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Sample records for function regulates drosophila

  1. Distinct functions of Crumbs regulating slit diaphragms and endocytosis in Drosophila nephrocytes.

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    Hochapfel, Florian; Denk, Lucia; Mendl, Gudrun; Schulze, Ulf; Maaßen, Christine; Zaytseva, Yulia; Pavenstädt, Hermann; Weide, Thomas; Rachel, Reinhard; Witzgall, Ralph; Krahn, Michael P

    2017-12-01

    Mammalian podocytes, the key determinants of the kidney's filtration barrier, differentiate from columnar epithelial cells and several key determinants of apical-basal polarity in the conventional epithelia have been shown to regulate podocyte morphogenesis and function. However, little is known about the role of Crumbs, a conserved polarity regulator in many epithelia, for slit-diaphragm formation and podocyte function. In this study, we used Drosophila nephrocytes as model system for mammalian podocytes and identified a conserved function of Crumbs proteins for cellular morphogenesis, nephrocyte diaphragm assembly/maintenance, and endocytosis. Nephrocyte-specific knock-down of Crumbs results in disturbed nephrocyte diaphragm assembly/maintenance and decreased endocytosis, which can be rescued by Drosophila Crumbs as well as human Crumbs2 and Crumbs3, which were both expressed in human podocytes. In contrast to the extracellular domain, which facilitates nephrocyte diaphragm assembly/maintenance, the intracellular FERM-interaction motif of Crumbs is essential for regulating endocytosis. Moreover, Moesin, which binds to the FERM-binding domain of Crumbs, is essential for efficient endocytosis. Thus, we describe here a new mechanism of nephrocyte development and function, which is likely to be conserved in mammalian podocytes.

  2. Rasputin functions as a positive regulator of orb in Drosophila oogenesis.

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    Alexandre Costa

    Full Text Available The determination of cell fate and the establishment of polarity axes during Drosophila oogenesis depend upon pathways that localize mRNAs within the egg chamber and control their on-site translation. One factor that plays a central role in regulating on-site translation of mRNAs is Orb. Orb is a founding member of the conserved CPEB family of RNA-binding proteins. These proteins bind to target sequences in 3' UTRs and regulate mRNA translation by modulating poly(A tail length. In addition to controlling the translation of axis-determining mRNAs like grk, fs(1K10, and osk, Orb protein autoregulates its own synthesis by binding to orb mRNA and activating its translation. We have previously shown that Rasputin (Rin, the Drosophila homologue of Ras-GAP SH3 Binding Protein (G3BP, associates with Orb in a messenger ribonucleoprotein (mRNP complex. Rin is an evolutionarily conserved RNA-binding protein believed to function as a link between Ras signaling and RNA metabolism. Here we show that Orb and Rin form a complex in the female germline. Characterization of a new rin allele shows that rin is essential for oogenesis. Co-localization studies suggest that Orb and Rin form a complex in the oocyte at different stages of oogenesis. This is supported by genetic and biochemical analyses showing that rin functions as a positive regulator in the orb autoregulatory pathway by increasing Orb protein expression. Tandem Mass Spectrometry analysis shows that several canonical stress granule proteins are associated with the Orb-Rin complex suggesting that a conserved mRNP complex regulates localized translation during oogenesis in Drosophila.

  3. Rasputin functions as a positive regulator of orb in Drosophila oogenesis.

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    Costa, Alexandre; Pazman, Cecilia; Sinsimer, Kristina S; Wong, Li Chin; McLeod, Ian; Yates, John; Haynes, Susan; Schedl, Paul

    2013-01-01

    The determination of cell fate and the establishment of polarity axes during Drosophila oogenesis depend upon pathways that localize mRNAs within the egg chamber and control their on-site translation. One factor that plays a central role in regulating on-site translation of mRNAs is Orb. Orb is a founding member of the conserved CPEB family of RNA-binding proteins. These proteins bind to target sequences in 3' UTRs and regulate mRNA translation by modulating poly(A) tail length. In addition to controlling the translation of axis-determining mRNAs like grk, fs(1)K10, and osk, Orb protein autoregulates its own synthesis by binding to orb mRNA and activating its translation. We have previously shown that Rasputin (Rin), the Drosophila homologue of Ras-GAP SH3 Binding Protein (G3BP), associates with Orb in a messenger ribonucleoprotein (mRNP) complex. Rin is an evolutionarily conserved RNA-binding protein believed to function as a link between Ras signaling and RNA metabolism. Here we show that Orb and Rin form a complex in the female germline. Characterization of a new rin allele shows that rin is essential for oogenesis. Co-localization studies suggest that Orb and Rin form a complex in the oocyte at different stages of oogenesis. This is supported by genetic and biochemical analyses showing that rin functions as a positive regulator in the orb autoregulatory pathway by increasing Orb protein expression. Tandem Mass Spectrometry analysis shows that several canonical stress granule proteins are associated with the Orb-Rin complex suggesting that a conserved mRNP complex regulates localized translation during oogenesis in Drosophila.

  4. Codon usage regulates protein structure and function by affecting translation elongation speed in Drosophila cells.

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    Zhao, Fangzhou; Yu, Chien-Hung; Liu, Yi

    2017-08-21

    Codon usage biases are found in all eukaryotic and prokaryotic genomes and have been proposed to regulate different aspects of translation process. Codon optimality has been shown to regulate translation elongation speed in fungal systems, but its effect on translation elongation speed in animal systems is not clear. In this study, we used a Drosophila cell-free translation system to directly compare the velocity of mRNA translation elongation. Our results demonstrate that optimal synonymous codons speed up translation elongation while non-optimal codons slow down translation. In addition, codon usage regulates ribosome movement and stalling on mRNA during translation. Finally, we show that codon usage affects protein structure and function in vitro and in Drosophila cells. Together, these results suggest that the effect of codon usage on translation elongation speed is a conserved mechanism from fungi to animals that can affect protein folding in eukaryotic organisms. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  5. Functional genomics identifies regulators of the phototransduction machinery in the Drosophila larval eye and adult ocelli.

