at the intermediate P:C ratios (1:2 and 1:4) stored a greater amount of lipids than those short-lived ones at the two imbalanced ratios (0:1 and 4:1). These findings provide a strong support to the notion that adequate dietary P:C balance is crucial for extending lifespan in D. melanogaster and offer new insights into how dietary P:C balance affects lifespan and ageing through its impacts on body composition.

Terhzaz, S., Teets, N. M., Cabrero, P., Henderson, L., Ritchie, M. G., Nachman, R. J., Dow, J. A., Denlinger, D. L. and Davies, S. A. (2015). Insect Capa neuropeptides impact desiccation and cold tolerance. Proc Natl Acad Sci U S A 112: 2882-2887. PubMed ID: 25730885
The success of insects is linked to their impressive tolerance to environmental stress, but little is known about how such responses are mediated by the neuroendocrine system. This study shows that the capability (capa) neuropeptide gene is a desiccation- and cold stress-responsive gene in diverse dipteran species. Using targeted in vivo gene silencing, physiological manipulations, stress-tolerance assays, and rationally designed neuropeptide analogs, this study demonstrate that the Drosophila melanogaster capa neuropeptide gene and its encoded peptides alter desiccation and cold tolerance. Knockdown of the capa gene increases desiccation tolerance but lengthens chill coma recovery time, and injection of Capa peptide analogs can reverse both phenotypes. Immunohistochemical staining suggests that Capa accumulates in the Capa-expressing Va neurons during desiccation and nonlethal cold stress but is not released until recovery from each stress. These results also suggest that regulation of cellular ion and water homeostasis mediated by Capa peptide signaling in the insect Malpighian (renal) tubules is a key physiological mechanism during recovery from desiccation and cold stress. This work augments understanding of how stress tolerance is mediated by neuroendocrine signaling and illustrates the use of rationally designed peptide analogs as agents for disrupting protective stress tolerance.

Hutchison, A. L., Maienschein-Cline, M., Chiang, A. H., Tabei, S. M., Gudjonson, H., Bahroos, N., Allada, R. and Dinner, A. R. (2015). Improved statistical methods enable greater sensitivity in rhythm detection for genome-wide data. PLoS Comput Biol 11: e1004094. PubMed ID: 25793520
Robust methods for identifying patterns of expression in genome-wide data are important for generating hypotheses regarding gene function. To this end, several analytic methods have been developed for detecting periodic patterns. One such method, JTK_CYCLE, was improved by explicitly calculating the null distribution such that it accounts for multiple hypothesis testing and by including non-sinusoidal reference waveforms. This method was termed empirical JTK_CYCLE with asymmetry search, and its performance was compared to JTK_CYCLE with Bonferroni and Benjamini-Hochberg multiple hypothesis testing correction, as well as to five other methods: cyclohedron test, address reduction, stable persistence, ANOVA, and F24. ANOVA, F24, and JTK_CYCLE were found to consistently outperform the other three methods when data are limited and noisy; empirical JTK_CYCLE with asymmetry search gives the greatest sensitivity while controlling for the false discovery rate. This analysis also provides insight into experimental design and, for a fixed number of samples, better sensitivity and specificity were achieved with higher numbers of replicates than with higher sampling density. Application of the methods to detecting circadian rhythms in a metadataset of microarrays that quantify time-dependent gene expression in whole heads of Drosophila melanogaster reveals annotations that are enriched among genes with highly asymmetric waveforms. These include a wide range of oxidation reduction and metabolic genes, as well as genes with transcripts that have multiple splice forms.

Wednesday, April 1st

Chambers, D.B., Androschuk, A., Rosenfelt, C., Langer, S., Harding, M. and Bolduc, F.V. (2015). Insulin signaling is acutely required for long-term memory in Drosophila. Front Neural Circuits 9: 8. PubMed ID: 25805973
Memory formation has been shown recently to be dependent on energy status in Drosophila. A well-established energy sensor is the insulin signaling (InS) pathway. Previous studies in various animal models including human have revealed the role of insulin levels in short-term memory but its role in long-term memory remains less clear. This study therefore investigated genetically the spatial and temporal role of InS using the olfactory learning and long-term memory model in Drosophila. InS was shown to be involved in both learning and memory. InS in the mushroom body was required for learning and long-term memory whereas long-term memory specifically was impaired after InS signaling disruption in the ellipsoid body, where it regulates the level of p70s6k, a downstream target of InS and a marker of protein synthesis. Finally, InS was shown to be acutely required for long-term memory formation in adult flies. This study shows that two key components, the Insulin receptor (InR) and the insulin receptor substrate (Chico) are required for normal learning and protein synthesis dependent memory but not for protein synthesis independent memory. InS in the ellipsoid body, a region highly connected and compared to the basal ganglia, is required for long-term memory and is one of the sites of protein synthesis in response to spaced training.

