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


Thursday, March 31st, 2016

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Patel, M.V., Zhu, J.Y., Jiang, Z., Richman, A., VanBerkum, M.F. and Han, Z. (2016). Gia/Mthl5 is an aorta specific GPCR required for Drosophila heart tube morphology and normal pericardial cell positioning. Dev Biol [Epub ahead of print]. PubMed ID: 26994946
G-protein signaling is known to be required for cell-cell contacts during the development of the Drosophila dorsal vessel. However, the identity of the G protein-coupled receptor (GPCR) that regulates this signaling pathway activity is unknown. This study describes the identification of a novel cardiac specific GPCR, called Gia, for "GPCR in aorta". Gia is the only heart-specific GPCR identified in Drosophila to date and it is specifically expressed in cardioblasts that fuse at the dorsal midline to become the aorta. Gia is the only Drosophila gene so far identified for which expression is entirely restricted to cells of the aorta. Deletion of Gia leads to a broken-hearted phenotype, characterized by pericardial cells dissociated from cardioblasts and abnormal distribution of cell junction proteins. Both phenotypes are similar to those observed in mutants of the heterotrimeric cardiac G proteins. Lack of Gia also led to defects in the alignment and fusion of cardioblasts in the aorta. Gia forms a protein complex with G-αo47A, the alpha subunit of the heterotrimeric cardiac G proteins and interacts genetically with G-αo47A during cardiac morphogenesis. Gia acts as an essential aorta-specific GPCR that functions upstream of cardiac heterotrimeric G proteins and is required for morphological integrity of the aorta during heart tube formation. These studies lead to a redefinition of the bro phenotype, to encompass morphological integrity of the heart tube as well as cardioblast-pericardial cell spatial interactions.

Konogami, T., Yang, Y., Ogihara, M. H., Hikiba, J., Kataoka, H. and Saito, K. (2016). Ligand-dependent responses of the silkworm prothoracicotropic hormone receptor, Torso, are maintained by unusual intermolecular disulfide bridges in the transmembrane region. Sci Rep 6: 22437. PubMed ID: 26928300
The insect membrane-protein, Torso, is a member of the receptor-tyrosine-kinase family, and is activated by its ligand, prothoracicotropic hormone (PTTH). Although PTTH is one of the most important regulators of insect development, the mechanism of Torso activation by the hormone has remained elusive. In this study, using heterologous expression in cultured Drosophila S2 cells, ligand-independent dimerization of silkworm Torso was observed, and the receptor molecules in the dimer were found to be linked by intermolecular disulfide bridges. By examining the oligomerization states of several truncation and substitution mutants of Torso, atypical cysteine residues in the transmembrane region were identified as being responsible for the intermolecular linkage in the dimer. The replacement of all of the cysteines in the region with phenylalanines abolished the disulfide-bond-mediated dimerization; however, non-covalent dimerization of the mutant was detected using a cross-linking reagent, both with and without ligand stimulation. This non-covalent dimerization caused apparent receptor autophosphorylation independently of the ligand stimulation, but did not promote the ERK phosphorylation in the downstream signaling pathway. The unique Torso structure with the intermolecular disulfide bridges in the transmembrane region is necessary to maintain the ligand-dependent receptor functions of autophosphorylation and downstream activation.

Wang, Z. H., Clark, C. and Geisbrecht, E. R. (2016). Drosophila Clueless is involved in Parkin-dependent mitophagy by promoting VCP-mediated Marf degradation. Hum Mol Genet [Epub ahead of print]. PubMed ID: 26931463
PINK1/Parkin-mediated mitochondrial quality control (MQC) requires valosin-containing protein (VCP)-dependent Mitofusin/Marf degradation to prevent damaged organelles from fusing with the healthy mitochondrial pool, facilitating mitochondrial clearance by autophagy. Drosophila clueless (clu) was found to interact genetically with PINK1 and parkin to regulate mitochondrial clustering in germ cells. However, whether Clu acts in MQC has not been investigated. This study shows that overexpression of Drosophila Clu complements PINK1, but not parkin, mutant muscles. Loss of clu leads to the recruitment of Parkin, VCP/p97, p62/Ref(2)P and Atg8a to depolarized swollen mitochondria. However, clearance of damaged mitochondria is impeded. This paradox is resolved by the findings that excessive mitochondrial fission or inhibition of fusion alleviates mitochondrial defects and impaired mitophagy caused by clu depletion. Furthermore, Clu is upstream of and binds to VCP in vivo and promotes VCP-dependent Marf degradation in vitro. Marf accumulates in whole muscle lysates of clu-deficient flies and is destabilized upon Clu overexpression. Thus, Clu is essential for mitochondrial homeostasis and functions in concert with Parkin and VCP for Marf degradation to promote damaged mitochondrial clearance.

Chung, H. L., Augustine, G. J. and Choi, K. W. (2016). Drosophila Schip1 links Expanded and Tao-1 to regulate Hippo signaling. Dev Cell 36: 511-524. PubMed ID: 26954546
Regulation of organ size is essential in animal development, and Hippo (Hpo) signaling is a major conserved mechanism for controlling organ growth. In Drosophila, Hpo and Warts kinases are core components of this pathway and function as tumor suppressors by inhibiting Yorkie (Yki). Expanded (Ex) is a regulator of the Hpo activity, but how they are linked is unknown. This study shows that Schip1, a Drosophila homolog of the mammalian Schwannomin interacting protein 1 (SCHIP1), provides a link between Ex and Hpo. Ex is required for apical localization of Schip1 in imaginal discs. Schip1 is necessary for promoting membrane localization and phosphorylation of Hpo by recruiting the Hpo kinase Tao-1. Taking these findings together, it is concluded that Schip1 directly links Ex to Hpo signaling by recruiting Tao-1. This study provides insights into the mechanism of Tao-1 regulation and a potential growth control function for SCHIP1 in mammals.

Wednesday, March 30th

Meissner, G. W., Luo, S. D., Dias, B. G., Texada, M. J. and Baker, B. S. (2016). Sex-specific regulation of Lgr3 in Drosophila neurons. Proc Natl Acad Sci U S A [Epub ahead of print]. PubMed ID: 26884206
The development of sexually dimorphic morphology and the potential for sexually dimorphic behavior in Drosophila are regulated by the Fruitless (Fru) and Doublesex (Dsx) transcription factors. Several direct targets of Dsx have been identified, but direct Fru targets have not been definitively identified. This study shows that Drosophila leucine-rich repeat G protein-coupled receptor 3 (Lgr3) is regulated by Fru and Dsx in separate populations of neurons. Lgr3 is a member of the relaxin-receptor family and a receptor for Dilp8, necessary for control of organ growth. Lgr3 expression in the anterior central brain of males is inhibited by the B isoform of Fru, whose DNA binding domain interacts with a short region of an Lgr3 intron. Fru A and C isoform mutants had no observed effect on Lgr3 expression. The female form of Dsx (DsxF) separately up- and down-regulates Lgr3 expression in distinct neurons in the abdominal ganglion through female- and male-specific Lgr3 enhancers. Excitation of neural activity in the DsxF-up-regulated abdominal ganglion neurons inhibits female receptivity, indicating the importance of these neurons for sexual behavior. Coordinated regulation of Lgr3 by Fru and Dsx marks a point of convergence of the two branches of the sex-determination hierarchy.

Kessler, R., Tisserand, J., Font-Burgada, J., Reina, O., Coch, L., Attolini, C. S., Garcia-Bassets, I. and Azorin, F. (2015). dDsk2 regulates H2Bub1 and RNA polymerase II pausing at dHP1c complex target genes. Nat Commun 6: 7049. PubMed ID: 25916810
dDsk2 is a conserved extraproteasomal ubiquitin receptor that targets ubiquitylated proteins for degradation. This study reports that dDsk2 plays a nonproteolytic function in transcription regulation. dDsk2 interacts with the dHP1c complex, localizes at promoters of developmental genes and is required for transcription. Through the ubiquitin-binding domain, dDsk2 interacts with H2Bub1, a modification that occurs at dHP1c complex-binding sites. H2Bub1 is not required for binding of the complex; however, dDsk2 depletion strongly reduces H2Bub1. Co-depletion of the H2Bub1 deubiquitylase dUbp8/Nonstop suppresses this reduction and rescues expression of target genes. RNA polymerase II is strongly paused at promoters of dHP1c complex target genes and dDsk2 depletion disrupts pausing. Altogether, these results suggest that dDsk2 prevents dUbp8/Nonstop-dependent H2Bub1 deubiquitylation at promoters of dHP1c complex target genes and regulates RNA polymerase II pausing. These results expand the catalogue of nonproteolytic functions of ubiquitin receptors to the epigenetic regulation of chromatin modifications.

Blick, A.J., Mayer-Hirshfeld, I., Malibiran, B.R., Cooper, M.A., Martino, P.A., Johnson, J.E. and Bateman, J.R. (2016). The capacity to act in trans varies among Drosophila enhancers. Genetics [Epub ahead of print]. PubMed ID: 26984057
The interphase nucleus is organized such that genomic segments interact in cis, on the same chromosome, and in trans, between different chromosomes. In Drosophila and other Dipterans, extensive interactions are observed between homologous chromosomes, which can permit enhancers and promoters to communicate in trans. Enhancer action in trans has been observed for a handful of genes in Drosophila, but it is as yet unclear whether this is a general property of all enhancers or specific to a few. This study tests a collection of well-characterized enhancers for the capacity to act in trans. Specifically, 18 enhancers that are active in either the eye or wing disc of third instar Drosophila larvae were tested, and, using two different assays, it was found that each enhancer can act in trans. However, the degree to which trans-action was supported varied greatly between enhancers. Quantitative analysis of enhancer activity supports a model wherein an enhancer's strength of transcriptional activation is a major determinant of its ability to act in trans, but that additional factors may also contribute to an enhancer's trans-activity. In sum, these data suggest that a capacity to activate a promoter on a paired chromosome is common among Drosophila enhancers.

Voutev, R. and Mann, R. S. (2016). Streamlined scanning for enhancer elements in Drosophila melanogaster. Biotechniques 60: 141-144. PubMed ID: 26956092
Enhancer elements in most eukaryotic organisms are often positioned at a great distance away from the transcription start site of the gene they regulate. Complex three-dimensional chromatin organization and insulators usually guide and limit the range of an enhancer's regulatory activity to a specific genetic locus. Rigorous testing of an entire genomic locus is often required in order to uncover the complete set of cis-regulatory modules (CRMs) regulating a gene, especially those with complex and dynamic expression patterns. This study reports a fast and efficient method for enhancer element identification by scanning large genomic regions using transgenic reporter genes. Because of their relatively small size, genomic libraries in the form of CHORI-322 clones can be efficiently inserted into the genome using the phiC31 system. Since the CHORI-322 library virtually tiles the entire genome, it was reasoned that even a large gene locus of ~200 kb (from one insulator to another) could be scanned with ~10 CHORI-322 clones. The method allows for rapid and efficient scanning of any Drosophila genomic region as a first step toward identifying enhancer elements of a gene of interest at any developmental stage and tissue. It will complement other genome-wide tiling projects and computational approaches, thus significantly improving the identification and verification of putative CRMs.

Tuesday, March 29th

Vourekas, A., Alexiou, P., Vrettos, N., Maragkakis, M. and Mourelatos, Z. (2016). Sequence-dependent but not sequence-specific piRNA adhesion traps mRNAs to the germ plasm. Nature 531: 390-394. PubMed ID: 26950602
The conserved Piwi family of proteins and piwi-interacting RNAs (piRNAs) have a central role in genomic stability, which is inextricably linked to germ-cell formation, by forming Piwi ribonucleoproteins (piRNPs) that silence transposable elements. In Drosophila melanogaster and other animals, primordial germ-cell specification in the developing embryo is driven by maternal messenger RNAs and proteins that assemble into specialized messenger ribonucleoproteins (mRNPs) localized in the germ (pole) plasm at the posterior of the oogenesis. Maternal piRNPs, especially those loaded on the Piwi protein Aubergine (Aub), are transmitted to the germ plasm to initiate transposon silencing in the offspring germ line. The transport of mRNAs to the oocyte by midoogenesis is an active, microtubule-dependent process; mRNAs necessary for primordial germ-cell formation are enriched in the germ plasm at late oogenesis via a diffusion and entrapment mechanism, the molecular identity of which remains unknown. Aub is a central component of germ granule RNPs, which house mRNAs in the germ plasm, and interactions between Aub and Tudor are essential for the formation of germ granules. This study shows that Aub-loaded piRNAs use partial base-pairing characteristics of Argonaute RNPs to bind mRNAs randomly in Drosophila, acting as an adhesive trap that captures mRNAs in the germ plasm, in a Tudor-dependent manner. Notably, germ plasm mRNAs in drosophilids are generally longer and more abundant than other mRNAs, suggesting that they provide more target sites for piRNAs to promote their preferential tethering in germ granules. Thus, complexes containing Tudor, Aub piRNPs and mRNAs couple piRNA inheritance with germline specification. These findings reveal an unexpected function for piRNP complexes in mRNA trapping that may be generally relevant to the function of animal germ granules.

Patel, P. H., Barbee, S. A. and Blankenship, J. T. (2016). GW-bodies and P-bodies constitute two separate pools of sequestered non-translating RNAs. PLoS One 11: e0150291. PubMed ID: 26930655
Non-translating RNAs that have undergone active translational repression are culled from the cytoplasm into P-bodies for decapping-dependent decay or for sequestration. Organisms that use microRNA-mediated RNA silencing have an additional pathway to remove RNAs from active translation. Consequently, proteins that govern microRNA-mediated silencing, such as GW182/Gw and AGO1, are often associated with the P-bodies of higher eukaryotic organisms. Due to the presence of Gw, these structures have been referred to as GW-bodies. However, several reports have indicated that GW-bodies have different dynamics to P-bodies. This study used live imaging to examine GW-body and P-body dynamics in the early Drosophila melanogaster embryo. While P-bodies are present throughout early embryonic development, cytoplasmic GW-bodies only form in significant numbers at the midblastula transition. Unlike P-bodies, which are predominantly cytoplasmic, GW-bodies are present in both nuclei and the cytoplasm. RNA decapping factors such as DCP1, Me31B, and Hpat are not associated with GW-bodies, indicating that P-bodies and GW-bodies are distinct structures. Furthermore, known Gw interactors such as AGO1 and the CCR4-NOT deadenylation complex, which have been shown to be important for Gw function, are also not present in GW-bodies. Use of translational inhibitors puromycin and cycloheximide, which respectively increase or decrease cellular pools of non-translating RNAs, alter GW-body size, underscoring that GW-bodies are composed of non-translating RNAs. Taken together, these data indicate that active translational silencing most likely does not occur in GW-bodies. Instead GW-bodies most likely function as repositories for translationally silenced RNAs. Finally, inhibition of zygotic gene transcription is unable to block the formation of either P-bodies or GW-bodies in the early embryo, suggesting that these structures are composed of maternal RNAs.

Besnard-Guerin, C., Jacquier, C., Pidoux, J., Deddouche, S. and Antoniewski, 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, this study showed that the Cricket Paralysis virus VSR CrPV1-A but not the Flock House virus B2 VSR abolishes silencing by miRNAs loaded into the Ago-2 protein. Thus, the results provide the first evidence that insect VSR have the potential to directly interfere with the miRNA silencing pathway.

Sridharan, V., Heimiller, J., Robida, M. D. and Singh, R. (2016). High throughput sequencing identifies misregulated genes in the Drosophila polypyrimidine tract-binding protein (hephaestus) mutant defective in spermatogenesis. PLoS One 11: e0150768. PubMed ID: 26942929
The Drosophila polypyrimidine tract-binding protein (dmPTB or Hephaestus) plays an important role during spermatogenesis. The heph2 mutation in this gene results in a specific defect in spermatogenesis, causing aberrant spermatid individualization and male sterility. However, the array of molecular defects in the mutant remains uncharacterized. This study has identified transcripts that are misregulated in this mutant. Aberrant transcripts show altered expression levels, exon skipping, and alternative 5' ends. This analysis shows misregulation of transcripts that have been connected to spermatogenesis, including components of the actomyosin cytoskeletal apparatus. It was shown, for example, that the Myosin light chain 1 (Mlc1) transcript is aberrantly spliced. Furthermore, bioinformatics analysis reveals that Mlc1 contains a high affinity binding site(s) for dmPTB/Hephaestus and that the site is conserved in many Drosophila species. Thus Mlc1 and other components of the actomyosin cytoskeletal apparatus offer important molecular links between the loss of dmPTB function and the observed developmental defect in spermatogenesis. This study provides the first comprehensive list of genes misregulated in vivo in the heph2 mutant in Drosophila and offers insight into the role of dmPTB during spermatogenesis.

Monday, March 28th

Yuan, K. and O'Farrell, P. H. (2016). TALE-light imaging reveals maternally guided, H3K9me2/3-independent emergence of functional heterochromatin in Drosophila embryos. Genes Dev 30: 579-593. PubMed ID: 26915820
Metazoans start embryogenesis with a relatively naive genome. The transcriptionally inert, late-replicating heterochromatic regions, including the constitutive heterochromatin on repetitive sequences near centromeres and telomeres, need to be re-established during development. To explore the events initiating heterochromatin formation and examine their temporal control, sequence specificity, and immediate regulatory consequence, a live imaging approach was established that enabled visualization of steps in heterochromatin emergence on specific satellite sequences during the mid-blastula transition (MBT) in Drosophila. Unexpectedly, only a subset of satellite sequences, including the 359-base-pair (bp) repeat sequence, recruited HP1a at the MBT. The recruitment of HP1a to the 359-bp repeat was dependent on HP1a's chromoshadow domain but not its chromodomain and was guided by maternally provided signals. HP1a recruitment to the 359-bp repeat was required for its programmed shift to later replication, and ectopic recruitment of HP1a was sufficient to delay replication timing of a different repeat. The results reveal that emergence of constitutive heterochromatin follows a stereotyped developmental program in which different repetitive sequences use distinct interactions and independent pathways to arrive at a heterochromatic state. This differential emergence of heterochromatin on various repetitive sequences changes their replication order and remodels the DNA replication schedule during embryonic development.

Jung, Y.L., Kang, H., Park, P.J. and Kuroda, M.I. (2016). Correspondence of Drosophila Polycomb Group proteins with broad H3K27me3 silent domains. Fly (Austin) [Epub ahead of print]. PubMed ID: 26940990
The Polycomb group (PcG) proteins are key conserved regulators of development, initially discovered in Drosophila and now strongly implicated in human disease. Nevertheless, differing silencing properties between the Drosophila and mammalian PcG systems have been observed. While specific DNA targeting sites for PcG proteins called Polycomb response elements (PREs) have been identified only in Drosophila, involvement of non-coding RNAs for PcG targeting has been favored in mammals. Another difference lies in the distribution patterns of PcG proteins. In mouse and human cells, PcG proteins show broad distributions, significantly overlapping with H3K27me3 domains. In contrast, only sharp peaks on PRE regions are observed for most PcG proteins in Drosophila, raising the question of how large domains of H3K27me3, up to many tens of kilobases, are formed and maintained in Drosophila. This study provides evidence that PcG distributions on silent chromatin in Drosophila are considerably broader than previously detected. Using BioTAP-XL, a chromatin crosslinking and tandem affinity purification approach, a broad, rather than PRE-limited overlap of PcG proteins with H3K27me3 was found, suggesting a conserved spreading mechanism for PcG in flies and mammals.

Kwok, R. S., Lam, V. H. and Chiu, J. C. (2016). Understanding the role of chromatin remodeling in the regulation of circadian transcription in Drosophila. Fly (Austin): [Epub ahead of print]. PubMed ID: 26926115
Circadian clocks enable organisms to anticipate daily changes in the environment and coordinate temporal rhythms in physiology and behavior with the 24-hour day-night cycle. The robust cycling of circadian gene expression is critical for proper timekeeping, and is regulated by transcription factor binding, RNA polymerase II (RNAPII) recruitment and elongation, and post-transcriptional mechanisms. Recently, it has become clear that dynamic alterations in chromatin landscape at the level of histone posttranslational modification and nucleosome density facilitate rhythms in transcription factor recruitment and RNAPII activity, and are essential for progression through activating and repressive phases of circadian transcription. This study discusses the characterization of the Brahma (Brm) chromatin-remodeling protein in Drosophila in the context of circadian clock regulation. By dissecting its catalytic vs. non-catalytic activities, a model is proposed in which the non-catalytic activity of Brm functions to recruit repressive factors to limit the transcriptional output of Clock (Clk) during the active phase of circadian transcription, while the primary function of the ATP-dependent catalytic activity is to tune and prevent over-recruitment of negative regulators by increasing nucleosome density. Finally, ongoing efforts and investigative directions towards a deeper mechanistic understanding of transcriptional regulation of circadian gene expression at the chromatin level are described.

Akan, I., Love, D. C., Harwood, K., Bond, M. R. and Hanover, J. A. (2016). Drosophila O-GlcNAcase deletion globally perturbs chromatin O-GlcNAcylation. J Biol Chem [Epub ahead of print]. PubMed ID: 26957542
O-GlcNAc Transferase (OGT/SXC) is essential for Polycomb repression suggesting that the O-GlcNAcylation of proteins plays a key role in regulating development. OGT transfers O-GlcNAc onto serine and threonine residues in intrinsically disordered domains of key transcriptional regulators; O-GlcNAcase (OGA) removes the modification. To pinpoint genomic regions that are regulated by O-GlcNAc levels, ChIP-chip and microarray analysis analysis was performed after OGT or OGA RNAi knockdown in S2 cells. After OGA RNAi, a genome-wide increase was observed in the intensity of most O-GlcNAc-occupied regions. In contrast, O-GlcNAc levels were strikingly insensitive to OGA RNAi at sites of polycomb repression. Microarray analysis suggested that altered O-GlcNAc cycling perturbed the expression of genes associated with morphogenesis and cell cycle regulation. A viable null allele of oga (ogadel.1) was produced in Drosophila allowing visualization of altered O-GlcNAc cycling on polytene chromosomes. Trithorax (Trx), Absent small or homeotic discs 1 (Ash1) and Compass member Set1 histone methyl-transferases were O-GlcNAc-modified in ogadel.1 mutants. The ogadel.1 mutants displayed altered expression of a distinct set of cell cycle related genes. These results show that the loss of Oga in Drosophila globally impacts the epigenetic machinery allowing O-GlcNAc accumulation on RNA Polymerase II and numerous chromatin factors including Trx, Ash1 and Set1.