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    Mishra, Abhishek Kumar; Bargmann, Bastiaan O R; Tsachaki, Maria; Fritsch, Cornelia; Sprecher, Simon G

    2016-02-15

    Sensory perception of light is mediated by specialized Photoreceptor neurons (PRs) in the eye. During development all PRs are genetically determined to express a specific Rhodopsin (Rh) gene and genes mediating a functional phototransduction pathway. While the genetic and molecular mechanisms of PR development is well described in the adult compound eye, it remains unclear how the expression of Rhodopsins and the phototransduction cascade is regulated in other visual organs in Drosophila, such as the larval eye and adult ocelli. Using transcriptome analysis of larval PR-subtypes and ocellar PRs we identify and study new regulators required during PR differentiation or necessary for the expression of specific signaling molecules of the functional phototransduction pathway. We found that the transcription factor Krüppel (Kr) is enriched in the larval eye and controls PR differentiation by promoting Rh5 and Rh6 expression. We also identified Camta, Lola, Dve and Hazy as key genes acting during ocellar PR differentiation. Further we show that these transcriptional regulators control gene expression of the phototransduction cascade in both larval eye and adult ocelli. Our results show that PR cell type-specific transcriptome profiling is a powerful tool to identify key transcriptional regulators involved during several aspects of PR development and differentiation. Our findings greatly contribute to the understanding of how combinatorial action of key transcriptional regulators control PR development and the regulation of a functional phototransduction pathway in both larval eye and adult ocelli. Copyright © 2015 Elsevier Inc. All rights reserved.

  6. Conserved properties of Drosophila Insomniac link sleep regulation and synaptic function.

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    Li, Qiuling; Kellner, David A; Hatch, Hayden A M; Yumita, Tomohiro; Sanchez, Sandrine; Machold, Robert P; Frank, C Andrew; Stavropoulos, Nicholas

    2017-05-01

    Sleep is an ancient animal behavior that is regulated similarly in species ranging from flies to humans. Various genes that regulate sleep have been identified in invertebrates, but whether the functions of these genes are conserved in mammals remains poorly explored. Drosophila insomniac (inc) mutants exhibit severely shortened and fragmented sleep. Inc protein physically associates with the Cullin-3 (Cul3) ubiquitin ligase, and neuronal depletion of Inc or Cul3 strongly curtails sleep, suggesting that Inc is a Cul3 adaptor that directs the ubiquitination of neuronal substrates that impact sleep. Three proteins similar to Inc exist in vertebrates-KCTD2, KCTD5, and KCTD17-but are uncharacterized within the nervous system and their functional conservation with Inc has not been addressed. Here we show that Inc and its mouse orthologs exhibit striking biochemical and functional interchangeability within Cul3 complexes. Remarkably, KCTD2 and KCTD5 restore sleep to inc mutants, indicating that they can substitute for Inc in vivo and engage its neuronal targets relevant to sleep. Inc and its orthologs localize similarly within fly and mammalian neurons and can traffic to synapses, suggesting that their substrates may include synaptic proteins. Consistent with such a mechanism, inc mutants exhibit defects in synaptic structure and physiology, indicating that Inc is essential for both sleep and synaptic function. Our findings reveal that molecular functions of Inc are conserved through ~600 million years of evolution and support the hypothesis that Inc and its orthologs participate in an evolutionarily conserved ubiquitination pathway that links synaptic function and sleep regulation.

  7. Two distinct E3 ubiquitin ligases have complementary functions in the regulation of delta and serrate signaling in Drosophila.

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    Roland Le Borgne

    2005-04-01

    Full Text Available Signaling by the Notch ligands Delta (Dl and Serrate (Ser regulates a wide variety of essential cell-fate decisions during animal development. Two distinct E3 ubiquitin ligases, Neuralized (Neur and Mind bomb (Mib, have been shown to regulate Dl signaling in Drosophila melanogaster and Danio rerio, respectively. While the neur and mib genes are evolutionarily conserved, their respective roles in the context of a single organism have not yet been examined. We show here that the Drosophila mind bomb (D-mib gene regulates a subset of Notch signaling events, including wing margin specification, leg segmentation, and vein determination, that are distinct from those events requiring neur activity. D-mib also modulates lateral inhibition, a neur- and Dl-dependent signaling event, suggesting that D-mib regulates Dl signaling. During wing development, expression of D-mib in dorsal cells appears to be necessary and sufficient for wing margin specification, indicating that D-mib also regulates Ser signaling. Moreover, the activity of the D-mib gene is required for the endocytosis of Ser in wing imaginal disc cells. Finally, ectopic expression of neur in D-mib mutant larvae rescues the wing D-mib phenotype, indicating that Neur can compensate for the lack of D-mib activity. We conclude that D-mib and Neur are two structurally distinct proteins that have similar molecular activities but distinct developmental functions in Drosophila.

  8. Cyclin G Functions as a Positive Regulator of Growth and Metabolism in Drosophila.

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    Patrick Fischer

    2015-08-01

    Full Text Available In multicellular organisms, growth and proliferation is adjusted to nutritional conditions by a complex signaling network. The Insulin receptor/target of rapamycin (InR/TOR signaling cascade plays a pivotal role in nutrient dependent growth regulation in Drosophila and mammals alike. Here we identify Cyclin G (CycG as a regulator of growth and metabolism in Drosophila. CycG mutants have a reduced body size and weight and show signs of starvation accompanied by a disturbed fat metabolism. InR/TOR signaling activity is impaired in cycG mutants, combined with a reduced phosphorylation status of the kinase Akt1 and the downstream factors S6-kinase and eukaryotic translation initiation factor 4E binding protein (4E-BP. Moreover, the expression and accumulation of Drosophila insulin like peptides (dILPs is disturbed in cycG mutant brains. Using a reporter assay, we show that the activity of one of the first effectors of InR signaling, Phosphoinositide 3-kinase (PI3K92E, is unaffected in cycG mutants. However, the metabolic defects and weight loss in cycG mutants were rescued by overexpression of Akt1 specifically in the fat body and by mutants in widerborst (wdb, the B'-subunit of the phosphatase PP2A, known to downregulate Akt1 by dephosphorylation. Together, our data suggest that CycG acts at the level of Akt1 to regulate growth and metabolism via PP2A in Drosophila.

  9. Hearing regulates Drosophila aggression.

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    Versteven, Marijke; Vanden Broeck, Lies; Geurten, Bart; Zwarts, Liesbeth; Decraecker, Lisse; Beelen, Melissa; Göpfert, Martin C; Heinrich, Ralf; Callaerts, Patrick

    2017-02-21

    Aggression is a universal social behavior important for the acquisition of food, mates, territory, and social status. Aggression in Drosophila is context-dependent and can thus be expected to involve inputs from multiple sensory modalities. Here, we use mechanical disruption and genetic approaches in Drosophila melanogaster to identify hearing as an important sensory modality in the context of intermale aggressive behavior. We demonstrate that neuronal silencing and targeted knockdown of hearing genes in the fly's auditory organ elicit abnormal aggression. Further, we show that exposure to courtship or aggression song has opposite effects on aggression. Our data define the importance of hearing in the control of Drosophila intermale aggression and open perspectives to decipher how hearing and other sensory modalities are integrated at the neural circuit level.