Lushchak, O. V., Carlsson, M. A. and Nassel, D. R. (2015). Food odors trigger an endocrine response that affects food ingestion and metabolism. Cell Mol Life Sci. PubMed ID: 25782410
This study demonstrated novel effects of food odors on food ingestion, metabolism and endocrine signaling in Drosophila melanogaster. Acute exposure to attractive vinegar odor triggers a rapid and transient increase in circulating glucose, and a rapid upregulation of genes encoding the glucagon-like hormone adipokinetic hormone (AKH), four insulin-like peptides (DILPs) and some target genes in peripheral tissues. Sustained exposure to food odors, however, decreases food intake. Hunger-induced strengthening of synaptic signaling from olfactory sensory neurons (OSNs) to brain neurons increases food-seeking behavior, and conversely fed flies display reduced food odor sensitivity and feeding. Increasing the strength of OSN signaling chronically by genetic manipulation of local peptide neuromodulation reduces feeding, elevates carbohydrates and diminishes lipids. Furthermore, constitutively strengthened odor sensitivity altered gene transcripts for AKH, DILPs and some of their targets. Thus, this study shows that food odor can induce a transient anticipatory endocrine response, and that boosted sensitivity to this odor affects food intake, as well as metabolism and hormonal signaling.

Klichko, V.I., Chow, E.S., Kotwica-Rolinska, J., Orr, W.C., Giebultowicz, J.M. and Radyuk, S.N. (2015). Aging alters circadian regulation of redox in Drosophila. Front Genet 6: 83. PubMed ID: 25806044
Circadian coordination of metabolism, physiology, and neural functions contributes to healthy aging and disease prevention. Clock genes govern the daily rhythmic expression of target genes whose activities underlie such broad physiological parameters as maintenance of redox homeostasis. Glutathione (GSH) biosynthesis has been shown to be controlled by the circadian system via effects of the clock genes on expression of the catalytic (Gclc) and modulatory (Gclm) subunits comprising the glutamate cysteine ligase (GCL) holoenzyme. The objective of this study was to determine whether and how aging, which leads to weakened circadian oscillations, affects the daily profiles of redox-active biomolecules. Fly aging was found to be associated with altered profiles of Gclc and Gclm expression at both the mRNA and protein levels. Analysis of free aminothiols and GCL activity revealed that aging abolishes daily oscillations in GSH levels and alters the activity of glutathione biosynthetic pathways. Unlike GSH, its precursors and products of catabolism, methionine, cysteine and cysteinyl-glycine, were not rhythmic in young or old flies, while rhythms of the glutathione oxidation product, GSSG, were detectable. It was concluded that the temporal regulation of GSH biosynthesis is altered in the aging organism and that age-related loss of circadian modulation of pathways involved in glutathione production is likely to impair temporal redox homeostasis.

Ma, L., Ma, J. and Xu, K. (2015). Effect of spaceflight on the circadian rhythm, lifespan and gene expression of Drosophila melanogaster. PLoS One 10: e0121600. PubMed ID: 25798821
Space travelers are reported to experience circadian rhythm disruption during spaceflight. However, how the space environment affects circadian rhythm is yet to be determined. The major focus of this study was to investigate the effect of spaceflight on the Drosophila circadian clock at both the behavioral and molecular level. China's Shenzhou-9 spaceship was used to carry Drosophila. After 13 days of spaceflight, behavior tests showed that the flies maintained normal locomotor activity rhythm and sleep pattern. The expression level and rhythm of major clock genes were also unaffected. However, expression profiling showed differentially regulated output genes of the circadian clock system between space flown and control flies, suggesting that spaceflight affected the circadian output pathway. Other physiological effects of spaceflight were also investigated such as lipid metabolism and lifespan, and genes were sought that were significantly affected by spaceflight using microarray analysis. These results provided new information on the effects of spaceflight on circadian rhythm, lipid metabolism and lifespan. Furthermore, studying the effect of spaceflight on gene expression using samples collected at different Zeitgeber time gave different results, suggesting the importance of appropriate sampling procedures in studies on the effects of spaceflight.

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