Sunday, March 27th

Svetec, N., Cridland, J. M., Zhao, L. and Begun, D. J. (2016). The adaptive significance of natural genetic variation in the DNA damage response of Drosophila melanogaster. PLoS Genet 12: e1005869. PubMed ID: 26950216
Despite decades of work, understanding of the distribution of fitness effects of segregating genetic variants in natural populations remains largely incomplete. One form of selection that can maintain genetic variation is spatially varying selection, such as that leading to latitudinal clines. While the introduction of population genomic approaches to understanding spatially varying selection has generated much excitement, little successful effort has been devoted to moving beyond genome scans for selection to experimental analysis of the relevant biology and the development of experimentally motivated hypotheses regarding the agents of selection; it remains an interesting question as to whether the vast majority of population genomic work will lead to satisfying biological insights. Motivated by population genomic results, this study investigated how spatially varying selection in the genetic model system, Drosophila melanogaster, has led to genetic differences between populations in several components of the DNA damage response. UVB (middle wave length ultraviolet light) incidence, which is negatively correlated with latitude, is an important agent of DNA damage. Sensitivity of early embryos to UVB exposure is strongly correlated with latitude such that low latitude populations show much lower sensitivity to UVB. It was then shown that lines with lower embryo UVB sensitivity also exhibit increased capacity for repair of damaged sperm DNA by the oocyte. A comparison of the early embryo transcriptome in high and low latitude embryos provides evidence that one mechanism of adaptive DNA repair differences between populations is the greater abundance of DNA repair transcripts in the eggs of low latitude females. Finally, population genomic comparisons of high and low latitude samples was used to reveal evidence that multiple components of the DNA damage response and both coding and non-coding variation likely contribute to adaptive differences in DNA repair between populations.

Nikhil, K. L., Ratna, K. and Sharma, V. K. (2016). Life-history traits of Drosophila melanogaster populations exhibiting early and late eclosion chronotypes. BMC Evol Biol 16: 46. PubMed ID: 26922082
The hypothesis that circadian clocks confer adaptive advantage to organisms has been proposed based on its ubiquity across almost all levels of complexity and organization of life-forms. This thought has received considerable attention, and studies employing diverse strategies have attempted to investigate it. However, only a handful of them have examined how selection for circadian clock controlled rhythmic behaviors influences life-history traits which are known to influence Darwinian fitness. The 'early' and 'late' chronotypes are amongst the most widely studied circadian phenotypes; however, life-history traits associated with these chronotypes, and their consequences on Darwinian fitness remain largely unexplored, primarily due to the lack of a suitable model system. Several life-history traits of Drosophila melanogaster populations were studied that were subjected to laboratory selection for morning (early) and evening (late) emergence. This paper reports that the late eclosion chronotypes evolved longer pre-adult duration as compared to the early eclosion chronotypes both under light/dark (LD) and constant dark (DD) conditions, and these differences appear to be mediated by both clock dependent and independent mechanisms. Furthermore, longer pre-adult duration in the late chronotypes does not lead to higher body-mass at pupariation or eclosion, but the late females were significantly more fecund and lived significantly shorter as compared to the early females. It is concluded that coevolution of multiple life-history traits in response to selection on timing of eclosion highlights correlations of the genetic architecture governing timing of eclosion with that of fitness components which suggests that timing ecologically relevant behaviors at specific time of the day might confer adaptive advantage.

Wu, X., Li, R., Li, Q., Bao, H. and Wu, C. (2016). Comparative transcriptome analysis among parental inbred and crosses reveals the role of dominance gene expression in heterosis in Drosophila melanogaster. Sci Rep 6: 21124. PubMed ID: 26928435
Heteroses for body weight was observed in Drosophila melanogaster after generating hybrids from three inbred lines. To better understand the mechanism for this phenomenon at the mRNA level, the mRNA profiles of the parental and hybrid lines were compared using high-throughput RNA-seq. A total of 5877 differentially expressed genes (DEGs) were found and about 92% of these exhibited parental expression level dominance. Genes in the dominance category were functionally characterized using the Kyoto Encyclopedia of Genes and Genomes (KEGG) and the gene classifications offered by the Gene Ontology (GO) Consortium. The analysis identified genes associated with crucial processes such as development and growth in all three crosses. Functional assignments involving aminoglycan metabolism, starch and sucrose metabolism, and galactose metabolism are significantly overrepresented amongst the 215 common dominance DEGs. It is concluded that dominance DEGs are important in heteroses in Drosophila melanogaster and contribute specifically to body weight heterosis.
Debelle, A., Ritchie, M. G. and Snook, R. R. (2016). Sexual selection and assortative mating: an experimental test. J Evol Biol [Epub ahead of print]. PubMed ID: 26970522
Mate choice and mate competition can both influence the evolution of sexual isolation between populations. Assortative mating may arise if traits and preferences diverge in step, and, alternatively, mate competition may counteract mating preferences and decrease assortative mating. This study examined potential assortative mating between populations of Drosophila pseudoobscura that have experimentally evolved under either increased ('polyandry') or decreased ('monogamy') sexual selection intensity for 100 generations. These populations have evolved differences in numerous traits, including a male signal and female preference traits. A 2 males: 1 female design was used, allowing both mate choice and competition to influence mating outcomes, to test for assortative mating between the populations. Mating latency shows subtle effects of male and female interactions, with females from the monogamous populations appearing reluctant to mate with males from the polyandrous populations. However, males from the polyandrous populations have a significantly higher probability of mating regardless of the female's population. These results suggest that if populations differ in the intensity of sexual selection, effects on mate competition may overcome mate choice.

Saturday, March 26th

Slaninova, V., Krafcikova, M., Perez-Gomez, R., Steffal, P., Trantirek, L., Bray, S. J. and Krejci, A. (2016). Notch stimulates growth by direct regulation of genes involved in the control of glycolysis and the tricarboxylic acid cycle. Open Biol 6. PubMed ID: 26887408
Glycolytic shift is a characteristic feature of rapidly proliferating cells, such as cells during development and during immune response or cancer cells, as well as of stem cells. It results in increased glycolysis uncoupled from mitochondrial respiration, also known as the Warburg effect. Notch signalling is active in contexts where cells undergo glycolytic shift. This study tested whether metabolic genes are direct transcriptional targets of Notch signalling and whether upregulation of metabolic genes can help Notch to induce tissue growth under physiological conditions and in conditions of Notch-induced hyperplasia. Genes mediating cellular metabolic changes towards the Warburg effect were shown to be direct transcriptional targets of Notch signalling. They include genes encoding proteins involved in glucose uptake, glycolysis, lactate to pyruvate conversion and repression of the tricarboxylic acid cycle. Even a short pulse of Notch activity is able to elicit long-lasting metabolic changes resembling the Warburg effect. Loss of Notch signalling in Drosophila wing discs as well as in human microvascular cells leads to downregulation of glycolytic genes. Notch-driven tissue overgrowth can be rescued by downregulation of genes for glucose metabolism. Notch activity is able to support growth of wing during nutrient-deprivation conditions, independent of the growth of the rest of the body. Notch is active in situations that involve metabolic reprogramming, and the direct regulation of metabolic genes may be a common mechanism that helps Notch to exert its effects in target tissues.

Cannell, E., Dornan, A. J., Halberg, K. A., Terhzaz, S., Dow, J. A. and Davies, S. A. (2016). The corticotropin-releasing factor-like diuretic hormone 44 (DH) and kinin neuropeptides modulate desiccation and starvation tolerance in Drosophila melanogaster. Peptides [Epub ahead of print]. PubMed ID: 26896569
Malpighian tubules are critical organs for epithelial fluid transport and stress tolerance in insects, and are under neuroendocrine control by multiple neuropeptides secreted by identified neurons. This study demonstrates roles for CRF-like diuretic hormone 44 (DH44) and Leukokinin (Lk) in desiccation and starvation tolerance. Gene expression and labelled DH44 ligand binding data, as well as highly selective knockdowns and/or neuronal ablations of DH44 in neurons of the pars intercerebralis and DH44 receptor (DH44-R2) in Malpighian tubule principal cells, indicate that suppression of DH44 signalling improves desiccation tolerance of the intact fly. Leucokinin receptor (Lkr), is expressed in DH44 neurons as well as in stellate cells of the Malpighian tubules. Lkr knockdown in DH44-expressing neurons reduces Malpighian tubule-specific Lkr, suggesting interactions between DH44 and LK signalling pathways. Finally, although a role for DK in desiccation tolerance was not defined, a novel role was demonstrated for Malpighian tubule cell-specific Lkr in starvation tolerance. Starvation increases gene expression of epithelial LKR. Also, Malpighian tubule stellate cell-specific knockdown of LKR significantly reduced starvation tolerance, demonstrating a role for neuropeptide signalling during starvation stress.

Chaturvedi, R., Luan, Z., Guo, P. and Li, H. S. (2016). Drosophila vision depends on carcinine uptake by an organic cation transporter. Cell Rep 14: 2076-2083. PubMed ID: 26923590
Recycling of neurotransmitters is essential for sustained neuronal signaling, yet recycling pathways for various transmitters, including histamine, remain poorly understood. In the first visual ganglion (lamina) of Drosophila, photoreceptor-released histamine is taken up into perisynaptic glia, converted to carcinine, and delivered back to the photoreceptor for histamine regeneration. This study identified an organic cation transporter, CarT (carcinine transporter), that transports carcinine into photoreceptors during histamine recycling. CarT mediated in vitro uptake of carcinine. Deletion of the CarT gene caused an accumulation of carcinine in laminar glia accompanied by a reduction in histamine, resulting in abolished photoreceptor signal transmission and blindness in behavioral assays. These defects were rescued by expression of CarT cDNA in photoreceptors, and they were reproduced by photoreceptor-specific CarT knockdown. These findings suggest a common role for the conserved family of CarT-like transporters in maintaining histamine homeostasis in both mammalian and fly brains.

Morrow, G., et al. (2016). Changes in Drosophila mitochondrial proteins following chaperone-mediated lifespan extension confirm a role of Hsp22 in mitochondrial UPR and reveal a mitochondrial localization for cathepsin D. Mech Ageing Dev 155: 36-47. PubMed ID: 26930296
Hsp22 is a small mitochondrial heat shock protein (sHSP) preferentially up-regulated during aging in Drosophila. Its developmental expression is strictly regulated and it is rapidly induced in conditions of stress. Hsp22 is one of the few sHSP to be localized inside mitochondria, and is the first sHSP to be involved in the mitochondrial unfolding protein response (UPRMT) together with Hsp60, mitochondrial Hsp70 and TRAP1. The UPRMT is a pro-longevity mechanism, and interestingly Hsp22 over-expression by-itself increases lifespan and resistance to stress. Among the proteins influenced by Hsp22 expression were proteins from the electron transport chain (ETC), the TCA cycle and mitochondrial Hsp70. Hsp22 co-migrates with ETC components and its over-expression is associated with an increase in mitochondrial protease activity. Interestingly, the only protease that showed significant changes upon Hsp22 over-expression was cathepsin D, which is localized in mitochondria in addition to lysosome in D. melanogaster as evidenced by cellular fractionation. Together the results are consistent with a role of Hsp22 in the UPRMT and in mitochondrial proteostasis.

Friday, March 25th

Andersen, D. and Horne-Badovinac, S. (2016). Influence of ovarian muscle contraction and oocyte growth on egg chamber elongation in Drosophila. Development [Epub ahead of print]. PubMed ID: 26952985
Organs are formed from multiple cell types that make distinct contributions to their shape. The Drosophila egg chamber provides a tractable model to dissect such contributions during morphogenesis. Egg chambers are comprised of 16 germ cells (GCs) surrounded by a somatic epithelium. Initially spherical, these structures elongate as they mature. This morphogenesis is thought to occur through a "molecular corset" mechanism, wherein structural elements within the epithelium become circumferentially organized perpendicular to the elongation axis and resist the expansive growth of the GCs to promote elongation. Whether this epithelial organization provides the hypothesized constraining force has been difficult to discern, however, and a role for GC growth has not been demonstrated. This study provides evidence for this mechanism by altering the contractile activity of the tubular muscle sheath that surrounds developing egg chambers. Muscle hypo-contraction indirectly reduces GC growth and shortens the egg, which demonstrates the necessity of GC growth for elongation. Conversely, muscle hyper-contraction enhances the elongation program. Although this is an abnormal function for this muscle, this observation suggests that a corset-like force from the egg chamber's exterior could promote its lengthening. These findings highlight how physical contributions from several cell types are integrated to shape an organ.

Monahan, A.J. and Starz-Gaiano, M. (2016). Apontic regulates somatic stem cell numbers in Drosophila testes. BMC Dev Biol 16: 5. PubMed ID: 26993259
Microenvironments called niches maintain resident stem cell populations by balancing self-renewal with differentiation, but the genetic regulation of this process is unclear. The niche of the Drosophila testis is well-characterized and genetically tractable, making it ideal for investigating the molecular regulation of stem cell biology. The JAK/STAT pathway, activated by signals from a niche component called the hub, maintains both germline and somatic stem cells. This study investigated the molecular regulation of the JAK/STAT pathway in the stem cells of the Drosophila testis. The transcriptional regulator Apontic (Apt) was found to act in the somatic (cyst) stem cells (CySCs) to balance differentiation and maintenance. Apt functions as a negative feedback inhibitor of STAT activity, which enables cyst cell maturation. Simultaneous loss of the STAT regulators apt and Socs36E, or the Stat92E-targeting microRNA miR-279, expanded the somatic stem cell-like population. Genetic analysis revealed that a conserved genetic regulatory network limits JAK/STAT activity in the somatic stem cells of Drosophila testis. In these cells, JAK/STAT signaling was found to promote apt expression. Then, Apt functions through Socs36E and miR-279 to attenuate pathway activation, which is required for timely CySC differentiation. The study proposes that Apt acts as a core component of a STAT-regulatory circuit to prevent stem cell overpopulation and allow stem cell maturation.

Aurich, F. and Dahmann, C. (2016). A mutation in fat2 uncouples tissue elongation from global tissue rotation. Cell Rep [Epub ahead of print]. PubMed ID: 26972006
Global tissue rotation was proposed as a morphogenetic mechanism controlling tissue elongation. In Drosophila ovaries, global tissue rotation of egg chambers coincides with egg chamber elongation. Egg chamber rotation has been put forward to result in circumferential alignment of extracellular fibers. These fibers serve as molecular corsets to restrain growth of egg chambers perpendicular to the anteroposterior axis, thereby leading to the preferential egg chamber elongation along this axis. The atypical cadherin Fat2 is required for egg chamber elongation, rotation, and the circumferential alignment of extracellular fibers. This study generated a truncated form of Fat2 that lacks the entire intracellular region. fat2 mutant egg chambers expressing this truncated protein fail to rotate yet display normal extracellular fiber alignment and properly elongate. These data suggest that global tissue rotation, even though coinciding with tissue elongation, is not a necessary prerequisite for elongation.

Hunter, C.M., Robinson, M.C., Aylor, D.L. and Singh, N.D. (2016). Genetic background, maternal age and interaction effects mediate rates of crossing over in Drosophila melanogaster females. G3 (Bethesda) [Epub ahead of print]. PubMed ID: 26994290
Meiotic recombination is a genetic process that is critical for proper chromosome segregation in many organisms. Despite being fundamental for organismal fitness, rates of crossing over vary greatly between taxa. Both genetic and environmental factors contribute to phenotypic variation in crossover frequency, as do genotype-environment interactions. This study tests the hypothesis that maternal age influences rates of crossing over in a genotypic-specific manner. Using classical genetic techniques, the study estimates rates of crossing over for individual Drosophila melanogaster females from five strains over their lifetime from a single mating event. It was found that both age and genetic background significantly contribute to observed variation in recombination frequency, as do genotype-age interactions. Further, there exist differences in the effect of age on recombination frequency in the two genomic regions surveyed. These results highlight the complexity of recombination rate variation and reveal a new role of genotype by maternal age interactions in mediating recombination rate.

Thursday, March 24th

Afonso, D. J., Machado, D. R. and Koh, K. (2016). Control of sleep by a network of cell cycle genes. Fly (Austin): [Epub ahead of print]. PubMed ID: 26925838
Sleep is essential for health and cognition, but the molecular and neural mechanisms of sleep regulation are not well understood. The identification of Taranis (Tara) has recently been reported as a sleep-promoting factor that acts in a previously unknown arousal center in Drosophila. tara mutants exhibit a dose-dependent reduction in sleep amount of up to approximately 60%. Tara and its mammalian homologs, the Trip-Br (Transcriptional Regulators Interacting with PHD zinc fingers and/or Bromodomains) family of proteins, are primarily known as a transcriptional coregulators involved in cell cycle progression, and contain a conserved Cyclin-A (CycA) binding homology domain. This study found that tara and CycA synergistically promote sleep, and CycA levels are reduced in tara mutants. Additional data demonstrated that Cyclin-dependent kinase 1 (Cdk1) antagonizes tara and CycA to promote wakefulness. Moreover, a subset of CycA expressing neurons was identified in the pars lateralis, a brain region proposed to be analogous to the mammalian hypothalamus, as an arousal center. This article reports further characterization of tara mutants and provides an extended discussion of future directions within the framework of a working model, in which a network of cell cycle genes, tara, CycA, and Cdk1, interact in an arousal center to regulate sleep.

Slade, J. D. and Staveley, B. E. (2016). Manipulation of components that control feeding behavior in Drosophila melanogaster increases sensitivity to amino acid starvation. Genet Mol Res 15. PubMed ID: 26909968
Feeding is a complex behavior that is regulated by several internal mechanisms. Neuropeptides are able to survey quantities of stored energy and inform the organism if nutrient intake is required. Neuropeptide F (NPF), a homolog of the mammalian neuropeptide Y, acts to induce feeding within the homeostatic regulation of this behavior. Drosophila and other insects bear a shorter form of NPF known as short NPF (sNPF) that can influence feeding. A neural hormone regulator, the dopamine transporter (DAT), works to clear dopamine from the synapses. This action may manipulate the post-feeding reward circuit in that lowered dopamine levels depress feeding, and excess dopamine levels encourage feeding. This study overexpressed and impaired the activities of NPF, sNPF, and DAT in Drosophila and examined their ability to survive during conditions of amino acid starvation. Too much or too little NPF or sNPF, which are key players in homeostatic feeding regulation, leads to increased sensitivity to amino acid starvation and diminished survivorship when compared to controls. When DAT, a member of the post-feeding reward system, is either overexpressed or reduced via mutation, Drosophila has increased sensitivity to amino acid starvation. Taken together, these results indicate that subtle variation in the expression of key components of these systems impacts survivorship during adverse nutrient conditions.

Min, S. and Chung, J. (2016). Identification of a neural pathway governing satiety in Drosophila. BMB Rep [Epub ahead of print]. PubMed ID: 26949022
Satiety cues a feeding animal to cease further ingestion of food for protecting from excessive energy gain. Impaired control of satiety is often associated with feeding-related disorders such as obesity. This paper reports the identification of a neural pathway that expresses myoinhibitory peptide (MIP) critical for satiety responses in Drosophila. Targeted silencing of MIP neuron activity strikingly increased body weight (BW) through elevated food intake. Similarly, genetic disruption of the gene encoding MIP also elevated feeding and BW. Suppressing MIP pathway behaviorally transformed satiated flies to feed like the starved ones with augmented sensitivity to food. Conversely, temporal activation of MIP neuron markedly reduced food intake and BW, and blunted the sensitivity of starved flies to food as if they have been satiated. Shortly after termination of MIP neuron activation, the reduced BW was reverted to normal level with a strong feeding rebound. These results consistently suggest the switch-like role of MIP pathway in feeding by controlling satiety.

Poudel, S. and Lee, Y. (2016). Gustatory receptors required for avoiding the toxic compound coumarin in Drosophila melanogaster. Mol Cells [Epub ahead of print]. PubMed ID: 26912085
Coumarin is a phenolic compound that mainly affects the liver due to its metabolization into a toxic compound. The deterrent and ovicidal activities of coumarin in insect models such as Drosophila melanogaster have been reported. This study explored the molecular mechanisms by which these insects protect themselves and their eggs from this toxic plant metabolite. Coumarin was fatal to the flies in a dosage-dependent manner. However, coumarin feeding could be inhibited through activation of the aversive gustatory receptor neurons (GRNs), but not the olfactory receptor neurons. Furthermore, three gustatory receptors, GR33a, GR66a, and GR93a, functioned together in coumarin detection by the proboscis. However, GR33a, but not GR66a and GR93a, was required to avoid coumarin during oviposition, with a choice of the same substrates provided as in binary food choice assay. Taken together, these findings suggest that anti-feeding activity and oviposition to avoid coumarin occur via separate mechanisms.