  10. Evidence for dynamic network regulation of Drosophila photoreceptor function from mutants lacking the neurotransmitter histamine

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    An eDau

    2016-03-01

    Full Text Available Synaptic feedback from interneurons to photoreceptors can help to optimize visual information flow by balancing its allocation on retinal pathways under changing light conditions. But little is known about how this critical network operation is regulated dynamically. Here, we investigate this question by comparing signaling properties and performance of wild-type Drosophila R1-R6 photoreceptors to those of the hdcJK910 mutant, which lacks the neurotransmitter histamine and therefore cannot transmit information to interneurons. Recordings show that hdcJK910 photoreceptors sample similar amounts of information from naturalistic stimulation to wild-type photoreceptors, but this information is packaged in smaller responses, especially under bright illumination. Analyses reveal how these altered dynamics primarily resulted from network overload that affected hdcJK910 photoreceptors in two ways. First, the missing inhibitory histamine input to interneurons almost certainly depolarized them irrevocably, which in turn increased their excitatory feedback to hdcJK910 R1-R6s. This tonic excitation depolarized the photoreceptors to artificially high potentials, reducing their operational range. Second, rescuing histamine input to interneurons in hdcJK910 mutant also restored their normal phasic feedback modulation to R1-R6s, causing photoreceptor output to accentuate dynamic intensity differences at bright illumination, similar to the wild-type. These results provide mechanistic explanations of how synaptic feedback connections optimize information packaging in photoreceptor output and novel insight into the operation and design of dynamic network regulation of sensory neurons.

  11. Equilibrative nucleoside transporter 2 regulates olfactory memory and synaptic function in Drosophila

    Czech Academy of Sciences Publication Activity Database

    Knight, D.; Harvey, P. J.; Iliadi, K. G.; Klose, M. K.; Iliadi, N.; Doleželová, Eva; Charlton, M. P.; Žurovec, Michal; Boulianne, G. L.

    2010-01-01

    Roč. 30, č. 14 (2010), s. 5047-5057 ISSN 0270-6474 R&D Projects: GA MŠk(CZ) LC06077 Grant - others:Canadian Institutes of Health Research(CA) MOP 14143; Canadian Institutes of Health Research(CA) MOP 67130 Institutional research plan: CEZ:AV0Z50070508 Keywords : Drosophila Subject RIV: ED - Physiology Impact factor: 7.271, year: 2010

  12. Cell-cycle regulation of non-enzymatic functions of the Drosophila methyltransferase PR-Set7.

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    Zouaz, Amel; Fernando, Céline; Perez, Yannick; Sardet, Claude; Julien, Eric; Grimaud, Charlotte

    2018-04-06

    Tight cell-cycle regulation of the histone H4-K20 methyltransferase PR-Set7 is essential for the maintenance of genome integrity. In mammals, this mainly involves the interaction of PR-Set7 with the replication factor PCNA, which triggers the degradation of the enzyme by the CRL4CDT2 E3 ubiquitin ligase. PR-Set7 is also targeted by the SCFβ-TRCP ligase, but the role of this additional regulatory pathway remains unclear. Here, we show that Drosophila PR-Set7 undergoes a cell-cycle proteolytic regulation, independently of its interaction with PCNA. Instead, Slimb, the ortholog of β-TRCP, is specifically required for the degradation of the nuclear pool of PR-Set7 prior to S phase. Consequently, inactivation of Slimb leads to nuclear accumulation of PR-Set7, which triggers aberrant chromatin compaction and G1/S arrest. Strikingly, these phenotypes result from non-enzymatic PR-Set7 functions that prevent proper histone H4 acetylation independently of H4K20 methylation. Altogether, these results identify the Slimb-mediated PR-Set7 proteolysis as a new critical regulatory mechanism required for proper interphase chromatin organization at G1/S transition.

  13. Functional analysis of the cell cycle regulator Rca1 in Drosophila melanogaster

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    Zielke, Norman

    2007-01-01

    Tight regulation of APC/C activity is essential for cell cycle progression. An important class of negative APC/C regulators are the Rca1/Emi1 family proteins. All members of the Rca1/Emi1 family share a conserved zinc binding region (ZBR) which is essential for their inhibitory activity. The Rca1/Emi1 proteins belong to the class of F-box proteins that are known to act as substrate recognition subunits in SCF-E3-ligase complexes. Emi1 and Rca1 interact in vitro with members of the Skp family ...

  14. Fermitins, the orthologs of mammalian Kindlins, regulate the development of a functional cardiac syncytium in Drosophila melanogaster.

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    James H Catterson

    Full Text Available The vertebrate Kindlins are an evolutionarily conserved family of proteins critical for integrin signalling and cell adhesion. Kindlin-2 (KIND2 is associated with intercalated discs in mice, suggesting a role in cardiac syncytium development; however, deficiency of Kind2 leads to embryonic lethality. Morpholino knock-down of Kind2 in zebrafish has a pleiotropic effect on development that includes the heart. It therefore remains unclear whether cardiomyocyte Kind2 expression is required for cardiomyocyte junction formation and the development of normal cardiac function. To address this question, the expression of Fermitin 1 and Fermitin 2 (Fit1, Fit2, the two Drosophila orthologs of Kind2, was silenced in Drosophila cardiomyocytes. Heart development was assessed in adult flies by immunological methods and videomicroscopy. Silencing both Fit1 and Fit2 led to a severe cardiomyopathy characterised by the failure of cardiomyocytes to develop as a functional syncytium and loss of synchrony between cardiomyocytes. A null allele of Fit1 was generated but this had no impact on the heart. Similarly, the silencing of Fit2 failed to affect heart function. In contrast, the silencing of Fit2 in the cardiomyocytes of Fit1 null flies disrupted syncytium development, leading to severe cardiomyopathy. The data definitively demonstrate a role for Fermitins in the development of a functional cardiac syncytium in Drosophila. The findings also show that the Fermitins can functionally compensate for each other in order to control syncytium development. These findings support the concept that abnormalities in cardiomyocyte KIND2 expression or function may contribute to cardiomyopathies in humans.

  15. Dynamical Analysis of bantam-Regulated Drosophila Circadian Rhythm Model

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    Li, Ying; Liu, Zengrong

    MicroRNAs (miRNAs) interact with 3‧untranslated region (UTR) elements of target genes to regulate mRNA stability or translation, and play a crucial role in regulating many different biological processes. bantam, a conserved miRNA, is involved in several functions, such as regulating Drosophila growth and circadian rhythm. Recently, it has been discovered that bantam plays a crucial role in the core circadian pacemaker. In this paper, based on experimental observations, a detailed dynamical model of bantam-regulated circadian clock system is developed to show the post-transcriptional behaviors in the modulation of Drosophila circadian rhythm, in which the regulation of bantam is incorporated into a classical model. The dynamical behaviors of the model are consistent with the experimental observations, which shows that bantam is an important regulator of Drosophila circadian rhythm. The sensitivity analysis of parameters demonstrates that with the regulation of bantam the system is more sensitive to perturbations, indicating that bantam regulation makes it easier for the organism to modulate its period against the environmental perturbations. The effectiveness in rescuing locomotor activity rhythms of mutated flies shows that bantam is necessary for strong and sustained rhythms. In addition, the biological mechanisms of bantam regulation are analyzed, which may help us more clearly understand Drosophila circadian rhythm regulated by other miRNAs.