Wednesday, March 23rd

Itoh, K., Akimoto, Y., Fuwa, T. J., Sato, C., Komatsu, A. and Nishihara, S. (2016). Mucin-type core 1 glycans regulate the localization of neuromuscular junctions and establishment of muscle cell architecture in Drosophila. Dev Biol [Epub ahead of print]. PubMed ID: 26896591
T antigen (Galβ1-3GalNAcα1-Ser/Thr), a core 1 mucin-type O-glycan structure, is synthesized by Drosophila core 1 β1,3-galactosyltrasferase 1 (dC1GalT1) and is expressed in various tissues. dC1GalT1 synthesizes T antigen expressed in hemocytes, lymph glands, and the central nervous system (CNS) and dC1GalT1 mutant larvae display decreased numbers of circulating hemocytes and excessive differentiation of hematopoietic stem cells in lymph glands. dC1GalT1 mutant larvae have also been shown to have morphological defects in the CNS. However, the functions of T antigen in other tissues remain largely unknown. This study found that glycans contributed to the localization of neuromuscular junction (NMJ) boutons. In dC1GalT1 mutant larvae, NMJs were ectopically formed in the cleft between muscles 6 and 7 and connected with these two muscles. dC1GalT1 synthesized T antigen, which was expressed at NMJs. In addition, the function of mucin-type O-glycans in muscle cells was determined. In dC1GalT1 mutant muscles, myofibers and basement membranes were disorganized. Moreover, ultrastructural defects in NMJs and accumulation of large endosome-like structures within both NMJ boutons and muscle cells were observed in dC1GalT1 mutants. Taken together, these results demonstrated that mucin-type O-glycans synthesized by dC1GalT1 were involved in the localization of NMJ boutons, synaptogenesis of NMJs, establishment of muscle cell architecture, and endocytosis.

Choudhury, S. D., Mushtaq, Z., Reddy-Alla, S., Balakrishnan, S. S., Thakur, R. S., Krishnan, K. S., Raghu, P., Ramaswami, M. and Kumar, V. (2016). σ2-adaptin facilitates basal synaptic transmission and is required for regenerating endo-exo cycling pool under high frequency nerve stimulation in Drosophila. Genetics [Epub ahead of print]. PubMed ID: 26920756
The functional requirement of AP2 complex (see AP-2α) in synaptic membrane retrieval by clathrin mediated endocytosis (CME) is not fully understood. This study isolated and functionally characterized a mutation that dramatically altered synaptic development. Based on the aberrant neuromuscular junction synapse, this mutation was named angur (a Hindi dialect meaning grapes). Loss-of-function alleles of angur show more than two-fold overgrowth in bouton numbers and dramatic decrease in bouton size. angur mutation was mapped to σ2-adaptin, the smallest subunit of the adapter complex 2. Reducing neuronal level of any of the subunits of AP2 complex or disrupting AP2 complex assembly in neurons phenocopied σ2-adaptin mutation. Genetic perturbation of σ2-adaptin in neurons leads to a reversible temperature sensitive paralysis at 38 degrees °C. Electrophysiological analysis of the mutants revealed reduced evoked junction potentials and quantal content. Interestingly, high frequency nerve stimulation caused prolonged synaptic fatigue at the NMJs. The synaptic level of subunits of AP2 complex and clathrin but not other endocytic proteins were reduced in the mutants. Moreover, the BMP/TGFβ signalling was altered in these mutants and was restored by normalizing σ2-adaptin in neurons. Thus, these data suggest that - 1) while σ2-adaptin facilitates SV recycling for basal synaptic transmission, its activity is also required for regenerating SV during high frequency nerve stimulation; and 2) σ2-adaptin regulates NMJ morphology by attenuating TGFβ signalling.

Summerville, J., Faust, J., Fan, E., Pendin, D., Daga, A., Formella, J., Stern, M. and McNew, J. A. (2016). The effects of ER morphology on synaptic structure and function in Drosophila melanogaster. J Cell Sci [Epub ahead of print]. PubMed ID: 26906425
Hereditary Spastic Paraplegia (HSP) is a set of genetic diseases caused by mutations in one of 72 genes that results in age-dependent corticospinal axon degeneration accompanied by spasticity and paralysis. Two genes implicated in HSPs encode proteins that regulate ER morphology. Atlastin (SPG3A) encodes an ER membrane fusion GTPase and Reticulon 2 (SPG12) helps shape ER tube formation. This study used a new fluorescent ER marker to show that the ER within wildtype Drosophila motor nerve terminals forms a network of tubules that is fragmented and made diffuse by atl loss. atl or Rtnl1 loss decreases evoked transmitter release and increases arborization. Similarly to other HSP genes, atl inhibits bone morphogenetic protein (BMP) signaling, and loss of atl causes age-dependent locomotor deficits in adults. These results demonstrate a critical role for ER in neuronal function and identify mechanistic links between ER morphology, neuronal function, BMP signaling, and adult behavior.

Uytterhoeven, V., Lauwers, E., Maes, I., Miskiewicz, K., Melo, M. N., Swerts, J., Kuenen, S., Wittocx, R., Corthout, N., Marrink, S. J., Munck, S. and Verstreken, P. (2015). Hsc70-4 deforms membranes to promote synaptic protein turnover by endosomal microautophagy. Neuron 88: 735-748. PubMed ID: 26590345
Synapses are often far from their cell bodies and must largely independently cope with dysfunctional proteins resulting from synaptic activity and stress. To identify membrane-associated machines that can engulf synaptic targets destined for degradation, a large-scale in vitro liposome-based screen was performed followed by functional studies. A presynaptically enriched chaperone Hsc70-4 was identified that bends membranes based on its ability to oligomerize. This activity promotes endosomal microautophagy and the turnover of specific synaptic proteins. Loss of microautophagy slows down neurotransmission while gain of microautophagy increases neurotransmission. Interestingly, Sgt, a cochaperone of Hsc70-4, is able to switch the activity of Hsc70-4 from synaptic endosomal microautophagy toward chaperone activity. Hence, Hsc70-4 controls rejuvenation of the synaptic protein pool in a dual way: either by refolding proteins together with Sgt, or by targeting them for degradation by facilitating endosomal microautophagy based on its membrane deforming activity.

Tuesday, March 22nd

Jin, M. and Mardon, G. (2016). Distinct biochemical activities of Eyes absent during Drosophila eye development. Sci Rep 6: 23228. PubMed ID: 26980695
Eyes absent (Eya) is a highly conserved transcriptional coactivator and protein phosphatase that plays vital roles in multiple developmental processes from Drosophila to humans. Eya proteins contain a PST (Proline-Serine-Threonine)-rich transactivation domain, a threonine phosphatase motif (TPM), and a tyrosine protein phosphatase domain. Using a genomic rescue system, this study finds that the PST domain is essential for Eya activity and Dac expression, and the TPM is required for full Eya function. The threonine phosphatase activity plays only a minor role during Drosophila eye development and the primary function of the PST and TPM domains is transactivation that can be largely substituted by the heterologous activation domain VP16. A primary function of Eya during Drosophila eye development is as a transcriptional coactivator. Moreover, the PST/TPM and the threonine phosphatase activity are not required for in vitro interaction between retinal determination factors. Finally, this work is the first report of an Eya-Ey physical interaction. These findings are particularly important because they highlight the need for an in vivo approach that accurately dissects protein function. 

Bernardo-Garcia, F.J., Fritsch, C. and Sprecher, S.G. (2016). The transcription factor glass links eye field specification with photoreceptor differentiation in Drosophila. Development [Epub ahead of print]. PubMed ID: 26952983
Eye development requires an evolutionarily conserved group of transcription factors, termed "retinal determination network" (RDN). However, little is known about the molecular mechanism by which the RDN instructs cells to differentiate into photoreceptors. This study shows that photoreceptor cell identity in Drosophila is critically regulated by the transcription factor Glass, which is primarily expressed in photoreceptors and whose role in this process was previously unknown. Glass is both required and sufficient for the expression of phototransduction proteins. Data demonstrate that the RDN member Sine oculis directly activates glass expression, and that Glass activates the expression of the transcription factors Hazy and Otd. Hazy was identified as a direct target of Glass. Induced expression of Hazy in the retina partially rescues the glass mutant phenotype. Together, these results provide a transcriptional link between eye field specification and photoreceptor differentiation in Drosophila, placing Glass at a central position in this developmental process.

Djabrayan, N.J. and Casanova, J. (2016). Snoo and Dpp act as spatial and temporal regulators respectively of adult progenitor cells in the Drosophila trachea. PLoS Genet 12: e1005909. PubMed ID: 26942411
Clusters of differentiated cells contributing to organ structures retain the potential to re-enter the cell cycle and replace cells lost during development or upon damage. To do so, they must be designated spatially and respond to proper activation cues. This study shows that in the case of Drosophila differentiated larval tracheal cells, progenitor potential is conferred by the spatially restricted activity of the Snoo transcription cofactor. Furthermore, Dpp signalling regulated by endocrine hormonal cues provides the temporal trigger for their activation. Finally, the study elucidates the genetic network elicited by Snoo and Dpp activity. These results illustrate a regulatory mechanism that translates intrinsic potential and extrinsic cues into the facultative stem cell features of differentiated progenitors.

Vollmer, J., Fried, P., Sánchez-Aragón, M., Lopes, C.S., Casares, F. and Iber, D. (2016). A quantitative analysis of growth control in the Drosophila eye disc. Development [Epub ahead of print]. PubMed ID: 26965369
The size and shape of organs is species-specific, and even in species in which organ size is strongly influenced by environmental cues, such as nutrition or temperature, it follows defined rules. Therefore, mechanisms must exist to ensure a tight control of organ size within a given species, while being flexible enough to allow for the evolution of different organ sizes in different species. This study combined computational modelling and quantitative measurements to analyse growth control in the Drosophila eye disc. It was found that the area growth rate declines inversely proportional to the increasing total eye disc area. Two growth laws were identified and found to be consistent with the growth data that would explain the extraordinary robustness and evolutionary plasticity of the growth process and thus of the final adult eye size. The study discusses how each of these laws constrains the set of candidate biological mechanisms for growth control in the Drosophila eye disc.

Monday, March 21st

Anvarian, Z., Nojima, H., van Kappel, E.C., Madl, T., Spit, M., Viertler, M., Jordens, I., Low, T.Y., van Scherpenzeel, R.C., Kuper, I., Richter, K., Heck, A.J., Boelens, R., Vincent, J.P., Rüdiger, S.G. and Maurice, M.M. (2016). Axin cancer mutants form nanoaggregates to rewire the Wnt signaling network. Nat Struct Mol Biol [Epub ahead of print]. PubMed ID: 26974125
Signaling cascades depend on scaffold proteins that regulate the assembly of multiprotein complexes. Missense mutations in scaffold proteins are frequent in human cancer, but their relevance and mode of action are poorly understood. This study shows that cancer point mutations in the scaffold protein Axin derail Wnt signaling and promote tumor growth in vivo through a gain-of-function mechanism. The effect is conserved for both the human and Drosophila proteins. Mutated Axin forms nonamyloid nanometer-scale aggregates decorated with disordered tentacles, which 'rewire' the Axin interactome. Importantly, the tumor-suppressor activity of both the human and Drosophila Axin cancer mutants is rescued by preventing aggregation of a single nonconserved segment. These findings establish a new paradigm for misregulation of signaling in cancer and show that targeting aggregation-prone stretches in mutated scaffolds holds attractive potential for cancer treatment.

Yamazaki, Y., Palmer, L., Alexandre, C., Kakugawa, S., Beckett, K., Gaugue, I., Palmer, R.H. and Vincent, J.P. (2016). Godzilla-dependent transcytosis promotes Wingless signalling in Drosophila wing imaginal discs. Nat Cell Biol [Epub ahead of print]. PubMed ID: 26974662
The apical and basolateral membranes of epithelia are insulated from each other, preventing the transfer of extracellular proteins from one side to the other. Thus, a signalling protein produced apically is not expected to reach basolateral receptors. Evidence suggests that Wingless, the main Drosophila Wnt, is secreted apically in the embryonic epidermis. However, in the wing imaginal disc epithelium, Wingless is mostly seen on the basolateral membrane where it spreads from secreting to receiving cells. This study examines the apico-basal movement of Wingless in Wingless-producing cells of wing imaginal discs. It was found that it is presented first on the apical surface before making its way to the basolateral surface, where it is released and allowed to interact with signalling receptors. Wingless transcytosis was shown to involve dynamin-dependent endocytosis from the apical surface. Subsequent trafficking from early apical endosomes to the basolateral surface requires Godzilla, a member of the RNF family of membrane-anchored E3 ubiquitin ligases. Without such transport, Wingless signalling is strongly reduced in this tissue.

Ristic, G., Tsou, W. L., Guzi, E., Kanack, A. J., Scaglione, K. M. and Todi, S. V. (2016). USP5 is dispensable for mono-ubiquitin maintenance in Drosophila. J Biol Chem [Epub ahead of print]. PubMed ID: 26917723
Ubiquitination is a post-translational modification that regulates most cellular pathways and processes, including degradation of proteins by the proteasome. Substrate ubiquitination is controlled at various stages, including through its reversal by deubiquitinases, or DUBs. A critical outcome of this process is the recycling of mono-ubiquitin. One DUB whose function has been proposed to include mono-ubiquitin recycling is Ubiquitin specific protease 5 (USP5). This study investigated whether Drosophila USP5 is important for maintaining mono-ubiquitin in vivo. The protein was found to have catalytic preferences similar to its human counterpart, and this DUB was found to be necessary during fly development. Biochemical and genetic experiments indicate that reduction of USP5 does not lead to mono-ubiquitin depletion in developing flies. Also, introduction of exogenous ubiquitin does not suppress developmental lethality caused by loss of endogenous USP5. This work indicates that a primary physiological role of USP5 is not to recycle mono-ubiquitin for reutilization, but that it may involve disassembly of conjugated ubiquitin to maintain proteasome function.

Mukunda, L., Miazzi, F., Sargsyan, V., Hansson, B. S. and Wicher, D. (2016). Calmodulin affects sensitization of Drosophila melanogaster odorant receptors. Front Cell Neurosci 10: 28. PubMed ID: 26903813
Odorant receptors (ORs) form heteromeric complexes of an odorant specific receptor protein (OrX) and a highly conserved co-receptor protein (Orco). The ORs form ligand gated ion channels that are tuned by intracellular signaling systems. Repetitive subthreshold odor stimulation of olfactory sensory neurons sensitizes insect ORs. This OR sensitization process requires Orco activity. This study first asked whether OR sensitization can be monitored with heterologously expressed OR proteins. Drosophila OR proteins expressed in CHO cells showed sensitization upon repeated weak stimulation. This was found for OR channels formed by Orco as well as by Or22a or Or56a and Orco. Moreover, inhibition of calmodulin (CaM) action on OR proteins, expressed in CHO cells, abolishes any sensitization. Finally, the sensitization phenomenon was investigated using an ex vivo preparation of olfactory sensory neurons (OSNs) expressing Or22a inside the fly's antenna. Using calcium imaging, sensitization was observed in the dendrites as well as in the soma. Inhibition of calmodulin with W7 disrupted the sensitization within the outer dendritic shaft, whereas the sensitization remained in the other OSN compartments. Taken together, these results suggest that CaM action is involved in sensitizing the OR complex and that this mechanisms accounts for the sensitization in the outer dendrites.

Sunday, March 20th

Parisky, K.M., Agosto Rivera, J.L., Donelson, N.C., Kotecha, S. and Griffith, L.C. (2016). Reorganization of sleep by temperature in Drosophila requires light, the homeostat, and the circadian clock. Curr Biol [Epub ahead of print]. PubMed ID: 26972320
Increasing ambient temperature reorganizes the Drosophila sleep pattern in a way similar to the human response to heat, increasing daytime sleep while decreasing nighttime sleep. Mutation of core circadian genes blocks the immediate increase in daytime sleep, but not the heat-stimulated decrease in nighttime sleep, when animals are in a light:dark cycle. The ability of per01 flies to increase daytime sleep in light:dark can be rescued by expression of PER in either LNv or DN1p clock cells and does not require rescue of locomotor rhythms. Prolonged heat exposure engages the homeostat to maintain daytime sleep in the face of nighttime sleep loss. In constant darkness, all genotypes show an immediate decrease in sleep in response to temperature shift during the subjective day, implying that the absence of light input uncovers a clock-independent pro-arousal effect of increased temperature. Interestingly, the effects of temperature on nighttime sleep are blunted in constant darkness and in cryOUT mutants in light:dark, suggesting that they are dependent on the presence of light the previous day. In contrast, flies of all genotypes kept in constant light sleep more at all times of day in response to high temperature, indicating that the presence of light can invert the normal nighttime response to increased temperature. The effect of temperature on sleep thus reflects coordinated regulation by light, the homeostat, and components of the clock, allowing animals to reorganize sleep patterns in response to high temperature with rough preservation of the total amount of sleep.

Saint-Charles, A., Michard-Vanhée, C., Alejevski, F., Chélot, E., Boivin, A. and Rouyer, F. (2016). Four of the six Drosophila rhodopsin-expressing photoreceptors can mediate circadian entrainment in low light. J Comp Neurol [Epub ahead of print]. PubMed ID: 26972685
Light is the major stimulus for the synchronization of circadian clocks with day-night cycles. The light-driven entrainment of the clock that controls rest-activity rhythms in Drosophila relies on different photoreceptive molecules. Cryptochrome (CRY) is expressed in most brain clock neurons whereas six different rhodopsins (RH) are present in the light-sensing organs. The compound eye includes outer photoreceptors that express RH1 and inner photoreceptors that each express one of the four RH3-6. In low light, the synchronization of behavioral rhythms relies on either CRY or the canonical rhodopsin phototransduction pathway, which requires the phospholipase C-β encoded by norpA (no receptor potential A). This study used norpAP24 cry02 double mutants that are circadianly blind in low light and restored NORPA function in each of the six types of photoreceptors, defined as expressing a particular rhodopsin. The NORPA pathway was less efficient than CRY for synchronizing rest-activity rhythms with delayed light-dark cycles but is important for proper phasing, whereas the two light-sensing pathways can mediate efficient adjustments to phase advances. Four of the six rhodopsin-expressing photoreceptors can mediate circadian entrainment and all are more efficient for advancing than for delaying the behavioral clock. These results thus reveal different contributions of rhodopsin-expressing photoreceptors and suggest the existence of several circuits for rhodopsin-dependent circadian entrainment.

Bartussek, J. and Lehmann, F. O. (2016). Proprioceptive feedback determines visuomotor gain in Drosophila. R Soc Open Sci 3: 150562. PubMed ID: 26909184
Multisensory integration is a prerequisite for effective locomotor control. In aerial performance of flies, continuous visual signalling from the compound eyes is fused with phasic proprioceptive feedback to ensure precise neural activation of wing steering muscles (WSM) within narrow temporal phase bands of the stroke cycle. This phase-locked activation relies on mechanoreceptors distributed over wings and gyroscopic halteres. This study investigated visual steering performance of tethered flying fruit flies with reduced haltere and wing feedback signalling. Using a flight simulator, visual object fixation behaviour, optomotor altitude control and saccadic escape reflexes were evaluated. The behavioural assays show an antagonistic effect of wing and haltere signalling on visuomotor gain during flight. Compared with controls, suppression of haltere feedback attenuates while suppression of wing feedback enhances the animal's wing steering range. The results suggest that the generation of motor commands owing to visual perception is dynamically controlled by proprioception. Collectively, the findings contribute to a general understanding how moving animals integrate sensory information with dynamically changing temporal structure.

Tuthill, J. C. and Wilson, R. I. (2016). Parallel transformation of tactile signals in central circuits of Drosophila. Cell 164: 1046-1059. PubMed ID: 26919434
To distinguish between complex somatosensory stimuli, central circuits must combine signals from multiple peripheral mechanoreceptor types, as well as mechanoreceptors at different sites in the body. This study investigated the first stages of somatosensory integration in Drosophila using in vivo recordings from genetically labeled central neurons in combination with mechanical and optogenetic stimulation of specific mechanoreceptor types. Three classes of central neurons were identified that process touch: one compares touch signals on different parts of the same limb, one compares touch signals on right and left limbs, and the third compares touch and proprioceptive signals. Each class encodes distinct features of somatosensory stimuli. The axon of an individual touch receptor neuron can diverge to synapse onto all three classes, meaning that these computations occur in parallel, not hierarchically. Representing a stimulus as a set of parallel comparisons is a fast and efficient way to deliver somatosensory signals to motor circuits.

Saturday, March 19th

Khodaei, Z. S., Barmchi, M. P., Gilbert, M. M., Samarasekera, G., Fulga, T. A., Van Vactor, D. and Auld, V. J. (2016). The tricellular junction protein Gliotactin auto-regulates mRNA levels via BMP signaling induction of miR-184. J Cell Sci [Epub ahead of print]. PubMed ID: 26906422
Epithelial bicellular and tricellular junctions are essential for establishing and maintaining permeability barriers. Tricellular junctions are formed by the convergence of three bicellular junctions at the corners of neighbouring epithelia. Gliotactin, a member of the Neuroligin family, is located to the Drosophila tricellular junction and is critical for the formation of tricellular and septate junctions and permeability barrier function. Gliotactin protein levels are tightly controlled by tyrosine phosphorylation and endocytosis. Blocking endocytosis or overexpression of Gliotactin triggers spread away from the tricellular junction, resulting in apoptosis, delamination and migration of epithelial cells. This study shows that Gliotactin levels are also regulated at the mRNA level by microRNA-mediated degradation targeted to a short region in the 3'UTR that includes a conserved miR-184 target site. miR-184 also targets a suite of septate junction proteins including Neurexin-IV, coracle and Mcr. miR-184 expression is triggered when Gliotactin is overexpressed leading to activation of the BMP signaling pathway. Gliotactin specifically interferes with Dad, an inhibitory SMAD, leading to activation of the Tkv type-I receptor, and Mad to elevate the biogenesis and expression of miR-184.
Bosveld, F., Markova, O., Guirao, B., Martin, C., Wang, Z., Pierre, A., Balakireva, M., Gaugue, I., Ainslie, A., Christophorou, N., Lubensky, D. K., Minc, N. and Bellaiche, Y. (2016). Epithelial tricellular junctions act as interphase cell shape sensors to orient mitosis. Nature 530: 495-498. PubMed ID: 26886796
The orientation of cell division along the long axis of the interphase cell--the century-old Hertwig's rule--has profound roles in tissue proliferation, morphogenesis, architecture and mechanics. In epithelial tissues, the shape of the interphase cell is influenced by cell adhesion, mechanical stress, neighbour topology, and planar polarity pathways. At mitosis, epithelial cells usually adopt a rounded shape to ensure faithful chromosome segregation and to promote morphogenesis. The mechanisms underlying interphase cell shape sensing in tissues are therefore unknown. This study shows that in Drosophila epithelia, tricellular junctions (TCJs) localize force generators, pulling on astral microtubules and orienting cell division via the Dynein-associated protein Mud independently of the classical Pins/Galphai pathway. Moreover, as cells round up during mitosis, TCJs serve as spatial landmarks, encoding information about interphase cell shape anisotropy to orient division in the rounded mitotic cell. Finally, experimental and simulation data show that shape and mechanical strain sensing by the TCJs emerge from a general geometric property of TCJ distributions in epithelial tissues. Thus, in addition to their function as epithelial barrier structures, TCJs serve as polarity cues promoting geometry and mechanical sensing in epithelial tissues.