  16. MicroRNA function in Drosophila melanogaster.

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    Carthew, Richard W; Agbu, Pamela; Giri, Ritika

    2017-05-01

    Over the last decade, microRNAs have emerged as critical regulators in the expression and function of animal genomes. This review article discusses the relationship between microRNA-mediated regulation and the biology of the fruit fly Drosophila melanogaster. We focus on the roles that microRNAs play in tissue growth, germ cell development, hormone action, and the development and activity of the central nervous system. We also discuss the ways in which microRNAs affect robustness. Many gene regulatory networks are robust; they are relatively insensitive to the precise values of reaction constants and concentrations of molecules acting within the networks. MicroRNAs involved in robustness appear to be nonessential under uniform conditions used in conventional laboratory experiments. However, the robust functions of microRNAs can be revealed when environmental or genetic variation otherwise has an impact on developmental outcomes. Copyright © 2016 Elsevier Ltd. All rights reserved.

  17. Drosophila VAMP7 regulates Wingless intracellular trafficking.

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    Gao, Han; He, Fang; Lin, Xinhua; Wu, Yihui

    2017-01-01

    Drosophila Wingless (Wg) is a morphogen that determines cell fate during development. Previous studies have shown that endocytic pathways regulate Wg trafficking and signaling. Here, we showed that loss of vamp7, a gene required for vesicle fusion, dramatically increased Wg levels and decreased Wg signaling. Interestingly, we found that levels of Dally-like (Dlp), a glypican that can interact with Wg to suppress Wg signaling at the dorsoventral boundary of the Drosophila wing, were also increased in vamp7 mutant cells. Moreover, Wg puncta in Rab4-dependent recycling endosomes were Dlp positive. We hypothesize that VAMP7 is required for Wg intracellular trafficking and the accumulation of Wg in Rab4-dependent recycling endosomes might affect Wg signaling.

  18. Regulation and functions of the lms homeobox gene during development of embryonic lateral transverse muscles and direct flight muscles in Drosophila.

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    Dominik Müller

    Full Text Available BACKGROUND: Patterning and differentiation of developing musculatures require elaborate networks of transcriptional regulation. In Drosophila, significant progress has been made into identifying the regulators of muscle development and defining their interactive networks. One major family of transcription factors involved in these processes consists of homeodomain proteins. In flies, several members of this family serve as muscle identity genes to specify the fates of individual muscles, or groups thereof, during embryonic and/or adult muscle development. Herein, we report on the expression and function of a new Drosophila homeobox gene during both embryonic and adult muscle development. METHODOLOGY/PRINCIPAL FINDINGS: The newly described homeobox gene, termed lateral muscles scarcer (lms, which has yet uncharacterized orthologs in other invertebrates and primitive chordates but not in vertebrates, is expressed exclusively in subsets of developing muscle tissues. In embryos, lms is expressed specifically in the four lateral transverse (LT muscles and their founder cells in each hemisegment, whereas in larval wing imaginal discs, it is expressed in myoblasts that develop into direct flight muscles (DFMs, which are important for proper wing positioning. We have analyzed the regulatory inputs of various other muscle identity genes with overlapping or complementary expression patterns towards the cell type specific regulation of lms expression. Further we demonstrate that lms null mutants exhibit reduced numbers of embryonic LT muscles, and null mutant adults feature held-out-wing phenotypes. We provide a detailed description of the pattern and morphology of the direct flight muscles in the wild type and lms mutant flies by using the recently-developed ultramicroscopy and show that, in the mutants, all DFMs are present and present normal morphologies. CONCLUSIONS/SIGNIFICANCE: We have identified the homeobox gene lms as a new muscle identity gene

  19. Comprehensive functional analysis of Rab GTPases in Drosophila nephrocytes.

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    Fu, Yulong; Zhu, Jun-Yi; Zhang, Fujian; Richman, Adam; Zhao, Zhanzheng; Han, Zhe

    2017-06-01

    The Drosophila nephrocyte is a critical component of the fly renal system and bears structural and functional homology to podocytes and proximal tubule cells of the mammalian kidney. Investigations of nephrocyte cell biological processes are fundamental to understanding the insect renal system. Nephrocytes are highly active in endocytosis and vesicle trafficking. Rab GTPases regulate endocytosis and trafficking but specific functions of nephrocyte Rabs remain undefined. We analyzed Rab GTPase expression and function in Drosophila nephrocytes and found that 11 out of 27 Drosophila Rabs were required for normal activity. Rabs 1, 5, 7, 11 and 35 were most important. Gene silencing of the nephrocyte-specific Rab5 eliminated all intracellular vesicles and the specialized plasma membrane structures essential for nephrocyte function. Rab7 silencing dramatically increased clear vacuoles and reduced lysosomes. Rab11 silencing increased lysosomes and reduced clear vacuoles. Our results suggest that Rab5 mediates endocytosis that is essential for the maintenance of functionally critical nephrocyte plasma membrane structures and that Rabs 7 and 11 mediate alternative downstream vesicle trafficking pathways leading to protein degradation and membrane recycling, respectively. Elucidating molecular pathways underlying nephrocyte function has the potential to yield important insights into human kidney cell physiology and mechanisms of cell injury that lead to disease. The Drosophila nephrocyte is emerging as a useful in vivo model system for molecular target identification and initial testing of therapeutic approaches in humans.

  20. Functional Neuroanatomy of "Drosophila" Olfactory Memory Formation

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    Guven-Ozkan, Tugba; Davis, Ronald L.

    2014-01-01

    New approaches, techniques and tools invented over the last decade and a half have revolutionized the functional dissection of neural circuitry underlying "Drosophila" learning. The new methodologies have been used aggressively by researchers attempting to answer three critical questions about olfactory memories formed with appetitive…

  1. Tet protein function during Drosophila development.

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    Fei Wang

    Full Text Available The TET (Ten-eleven translocation 1, 2 and 3 proteins have been shown to function as DNA hydroxymethylases in vertebrates and their requirements have been documented extensively. Recently, the Tet proteins have been shown to also hydroxylate 5-methylcytosine in RNA. 5-hydroxymethylcytosine (5hmrC is enriched in messenger RNA but the function of this modification has yet to be elucidated. Because Cytosine methylation in DNA is barely detectable in Drosophila, it serves as an ideal model to study the biological function of 5hmrC. Here, we characterized the temporal and spatial expression and requirement of Tet throughout Drosophila development. We show that Tet is essential for viability as Tet complete loss-of-function animals die at the late pupal stage. Tet is highly expressed in neuronal tissues and at more moderate levels in somatic muscle precursors in embryos and larvae. Depletion of Tet in muscle precursors at early embryonic stages leads to defects in larval locomotion and late pupal lethality. Although Tet knock-down in neuronal tissue does not cause lethality, it is essential for neuronal function during development through its affects upon locomotion in larvae and the circadian rhythm of adult flies. Further, we report the function of Tet in ovarian morphogenesis. Together, our findings provide basic insights into the biological function of Tet in Drosophila, and may illuminate observed neuronal and muscle phenotypes observed in vertebrates.

  2. Human Intellectual Disability Genes Form Conserved Functional Modules in Drosophila

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    Oortveld, Merel A. W.; Keerthikumar, Shivakumar; Oti, Martin; Nijhof, Bonnie; Fernandes, Ana Clara; Kochinke, Korinna; Castells-Nobau, Anna; van Engelen, Eva; Ellenkamp, Thijs; Eshuis, Lilian; Galy, Anne; van Bokhoven, Hans; Habermann, Bianca; Brunner, Han G.; Zweier, Christiane; Verstreken, Patrik; Huynen, Martijn A.; Schenck, Annette

    2013-01-01

    Intellectual Disability (ID) disorders, defined by an IQ below 70, are genetically and phenotypically highly heterogeneous. Identification of common molecular pathways underlying these disorders is crucial for understanding the molecular basis of cognition and for the development of therapeutic intervention strategies. To systematically establish their functional connectivity, we used transgenic RNAi to target 270 ID gene orthologs in the Drosophila eye. Assessment of neuronal function in behavioral and electrophysiological assays and multiparametric morphological analysis identified phenotypes associated with knockdown of 180 ID gene orthologs. Most of these genotype-phenotype associations were novel. For example, we uncovered 16 genes that are required for basal neurotransmission and have not previously been implicated in this process in any system or organism. ID gene orthologs with morphological eye phenotypes, in contrast to genes without phenotypes, are relatively highly expressed in the human nervous system and are enriched for neuronal functions, suggesting that eye phenotyping can distinguish different classes of ID genes. Indeed, grouping genes by Drosophila phenotype uncovered 26 connected functional modules. Novel links between ID genes successfully predicted that MYCN, PIGV and UPF3B regulate synapse development. Drosophila phenotype groups show, in addition to ID, significant phenotypic similarity also in humans, indicating that functional modules are conserved. The combined data indicate that ID disorders, despite their extreme genetic diversity, are caused by disruption of a limited number of highly connected functional modules. PMID:24204314

  3. The Drosophila homolog of the mammalian imprint regulator, CTCF, maintains the maternal genomic imprint in Drosophila melanogaster

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    Rasheva Vanya

    2010-07-01

    Full Text Available Abstract Background CTCF is a versatile zinc finger DNA-binding protein that functions as a highly conserved epigenetic transcriptional regulator. CTCF is known to act as a chromosomal insulator, bind promoter regions, and facilitate long-range chromatin interactions. In mammals, CTCF is active in the regulatory regions of some genes that exhibit genomic imprinting, acting as insulator on only one parental allele to facilitate parent-specific expression. In Drosophila, CTCF acts as a chromatin insulator and is thought to be actively involved in the global organization of the genome. Results To determine whether CTCF regulates imprinting in Drosophila, we generated CTCF mutant alleles and assayed gene expression from the imprinted Dp(1;fLJ9 mini-X chromosome in the presence of reduced CTCF expression. We observed disruption of the maternal imprint when CTCF levels were reduced, but no effect was observed on the paternal imprint. The effect was restricted to maintenance of the imprint and was specific for the Dp(1;fLJ9 mini-X chromosome. Conclusions CTCF in Drosophila functions in maintaining parent-specific expression from an imprinted domain as it does in mammals. We propose that Drosophila CTCF maintains an insulator boundary on the maternal X chromosome, shielding genes from the imprint-induced silencing that occurs on the paternally inherited X chromosome. See commentary: http://www.biomedcentral.com/1741-7007/8/104

  4. Functional neuroanatomy of Drosophila olfactory memory formation

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    Guven-Ozkan, Tugba; Davis, Ronald L.

    2014-01-01

    New approaches, techniques and tools invented over the last decade and a half have revolutionized the functional dissection of neural circuitry underlying Drosophila learning. The new methodologies have been used aggressively by researchers attempting to answer three critical questions about olfactory memories formed with appetitive and aversive reinforcers: (1) Which neurons within the olfactory nervous system mediate the acquisition of memory? (2) What is the complete neural circuitry exten...

  5. Drosophila Glypicans Regulate Follicle Stem Cell Maintenance and Niche Competition.

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    Su, Tsu-Yi; Nakato, Eriko; Choi, Pui Yee; Nakato, Hiroshi

    2018-04-09

    Adult stem cells reside in specialized microenvironments, called niches, which provide signals for stem cells to maintain their undifferentiated and self-renewing state. To maintain stem cell quality, several types of stem cells are known to be regularly replaced by progenitor cells through niche competition. However, the cellular and molecular bases for stem cell competition for niche occupancy are largely unknown. Here, we show that two Drosophila members of the glypican family of heparan sulfate proteoglycans (HSPGs), Dally and Dally-like (Dlp), differentially regulate follicle stem cell (FSC) maintenance and FSC competitiveness for niche occupancy. Lineage analyses of glypican mutant FSC clones showed that dally is essential for normal FSC maintenance. In contrast, dlp is a hyper-competitive mutation: dlp mutant FSC progenitors often eventually occupy the entire epithelial sheet. RNAi knockdown experiments showed that Dally and Dlp play both partially redundant and distinct roles in regulating Jak/Stat, Wg and Hh signaling in FSCs. The Drosophila FSC system offers a powerful genetic model to study the mechanisms by which HSPGs exert specific functions in stem cell replacement and competition. Copyright © 2018, Genetics.

  6. JAK/Stat signaling regulates heart precursor diversification in Drosophila

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    Johnson, Aaron N.; Mokalled, Mayssa H.; Haden, Tom N.; Olson, Eric N.