Valencia-Exposito, A., Grosheva, I., Miguez, D. G., Gonzalez-Reyes, A. and Martin-Bermudo, M. D. (2016). Myosin light-chain phosphatase regulates basal actomyosin oscillations during morphogenesis. Nat Commun 7: 10746. PubMed ID: 26888436
Contractile actomyosin networks generate forces that drive tissue morphogenesis. Actomyosin contractility is controlled primarily by reversible phosphorylation of the myosin-II regulatory light chain through the action of myosin kinases and phosphatases. While the role of myosin light-chain kinase in regulating contractility during morphogenesis has been largely characterized, there is surprisingly little information on myosin light-chain phosphatase (MLCP) function in this context. This study used live imaging of Drosophila follicle cells combined with mathematical modelling to demonstrate that the MLCP subunit Flapwing (Flw) is a key regulator of basal myosin oscillations and cell contractions underlying egg chamber elongation. Flw expression decreases specifically on the basal side of follicle cells at the onset of contraction and flw controls the initiation and periodicity of basal actomyosin oscillations. Contrary to previous reports, basal F-actin pulsates similarly to myosin. Finally, a quantitative model is proposed in which periodic basal actomyosin oscillations arise in a cell-autonomous fashion from intrinsic properties of motor assemblies.

Arora, G. K., Tran, S. L., Rizzo, N., Jain, A. and Welte, M. A. (2016). Temporal control of bidirectional lipid-droplet motion depends on the ratio of kinesin-1 and its cofactor Halo. J Cell Sci [Epub ahead of print]. PubMed ID: 26906417
During bidirectional transport, individual cargoes move continuously back and forth along microtubule tracks, yet the cargo population overall displays directed net transport. How such transport is controlled temporally is not well understood. This issue was analyzed for bidirectionally moving lipid droplets in Drosophila embryos, a system in which net transport direction is developmentally controlled. By quantitating how the droplet distribution changes as embryos develop, temporal transitions were characterized in net droplet transport, and the critical contribution was identified of the previously identified, but poorly characterized transacting regulator Halo. In particular, Halo was found to be transiently expressed; rising and falling Halo levels control the switches in global distribution. Rising Halo levels have to pass a threshold before net plus-end transport is initiated. This threshold level depends on the amount of the motor kinesin-1: the more kinesin-1 is present, the more Halo is needed before net plus-end transport commences. Because Halo and kinesin-1 are present in common protein complexes, it is proposed that Halo acts as a rate-limiting co-factor of kinesin-1.

Friday, March 18th

Pushpavalli, S. N., Sarkar, A., Ramaiah, M. J., Koteswara Rao, G., Bag, I., Bhadra, U. and Pal-Bhadra, M. (2015). Drosophila MOF regulates DIAP1 and induces apoptosis in a JNK dependent pathway. Apoptosis [Epub ahead of print]. PubMed ID: 26711898
Histone modulations have been implicated in various cellular and developmental processes where in Drosophila Mof is involved in acetylation of H4K16. Reduction in the size of larval imaginal discs is observed in the null mutants of mof with increased apoptosis. Deficiency involving Hid, Reaper and Grim [H99] alleviated mof RNAi induced apoptosis in the eye discs. mof RNAi induced apoptosis leads to activation of caspases which is suppressed by over expression of caspase inhibitors like P35 and Diap1 clearly depicting the role of caspases in programmed cell death. Also apoptosis induced by knockdown of mof is rescued by JNK mutants of bsk and tak1 indicating the role of JNK in mof RNAi induced apoptosis. The adult eye ablation phenotype produced by ectopic expression of Hid, Rpr and Grim, was restored by over expression of Mof. Accumulation of Mof at the Diap1 promoter 800 bp upstream of the transcription start site in wild type larvae is significantly higher (up to twofolds) compared to mof mutants. This enrichment coincides with modification of histone H4K16Ac indicating an induction of direct transcriptional up regulation of Diap1 by Mof. Based on these results it is proposed that apoptosis triggered by mof RNAi proceeds through a caspase-dependent and JNK mediated pathway.

Bouche, V., Perez Espinosa, A., Leone, L., Sardiello, M., Ballabio, A. and Botas, J. (2016). Drosophila Mitf regulates the V-ATPase and the lysosomal-autophagic pathway. Autophagy 12(3):484-98. PubMed ID: 26761346
An evolutionary conserved gene network regulates the expression of genes involved in lysosome biogenesis, autophagy and lipid metabolism. This study reports that the lysosomal-autophagy pathway is controlled by Mitf gene in Drosophila. Mitf regulates the expression of genes encoding V-ATPase subunits as well as many additional genes involved in the lysosomal-autophagy pathway. Reduction of Mitf function leads to abnormal lysosomes and impairs autophagosome fusion and lipid breakdown during the response to starvation. In contrast, elevated Mitf levels increase the number of lysosomes, autophagosomes and autolysosomes, and decrease the size of lipid droplets. Inhibition of Drosophila MTORC1 induces Mitf translocation to the nucleus, underscoring conserved regulatory mechanisms between Drosophila and mammalian systems. Furthermore, Mitf-mediated clearance of cytosolic and nuclear expanded ATXN1 (ataxin 1) was demonstrated in a cellular model of spinocerebellar ataxia type 1 (SCA1). This remarkable observation illustrates the potential of the lysosomal-autophagy system to prevent toxic protein aggregation in both the cytoplasmic and nuclear compartments. It is anticipated that the genetics of the Drosophila model and the absence of redundant MIT transcription factors will be exploited to investigate the regulation and function of the lysosomal-autophagy gene network.

Mauvezin, C., Neisch, A. L., Ayala, C. I., Kim, J., Beltrame, A., Braden, C. R., Gardner, M. K., Hays, T. S. and Neufeld, T. P. (2016). Coordination of autophagosome-lysosome fusion and transport by a Klp98A-Rab14 complex.J Cell Sci [Epub ahead of print]. PubMed ID: 26763909
Degradation of cellular material by autophagy is essential for cell survival and homeostasis, and requires intracellular transport of autophagosomes to encounter acidic lysosomes through unknown mechanisms. This study identified the PX domain-containing kinesin Klp98A as a novel regulator of autophagosome formation, transport and maturation in Drosophila. Depletion of Klp98A caused abnormal clustering of autophagosomes and lysosomes at the cell center and reduced the formation of starvation-induced autophagic vesicles. Reciprocally, overexpression of Klp98A redistributed autophagic vesicles toward the cell periphery. These effects were accompanied by reduced autophagosome-lysosome fusion and autophagic degradation. In contrast, depletion of the conventional kinesin heavy chain caused a similar mislocalization of autophagosomes without perturbing their fusion with lysosomes, indicating that vesicle fusion and localization are separable, independent events. Klp98A-mediated fusion required the endolysosomal GTPase Rab14, which interacted and colocalized with Klp98A and required Klp98A for normal localization. Thus, Klp98A coordinates the movement and fusion of autophagic vesicles by regulating their positioning and interaction with the endolysosomal compartment.

Timmons, A. K., Mondragon, A. A., Schenkel, C. E., Yalonetskaya, A., Taylor, J. D., Moynihan, K. E., Etchegaray, J. I., Meehan, T. L. and McCall, K. (2016). Phagocytosis genes nonautonomously promote developmental cell death in the Drosophila ovary. Proc Natl Acad Sci U S A. PubMed ID: 26884181
Programmed cell death (PCD) is usually considered a cell-autonomous suicide program, synonymous with apoptosis. Recent research has revealed that PCD is complex, with at least a dozen cell death modalities. This study demonstrates that the large-scale nonapoptotic developmental PCD in the Drosophila ovary occurs by an alternative cell death program where the surrounding follicle cells nonautonomously promote death of the germ line. The phagocytic machinery of the follicle cells, including Draper, cell death abnormality (Ced)-12, and c-Jun N-terminal kinase (JNK), is essential for the death and removal of germ-line-derived nurse cells during late oogenesis. Cell death events including acidification, nuclear envelope permeabilization, and DNA fragmentation of the nurse cells are impaired when phagocytosis is inhibited. Moreover, elimination of a small subset of follicle cells prevents nurse cell death and cytoplasmic dumping. Developmental PCD in the Drosophila ovary is an intriguing example of nonapoptotic, nonautonomous PCD, providing insight on the diversity of cell death mechanisms.

Thursday, March 17th

Zhang, S., Guo, X., Chen, C., Chen, Y., Li, J., Sun, Y., Wu, C., Yang, Y., Jiang, C., Li, W. and Xue, L. (2016). dFoxO promotes Wingless signaling in Drosophila. Sci Rep 6: 22348. PubMed ID: 26936649
The Wnt/β-catenin signaling is an evolutionarily conserved pathway that regulates a wide range of physiological functions, including embryogenesis, organ maintenance, cell proliferation and cell fate decision. Dysregulation of Wnt/β-catenin signaling has been implicated in various cancers, but its role in cell death has not yet been fully elucidated. This study shows that activation of Wg signaling induces cell death in Drosophila eyes and wings, which depends on dFoxO, a transcription factor known to be involved in cell death. In addition, dFoxO is required for ectopic and endogenous Wg signaling to regulate wing patterning. Moreover, dFoxO is necessary for activated Wg signaling-induced target genes expression. Furthermore, Arm is reciprocally required for dFoxO-induced cell death. Finally, dFoxO physically interacts with Arm both in vitro and in vivo. Thus, this study characterizes a previously unknown role of dFoxO in promoting Wg signaling, and shows that a dFoxO-Arm complex is likely involved in their mutual functions, e.g. cell death.

Payankaulam, S., Yeung, K., McNeill, H., Henry, R.W. and Arnosti, D.N. (2016). Regulation of cell polarity determinants by the Retinoblastoma tumor suppressor protein. Sci Rep 6: 22879. PubMed ID: 26971715
In addition to their canonical roles in the cell cycle, RB family proteins regulate numerous developmental pathways, although the mechanisms remain obscure. This study found that Drosophila Rbf1 associates with genes encoding components of the highly conserved apical-basal and planar cell polarity pathways, suggesting a possible regulatory role. It was shown that depletion of Rbf1 in Drosophila tissues is indeed associated with polarity defects in the wing and eye. Key polarity genes aPKC, par6, vang, pk, and fmi are upregulated, and an aPKC mutation suppresses the Rbf1-induced phenotypes. RB control of cell polarity may be an evolutionarily conserved function, with important implications in cancer metastasis.

Barron, D.A. and Moberg, K. (2016). Inverse regulation of two classic Hippo pathway target genes in Drosophila by the dimerization hub protein Ctp. Sci Rep 6: 22726. PubMed ID: 26972460
The LC8 family of small ~8 kD proteins are highly conserved and interact with multiple protein partners in eukaryotic cells. LC8-binding modulates target protein activity, often through induced dimerization via LC8:LC8 homodimers. Although many LC8-interactors have roles in signaling cascades, LC8's role in developing epithelia is poorly understood. Using the Drosophila wing as a developmental model, this study found that the LC8 family member Cut up (Ctp) is primarily required to promote epithelial growth, which correlates with effects on the pro-growth factor dMyc and two genes, diap1 and bantam, that are classic targets of the Hippo pathway coactivator Yorkie. Genetic tests confirm that Ctp supports Yorkie-driven tissue overgrowth and indicate that Ctp acts through Yorkie to control bantam (ban) and diap1 transcription. Quite unexpectedly however, Ctp loss has inverse effects on ban and diap1: it elevates ban expression but reduces diap1 expression. In both cases these transcriptional changes map to small segments of these promoters that recruit Yorkie. Although LC8 complexes with Yap1, a Yorkie homolog, in human cells, an orthologous interaction was not detected in Drosophila cells. Collectively these findings reveal that that Drosophila Ctp is a required regulator of Yorkie-target genes in vivo and suggest that Ctp may interact with a Hippo pathway protein(s) to exert inverse transcriptional effects on Yorkie-target genes.

Chan, P., Han, X., Zheng, B., DeRan, M., Yu, J., Jarugumilli, G. K., Deng, H., Pan, D., Luo, X. and Wu, X. (2016). Autopalmitoylation of TEAD proteins regulates transcriptional output of the Hippo pathway. Nat Chem Biol [Epub ahead of print]. PubMed ID: 26900866
TEA domain (TEAD) transcription factors bind to the coactivators YAP and TAZ (homologs of Drosophila Yorkie) and regulate the transcriptional output of the Hippo pathway, playing critical roles in organ size control and tumorigenesis. Protein S-palmitoylation attaches a fatty acid, palmitate, to cysteine residues and regulates protein trafficking, membrane localization and signaling activities. Using activity-based chemical probes, this study discovered that human TEADs possess intrinsic palmitoylating enzyme-like activities and undergo autopalmitoylation at evolutionarily conserved cysteine residues under physiological conditions. The crystal structures of lipid-bound TEADs were determined, and the lipid chain of palmitate was found to insert into a conserved deep hydrophobic pocket. Strikingly, palmitoylation did not alter TEAD's localization, but it was required for TEAD's binding to YAP and TAZ and was dispensable for its binding to the Vgll4 tumor suppressor. Moreover, palmitoylation-deficient TEAD mutants impaired TAZ-mediated muscle differentiation in vitro and tissue overgrowth mediated by the Drosophila YAP homolog Yorkie in vivo. This study study directly links autopalmitoylation to the transcriptional regulation of the Hippo pathway.

Wednesday, March 16th

Sandler, J. E. and Stathopoulos, A. (2016). Quantitative single-embryo profile of Drosophila genome activation and the dorsal-ventral patterning network. Genetics [Epub ahead of print]. PubMed ID: 26896327
During embryonic development of Drosophila melanogaster, the Maternal to Zygotic Transition (MZT) marks a significant and rapid turning point when zygotic transcription begins and control of development is transferred from maternally deposited transcripts. Characterizing the sequential activation of the genome during the MZT requires precise timing and a sensitive assay to measure changes in expression. This study used the NanoString nCounter instrument, which directly counts mRNA transcripts without reverse transcription or amplification, to study over 70 genes expressed along the dorsal-ventral (DV) axis of early Drosophila embryos, dividing the MZT into 10 time points. Transcripts were quantified for every gene studied at all time points, providing the first data set of absolute numbers of transcripts during Drosophila development. Gene expression was found to change quickly during the MZT, with early Nuclear Cycle (NC) 14 the most dynamic time for the embryo. twist is one of the most abundant genes in the entire embryo and mutants were used to quantitatively demonstrate how it cooperates with Dorsal to activate transcription and is responsible for some of the rapid changes in transcription observed during early NC14. Elements within the gene regulatory network were uncovered that maintain precise transcript levels for sets of genes that are spatiotemporally co-transcribed within the presumptive mesoderm or dorsal ectoderm. Using this new data, it was shown that a fine-scale, quantitative analysis of temporal gene expression can provide new insights into developmental biology by uncovering trends in gene networks including coregulation of target genes and specific temporal input by transcription factors.

Mossman, J.A., Biancani, L.M. and Rand, D.M. (2016). Mitonuclear epistasis for development time and its modification by diet in Drosophila. Genetics [Epub ahead of print]. PubMed ID: 26966258
Mitochondrial (mtDNA) and nuclear genes have to operate in a coordinated manner to maintain organismal function, and the regulation of this homeostasis presents a substantial source of potential epistatic (G x G) interactions. How these interactions shape the fitness landscape is poorly understood. This study developed a novel mitonuclear epistasis model using selected strains of the Drosophila Genetic Reference Panel (DGRP) and mitochondrial genomes from within Drosophila melanogaster and D. simulans to test the hypothesis that mtDNA x nDNA interactions influence fitness. In total 72 genotypes (12 nuclear backgrounds x 6 mtDNA haplotypes, with 3 from each species) were built to dissect the relationship between genotype and phenotype. Each genotype was assayed on four food environments. Considerable variation was found in several phenotypes including development time and egg-to-adult viability, and this variation was partitioned into genetic (G), environmental (E) and higher order (G x G, G x E, and G x G x E) components. Food type has a significant impact on development time and also modifies mitonuclear epistases, evidencing a broad spectrum of G x G x E across these genotypes. Nuclear background effects were substantial, followed by mtDNA effects and their G x G interaction. The species of mtDNA haplotype has negligible effects on phenotypic variation and there was no evidence that mtDNA variation has different effects on male and female fitness traits. These results demonstrate that mitonuclear epistases are context-dependent; suggesting the selective pressure acting on mitonuclear genotypes may vary with food environment in a genotype-specific manner.

Ali-Murthy, Z. and Kornberg, T. B. (2016). Bicoid gradient formation and function in the Drosophila pre-syncytial blastoderm. Elife 5. PubMed ID: 26883601
Bicoid (Bcd) protein distributes in a concentration gradient that organizes the anterior/posterior axis of the Drosophila embryo. It has been understood that bcd RNA is sequestered at the anterior pole during oogenesis, is not translated until fertilization, and produces a protein gradient that functions in the syncytial blastoderm after 9-10 nuclear divisions. However, technical issues limited the sensitivity of analysis of pre-syncytial blastoderm embryos and precluded studies of oocytes after stage 13. This study developed methods to analyze stage 14 oocytes and pre-syncytial blastoderm embryos, and found that stage 14 oocytes make Bcd protein, that bcd RNA and Bcd protein distribute in matching concentration gradients in the interior of nuclear cycle 2-6 embryos, and that Bcd regulation of target gene expression is apparent at nuclear cycle 7, two cycles prior to syncytial blastoderm. The implications are discussed for the generation and function of the Bcd gradient.

Ozturk-Colak, A., Moussian, B., Araujo, S. J. and Casanova, J. (2016). A feedback mechanism converts individual cell features into a supracellular ECM structure in trachea. Elife 5. PubMed ID: 26836303
The extracellular matrix (ECM), a structure contributed to and commonly shared by many cells in an organism, plays an active role during morphogenesis. This study used the Drosophila tracheal system to study the complex relationship between the ECM and epithelial cells during development. An active feedback mechanism was demonstrated between the apical ECM (aECM) and the apical F-actin in tracheal cells. Furthermore, it was revealed that cell-cell junctions are key players in this aECM patterning and organisation and that individual cells contribute autonomously to their aECM. Strikingly, changes in the aECM influence the levels of phosphorylated Src42A (pSrc) at cell junctions. Therefore, it is proposed that Src42A phosphorylation levels provide a link for the extracellular matrix environment to ensure proper cytoskeletal organisation.

Rahimi, N., Averbukh, I., Haskel-Ittah, M., Degani, N., Schejter, E. D., Barkai, N. and Shilo, B. Z. (2016). A WntD-dependent integral feedback loop attenuates variability in Drosophila toll signaling. Dev Cell 36: 401-414. PubMed ID: 26906736
Patterning by morphogen gradients relies on the capacity to generate reproducible distribution profiles. Morphogen spread depends on kinetic parameters, including diffusion and degradation rates, which vary between embryos, raising the question of how variability is controlled. This was examined in the context of Toll-dependent dorsoventral (DV) patterning of the Drosophila embryo. Low embryo-to-embryo variability in DV patterning was found to relies on wntD, a Toll-target gene expressed initially at the posterior pole. WntD protein is secreted and disperses in the extracellular milieu, associates with its receptor Frizzled4, and inhibits the Toll pathway by blocking the Toll extracellular domain. Mathematical modeling predicts that WntD accumulates until the Toll gradient narrows to its desired spread, and this feedback was supported experimentally. This circuit exemplifies a broadly applicable induction-contraction mechanism, which reduces patterning variability through a restricted morphogen-dependent expression of a secreted diffusible inhibitor.

Lei, L. and Spradling, A. C. (2016). Mouse oocytes differentiate through organelle enrichment from sister cyst germ cells. Science. PubMed ID: 26917595
Oocytes differentiate in diverse species by receiving organelles and cytoplasm from sister germ cells while joined in germline cysts or syncytia. Mouse primordial germ cells form germline cysts but the role of cysts in oogenesis is unknown. This study found that mouse germ cells receive organelles from neighboring cyst cells and build a Balbiani body to become oocytes, whereas nurse-like germ cells die. Organelle movement, Balbiani body formation, and oocyte fate determination are selectively blocked by low levels of microtubule-dependent transport inhibitors. Membrane breakdown within the cyst and an apoptosis-like process are associated with organelle transfer into the oocyte, events reminiscent of nurse cell dumping in Drosophila. It is proposed that cytoplasmic and organelle transport plays an evolutionarily conserved and functionally important role in mammalian oocyte differentiation.

Tuesday, March 15th

Adlesic, M., Frei, C. and Frew, I.J. (2016). Cdk4 functions in multiple cell types to control Drosophila intestinal stem cell proliferation and differentiation. Biol Open [Epub ahead of print]. PubMed ID: 26879465
The proliferation of intestinal stem cells (ISCs) and differentiation of enteroblasts to form mature enteroendocrine cells and enterocytes in the Drosophila intestinal epithelium must be tightly regulated to maintain homeostasis. This study shows that genetic modulation of CyclinD/Cdk4 activity or mTOR-dependent signalling cell-autonomously regulates enterocyte growth, which influences ISC proliferation and enteroblast differentiation. Increased enterocyte growth results in higher numbers of ISCs and defective enterocyte growth reduces ISC abundance and proliferation in the midgut. Adult midguts deficient for Cdk4 show severe disruption of intestinal homeostasis characterised by decreased ISC self-renewal, enteroblast differentiation defects and low enteroendocrine cell and enterocyte numbers. The ISC/enteroblast phenotypes result from a combination of cell autonomous and non-autonomous requirements for Cdk4 function. One non-autonomous consequence of Cdk4-dependent deficient enterocyte growth is high expression of Delta in ISCs and Delta retention in enteroblasts. The study postulates that aberrant activation of the Delta-Notch pathway is a possible partial cause of lost ISC stemness. These results support the idea that enterocytes contribute to a putative stem cell niche that maintains intestinal homeostasis in the Drosophila anterior midgut.

Billmann, M., Horn, T., Fischer, B., Sandmann, T., Huber, W. and Boutros, M. (2016). A genetic interaction map of cell cycle regulators. Mol Biol Cell [Epub ahead of print]. PubMed ID: 26912791
Cell based RNAi is a powerful approach to screen for modulators of many cellular processes. However, resulting candidate gene lists from cell-based assays comprise diverse effectors, both direct and indirect, and further dissecting their functions can be challenging. This study screened a genome-wide RNAi library for modulators of mitosis and cytokinesis in Drosophila S2 cells. The screen identified many previously known genes as well as modulators that have previously not been connected to cell cycle control. Approximately 300 candidate modifiers where characterized further by genetic interaction analysis using double RNAi and a multiparametric, imaging-based assay. Analyzing cell-cycle relevant phenotypes increased the sensitivity for associating novel gene function. Genetic interaction maps based on mitotic index and nuclear size grouped candidates into known regulatory complexes of mitosis or cytokinesis, respectively, and predicted previously uncharacterized components of known processes. For example, a role was confirmed for the Drosophila CCR4 mRNA processing complex component l(2)NC136 during the mitotic exit. These results show that the combination of genome-scale RNAi screening and genetic interaction analysis using process-directed phenotypes provides a powerful two-step approach to assign components to specific pathways and complexes.

Clavier, A., Rincheval-Arnold, A., Baillet, A., Mignotte, B. and Guenal, I. (2016). Two different specific JNK activators are required to trigger apoptosis or compensatory proliferation in response to Rbf1 in Drosophila. Cell Cycle 15: 283-294. PubMed ID: 26825229
The Jun Kinase (JNK) signaling pathway responds to diverse stimuli by appropriate and specific cellular responses such as apoptosis, differentiation or proliferation. The mechanisms that mediate this specificity remain largely unknown. The core of this signaling pathway, composed of a JNK protein and a JNK kinase (JNKK), can be activated by various putative JNKK kinases (JNKKK) which are themselves downstream of different adaptor proteins. A proposed hypothesis is that the JNK pathway specific response lies in the combination of a JNKKK and an adaptor protein upstream of the JNKK. Previous studies have showed that the Drosophila homolog of pRb (Rbf1) and a mutant form of Rbf1 (Rbf1D253A) have JNK-dependent pro-apoptotic properties. Rbf1D253A is also able to induce a JNK-dependent abnormal proliferation. This study shows that Rbf1-induced apoptosis triggers proliferation which depends on the JNK pathway activation. Taking advantage of these phenotypes, this study investigated the JNK signaling involved in either Rbf1-induced apoptosis or in proliferation in response to Rbf1-induced apoptosis. Two different JNK pathways involving different adaptor proteins and kinases were shown to be involved in Rbf1-apoptosis (i.e. Rac1-dTak1-dMekk1-JNK pathway) and in proliferation in response to Rbf1-induced apoptosis (i.e., dTRAF1-Slipper-JNK pathway). Using a transient induction of rbf1, this study shows that Rbf1-induced apoptosis activates a compensatory proliferation mechanism which also depends on Slipper and dTRAF1. Thus, these 2 proteins seem to be key players of compensatory proliferation in Drosophila.

Blattner, A. C., Aguilar-Rodriguez, J., Kranzlin, M., Wagner, A. and Lehner, C. F. (2016). Drosophila Nnf1 paralogs are partially redundant for somatic and germ line kinetochore function. Chromosoma [Epub ahead of print]. PubMed ID: 26892014
Kinetochores allow attachment of chromosomes to spindle microtubules. Moreover, they host proteins that permit correction of erroneous attachments and prevent premature anaphase onset before bi-orientation of all chromosomes in metaphase has been achieved. Kinetochores are assembled from subcomplexes. Kinetochore proteins as well as the underlying centromere proteins and the centromeric DNA sequences evolve rapidly despite their fundamental importance for faithful chromosome segregation during mitotic and meiotic divisions. During evolution of Drosophila melanogaster, several centromere proteins were lost and a recent gene duplication has resulted in two Nnf1 paralogs, Nnf1a and Nnf1b, which code for alternative forms of a Mis12 kinetochore complex component. The rapid evolutionary divergence of centromere/kinetochore constituents in animals and plants has been proposed to be driven by an intragenome conflict resulting from centromere drive during female meiosis. Thus, a female meiosis-specific paralog might be expected to evolve rapidly under positive selection. While this characterization of the D. melanogaster Nnf1 paralogs hints at some partial functional specialization of Nnf1b for meiosis, no evidence was detected for positive selection in the analysis of Nnf1 sequence evolution in the Drosophilid lineage. Neither paralog is essential, even though some clear differences were found in subcellular localization and expression during development. Loss of both paralogs results in developmental lethality. It is therefore concluded that the two paralogs are still in early stages of differentiation.

Monday, March 14th

Min, S., Chae, H.S., Jang, Y.H., Choi, S., Lee, S., Jeong, Y.T., Jones, W.D., Moon, S.J., Kim, Y.J. and Chung, J. (2016). Identification of a peptidergic pathway critical to satiety responses in Drosophila. Curr Biol [Epub ahead of print]. PubMed ID: 26948873
Although several neural pathways have been implicated in feeding behaviors in mammals, it remains unclear how the brain coordinates feeding motivations to maintain a constant body weight (BW). This study identified a neuropeptide pathway important for the satiety and BW control in Drosophila. Silencing of myoinhibitory peptide (MIP) neurons significantly increases BW through augmented food intake and fat storage. Likewise, the loss-of-function mutation of mip also increases feeding and BW. Suppressing the MIP pathway induces satiated flies to behave like starved ones, with elevated sensitivity toward food. Conversely, activating MIP neurons greatly decreases food intake and BW and markedly blunts the sensitivity of starved flies toward food. Upon terminating the activation protocol of MIP neurons, the decreased BW reverts rapidly to the normal level through a strong feeding rebound, indicating the switch-like role of MIP pathway in feeding. Surprisingly, the MIP-mediated BW decrease occurs independently of sex peptide receptor (SPR), the only known receptor for MIP, suggesting the presence of a yet-unknown MIP receptor. Together, these results reveal a novel anorexigenic pathway that controls satiety in Drosophila and provide a new avenue to study how the brain actively maintains a constant BW.

King, L. B., Koch, M., Murphy, K., Velazquez, Y., Ja, W. W. and Tomchik, S. M. (2016). Neurofibromin loss of function drives excessive grooming in Drosophila. G3 (Bethesda) [Epub ahead of print] PubMed ID: 26896440
Neurofibromatosis I is a common genetic disorder that results in tumor formation and predisposes individuals to a range of cognitive/behavioral symptoms, including deficits in attention, visuospatial skills, learning, language development, sleep, and autism spectrum disorder-like traits. The nf1-encoded neurofibromin protein (Nf1) exhibits high conservation, from the common fruit fly, Drosophila melanogaster, to humans. Drosophila provide a powerful platform to investigate the signaling cascades upstream and downstream of Nf1, and the fly model exhibits similar behavioral phenotypes to mammalian models. In order to understand how loss of Nf1 affects motor behavior in flies, traditional activity monitoring was combined with video analysis of grooming behavior. In nf1 mutants, spontaneous grooming was increased up to 7x. This increase in activity was distinct from previously-described dopamine-dependent hyperactivity, as dopamine transporter mutants exhibited slightly decreased grooming. Finally, relative grooming frequencies can be compared in standard activity monitors that measure infrared beam breaks, enabling the use of activity monitors as an automated method to screen for grooming phenotypes. Overall, these data suggest that loss of nf1 produces excessive activity that is manifested as increased grooming, providing a platform to dissect the molecular genetics of neurofibromin signaling across neuronal circuits.

Lone, S.R., Potdar, S., Srivastava, M. and Sharma, V.K. (2016). Social experience is sufficient to modulate sleep need of Drosophila without increasing wakefulness. PLoS One 11: e0150596. PubMed ID: 26938057
Fruit flies exposed to highly enriched social environment are found to show increased synaptic connections and a corresponding increase in sleep. This study investigated if social environment comprising a pair of same-sex individuals could enhance sleep in the participating individuals. Same-sex pairs were maintained for a period of 1 to 4 days, and after separation, sleep of the previously socialized and solitary individuals was monitored under similar conditions. Males maintained in pairs for 3 or more days were found to sleep significantly more during daytime and showed a tendency to fall asleep sooner as compared to solitary controls (both measures together are henceforth referred to as "sleep-enhancement"). Sleep-enhancement was found to occur without any significant increase in wakefulness. Furthermore, while sleep-enhancement due to group-wise social interaction requires Pigment Dispersing Factor (PDF) positive neurons; PDF positive and Cryptochrome (CRY) positive circadian clock neurons and the core circadian clock genes are not required for sleep-enhancement to occur when males interact in pairs. Pair-wise social interaction mediated sleep-enhancement requires dopamine and olfactory signaling, while visual and gustatory signaling systems seem to be dispensable. These results suggest that socialization alone (without any change in wakefulness) is sufficient to cause sleep-enhancement in fruit fly D. melanogaster males, and that its neuronal control is context-specific. 

Pasquaretta, C., Battesti, M., Klenschi, E., Bousquet, C.A., Sueur, C. and Mery, F. (2016). How social network structure affects decision-making in Drosophila melanogaster. Proc Biol Sci 283(1826). PubMed ID: 26936247
Animals use a number of different mechanisms to acquire crucial information. During social encounters, animals can pass information from one to another but, ideally, they would only use information that benefits survival and reproduction. Therefore, individuals need to be able to determine the value of the information they receive. One cue can come from the behaviour of other individuals that are already using the information. Using a previous extended dataset, this study analyzed how individual decision-making is influenced by the behaviour of conspecifics in Drosophila melanogaster. How uninformed flies acquire and later use information about oviposition site choice they learn from informed flies was investigated. Data suggest that uninformed flies adjust their future choices based on how coordinated the behaviours of the informed individuals they encounter are. Following social interaction, uninformed flies tend either to collectively follow the choice of the informed flies or to avoid it. Using social network analysis, it was shown that this selective information use seems to be based on the level of homogeneity of the social network. In particular, the variance of individual centrality parameters among informed flies is lower in the case of a 'follow' outcome compared with the case of an 'avoid' outcome. 

Sunday, March 13th

Kapun, M., Schmidt, C., Durmaz, E., Schmidt, P. S. and Flatt, T. (2016). Parallel effects of the inversion In(3R)Payne on body size across the North American and Australian clines in Drosophila melanogaster. J Evol Biol [Epub ahead of print]. PubMed ID: 26881839
Chromosomal inversions are thought to play a major role in climatic adaptation. In D. melanogaster, the cosmopolitan inversion In(3R)Payne exhibits latitudinal clines on multiple continents. Since many fitness traits show similar clines, it is tempting to hypothesize that In(3R)P underlies observed clinal patterns for some of these traits. In support of this idea, previous work in Australian populations has demonstrated that In(3R)P affects body size but not development time or cold resistance. However, similar data from other clines of this inversion are largely lacking; finding parallel effects of In(3R)P across multiple clines would considerably strengthen the case for clinal selection. This study analyzed phenotypic effects of In(3R)P in populations originating from the endpoints of the latitudinal cline along the North American east coast. Development time, egg-to-adult survival, several size-related traits (femur and tibia length, wing area and shape), chill coma recovery, oxidative stress resistance and triglyceride content in were measured in homokaryon lines carrying In(3R)P or the standard arrangement. The central finding is that the effects of In(3R)P along the North American cline match those observed in Australia: standard arrangement lines were larger than inverted lines, but the inversion did not influence development time or cold resistance. Similarly, In(3R)P did not affect egg-to-adult survival, oxidative stress resistance and lipid content. In(3R)P thus seems to specifically affect size traits in populations from both continents. This parallelism strongly suggests an adaptive pattern, whereby the inversion has captured alleles associated with growth regulation and clinal selection acts on size across both continents.

Miller, D. E., Cook, K. R., Yeganeh Kazemi, N., Smith, C. B., Cockrell, A. J., Hawley, R. S. and Bergman, C. M. (2016). Rare recombination events generate sequence diversity among balancer chromosomes in Drosophila melanogaster. Proc Natl Acad Sci U S A [Epub ahead of print]. PubMed ID: 26903656
Multiply inverted balancer chromosomes that suppress exchange with their homologs are an essential part of the Drosophila melanogaster genetic toolkit. Despite their widespread use, the organization of balancer chromosomes has not been characterized at the molecular level, and the degree of sequence variation among copies of balancer chromosomes is unknown. To map inversion breakpoints and study potential diversity in descendants of a structurally identical balancer chromosome, a panel of laboratory stocks containing the most widely used X chromosome balancer, First Multiple 7 (FM7) were sequenced. The locations of FM7 breakpoints were mapped to precise euchromatic coordinates, and the flanking sequence of breakpoints were identified in heterochromatic regions. Analysis of SNP variation revealed megabase-scale blocks of sequence divergence among currently used FM7 stocks. Evidence is presented that this divergence arose through rare double-crossover events that replaced a female-sterile allele of the singed gene (snX2) on FM7c with a sequence from balanced chromosomes. It is proposed that although double-crossover events are rare in individual crosses, many FM7c chromosomes in the Bloomington Drosophila Stock Center have lost snX2 by this mechanism on a historical timescale. Finally, the original allele of the Bar gene (B1) that is carried on FM7 was characterized, and the hypothesis was validated that the origin and subsequent reversion of the B1 duplication are mediated by unequal exchange. These results reject a simple nonrecombining, clonal mode for the laboratory evolution of balancer chromosomes and have implications for how balancer chromosomes should be used in the design and interpretation of genetic experiments in Drosophila.

de Mendoza, A., Jones, J.W. and Friedrich, M. (2016). Methuselah/Methuselah-like G protein-coupled receptors constitute an ancient metazoan gene family. Sci Rep 6: 21801. PubMed ID: 26915348
Inconsistent conclusions have been drawn regarding the phylogenetic age of the Methuselah/Methuselah-like (Mth/Mthl) gene family of G protein-coupled receptors, the founding member of which regulates development and lifespan in Drosophila. This study reports the results from a targeted homolog search of 39 holozoan genomes and phylogenetic analysis of the conserved seven transmembrane domain. It was found that the Mth/Mthl gene family is ancient, has experienced numerous extinction and expansion events during metazoan evolution, and acquired the current definition of the Methuselah ectodomain during its exceptional expansion in arthropods. In addition, Mthl1, Mthl5, Mthl14, and Mthl15 are the oldest Mth/Mthl gene family paralogs in Drosophila. Future studies of these genes have the potential to define ancestral functions of the Mth/Mthl gene family.

Zajitschek, F., Zajitschek, S. R., Canton, C., Georgolopoulos, G., Friberg, U. and Maklakov, A. A. (2016). Evolution under dietary restriction increases male reproductive performance without survival cost. Proc Biol Sci 283. PubMed ID: 26911958
The evolutionary underpinnings of the Dietary restriction (DR) effect on lifespan are still widely debated. The leading theory suggests that this effect is adaptive and results from reallocation of resources from reproduction to somatic maintenance, in order to survive periods of famine in nature. However, such response would cease to be adaptive when DR is chronic and animals are selected to allocate more resources to reproduction. This study evolved Drosophila melanogaster fruit flies on 'DR', 'standard' and 'high' adult diets in replicate populations with overlapping generations. After approximately 25 generations of experimental evolution, male 'DR' flies had higher fitness than males from 'standard' and 'high' populations. Strikingly, this increase in reproductive success did not come at a cost to survival. These results suggest that sustained DR selects for more robust male genotypes that are overall better in converting resources into energy, which they allocate mostly to reproduction.

Saturday, March 12th

Kari, B., Csordás, G., Honti, V., Cinege, G., Williams, M.J., Andó, I. and Kurucz, É. (2016). The raspberry gene is involved in the regulation of the cellular immune response in Drosophila melanogaster. PLoS One 11: e0150910. PubMed ID: 26942456
Drosophila is an extremely useful model organism for understanding how innate immune mechanisms defend against microbes and parasitoids. Large foreign objects trigger a potent cellular immune response in Drosophila larva. In the case of endoparasitoid wasp eggs, this response includes hemocyte proliferation, lamellocyte differentiation and eventual encapsulation of the egg. The encapsulation reaction involves the attachment and spreading of hemocytes around the egg, which requires cytoskeletal rearrangements, changes in adhesion properties and cell shape, as well as melanization of the capsule. Guanine nucleotide metabolism has an essential role in the regulation of pathways necessary for this encapsulation response. This study shows that the Drosophila inosine 5'-monophosphate dehydrogenase (IMPDH), encoded by raspberry (ras), is centrally important for a proper cellular immune response against eggs from the parasitoid wasp Leptopilina boulardi. Notably, hemocyte attachment to the egg and subsequent melanization of the capsule are deficient in hypomorphic ras mutant larvae, which results in a compromised cellular immune response and increased survival of the parasitoid. 

Zhong, X., Rao, X. J., Yi, H. Y., Lin, X. Y., Huang, X. H. and Yu, X. Q. (2016). Co-expression of Dorsal and Rel2 negatively regulates antimicrobial peptide expression in the tobacco hornworm Manduca sexta. Sci Rep 6: 20654. PubMed ID: 26847920
Nuclear factor κB (NF-κB) plays an essential role in regulation of innate immunity. In mammals, NF-κB factors can form homodimers and heterodimers to activate gene expression. In insects, three NF-κB factors, Dorsal, Dif and Relish, have been identified to activate antimicrobial peptide (AMP) gene expression. However, it is not clear whether Dorsal (or Dif) and Relish can form heterodimers. This study reports the identification and functional analysis of a Dorsal homologue (MsDorsal) and two Relish short isoforms (MsRel2A and MsRel2B) from the tobacco hornworm, Manduca sexta. Both MsRel2A and MsRel2B contain only a Rel homology domain (RHD) and lack the ankyrin-repeat inhibitory domain. Overexpression of the RHD domains of MsDorsal and MsRel2 in Drosophila S2 and Spodoptera frugiperda Sf9 cells can activate AMP gene promoters from M. sexta and D. melanogaster. This study confirmed the interaction between MsDorsal-RHD and MsRel2-RHD, and suggest that Dorsal and Rel2 may form heterodimers. More importantly, co-expression of MsDorsal-RHD with MsRel2-RHD suppressed activation of several M. sexta AMP gene promoters. These results suggest that the short MsRel2 isoforms may form heterodimers with MsDorsal as a novel mechanism to prevent over-activation of antimicrobial peptides.

Cao, C., Magwire, M. M., Bayer, F. and Jiggins, F. M. (2016). A polymorphism in the processing body component Ge-1 controls resistance to a naturally occurring ehabdovirus in Drosophila. PLoS Pathog 12: e1005387. PubMed ID: 26799957
Hosts encounter an ever-changing array of pathogens, so there is continual selection for novel ways to resist infection. A powerful way to understand how hosts evolve resistance is to identify the genes that cause variation in susceptibility to infection. Using high-resolution genetic mapping this study has identified a naturally occurring polymorphism in a gene called Ge-1 that makes Drosophila highly resistant to its natural pathogen Drosophila melanogaster sigma virus (DMelSV). By modifying the sequence of the gene in transgenic flies, 26 amino acid deletion in the serine-rich linker region of Ge-1 was identified that is causing the resistance. Knocking down the expression of the susceptible allele leads to a decrease in viral titre in infected flies, indicating that Ge-1 is an existing restriction factor whose antiviral effects have been increased by the deletion. Ge-1 plays a central role in RNA degradation and the formation of processing bodies (P bodies). A key effector in antiviral immunity, the RNAi induced silencing complex (RISC), localises to P bodies, but this study found that Ge-1-based resistance is not dependent on the small interfering RNA (siRNA) pathway. However, Decapping protein 1 (DCP1) was found to protect flies against sigma virus. This protein interacts with Ge-1 and commits mRNA for degradation by removing the 5' cap, suggesting that resistance may rely on this RNA degradation pathway. The serine-rich linker domain of Ge-1 has experienced strong selection during the evolution of Drosophila, suggesting that this gene may be under long-term selection by viruses. These findings demonstrate that studying naturally occurring polymorphisms that increase resistance to infections enables identification of novel forms of antiviral defence, and support a pattern of major effect polymorphisms controlling resistance to viruses in Drosophila.