    2011-01-01

    Intercellular signal transduction pathways regulate the NK-2 family of transcription factors in a conserved gene regulatory network that directs cardiogenesis in both flies and mammals. The Drosophila NK-2 protein Tinman (Tin) was recently shown to regulate Stat92E, the Janus kinase (JAK) and Signal transducer and activator of transcription (Stat) pathway effector, in the developing mesoderm. To understand whether the JAK/Stat pathway also regulates cardiogenesis, we performed a systematic characterization of JAK/Stat signaling during mesoderm development. Drosophila embryos with mutations in the JAK/Stat ligand upd or in Stat92E have non-functional hearts with luminal defects and inappropriate cell aggregations. Using strong Stat92E loss-of-function alleles, we show that the JAK/Stat pathway regulates tin expression prior to heart precursor cell diversification. tin expression can be subdivided into four phases and, in Stat92E mutant embryos, the broad phase 2 expression pattern in the dorsal mesoderm does not restrict to the constrained phase 3 pattern. These embryos also have an expanded pericardial cell domain. We show the E(spl)-C gene HLHm5 is expressed in a pattern complementary to tin during phase 3 and that this expression is JAK/Stat dependent. In addition, E(spl)-C mutant embryos phenocopy the cardiac defects of Stat92E embryos. Mechanistically, JAK/Stat signals activate E(spl)-C genes to restrict Tin expression and the subsequent expression of the T-box transcription factor H15 to direct heart precursor diversification. This study is the first to characterize a role for the JAK/Stat pathway during cardiogenesis and identifies an autoregulatory circuit in which tin limits its own expression domain. PMID:21965617

  7. The NSL Complex Regulates Housekeeping Genes in Drosophila

    Science.gov (United States)

    Raja, Sunil Jayaramaiah; Holz, Herbert; Luscombe, Nicholas M.; Manke, Thomas; Akhtar, Asifa

    2012-01-01

    MOF is the major histone H4 lysine 16-specific (H4K16) acetyltransferase in mammals and Drosophila. In flies, it is involved in the regulation of X-chromosomal and autosomal genes as part of the MSL and the NSL complexes, respectively. While the function of the MSL complex as a dosage compensation regulator is fairly well understood, the role of the NSL complex in gene regulation is still poorly characterized. Here we report a comprehensive ChIP–seq analysis of four NSL complex members (NSL1, NSL3, MBD-R2, and MCRS2) throughout the Drosophila melanogaster genome. Strikingly, the majority (85.5%) of NSL-bound genes are constitutively expressed across different cell types. We find that an increased abundance of the histone modifications H4K16ac, H3K4me2, H3K4me3, and H3K9ac in gene promoter regions is characteristic of NSL-targeted genes. Furthermore, we show that these genes have a well-defined nucleosome free region and broad transcription initiation patterns. Finally, by performing ChIP–seq analyses of RNA polymerase II (Pol II) in NSL1- and NSL3-depleted cells, we demonstrate that both NSL proteins are required for efficient recruitment of Pol II to NSL target gene promoters. The observed Pol II reduction coincides with compromised binding of TBP and TFIIB to target promoters, indicating that the NSL complex is required for optimal recruitment of the pre-initiation complex on target genes. Moreover, genes that undergo the most dramatic loss of Pol II upon NSL knockdowns tend to be enriched in DNA Replication–related Element (DRE). Taken together, our findings show that the MOF-containing NSL complex acts as a major regulator of housekeeping genes in flies by modulating initiation of Pol II transcription. PMID:22723752

  8. Functions of the Drosophila JAK-STAT pathway

    Science.gov (United States)

    Amoyel, Marc; Bach, Erika A.

    2012-01-01

    JAK-STAT signaling has been proposed to act in numerous stem cells in a variety of organisms. Here we provide an overview of its roles in three well characterized stem cell populations in Drosophila, in the intestine, lymph gland and testis. In flies, there is a single JAK and a single STAT, which has made the genetic dissection of pathway function considerably easier and facilitated the analysis of communication between stem cells, their niches and offspring. Studies in flies have revealed roles for this pathway as diverse as regulating bona fide intrinsic self-renewal, integrating response to environmental cues that control quiescence and promoting mitogenic responses to stress. PMID:24058767

  9. RNAi-Mediated Reverse Genetic Screen Identified Drosophila Chaperones Regulating Eye and Neuromuscular Junction Morphology

    Directory of Open Access Journals (Sweden)

    Sandeep Raut

    2017-07-01

    Full Text Available Accumulation of toxic proteins in neurons has been linked with the onset of neurodegenerative diseases, which in many cases are characterized by altered neuronal function and synapse loss. Molecular chaperones help protein folding and the resolubilization of unfolded proteins, thereby reducing the protein aggregation stress. While most of the chaperones are expressed in neurons, their functional relevance remains largely unknown. Here, using bioinformatics analysis, we identified 95 Drosophila chaperones and classified them into seven different classes. Ubiquitous actin5C-Gal4-mediated RNAi knockdown revealed that ∼50% of the chaperones are essential in Drosophila. Knocking down these genes in eyes revealed that ∼30% of the essential chaperones are crucial for eye development. Using neuron-specific knockdown, immunocytochemistry, and robust behavioral assays, we identified a new set of chaperones that play critical roles in the regulation of Drosophila NMJ structural organization. Together, our data present the first classification and comprehensive analysis of Drosophila chaperones. Our screen identified a new set of chaperones that regulate eye and NMJ morphogenesis. The outcome of the screen reported here provides a useful resource for further elucidating the role of individual chaperones in Drosophila eye morphogenesis and synaptic development.

  10. Global identification of bursicon-regulated genes in Drosophila melanogaster

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    Beerntsen Brenda

    2008-09-01

    Full Text Available Abstract Background Bursicon is a heterodimer neuropeptide responsible for regulating cuticle sclerotization and wing expansion in several insect species. Recent studies indicate that the action of bursicon is mediated by a specific G protein-coupled receptor DLGR2 and the cAMP/PKA signaling pathway. However, little is known regarding the genes that are regulated by bursicon. The identification of bursicon-regulated genes is the focus of this investigation. Results We used DNA microarray analysis to identify bursicon-regulated genes in neck-ligated flies (Drosophila melanogaster that received recombinant bursicon (r-bursicon. Fifty four genes were found to be regulated by bursicon 1 h post r-bursicon injection, 52 being up-regulated and 2 down-regulated while 33 genes were influenced by r-bursicon 3 h post-injection (24 up-regulated and 9 down-regulated genes. Analysis of these genes by inference from the fly database http://flybase.bio.indiana.edu revealed that these genes encode proteins with diverse functions, including cell signaling, gene transcription, DNA/RNA binding, ion trafficking, proteolysis-peptidolysis, metabolism, cytoskeleton formation, immune response and cell-adhesion. Twenty eight genes randomly selected from the microarray-identified list were verified by real time PCR (qPCR which supported the microarray data. Temporal response studies of 13 identified and verified genes by qPCR revealed that the temporal expression patterns of these genes are consistent with the microarray data. Conclusion Using r-bursicon, we identified 87 genes that are regulated by bursicon, 30 of which have no previously known function. Most importantly, all genes randomly selected from the microarray-identified list were verified by real time PCR. Temporal analysis of 13 verified genes revealed that the expression of these genes was indeed induced by bursicon and correlated well with the cuticle sclerotization process. The composite data suggest that