Shokal, U., Yadav, S., Atri, J., Accetta, J., Kenney, E., Banks, K., Katakam, A., Jaenike, J. and Eleftherianos, I. (2016). Effects of co-occurring Wolbachia and Spiroplasma endosymbionts on the Drosophila immune response against insect pathogenic and non-pathogenic bacteria. BMC Microbiol 16: 16. PubMed ID: 26862076
Symbiotic interactions between microbes and animals are common in nature. Symbiotic organisms are particularly common in insects and, in some cases, they may protect their hosts from pathogenic infections. Wolbachia and Spiroplasma endosymbionts naturally inhabit various insects including Drosophila fruit flies. Therefore, this symbiotic association is considered an excellent model to investigate whether endosymbiotic bacteria participate in host immune processes against certain pathogens. This study investigated whether the presence of Wolbachia alone or together with Spiroplasma endosymbionts in D. melanogaster adult flies affects the immune response against the virulent insect pathogen Photorhabdus luminescens and against non-pathogenic Escherichia coli bacteria. Drosophila flies carrying no endosymbionts, those carrying both Wolbachia and Spiroplasma, and those containing Wolbachia only had similar survival rates after infection with P. luminescens or Escherichia coli bacteria. However, flies carrying both endosymbionts or Wolbachia only contained higher numbers of E. coli cells at early time-points post infection than flies without endosymbiotic bacteria. Interestingly, flies containing Wolbachia only had lower titers of this endosymbiont upon infection with the pathogen P. luminescens than uninfected flies of the same strain. It was further found that the presence of Wolbachia and Spiroplasma in D. melanogaster up-regulated certain immune-related genes upon infection with P. luminescens or E. coli bacteria, but it failed to alter the phagocytic ability of the flies toward E. coli inactive bioparticles. These results suggest that the presence of Wolbachia and Spiroplasma in D. melanogaster can modulate immune signaling against infection by certain insect pathogenic and non-pathogenic bacteria. Results from such studies are important for understanding the molecular basis of the interactions between endosymbiotic bacteria of insects and exogenous microbes.

Friday, March 11th

Lo, P. K., Huang, Y. C., Poulton, J. S., Leake, N., Palmer, W. H., Vera, D., Xie, G., Klusza, S. and Deng, W. M. (2016). RNA helicase Belle/DDX3 regulates transgene expression in Drosophila. Dev Biol [Epub ahead of print]. PubMed ID: 26900887
Belle (Bel), the Drosophila homolog of the yeast DEAD-box RNA helicase DED1 and human DDX3, has been shown to be required for oogenesis and female fertility. This study reports a novel role of Bel in regulating the expression of transgenes. Abrogation of Bel by mutations or RNAi induces silencing of a variety of P-element-derived transgenes. This silencing effect depends on downregulation of their RNA levels. These genetic studies have revealed that the RNA helicase Spindle-E (Spn-E), a nuage RNA helicase that plays a crucial role in regulating RNA processing and PIWI-interacting RNA (piRNA) biogenesis in germline cells, is required for loss-of-bel-induced transgene silencing. Conversely, Bel abrogation alleviates the nuage-protein mislocalization phenotype in spn-E mutants, suggesting a competitive relationship between these two RNA helicases. Additionally, disruption of the chromatin remodeling factor Mod(mdg4) or the microRNA biogenesis enzyme Dicer-1 (Dcr-1) also alleviates the transgene-silencing phenotypes in bel mutants, suggesting the involvement of chromatin remodeling and microRNA biogenesis in loss-of-bel-induced transgene silencing. Finally, genetic inhibition of Bel function was shown to lead to de novo generation of piRNAs from the transgene region inserted in the genome, suggesting a potential piRNA-dependent mechanism that may mediate transgene silencing as Bel function is inhibited.

Romano, M., Feiguin, F. and Buratti, E. (2016). TBPH/TDP-43 modulates translation of Drosophila futsch mRNA through an UG-rich sequence within its 5'UTR. Brain Res [Epub ahead of print]. PubMed ID: 26902497
Nuclear factor TDP-43 is an evolutionarily conserved multifunctional RNA-binding protein associated with Frontotemporal lobar degeneration (FTLD) and amyotrophic lateral sclerosis (ALS). In recent years, Drosophila models of ALS based on TDP-43 knockdown/overexpression have allowed to find several connections with disease. Among these, previous studies have described that silencing the expression of its fly ortholog (TBPH) can alter the expression of the neuronal microtubule-associated protein Futsch leading to alterations of neuromuscular junction (NMJ) organization. In particular, TBPH knocked out flies displayed a significant reduction of Futsch protein levels, although minimal variation in the futsch mRNA content was observed. These conclusions were recently validated in an independent study. Together, these observations strongly support the hypothesis that TBPH might regulate the translation of futsch mRNA. However, the mechanism of TBPH interference in futsch mRNA translation is still unknown. This work used EMSA experiments coupled with RNA-protein co-immunprecipitations and luciferase assays to show that TBPH interacts with a stretch of UG within the 5'UTR of futsch mRNA and translation is positively modulated by this binding. Most importantly, this function is also conserved in human TDP-43. This result can therefore represent the first step in elucidating the relationship between TDP-43, protein translation, and eventual disease onset or progression.

Bolin, K., Rachmaninoff, N., Moncada, K., Pula, K., Kennell, J. and Buttitta, L. (2016). miR-8 modulates cytoskeletal regulators to influence cell survival and epithelial organization in Drosophila wings. Dev Biol [Epub ahead of print]. PubMed ID: 26902111
The miR-200 microRNA family plays important tumor suppressive roles. The sole Drosophila miR-200 ortholog, miR-8 plays conserved roles in Wingless, Notch and Insulin signaling - pathways linked to tumorigenesis, yet homozygous null animals are viable and often appear morphologically normal. This study observed that wing tissues mosaic for miR-8 levels by genetic loss or gain of function exhibited patterns of cell death consistent with a role for miR-8 in modulating cell survival in vivo. This study shows that miR-8 levels impact several actin cytoskeletal regulators that can affect cell survival and epithelial organization. Loss of miR-8 can confer resistance to apoptosis independent of an epithelial to mesenchymal transition while the persistence of cells expressing high levels of miR-8 in the wing epithelium leads to increased JNK signaling, aberrant expression of extracellular matrix remodeling proteins and disruption of proper wing epithelial organization. Altogether these results suggest that very low as well as very high levels of miR-8 can contribute to hallmarks associated with cancer, suggesting approaches to increase miR-200 microRNAs in cancer treatment should be moderate.
Guven-Ozkan, T., Busto, G. U., Schutte, S. S., Cervantes-Sandoval, I., O'Dowd, D. K. and Davis, R. L. (2016). miR-980 is a memory suppressor microRNA that regulates the autism-susceptibility gene A2bp1. Cell Rep 14: 1698-1709. PubMed ID: 26876166
MicroRNAs have been associated with many different biological functions, but little is known about their roles in conditioned behavior. This study demonstrates that Drosophila miR-980 is a memory suppressor gene functioning in multiple regions of the adult brain. Memory acquisition and stability were both increased by miR-980 inhibition. Whole cell recordings and functional imaging experiments indicated that miR-980 regulates neuronal excitability. This study identified the autism susceptibility gene, A2bp1, as an mRNA target for miR-980. A2bp1 levels varied inversely with miR-980 expression; memory performance was directly related to A2bp1 levels. In addition, A2bp1 knockdown reversed the memory gains produced by miR-980 inhibition, consistent with A2bp1 being a downstream target of miR-980 responsible for the memory phenotypes. These results indicate that miR-980 represses A2bp1 expression to tune the excitable state of neurons, and the overall state of excitability translates to memory impairment or improvement.

Thursday, March 10th

Iwanami, N., Nakamura, Y., Satoh, T., Liu, Z. and Satoh, A. K. (2016). Rab6 is required for multiple apical transport pathways but not the basolateral transport pathway in Drosophila photoreceptors. PLoS Genet 12: e1005828. PubMed ID: 26890939
Polarized membrane trafficking is essential for the construction and maintenance of multiple plasma membrane domains of cells. Highly polarized Drosophila photoreceptors are an excellent model for studying polarized transport. A single cross-section of Drosophila retina contains many photoreceptors with 3 clearly differentiated plasma membrane domains: a rhabdomere, stalk, and basolateral membrane. Genome-wide high-throughput ethyl methanesulfonate screening followed by precise immunohistochemical analysis identified a mutant with a rare phenotype characterized by a loss of 2 apical transport pathways with normal basolateral transport. Rapid gene identification using whole-genome resequencing and single nucleotide polymorphism mapping identified a nonsense mutation of Rab6 responsible for the apical-specific transport deficiency. Detailed analysis of the trafficking of a major rhabdomere protein Rh1 using blue light-induced chromophore supply identified Rab6 as essential for Rh1 to exit the Golgi units. Rab6 is mostly distributed from the trans-Golgi network to a Golgi-associated Rab11-positive compartment that likely recycles endosomes or transport vesicles going to recycling endosomes. Furthermore, the Rab6 effector, Rich, is required for Rab6 recruitment in the trans-Golgi network. Moreover, a Rich null mutation phenocopies the Rab6 null mutant, indicating that Rich functions as a guanine nucleotide exchange factor for Rab6. The results collectively indicate that Rab6 and Rich are essential for the trans-Golgi network-recycling endosome transport of cargoes destined for 2 apical domains. However, basolateral cargos are sorted and exported from the trans-Golgi network in a Rab6-independent manner.

Spring, A. M., Brusich, D. J. and Frank, C. A. (2016). C-terminal Src kinase gates homeostatic synaptic plasticity and regulates Fasciclin II expression at the Drosophila neuromuscular junction. PLoS Genet 12: e1005886. PubMed ID: 26901416
Forms of homeostatic plasticity stabilize neuronal outputs and promote physiologically favorable synapse function. At the Drosophila neuromuscular junction (NMJ) impairment of postsynaptic glutamate receptor activity is offset by a compensatory increase in presynaptic neurotransmitter release. This study aimed to elucidate how this process operates on a molecular level and is preserved throughout development. A tyrosine kinase-driven signaling system was identified that sustains homeostatic control of NMJ function. C-terminal Src Kinase (Csk) was identified as a potential regulator of synaptic homeostasis through an RNAi- and electrophysiology-based genetic screen. Csk loss-of-function mutations impair the sustained expression of homeostatic plasticity at the NMJ, without drastically altering synapse growth or baseline neurotransmission. Muscle-specific overexpression of Src Family Kinase (SFK) substrates that are negatively regulated by Csk also impaired NMJ homeostasis. Surprisingly, it was found that transgenic Csk-YFP can support homeostatic plasticity at the NMJ when expressed either in the muscle or in the nerve. However, only muscle-expressed Csk-YFP was able to localize to NMJ structures. By immunostaining, it was found that Csk mutant NMJs had dysregulated expression of the Neural Cell Adhesion Molecule homolog Fasciclin II (FasII). By immunoblotting, it was found that levels of a specific isoform of FasII were decreased in homeostatically challenged GluRIIA mutant animals-but markedly increased in Csk mutant animals. Additionally, it was found that postsynaptic overexpression of FasII from its endogenous locus was sufficient to impair synaptic homeostasis, and genetically reducing FasII levels in Csk mutants fully restored synaptic homeostasis. Based on these data, it is propose that Csk and its SFK substrates impinge upon homeostatic control of NMJ function by regulating downstream expression or localization of FasII.

Zhang, P., Zhou, L., Pei, C., Lin, X. and Yuan, Z. (2016). Dysfunction of Wntless triggers the retrograde Golgi-to-ER transport of Wingless and induces ER stress. Sci Rep 6: 19418. PubMed ID: 26887613
Secreted Wnts play diverse roles in a non-cell-autonomous fashion. However, the cell-autonomous effect of unsecreted Wnts remains unknown. Endoplasmic reticulum (ER) stress is observed in specialized secretory cells and participates in pathophysiological processes. The correlation between Wnt secretion and ER stress remains poorly understood. This study demonstrate that Drosophila miR-307a initiates ER stress specifically in wingless (wg)-expressing cells through targeting wntless (wls/evi). This phenotype could be mimicked by retromer loss-of-function or porcupine (porc) depletion, and rescued by wg knockdown, arguing that unsecreted Wg triggers ER stress. Consistently, it was found that disrupting the secretion of human Wnt5a also induced ER stress in mammalian cells. Furthermore, it was shown that a C-terminal KKVY-motif of Wg is required for its retrograde Golgi-to-ER transport, thus inducing ER stress. Next, it was investigated if COPI, the regulator of retrograde transport, is responsible for unsecreted Wg to induce ER stress. Surprisingly, it was found that COPI acts as a novel regulator of Wg secretion. Taken together, this study reveals a previously unknown Golgi-to-ER retrograde route of Wg, and elucidates a correlation between Wnt secretion and ER stress during development.

Freyberg, Z., et al. (2016). Mechanisms of amphetamine action illuminated through optical monitoring of dopamine synaptic vesicles in Drosophila brain. Nat Commun 7: 10652. PubMed ID: 26879809
Amphetamines elevate extracellular dopamine, but the underlying mechanisms remain uncertain. This study shows in rodents that acute pharmacological inhibition of the vesicular monoamine transporter (VMAT) blocks amphetamine-induced locomotion and self-administration without impacting cocaine-induced behaviours. To study VMAT's role in mediating amphetamine action in dopamine neurons, novel genetic, pharmacological and optical approaches were used in Drosophila. In an ex vivo whole-brain preparation, fluorescent reporters of vesicular cargo and of vesicular pH reveal that amphetamine redistributes vesicle contents and diminishes the vesicle pH-gradient responsible for dopamine uptake and retention. This amphetamine-induced deacidification requires VMAT function and results from net H(+) antiport by VMAT out of the vesicle lumen coupled to inward amphetamine transport. Amphetamine-induced vesicle deacidification also requires functional dopamine transporter (DAT) at the plasma membrane. Thus, this study found that at pharmacologically relevant concentrations, amphetamines must be actively transported by DAT and VMAT in tandem to produce psychostimulant effects.

Baron, M. N., Klinger, C. M., Rachubinski, R. A. and Simmonds, A. J. (2016). A systematic cell-based analysis of localization of predicted Drosophila peroxisomal proteins. Traffic [Epub ahead of print]. PubMed ID: 26865094
Peroxisomes are membrane-bound organelles found in almost all eukaryotic cells. They perform specialized biochemical functions that vary with organism, tissue or cell type. Mutations in human genes required for the assembly of peroxisomes result in a spectrum of diseases called the peroxisome biogenesis disorders. This study tested systematically the localization and selected functions of epitope-tagged proteins in Drosophila Schneider 2 cells to determine the subcellular localization of 82 potential Drosophila peroxisomal protein homologues. Excluding the Pex proteins, 34 proteins localized primarily to the peroxisome, 8 showed dual localization to the peroxisome and other structures, and 26 localized exclusively to organelles other than the peroxisome. Drosophila is a well-developed laboratory animal often used for discovery of gene pathways, including those linked to human disease. This work establishes a basic understanding of peroxisome protein localization in Drosophila. This will facilitate use of Drosophila as a genetically tractable, multicellular model system for studying key aspects of human peroxisome disease.

Hutchison, J. B., Karunanayake Mudiyanselage, A. P., Weis, R. M. and Dinsmore, A. D. (2016). Osmotically-induced tension and the binding of N-BAR protein to lipid vesicles. Soft Matter [Epub ahead of print]. PubMed ID: 26822233

The binding affinity of a curvature-sensing protein domain (N-BAR) is measured as a function of applied osmotic stress while the membrane curvature is nearly constant. Varying the osmotic stress allows control of membrane tension, which provides a probe of the mechanism of binding. The N-BAR domain of the Drosophila amphiphysin was studied, and its binding on 50 nm-radius vesicles composed of 90 mol% DOPC and 10 mol% PIP was monitored. The bound fraction of N-BAR is enhanced by a factor of approximately 6.5 when the tension increases from zero to 2.6 mN m-1. This tension-induced response can be explained by the hydrophobic insertion mechanism. From the data a hydrophobic domain area was extracted that is consistent with known structure. These results indicate that membrane stress and strain could play a major role in the previously reported curvature-affinity of N-BAR.

Wednesday, March 9th

Li, X., Chatterjee, N., Spirohn, K., Boutros, M. and Bohmann, D. (2016). Cdk12 is a gene-selective RNA polymerase II kinase that regulates a subset of the transcriptome, including Nrf2 target genes. Sci Rep 6: 21455. PubMed ID: 26911346
The Nrf2 transcription factor is well conserved throughout metazoan evolution and serves as a central regulator of adaptive cellular responses to oxidative stress. This study carried out an RNAi screen in Drosophila S2 cells to better understand the regulatory mechanisms governing Nrf2 target gene expression. This paper describes the identification and characterization of the RNA polymerase II (Pol II) kinase Cdk12 as a factor that is required for Nrf2 target gene expression in cell culture and in vivo. Cdk12 is, however, not essential for bulk mRNA transcription and cells lacking CDK12 function are viable and able to proliferate. Consistent with previous findings on the DNA damage and heat shock responses, it emerges that Cdk12 may be specifically required for stress activated gene expression. Transcriptome analysis revealed that antioxidant gene expression is compromised in flies with reduced Cdk12 function, which makes them oxidative stress sensitive. In addition to supporting Reactive Oxygen Species (ROS) induced gene activation, Cdk12 suppresses genes that support metabolic functions in stressed conditions. The study suggests that Cdk12 acts as a gene-selective Pol II kinase that engages a global shift in gene expression to switch cells from a metabolically active state to "stress-defence mode" when challenged by external stress.

Lopes, C. A., Jana, S. C., Cunha-Ferreira, I., Zitouni, S., Bento, I., Duarte, P., Gilberto, S., Freixo, F., Guerrero, A., Francia, M., Lince-Faria, M., Carneiro, J. and Bettencourt-Dias, M. (2015). PLK4 trans-autoactivation controls centriole biogenesis in space. Dev Cell 35: 222-235. PubMed ID: 26481051
Centrioles are essential for cilia and centrosome assembly. In centriole-containing cells, centrioles always form juxtaposed to pre-existing ones, motivating a century-old debate on centriole biogenesis control. This study shows that trans-autoactivation of Polo-like kinase 4 (PLK4), the trigger of centriole biogenesis, is a critical event in the spatial control of that process. Centrioles promote PLK4 activation through its recruitment and local accumulation. Though centriole removal reduces the proportion of active PLK4, this is rescued by concentrating PLK4 to the peroxisome lumen. Moreover, while mild overexpression of PLK4 only triggers centriole amplification at the existing centriole, higher PLK4 levels trigger both centriolar and cytoplasmatic (de novo) biogenesis. Hence, centrioles promote their assembly locally and disfavor de novo synthesis. Similar mechanisms enforcing the local concentration and/or activity of other centriole components are likely to contribute to the spatial control of centriole biogenesis under physiological conditions.

Bailey, M. J. and Prehoda, K. E. (2015). Establishment of par-polarized cortical domains via phosphoregulated membrane motifs. Dev Cell 35: 199-210. PubMed ID: 26481050
The Par polarity complex creates mutually exclusive cortical domains in diverse animal cells. Activity of the atypical protein kinase C (aPKC) is a key output of the Par complex as phosphorylation removes substrates from the Par domain. This study investigate how diverse, apparently unrelated Par substrates couple phosphorylation to cortical displacement. Each protein contains a basic and hydrophobic (BH) motif that interacts directly with phospholipids and also overlaps with aPKC phosphorylation sites. Phosphorylation alters the electrostatic character of the sequence, inhibiting interaction with phospholipids and the cell cortex. Overlapping BH and aPKC phosphorylation site motifs (i.e., putative phosphoregulated BH motifs) were sought in several animal proteomes. Candidate proteins with strong PRBH signals associated with the cell cortex but were displaced into the cytoplasm by aPKC. These findings demonstrate a potentially general mechanism for exclusion of proteins from the Par cortical domain in polarized cells.

Ye, F., Liu, W., Shang, Y. and Zhang, M. (2016). An exquisitely specific PDZ/target recognition revealed by the structure of INAD PDZ3 in complex with TRP channel tail. Structure [Epub ahead of print]. PubMed ID: 26853938
The vast majority of PDZ domains are known to bind to a few C-terminal tail residues of target proteins with modest binding affinities and specificities. Such promiscuous PDZ/target interactions are not compatible with highly specific physiological functions of PDZ domain proteins and their targets. This study reports an unexpected PDZ/target binding occurring between the scaffold protein Inactivation no afterpotential D (INAD) and Transient receptor potential (TRP) channel in Drosophila photoreceptors. The C-terminal 15 residues of TRP are required for the specific interaction with INAD PDZ3. The INAD PDZ3/TRP peptide complex structure reveals that only the extreme C-terminal Leu of TRP binds to the canonical αB/βB groove of INAD PDZ3. The rest of the TRP peptide, by forming a β hairpin structure, binds to a surface away from the αB/βB groove of PDZ3 and contributes to the majority of the binding energy. Thus, the INAD PDZ3/TRP channel interaction is exquisitely specific and represents a new mode of PDZ/target recognitions.

Tuesday, March 8th

Saavedra, P., Brittle, A., Palacios, I.M., Strutt, D., Casal, J. and Lawrence, P.A. (2016). Planar cell polarity: the Dachsous/Fat system contributes differently to the embryonic and larval stages of Drosophila. Biol Open [Epub ahead of print]. PubMed ID: 26935392
The epidermal patterns of all three larval instars (L1-L3) of Drosophila are made by one unchanging set of cells. The seven rows of cuticular denticles of all larval stages are consistently planar polarised, some pointing forwards, others backwards. In L1 all the predenticles originate at the back of the cells but, in L2 and L3, they form at the front or the back of the cell depending on the polarity of the forthcoming denticles. This study finds that, to polarise all rows, the Dachsous/Fat system is differentially utilised; in L1 it is active in the placement of the actin-based predenticles but is not crucial for the final orientation of the cuticular denticles, in L2 and L3 it is needed for placement and polarity. Four-jointed is strongly expressed in the tendon cells and this might explain the orientation of all seven rows. Unexpectedly, it was found that L3 that lack Dachsous differ from larvae lacking Fat and this is due to differently mislocalised Dachs.

Zappia, M.P. and Frolov, M.V. (2016). E2F function in muscle growth is necessary and sufficient for viability in Drosophila. Nat Commun 7: 10509. PubMed ID: 26823289
The E2F transcription factor is a key cell cycle regulator. However, the inactivation of the entire E2F family in Drosophila is permissive throughout most of animal development until pupation when lethality occurs. This study shows that E2F function in the adult skeletal muscle is essential for animal viability since providing E2F function in muscles rescues the lethality of the whole-body E2F-deficient animals. Muscle-specific loss of E2F results in a significant reduction in muscle mass and thinner myofibrils. It was demonstrated that E2F is dispensable for proliferation of muscle progenitor cells, but is required during late myogenesis to directly control the expression of a set of muscle-specific genes. Interestingly, E2f1 provides a major contribution to the regulation of myogenic function, while E2f2 appears to be less important. These findings identify a key function of E2F in skeletal muscle required for animal viability, and illustrate how the cell cycle regulator is repurposed in post-mitotic cells. 

Pesch, Y. Y., Riedel, D., Patil, K. R., Loch, G. and Behr, M. (2016). Chitinases and imaginal disc growth factors organize the extracellular matrix formation at barrier tissues in insects. Sci Rep 6: 18340. PubMed ID: 26838602
The cuticle forms an apical extracellular-matrix (ECM) that covers exposed organs, such as epidermis, trachea and gut, for organizing morphogenesis and protection of insects. This study investigated the poorly described glyco-18-domain hydrolase family in Drosophila and identified the Chitinases (Chts; Cht2, Cht4, Cht5, Cht6, Cht7, Cht8, Cht9, Cht11, Cht10, Cht12) and imaginal-disc-growth-factors (Idgfs; idgf1, idgf2, idgf3, digf4, idgf5 and idgf6) that are essential for larval and adult molting. Cht and idgf depletion results in deformed cuticles, larval and adult molting defects, and insufficient protection against wounding and bacterial infection, which altogether leads to early lethality. Cht2/Cht5/Cht7/Cht9/Cht12 and idgf1/idgf3/idgf4/idgf5/idgf6 are needed for organizing proteins and chitin-matrix at the apical cell surface. The data indicate that normal ECM formation requires Chts, which potentially hydrolyze chitin-polymers. It is furtherer suggest that the non-enzymatic idgfs act as structural proteins to maintain the ECM scaffold against chitinolytic degradation. Conservation of Chts and Idgfs proposes analogous roles in ECM dynamics across the insect taxa, indicating that Chts/Idgfs are new targets for species specific pest control.

Gavish, A., Shwartz, A., Weizman, A., Schejter, E., Shilo, B. Z. and Barkai, N. (2016). Periodic patterning of the Drosophila eye is stabilized by the diffusible activator Scabrous. Nat Commun 7: 10461. PubMed ID: 26876750
Generation of periodic patterns is fundamental to the differentiation of multiple tissues during development. How such patterns form robustly is still unclear. The Drosophila eye comprises approximately 750 units, whose crystalline order is set during differentiation of the eye imaginal disc: an activation wave sweeping across the disc is coupled to lateral inhibition, sequentially selecting pro-neural cells. Using mathematical modelling, this study shows that this template-based lateral inhibition is highly sensitive to spatial variations in biochemical parameters and cell sizes. The basis of this sensitivity is revealed, and it suggested that the sensitivity can be overcome by assuming a short-range diffusible activator. Clonal experiments identify Scabrous, a previously implicated inhibitor, as the predicted activator. These results reveal the mechanism by which periodic patterning in the fly eye is stabilized against spatial variations, highlighting how the need to maintain robustness shapes the design of patterning circuits.

Monday, March 7th

Syed, D.S., Bm, S., Reddy, O.V., Reichert, H. and VijayRaghavan, K. (2016). Glial and neuronal Semaphorin signaling instruct the development of a functional myotopic map for Drosophila walking. Elife [Epub ahead of print]. PubMed ID: 26926907
Motoneurons developmentally acquire appropriate cellular architectures that ensure connections with postsynaptic muscles and presynaptic neurons. In Drosophila, leg motoneurons are organized as a myotopic map, where their dendritic domains represent the muscle field. This study investigates mechanisms underlying development of aspects of this myotopic map, required for walking. A behavioral screen identified roles for Semaphorins (Sema) and Plexins (Plex) in walking behavior. Deciphering this phenotype, it was shown that PlexA/Sema1a mediates motoneuron axon branching in ways that differ in the proximal femur and distal tibia, based on motoneuronal birth order. Importantly, glia plays a role in positioning dendrites of specific motoneurons; PlexB/Sema2a is required for dendritic positioning of late-born motoneurons but not early-born motoneurons. These findings indicate that communication within motoneurons and between glia and motoneurons, mediated by the combined action of different Plexin/Semaphorin signaling systems, are required for the formation of a functional myotopic map.

Leonhardt, A., Ammer, G., Meier, M., Serbe, E., Bahl, A. and Borst, A. (2016). Asymmetry of Drosophila ON and OFF motion detectors enhances real-world velocity estimation. Nat Neurosci [Epub ahead of print]. PubMed ID: 26928063
The reliable estimation of motion across varied surroundings represents a survival-critical task for sighted animals. How neural circuits have adapted to the particular demands of natural environments, however, is not well understood. This study explored this question in the visual system of Drosophila melanogaster. In this system, as in many mammalian retinas, motion is computed in parallel streams for brightness increments (ON) and decrements (OFF). When genetically isolated, ON and OFF pathways prove equally capable of accurately matching walking responses to realistic motion. Detailed characterization of their functional tuning properties through in vivo calcium imaging and electrophysiology reveals stark differences in temporal tuning between ON and OFF channels. By training an in silico motion estimation model on natural scenes, it was discovered that the optimized detector exhibits differences similar to those of the biological system. Thus, functional ON-OFF asymmetries in fly visual circuitry may reflect ON-OFF asymmetries in natural environments.

Shepherd, D., Harris, R., Williams, D. and Truman, J. W. (2016). . Postembryonic Lineages of the Drosophila Ventral Nervous System: Neuroglian expression reveals the adult hemilineage associated fiber tracts in the adult thoracic neuromeres. J Comp Neurol [Epub ahead of print]. PubMed ID: 26878258
During larval life most of the thoracic neuroblasts (NBs) in Drosophila undergo a second phase of neurogenesis to generate adult-specific neurons that remain in an immature, developmentally stalled state until pupation. Using a combination of MARCM and immunostaining with a neurotactin antibody 24 adult specific NB lineages have been identified within each thoracic hemineuromere of the larval ventral nervous system (VNS) but because the neurotactin labeling of lineage tracts disappearing early in metamorphosis it was not possible to extend the identification of the these lineages into the adult. This study shows that immunostaining with an antibody against the cell adhesion molecule Neuroglian reveals the same larval secondary lineage projections through metamorphosis and by identifying each neuroglian positive tract at selected stages the larval hemilineage tracts for all three thoracic neuromeres were traced through metamorphosis into the adult. To validate tract identifications a genetic toolkit was used to preserve hemilineage specific GAL4 expression patterns from larval into the adult stage. The immortalized expression proved a powerful confirmation of the analysis of the neuroglian scaffold. This work has enabled direct link ing of the secondary, larval NB lineages to their adult counterparts. The data provide an anatomical framework that 1) makes it possible to assign most neurons to their parent lineage and 2) allows more precise definitions of the neuronal organization of the adult VNS based in developmental units/rules.

Rohwedder, A., Wenz, N. L., Stehle, B., Huser, A., Yamagata, N., Zlatic, M., Truman, J. W., Tanimoto, H., Saumweber, T., Gerber, B. and Thum, A. S. (2016). Four individually identified paired dopamine neurons signal reward in larval Drosophila. Curr Biol [Epub ahead of print]. PubMed ID: 26877086
Dopaminergic neurons serve multiple functions, including reinforcement processing during associative learning. It is thus warranted to understand which dopaminergic neurons mediate which function. Larval Drosophila were used, in which only approximately 120 of a total of 10,000 neurons are dopaminergic, as judged by the expression of tyrosine hydroxylase (TH), the rate-limiting enzyme of dopamine biosynthesis. Dopaminergic neurons mediating reinforcement in insect olfactory learning target the mushroom bodies, a higher-order "cortical" brain region. Four previously undescribed paired neurons, the primary protocerebral anterior medial (pPAM) neurons, were discovered. These neurons are TH positive and subdivide the medial lobe of the mushroom body into four distinct subunits. These pPAM neurons are acutely necessary for odor-sugar reward learning and require intact TH function in this process. However, they are dispensable for aversive learning and innate behavior toward the odors and sugars employed. Optogenetical activation of pPAM neurons is sufficient as a reward. Thus, the pPAM neurons convey a likely dopaminergic reward signal. In contrast, DL1 cluster neurons convey a corresponding punishment signal, suggesting a cellular division of labor to convey dopaminergic reward and punishment signals. On the level of individually identified neurons, this uncovers an organizational principle shared with adult Drosophila and mammals. The numerical simplicity and connectomic tractability of the larval nervous system now offers a prospect for studying circuit principles of dopamine function at unprecedented resolution.

Sunday, March 6th

Sadasivam, D.A. and Huang, D.H. (2016). Maintenance of tissue pluripotency by epigenetic factors acting at multiple levels. PLoS Genet 12: e1005897. PubMed ID: 26926299
Pluripotent stem cells often adopt a unique developmental program while retaining certain flexibility. The molecular basis of such properties remains unclear. Using differentiation of pluripotent Drosophila imaginal tissues as assays, this study examined the contribution of epigenetic factors in ectopic activation of Hox genes. It was found that over-expression of Trithorax H3K4 methyltransferase can induce ectopic adult appendages by selectively activating the Hox genes Ultrabithorax and Sex comb reduced in wing and leg discs, respectively. This tissue-specific inducibility correlates with the presence of paused RNA polymerase II in the promoter-proximal region of these genes. Although the Antennapedia promoter is paused in eye-antenna discs, it cannot be induced by Trx without a reduction in histone variants or their chaperones, suggesting additional control by the nucleosomal architecture. Lineage tracing and pulse-chase experiments revealed that the active state of Hox genes is maintained substantially longer in mutants deficient for HIRA, a chaperone for the H3.3 variant. In addition, both HIRA and H3.3 appear to act cooperatively with the Polycomb group of epigenetic repressors. These results support the involvement of H3.3-mediated nucleosome turnover in restoring the repressed state. The study proposes a regulatory framework integrating transcriptional pausing, histone modification, nucleosome architecture and turnover for cell lineage maintenance.

Saha, N., Liu, M., Gajan, A. and Pile, L. A. (2016). Genome-wide studies reveal novel and distinct biological pathways regulated by SIN3 isoforms. BMC Genomics 17: 111. PubMed ID: 26872827
The multisubunit SIN3 complex is a global transcriptional regulator. In Drosophila, a single Sin3A gene encodes different isoforms of SIN3, of which SIN3 187 and SIN3 220 are the major isoforms. Previous studies have demonstrated functional non-redundancy of SIN3 isoforms. The role of SIN3 isoforms in regulating distinct biological processes, however, is not well characterized. This study established a Drosophila S2 cell culture model system in which cells predominantly express either SIN3 187 or SIN3 220. To identify genomic targets of SIN3 isoforms, chromatin immunoprecipitation was performed followed by deep sequencing. The data demonstrate that upon overexpression of SIN3 187, the level of SIN3 220 decreased and the large majority of genomic sites bound by SIN3 220 were instead bound by SIN3 187. RNA-seq was used to identify genes regulated by the expression of one isoform or the other. In S2 cells, which predominantly express SIN3 220, it was found that SIN3 220 directly regulates genes involved in metabolism and cell proliferation. It was also determined that SIN3 187 regulates a unique set of genes and likely modulates expression of many genes also regulated by SIN3 220. Interestingly, biological pathways enriched for genes specifically regulated by SIN3 187 strongly suggest that this isoform plays an important role during the transition from the embryonic to the larval stage of development. These data establish the role of SIN3 isoforms in regulating distinct biological processes. This study substantially contributes to understanding of the complexity of gene regulation by SIN3.

Doiguchi, M., et al. (2016). SMARCAD1 is an ATP-dependent stimulator of nucleosomal H2A acetylation via CBP, resulting in transcriptional regulation. Sci Rep 6: 20179. PubMed ID: 26888216
This study discovered an ATP-dependent histone H2A acetylation activity in Drosophila nuclear extracts. This activity was column purified and demonstrated to be composed of the enzymatic activities of CREB-binding protein (CBP) and SMARCAD1, which belongs to the Etl1 subfamily of the Snf2 family of helicase-related proteins. SMARCAD1 enhanced acetylation by CBP of H2A K5 and K8 in nucleosomes in an ATP-dependent fashion. Expression array analysis of S2 cells having ectopically expressed SMARCAD1 revealed up-regulated genes. Using native genome templates of these up-regulated genes, it was found that SMARCAD1 activates their transcription in vitro. Knockdown analysis of SMARCAD1 and CBP indicated overlapping gene control, and ChIP-seq analysis of these commonly controlled genes showed that CBP is recruited to the promoter prior to SMARCAD1. Moreover, Drosophila genetic experiments demonstrated interaction between SMARCAD1/Etl1 and CBP/nej during development. The interplay between the remodeling activity of SMARCAD1 and histone acetylation by CBP sheds light on the function of chromatin and the genome-integrity network.

Uchino, R., Sugiyama, S., Katagiri, M., Chuman, Y. and Furukawa, K. (2016). Non-farnesylated B-type lamin can tether chromatin inside the nucleus and its chromatin interaction requires the Ig-fold region. Chromosoma [Epub ahead of print]. PubMed ID: 26892013
Lamins are thought to direct heterochromatin to the nuclear lamina (NL); however, this function of lamin has not been clearly demonstrated in vivo. To address this, polytene chromosome morphology were analyzed when artificial lamin variants were expressed in Drosophila endoreplicating cells. The CaaX-motif-deleted B-type lamin Dm0, but not A-type lamin C, was able to form a nuclear envelope-independent layer that was closely associated with chromatin. Other nuclear envelope proteins were not detected in this "ectopic lamina," and the associated chromatin showed a repressive histone modification marker but not a permissive histone modification marker nor RNA polymerase II proteins. Furthermore, deletion of the C-terminal lamin-Ig-fold domain prevents chromatin association with this ectopic lamina. Thus, non-farnesylated B-type lamin Dm0 can form an ectopic lamina and induce changes to chromatin structure and status inside the interphase nucleus.

Saturday, March 4th

Ong, C., Lee, Q.Y., Cai, Y., Liu, X., Ding, J., Yung, L.Y., Bay, B.H. and Baeg, G.H. (2016). Silver nanoparticles disrupt germline stem cell maintenance in the Drosophila testis. Sci Rep 6: 20632. PubMed ID: 26847594
Silver nanoparticles (AgNPs), one of the most popular nanomaterials, are commonly used in consumer products and biomedical devices, despite their potential toxicity. Recently, AgNP exposure has been reported to be associated with male reproductive toxicity in mammalian models. However, there is still a limited understanding of the effects of AgNPs on spermatogenesis. The fruit fly Drosophila testis is an excellent in vivo model to elucidate the mechanisms underlying AgNP-induced defects in spermatogenesis, as germ lineages can be easily identified and imaged. This study evaluated AgNP-mediated toxicity on spermatogenesis by feeding Drosophila with AgNPs at various concentrations. A dose-dependent uptake of AgNPs was observed in vivo. Concomitantly, AgNP exposure causes a significant decrease in the viability and delay in the development of Drosophila in a dose-dependent manner. Furthermore, AgNP-treated male flies show a reduction in fecundity, and the resulting testes contain a decreased number of germline stem cells (GSCs) compared to controls. Interestingly, testes exposed to AgNPs exhibit a dramatic increase in reactive oxygen species levels and show precocious GSC differentiation. Taken together, this study suggests that AgNP exposure may increase ROS levels in the Drosophila testis, leading to a reduction of GSC number by promoting premature GSC differentiation. 

Börner, K., Jain, D., Vazquez-Pianzola, P., Vengadasalam, S., Steffen, N., Fyodorov, D.V., Tomancak, P., Konev, A., Suter, B. and Becker, P.B. (2016). A role for tuned levels of nucleosome remodeler subunit ACF1 during Drosophila oogenesis. Dev Biol [Epub ahead of print]. PubMed ID: 26851213
The Chromatin Accessibility Complex (CHRAC) consists of the ATPase ISWI, the large ACF1 subunit and a pair of small histone-like proteins, CHRAC-14 and CHRAC-16. CHRAC is a prototypical nucleosome sliding factor that mobilizes nucleosomes to improve the regularity and integrity of the chromatin fiber. This may facilitate the formation of repressive chromatin. This study explored roles for ACF1 during Drosophila oogenesis. ACF1 is expressed in somatic and germline cells, with notable enrichment in germline stem cells and oocytes. The asymmetrical localization of ACF1 to these cells depends on the transport of the Acf1 mRNA by the Bicaudal-D/Egalitarian complex. Loss of ACF1 function in the novel Acf17 allele leads to defective egg chambers and their elimination through apoptosis. In addition, a variety of unusual 16-cell cyst packaging phenotypes were found in the previously known Acf11 allele, with a striking prevalence of egg chambers with two functional oocytes at opposite poles. Surprisingly, Acf11 deletion - despite disruption of the Acf1 reading frame - expresses low levels of a PHD-bromodomain module from the C-terminus of ACF1 that becomes enriched in oocytes. Expression of this module from the Acf1 genomic locus leads to packaging defects in the absence of functional ACF1, suggesting competitive interactions with unknown target molecules. Remarkably, a two-fold overexpression of CHRAC (ACF1 and CHRAC-16) leads to increased apoptosis and packaging defects. Evidently, finely tuned CHRAC levels are required for proper oogenesis.

Aughey, G. N., Grice, S. J. and Liu, J. L. (2016). The interplay between Myc and CTP synthase in Drosophila. PLoS Genet 12: e1005867. PubMed ID: 26889675
CTP synthase (CTPsyn) is essential for the biosynthesis of pyrimidine nucleotides. Previous studies have shown that CTPsyn is incorporated into a novel cytoplasmic structure which has been termed the cytoophidium. This study reports that Myc regulates cytoophidium formation during Drosophila oogenesis. Myc protein levels correlate with cytoophidium abundance in follicle epithelia. Reducing Myc levels results in cytoophidium loss and small nuclear size in follicle cells, while overexpression of Myc increases the length of cytoophidia and the nuclear size of follicle cells. Ectopic expression of Myc induces cytoophidium formation in late stage follicle cells. Furthermore, knock-down of CTPsyn is sufficient to suppress the overgrowth phenotype induced by Myc overexpression, suggesting CTPsyn acts downstream of Myc and is required for Myc-mediated cell size control. Taken together, these data suggest a functional link between Myc, a renowned oncogene, and the essential nucleotide biosynthetic enzyme CTPsyn.

Lee, J., Lee, S., Chen, C., Shim, H. and Kim-Ha, J. (2016). shot regulates the microtubule reorganization required for localization of axis-determining mRNAs during oogenesis. FEBS Lett [Epub ahead of print]. PubMed ID: 26832192
The Drosophila mid-oogenesis stages are notable as the time when most maternal mRNAs become localized at discrete regions of the oocyte. Microtubule rearrangement occurs during this period and is critical for the localization of axis-determining maternal mRNAs. This study has identified the cytoskeletal cross-linker protein short stop (shot) as a key player in establishing the cytoskeletal arrangement required for the spatial localization of axis-determining maternal mRNAs. The spatial distribution of the Shot protein was found to be regulated by its mRNA localization. These results suggest that the RNA localization mechanism is used not only for restricted accumulation of patterning molecules but also for the microtubule organization that leads to the initial development of oocyte polarity.

Friday, March 4th

Terada, S. I., Matsubara, D., Onodera, K., Matsuzaki, M., Uemura, T. and Usui, T. (2016). Neuronal processing of noxious thermal stimuli mediated by dendritic Ca influx in somatosensory neurons. Elife 5. PubMed ID: 26880554
Adequate responses to noxious stimuli causing tissue damages are essential for organismal survival. Class IV neurons in Drosophila larvae are polymodal nociceptors responsible for thermal, mechanical, and light sensation. Importantly, activation of Class IV provoked distinct avoidance behaviors, depending on the inputs. Noxious thermal stimuli, but not blue light stimulation, was shown to cause a unique pattern of Class IV, which were composed of pauses after high frequency spike trains and a large Ca2+ rise in the dendrite (the Ca2+ transient). Both of these responses depended on two TRPA channels and the L-type voltage-gated calcium channel (L-VGCC), showing that the thermosensation provokes Ca2+ influx. The precipitous fluctuation of firing rate in Class IV neurons enhanced the robust heat avoidance. It is hypothesized that the Ca2+ influx can be a key signal encoding a specific modality.