  11. Mitochondrial apoptotic pathways induced by Drosophila programmed cell death regulators

    International Nuclear Information System (INIS)

    Claveria, Cristina; Torres, Miguel

    2003-01-01

    Multicellular organisms eliminate unwanted or damaged cells by cell death, a process essential to the maintenance of tissue homeostasis. Cell death is a tightly regulated event, whose alteration by excess or defect is involved in the pathogenesis of many diseases such as cancer, autoimmune syndromes, and neurodegenerative processes. Studies in model organisms, especially in the nematode Caenorhabditis elegans, have been crucial in identifying the key molecules implicated in the regulation and execution of programmed cell death. In contrast, the study of cell death in Drosophila melanogaster, often an excellent model organism, has identified regulators and mechanisms not obviously conserved in other metazoans. Recent molecular and cellular analyses suggest, however, that the mechanisms of action of the main programmed cell death regulators in Drosophila include a canonical mitochondrial pathway

  12. Functional analysis of PGRP-LA in Drosophila immunity.

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    Mathilde Gendrin

    Full Text Available PeptidoGlycan Recognition Proteins (PGRPs are key regulators of the insect innate antibacterial response. Even if they have been intensively studied, some of them have yet unknown functions. Here, we present a functional analysis of PGRP-LA, an as yet uncharacterized Drosophila PGRP. The PGRP-LA gene is located in cluster with PGRP-LC and PGRP-LF, which encode a receptor and a negative regulator of the Imd pathway, respectively. Structure predictions indicate that PGRP-LA would not bind to peptidoglycan, pointing to a regulatory role of this PGRP. PGRP-LA expression was enriched in barrier epithelia, but low in the fat body. Use of a newly generated PGRP-LA deficient mutant indicates that PGRP-LA is not required for the production of antimicrobial peptides by the fat body in response to a systemic infection. Focusing on the respiratory tract, where PGRP-LA is strongly expressed, we conducted a genome-wide microarray analysis of the tracheal immune response of wild-type, Relish, and PGRP-LA mutant larvae. Comparing our data to previous microarray studies, we report that a majority of genes regulated in the trachea upon infection differ from those induced in the gut or the fat body. Importantly, antimicrobial peptide gene expression was reduced in the tracheae of larvae and in the adult gut of PGRP-LA-deficient Drosophila upon oral bacterial infection. Together, our results suggest that PGRP-LA positively regulates the Imd pathway in barrier epithelia.

  13. Molecular mechanisms of aging and immune system regulation in Drosophila.

    Science.gov (United States)

    Eleftherianos, Ioannis; Castillo, Julio Cesar

    2012-01-01

    Aging is a complex process that involves the accumulation of deleterious changes resulting in overall decline in several vital functions, leading to the progressive deterioration in physiological condition of the organism and eventually causing disease and death. The immune system is the most important host-defense mechanism in humans and is also highly conserved in insects. Extensive research in vertebrates has concluded that aging of the immune function results in increased susceptibility to infectious disease and chronic inflammation. Over the years, interest has grown in studying the molecular interaction between aging and the immune response to pathogenic infections. The fruit fly Drosophila melanogaster is an excellent model system for dissecting the genetic and genomic basis of important biological processes, such as aging and the innate immune system, and deciphering parallel mechanisms in vertebrate animals. Here, we review the recent advances in the identification of key players modulating the relationship between molecular aging networks and immune signal transduction pathways in the fly. Understanding the details of the molecular events involved in aging and immune system regulation will potentially lead to the development of strategies for decreasing the impact of age-related diseases, thus improving human health and life span.

  14. Functional Analysis of Drosophila NF1

    National Research Council Canada - National Science Library

    Bernards, Andre

    2005-01-01

    ...) for Ras, yet homozygous loss of a highly conserved Drosophila NF1 ortholog results in several phenotypes that are insensitive to manipulating Ras signal transduction, but rescued by increasing...

  15. Transcriptional regulation of Drosophila gonad formation.

    Science.gov (United States)

    Tripathy, Ratna; Kunwar, Prabhat S; Sano, Hiroko; Renault, Andrew D

    2014-08-15

    The formation of the Drosophila embryonic gonad, involving the fusion of clusters of somatic gonadal precursor cells (SGPs) and their ensheathment of germ cells, provides a simple and genetically tractable model for the interplay between cells during organ formation. In a screen for mutants affecting gonad formation we identified a SGP cell autonomous role for Midline (Mid) and Longitudinals lacking (Lola). These transcriptional factors are required for multiple aspects of SGP behaviour including SGP cluster fusion, germ cell ensheathment and gonad compaction. The lola locus encodes more than 25 differentially spliced isoforms and we have identified an isoform specific requirement for lola in the gonad which is distinct from that in nervous system development. Mid and Lola work in parallel in gonad formation and surprisingly Mid overexpression in a lola background leads to additional SGPs at the expense of fat body cells. Our findings support the idea that although the transcription factors required by SGPs can ostensibly be assigned to those being required for either SGP specification or behaviour, they can also interact to impinge on both processes. Copyright © 2014 Elsevier Inc. All rights reserved.

  16. Identification of functional elements and regulatory circuits by Drosophila modENCODE.

    Science.gov (United States)

    Roy, Sushmita; Ernst, Jason; Kharchenko, Peter V; Kheradpour, Pouya; Negre, Nicolas; Eaton, Matthew L; Landolin, Jane M; Bristow, Christopher A; Ma, Lijia; Lin, Michael F; Washietl, Stefan; Arshinoff, Bradley I; Ay, Ferhat; Meyer, Patrick E; Robine, Nicolas; Washington, Nicole L; Di Stefano, Luisa; Berezikov, Eugene; Brown, Christopher D; Candeias, Rogerio; Carlson, Joseph W; Carr, Adrian; Jungreis, Irwin; Marbach, Daniel; Sealfon, Rachel; Tolstorukov, Michael Y; Will, Sebastian; Alekseyenko, Artyom A; Artieri, Carlo; Booth, Benjamin W; Brooks, Angela N; Dai, Qi; Davis, Carrie A; Duff, Michael O; Feng, Xin; Gorchakov, Andrey A; Gu, Tingting; Henikoff, Jorja G; Kapranov, Philipp; Li, Renhua; MacAlpine, Heather K; Malone, John; Minoda, Aki; Nordman, Jared; Okamura, Katsutomo; Perry, Marc; Powell, Sara K; Riddle, Nicole C; Sakai, Akiko; Samsonova, Anastasia; Sandler, Jeremy E; Schwartz, Yuri B; Sher, Noa; Spokony, Rebecca; Sturgill, David; van Baren, Marijke; Wan, Kenneth H; Yang, Li; Yu, Charles; Feingold, Elise; Good, Peter; Guyer, Mark; Lowdon, Rebecca; Ahmad, Kami; Andrews, Justen; Berger, Bonnie; Brenner, Steven E; Brent, Michael R; Cherbas, Lucy; Elgin, Sarah C R; Gingeras, Thomas R; Grossman, Robert; Hoskins, Roger A; Kaufman, Thomas C; Kent, William; Kuroda, Mitzi I; Orr-Weaver, Terry; Perrimon, Norbert; Pirrotta, Vincenzo; Posakony, James W; Ren, Bing; Russell, Steven; Cherbas, Peter; Graveley, Brenton R; Lewis, Suzanna; Micklem, Gos; Oliver, Brian; Park, Peter J; Celniker, Susan E; Henikoff, Steven; Karpen, Gary H; Lai, Eric C; MacAlpine, David M; Stein, Lincoln D; White, Kevin P; Kellis, Manolis