Fushiki, A., Zwart, M. F., Kohsaka, H., Fetter, R. D., Cardona, A. and Nose, A. (2016). A circuit mechanism for the propagation of waves of muscle contraction in Drosophila. Elife 5. PubMed ID: 26880545
Animals move by adaptively coordinating the sequential activation of muscles. The circuit mechanisms underlying coordinated locomotion are poorly understood. This study reports on a novel circuit for propagation of waves of muscle contraction, using the peristaltic locomotion of Drosophila larvae as a model system. An intersegmental chain of synaptically connected neurons, alternating excitatory and inhibitory, was found to be necessary for wave propagation and active in phase with the wave. The excitatory neurons (A27h) are premotor and necessary only for forward locomotion, and are modulated by stretch receptors and descending inputs. The inhibitory neurons (GDL) are necessary for both forward and backward locomotion, suggestive of different yet coupled central pattern generators, and its inhibition is necessary for wave propagation. The circuit structure and functional imaging indicated that the commands to contract one segment promote the relaxation of the next segment, revealing a mechanism for wave propagation in peristaltic locomotion.

Ray, R.P., Nakata, T., Henningsson, P. and Bomphrey, R.J. (2016). Enhanced flight performance by genetic manipulation of wing shape in Drosophila. Nat Commun 7: 10851. PubMed ID: 26926954
Insect wing shapes are remarkably diverse and the combination of shape and kinematics determines both aerial capabilities and power requirements. However, the contribution of any specific morphological feature to performance is not known. Using targeted RNA interference to modify wing shape far beyond the natural variation found within the population of a single species, this study shows a direct effect on flight performance that can be explained by physical modelling of the novel wing geometry. Data show that altering the expression of a single gene (narrow) can significantly enhance aerial agility and that the Drosophila wing shape is not, therefore, optimized for certain flight performance characteristics that are known to be important. This technique points in a new direction for experiments on the evolution of performance specialities in animals.

Delventhal, R. and Carlson, J. R. (2016). Bitter taste receptors confer diverse functions to neurons. Elife 5. PubMed ID: 26880560
Bitter compounds elicit an aversive response. In Drosophila, bitter-sensitive taste neurons coexpress many members of the Gr family of taste receptors. However, the molecular logic of bitter signaling is unknown. This study used an in vivo expression approach to analyze the logic of bitter taste signaling. Ectopic or overexpression of bitter Grs increased endogenous responses or conferred novel responses. Surprisingly, expression of Grs also suppressed many endogenous bitter responses. Conversely, deletion of an endogenous Gr led to novel responses. Expression of individual Grs conferred strikingly different effects in different neurons. The results support a model in which bitter Grs interact, exhibiting competition, inhibition, or activation. The results have broad implications for the problem of how taste systems evolve to detect new environmental dangers.

Tuthill, J.C. and Wilson, R.I. (2016). Parallel transformation of tactile signals in central circuits of Drosophila. Cell 164: 1046-1059. PubMed ID: 26919434
To distinguish between complex somatosensory stimuli, central circuits must combine signals from multiple peripheral mechanoreceptor types, as well as mechanoreceptors at different sites in the body. This study investigates the first stages of somatosensory integration in Drosophila using in vivo recordings from genetically labeled central neurons in combination with mechanical and optogenetic stimulation of specific mechanoreceptor types. Three classes of central neurons that process touch were identified: one compares touch signals on different parts of the same limb, one compares touch signals on right and left limbs, and the third compares touch and proprioceptive signals. Each class encodes distinct features of somatosensory stimuli. The axon of an individual touch receptor neuron can diverge to synapse onto all three classes, meaning that these computations occur in parallel, not hierarchically. Representing a stimulus as a set of parallel comparisons is a fast and efficient way to deliver somatosensory signals to motor circuits. 

Ardin, P., Peng, F., Mangan, M., Lagogiannis, K. and Webb, B. (2016). Using an insect mushroom body circuit to encode route memory in complex natural environments. PLoS Comput Biol 12: e1004683. PubMed ID: 26866692
Ants, like many other animals, use visual memory to follow extended routes through complex environments, but it is unknown how their small brains implement this capability. The mushroom body neuropils have been identified as a crucial memory circuit in the insect brain, but their function has mostly been explored for simple olfactory association tasks. This study shows that a spiking neural model of this circuit originally developed to describe fruitfly (Drosophila melanogaster) olfactory association, can also account for the ability of desert ants (Cataglyphis velox) to rapidly learn visual routes through complex natural environments. It was further demonstrated that abstracting the key computational principles of this circuit, which include one-shot learning of sparse codes, enables the theoretical storage capacity of the ant mushroom body to be estimated at hundreds of independent images.

Thursday, March 3rd

Sivachenko, A., Gordon, H. B., Kimball, S. S., Gavin, E. J., Bonkowsky, J. L. and Letsou, A. (2016). Neurodegeneration in a Drosophila model of Adrenoleukodystrophy: the roles of the bubblegum and double bubble acyl-CoA synthetases. Dis Model Mech [Epub ahead of print]. PubMed ID: 26893370
Debilitating neurodegenerative conditions with metabolic origins affect millions of individuals worldwide. Still, for most of these neurometabolic disorders there are neither cures nor disease- modifying therapies, and novel animal models are needed for elucidation of disease pathology and identification of potential therapeutic agents. This study presents the first analysis of a very long chain acyl-CoA synthetase double mutant. The Drosophila bubblegum (bgm) and double bubble (dbb) genes have overlapping functions, and the consequences of bubblegum double bubble double knockout in the fly brain are profound, affecting behavior and brain morphology, and providing the best paradigm to date for an animal model of Adrenoleukodystrophy (ALD), a fatal childhood neurodegenerative disease associated with the accumulation of very long chain fatty acids. Using this more fully penetrant model of disease to interrogate brain morphology at the level of electron microscopy, this study shows that dysregulation of fatty acid metabolism via disruption of ACS function in vivo is causal of neurodegenerative pathologies evident in both neuronal cells and their support cell populations, and leads ultimately to lytic cell death in affected areas of the brain. Finally, in an extension of the model system to the study of human disease, identification of a leukodystrophy patient who harbors a rare mutation in a human homologue of Bgm and Dbb was found: the SLC27a6-encoded very-long-chain acyl-CoA synthetase.

Zhang, C. W., Adeline, H. B., Chai, B. H., Hong, E. T., Ng, C. H. and Lim, K. L. (2016). Pharmacological or Genetic Activation of Hsp70 Protects against Loss of Parkin Function. Neurodegener Dis. PubMed ID: 26886023
Mutations of parkin are a prevalent genetic contributor to familial Parkinson's disease (PD). As a key regulator of protein and mitochondrial homeostasis, parkin plays a pivotal role in maintaining dopaminergic neuronal survival. However, whereas Drosophila parkin null mutants exhibit prominent parkinsonian features, parkin-deficient mice generally lack an overt phenotype. This study found that the expression of Hsp70 along with several other members of the chaperone family is elevated in parkin null mice, suggesting a possible compensatory mechanism for the loss of parkin function in these mice that could have masked their phenotype. Supporting this, it was demonstrated that the enhancement of chaperone function induced either pharmacologically via 17-AAG treatment or genetically via Hsp70 overexpression can protect cells against proteolytic and mitochondrial stress in a manner that is similar to that brought about by parkin overexpression. Importantly, it was further shown that enhanced chaperone activity can ameliorate the pathological phenotypes in Drosophila parkin null mutants, which suggests the ability of chaperones to phenocopy parkin function. Taken together, these results suggest that Hsp members may act as compensatory factors for parkin loss of function and that the exploitation of these factors may be of potential therapeutic value.

Troutwine, B.R., Ghezzi, A., Pietrzykowski, A.Z. and Atkinson, N.S. (2016). Alcohol resistance in Drosophila is modulated by the Toll innate immune pathway. Genes Brain Behav [Epub ahead of print]. PubMed ID: 26916032
This study shows that the Toll innate immune signaling pathway modulates the level of alcohol resistance in Drosophila. In humans, a low level of response to alcohol is correlated with increased risk of developing an alcohol use disorder. The Toll signaling pathway was originally discovered in, and has been extensively studied in Drosophila. The Toll pathway is a major regulator of innate immunity in Drosophila, and mammalian Toll-like receptor signaling has been implicated in alcohol responses. This study uses Drosophila-specific genetic tools to test eight genes in the Toll signaling pathway for effects on the level of response to ethanol. It was shown that increasing the activity of the pathway increases ethanol resistance while decreasing pathway activity reduces ethanol resistance. Furthermore, gene products known to be outputs of innate immune signaling are rapidly induced following ethanol exposure. The interaction between the Toll signaling pathway and ethanol is rooted in the natural history of Drosophila melanogaster.

Nagy, P., Kovács, L., Sándor, G.O. and Juhász, G. (2016). Stem cell-specific endocytic degradation defects lead to intestinal dysplasia in Drosophila. Dis Model Mech [Epub ahead of print]. PubMed ID: 26921396
UVRAG is a tumor suppressor involved in autophagy, endocytosis and DNA damage repair, but how its loss contributes to colorectal cancer is poorly understood. This study shows that UVRAG deficiency in Drosophila intestinal stem cells leads to uncontrolled proliferation and impaired differentiation without preventing autophagy. As a result, affected animals suffer from gut dysfunction and short lifespan. Dysplasia upon loss of UVRAG is characterized by the accumulation of endocytosed ligands and sustained activation of STAT and JNK signaling, and attenuation of these pathways suppresses stem cell hyperproliferation. Importantly, the inhibition of early (dynamin-dependent) or late (Rab7-dependent) steps of endocytosis in intestinal stem cells also induces hyperproliferation and dysplasia. These data raise the possibility that endocytic but not autophagic defects contribute to UVRAG deficient colorectal cancer development in human patients.

Zhou, S., Morozova, T. V., Hussain, Y. N., Luoma, S. E., McCoy, L., Yamamoto, A., Mackay, T. F. and Anholt, R. R. (2016). The genetic basis for variation in sensitivity to lead toxicity in Drosophila. Environ Health Perspect. PubMed ID: 26859824
The goal of this study was to use Drosophila melanogaster to identify evolutionarily conserved candidate genes associated with individual variation in susceptibility to lead exposure. To identify candidate genes associated with variation in susceptibility to lead toxicity effects of lead exposure were measured on development time, viability and adult activity in the Drosophila Genetic Reference Panel (DGRP)' genome-wide association analyses was performed to identify candidate genes. Mutants were used to assess functional causality of candidate genes, and a genetic network associated with variation in sensitivity to lead exposure was constructed, on which human orthologs could be superimpose. Substantial heritabilities were found for all three traits, and candidate genes associated with variation in susceptibility to lead exposure were identified for each phenotype. The genetic architectures that determine variation in sensitivity to lead exposure are highly polygenic. Gene ontology and network analyses showed enrichment of genes associated with early development and function of the nervous system. Drosophila presents an advantageous model to study the genetic underpinnings of variation in susceptibility to lead toxicity. Evolutionary conservation of cellular pathways that respond to toxic exposure allows predictions regarding orthologous genes and pathways across phyla. Thus, studies in the Drosophila model system can identify candidate susceptibility genes to guide subsequent studies in human populations.

Harvanek, Z. M., Mourao, M. A., Schnell, S. and Pletcher, S. D. (2016). A computational approach to studying ageing at the individual level. Proc Biol Sci 283. PubMed ID: 26865300
The ageing process is actively regulated throughout an organism's life, but studying the rate of ageing in individuals is difficult with conventional methods. Consequently, ageing studies typically make biological inference based on population mortality rates, which often do not accurately reflect the probabilities of death at the individual level. To study the relationship between individual and population mortality rates, in vivo switch experiments were integrated with in silico stochastic simulations to elucidate how carefully designed experiments allow key aspects of individual ageing to be deduced from group mortality measurements. As this case study, the recent report was used demonstrating that pheromones of the opposite sex decrease lifespan in Drosophila melanogaster by reversibly increasing population mortality rates. The population mortality reversal following pheromone removal was shown to be almost surely occurring in individuals, albeit more slowly than suggested by population measures. Furthermore, heterogeneity among individuals due to the inherent stochasticity of behavioural interactions skewed population mortality rates in middle-age away from the individual-level trajectories of which they are comprised. This article exemplifies how computational models function as important predictive tools for designing wet-laboratory experiments to use population mortality rates to understand how genetic and environmental manipulations affect ageing in the individual.

Wednesday, March 2nd

Peco, E., Davla, S., Camp, D., Stacey, S., Landgraf, M. and van Meyel, D. (2016). Drosophila astrocytes cover specific territories of CNS neuropil and are instructed to differentiate by Prospero, a key effector of Notch. Development [Epub ahead of print]. PubMed ID: 26893340
Astrocytes are recognized as critical elements in the formation, fine-tuning, function and plasticity of neural circuits in the central nervous system. However, important questions remain unanswered about the mechanisms instructing astrocyte cell fate. This paper describes a study of astrogenesis in the ventral nerve cord of Drosophila larvae, where astrocytes have remarkable morphological and molecular similarities to astrocytes in mammals. The births of larval astrocytes from a multi-glial lineage are described, their allocation to reproducible positions, and their deployment of ramified arbors to cover specific neuropil territories to form a stereotyped astroglial map. Finally, a molecular pathway was unraveled for astrocyte differentiation in which the Ets protein Pointed and Notch signaling pathway are required for astrogenesis; however, only Notch is sufficient to direct non-astrocytic progenitors toward astrocytic fate. Prospero was found to be a key effector of Notch in this process. These data identify an instructive astrogenic program that acts as a binary switch to distinguish astrocytes from other glial cells.

Semmens, D. C., Mirabeau, O., Moghul, I., Pancholi, M. R., Wurm, Y. and Elphick, M. R. (2016). Transcriptomic identification of starfish neuropeptide precursors yields new insights into neuropeptide evolution. Open Biol 6. PubMed ID: 26865025
This study has identified 40 neuropeptide precursors in the starfish Asterias rubens, a deuterostomian invertebrate from the phylum Echinodermata. Importantly, these include kisspeptin-type and melanin-concentrating hormone-type precursors, which are the first to be discovered in a non-chordate species. Starfish tachykinin-type, somatostatin-type, pigment-dispersing factor-type and corticotropin-releasing hormone-type precursors are the first to be discovered in the echinoderm/ambulacrarian clade of the animal kingdom. Other precursors identified include vasopressin/oxytocin-type, gonadotropin-releasing hormone-type, thyrotropin-releasing hormone-type, calcitonin-type, cholecystokinin/gastrin-type, orexin-type, luqin-type, pedal peptide/orcokinin-type, glycoprotein hormone-type, bursicon-type, relaxin-type and insulin-like growth factor-type precursors. This is the most comprehensive identification of neuropeptide precursor proteins in an echinoderm to date, yielding new insights into the evolution of neuropeptide signalling systems. Furthermore, these data provide a basis for experimental analysis of neuropeptide function in the unique context of the decentralized, pentaradial echinoderm bauplan.

Thoma, V., Knapek, S., Arai, S., Hartl, M., Kohsaka, H., Sirigrivatanawong, P., Abe, A., Hashimoto, K. and Tanimoto, H. (2016). Functional dissociation in sweet taste receptor neurons between and within taste organs of Drosophila. Nat Commun 7: 10678. PubMed ID: 26893070
Finding food sources is essential for survival. Insects detect nutrients with external taste receptor neurons. Drosophila possesses multiple taste organs that are distributed throughout its body. However, the role of different taste organs in feeding remains poorly understood. By blocking subsets of sweet taste receptor neurons, receptor neurons in the legs were shown to be required for immediate sugar choice. Furthermore, two anatomically distinct classes of sweet taste receptor neurons were identified in the leg. The axonal projections of one class terminate in the thoracic ganglia, whereas the other projects directly to the brain. These two classes are functionally distinct: the brain-projecting neurons are involved in feeding initiation, whereas the thoracic ganglia-projecting neurons play a role in sugar-dependent suppression of locomotion. Distinct receptor neurons for the same taste quality may coordinate early appetitive responses, taking advantage of the legs as the first appendages to contact food.

Liang, X., Holy, T.E. and Taghert, P.H. (2016). Synchronous Drosophila circadian pacemakers display nonsynchronous Ca²⁺ rhythms in vivo. Science 351: 976-981. PubMed ID: 26917772
In Drosophila, molecular clocks control circadian rhythmic behavior through a network of ~150 pacemaker neurons. To explain how the network's neuronal properties encode time, this study performed brainwide calcium imaging of groups of pacemaker neurons in vivo for 24 hours. Pacemakers exhibit daily rhythmic changes in intracellular Ca(2+) that are entrained by environmental cues and timed by molecular clocks. However, these rhythms are not synchronous, as each group exhibits its own phase of activation. Ca(2+) rhythms displayed by pacemaker groups that are associated with the morning or evening locomotor activities occur ~4 hours before their respective behaviors. Loss of the receptor for the neuropeptide PDF promotes synchrony of Ca(2+) waves. Thus, neuropeptide modulation is required to sequentially time outputs from a network of synchronous molecular pacemakers.

Tuesday, March 1st

Fujioka, M., Mistry, H., Schedl, P. and Jaynes, J. B. (2016). Determinants of chromosome architecture: Insulator pairing in cis and in trans. PLoS Genet 12: e1005889. PubMed ID: 26910731
To elucidate the principles governing insulator architectural functions, this study used two insulators, Homie and Nhomie, that flank the Drosophila even-skipped locus. It was shown that homologous insulator interactions in trans, between Homie on one homolog and Homie on the other, or between Nhomie on one homolog and Nhomie on the other, mediate transvection. Critically, these homologous insulator:insulator interactions are orientation-dependent. Consistent with a role in the alignment and pairing of homologs, self-pairing in trans is head-to-head. Head-to-head self-interactions in cis have been reported for other fly insulators, suggesting that this is a general principle of self-pairing. Homie and Nhomie not only pair with themselves, but with each other. Heterologous Homie-Nhomie interactions occur in cis, and they serve to delimit a looped chromosomal domain that contains the even skipped transcription unit and its associated enhancers. The topology of this loop is defined by the heterologous pairing properties of Homie and Nhomie. Instead of being head-to-head, which would generate a circular loop, Homie-Nhomie pairing is head-to-tail. Head-to-tail pairing in cis generates a stem-loop, a configuration much like that observed in classical lampbrush chromosomes. These pairing principles provide a mechanistic underpinning for the observed topologies within and between chromosomes.

Heseding, C., Saumweber, H., Rathke, C. and Ehrenhofer-Murray, A. E. (2016). Widespread colocalization of the Drosophila histone acetyltransferase homolog MYST5 with DREF and insulator proteins at active genes. Chromosoma [Epub ahead of print]. PubMed ID: 26894919
MYST family histone acetyltransferases play important roles in gene regulation. This study has characterized the Drosophila MYST histone acetyltransferase (HAT) encoded by CG1894, whose closest homolog is Drosophila MOF, and which was termed MYST5. It localized to a large number of interbands as well as to the telomeres of polytene chromosomes, and it showed strong colocalization with the interband protein Z4/Putzig and RNA polymerase II. Accordingly, genome-wide location analysis by ChIP-seq showed co-occurrence of MYST5 with the Z4-interacting partner Chriz/Chromator. Interestingly, MYST5 bound to the promoter of actively transcribed genes, and about half of MYST5 sites colocalized with the transcription factor DNA replication-related element-binding factor (DREF), indicating a role for MYST5 in gene expression. Moreover, substantial overlap of MYST5 binding was observed with that of the insulator proteins CP190, dCTCF, and BEAF-32, which mediate the organization of the genome into functionally distinct topological domains. Altogether, these data suggest a broad role for MYST5 both in gene-specific transcriptional regulation and in the organization of the genome into chromatin domains, with the two roles possibly being functionally interconnected.

Shinoda, N., Obata, F., Zhang, L. and Miura, M. (2016). Drosophila SETDB1 and caspase cooperatively fine-tune cell fate determination of sensory organ precursor. Genes Cells [Epub ahead of print]. PubMed ID: 26914287
Drosophila produce a constant number of mechanosensory bristles called macrochaetae (MC), which develop from sensory organ precursor (SOP) cells within a proneural cluster (PNC). However, what ensures the precise determination of SOP cells remains to be elucidated. This study conducted RNAi screening in PNC for genes involved in epigenetic regulation. A H3K9 histone methyltransferase, SETDB1/eggless, was identified as a regulator of SOP development. Knockdown of SETDB1 in PNC leads to additional SOPs. The relationship between SETDB1 and non-apoptotic function of caspase was tested on SOP development. Reinforcing caspase activation by heterozygous Drosophila inhibitor of apoptosis protein 1 (DIAP1) mutation rescues ectopic SOP development caused by SETDB1 knockdown. Knockdown of SETDB1, however, was found to have little effect on caspase activity. Simultaneous loss of SETDB1 and caspase activity results in further increase in MC, indicating that the two components work cooperatively. These results exhibit the fine-tuning mechanisms for SOP development by epigenetic methyltransferase and non-apoptotic caspase function.

Zhou, J. and Troyanskaya, O. G. (2016). Probabilistic modelling of chromatin code landscape reveals functional diversity of enhancer-like chromatin states. Nat Commun 7: 10528. PubMed ID: 26841971
Interpreting the functional state of chromatin from the combinatorial binding patterns of chromatin factors, that is, the chromatin codes, is crucial for decoding the epigenetic state of the cell. This study presents a systematic map of Drosophila chromatin states derived from data-driven probabilistic modelling of dependencies between chromatin factors. The model not only recapitulates enhancer-like chromatin states as indicated by widely used enhancer marks but also divides these states into three functionally distinct groups, of which only one specific group possesses active enhancer activity. Moreover, a strong association was discovered between one specific enhancer state and RNA Polymerase II pausing, linking transcription regulatory potential and chromatin organization. With the exception of long-intron genes, chromatin state transition positions in transcriptionally active genes were also observed to align with an absolute distance to their corresponding transcription start site, regardless of gene length. Using this method, a resource is provided that helps elucidate the functional and spatial organization of the chromatin code landscape.

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