    2010-12-24

    To gain insight into how genomic information is translated into cellular and developmental programs, the Drosophila model organism Encyclopedia of DNA Elements (modENCODE) project is comprehensively mapping transcripts, histone modifications, chromosomal proteins, transcription factors, replication proteins and intermediates, and nucleosome properties across a developmental time course and in multiple cell lines. We have generated more than 700 data sets and discovered protein-coding, noncoding, RNA regulatory, replication, and chromatin elements, more than tripling the annotated portion of the Drosophila genome. Correlated activity patterns of these elements reveal a functional regulatory network, which predicts putative new functions for genes, reveals stage- and tissue-specific regulators, and enables gene-expression prediction. Our results provide a foundation for directed experimental and computational studies in Drosophila and related species and also a model for systematic data integration toward comprehensive genomic and functional annotation.

  17. Functional neuroanatomy of Drosophila olfactory memory formation.

    Science.gov (United States)

    Guven-Ozkan, Tugba; Davis, Ronald L

    2014-10-01

    New approaches, techniques and tools invented over the last decade and a half have revolutionized the functional dissection of neural circuitry underlying Drosophila learning. The new methodologies have been used aggressively by researchers attempting to answer three critical questions about olfactory memories formed with appetitive and aversive reinforcers: (1) Which neurons within the olfactory nervous system mediate the acquisition of memory? (2) What is the complete neural circuitry extending from the site(s) of acquisition to the site(s) controlling memory expression? (3) How is information processed across this circuit to consolidate early-forming, disruptable memories to stable, late memories? Much progress has been made and a few strong conclusions have emerged: (1) Acquisition occurs at multiple sites within the olfactory nervous system but is mediated predominantly by the γ mushroom body neurons. (2) The expression of long-term memory is completely dependent on the synaptic output of α/β mushroom body neurons. (3) Consolidation occurs, in part, through circuit interactions between mushroom body and dorsal paired medial neurons. Despite this progress, a complete and unified model that details the pathway from acquisition to memory expression remains elusive. © 2014 Guven-Ozkan and Davis; Published by Cold Spring Harbor Laboratory Press.

  18. The Drosophila DHR96 nuclear receptor binds cholesterol and regulates cholesterol homeostasis

    OpenAIRE

    Horner, Michael A.; Pardee, Keith; Liu, Suya; King-Jones, Kirst; Lajoie, Gilles; Edwards, Aled; Krause, Henry M.; Thummel, Carl S.

    2009-01-01

    Cholesterol homeostasis is required to maintain normal cellular function and avoid the deleterious effects of hypercholesterolemia. Here we show that the Drosophila DHR96 nuclear receptor binds cholesterol and is required for the coordinate transcriptional response of genes that are regulated by cholesterol and involved in cholesterol uptake, trafficking, and storage. DHR96 mutants die when grown on low levels of cholesterol and accumulate excess cholesterol when maintained on a high-choleste...

  19. Hox gene regulation in the central nervous system of Drosophila

    Directory of Open Access Journals (Sweden)

    Maheshwar eGummalla

    2014-04-01

    Full Text Available Hox genes specify the structures that form along the anteroposterior (AP axis of bilateria. Within the genome, they often form clusters where, remarkably enough, their position within the clusters reflects the relative positions of the structures they specify along the AP axis. This correspondence between genomic organization and gene expression pattern has been conserved through evolution and provides a unique opportunity to study how chromosomal context affects gene regulation. In Drosophila, a general rule, often called posterior dominance, states that Hox genes specifying more posterior structures repress the expression of more anterior Hox genes. This rule explains the apparent spatial complementarity of Hox gene expression patterns in Drosophila. Here we review a noticeable exception to this rule where the more-posteriorly expressed Abd-B hox gene fails to repress the more-anterior abd-A gene in cells of the central nervous system (CNS. While Abd-B is required to repress ectopic expression of abd-A in the posterior epidermis, abd-A repression in the posterior CNS is accomplished by a different mechanism that involves a large 92kb long non-coding RNA (lncRNA encoded by the intergenic region separating abd-A and Abd-B (the iab8ncRNA. Dissection of this lncRNA revealed that abd-A is repressed by the lncRNA using two redundant mechanisms. The 1st mechanism is mediated by a microRNA (mir-iab-8 encoded by intronic sequence within the large iab8-ncRNA. Meanwhile, the second mechanism seems to involve transcriptional interference by the long iab-8 ncRNA on the abd-A promoter. Recent work demonstrating CNS-specific regulation of genes by ncRNAs in Drosophila, seem to highlight a potential role for the iab-8-ncRNA in the evolution of the Drosophila hox complexes

  20. Dynamic regulation of Drosophila nuclear receptor activity in vivo.

    Science.gov (United States)

    Palanker, Laura; Necakov, Aleksandar S; Sampson, Heidi M; Ni, Ruoyu; Hu, Chun; Thummel, Carl S; Krause, Henry M

    2006-09-01

    Nuclear receptors are a large family of transcription factors that play major roles in development, metamorphosis, metabolism and disease. To determine how, where and when nuclear receptors are regulated by small chemical ligands and/or protein partners, we have used a 'ligand sensor' system to visualize spatial activity patterns for each of the 18 Drosophila nuclear receptors in live developing animals. Transgenic lines were established that express the ligand binding domain of each nuclear receptor fused to the DNA-binding domain of yeast GAL4. When combined with a GAL4-responsive reporter gene, the fusion proteins show tissue- and stage-specific patterns of activation. We show that these responses accurately reflect the presence of endogenous and exogenously added hormone, and that they can be modulated by nuclear receptor partner proteins. The amnioserosa, yolk, midgut and fat body, which play major roles in lipid storage, metabolism and developmental timing, were identified as frequent sites of nuclear receptor activity. We also see dynamic changes in activation that are indicative of sweeping changes in ligand and/or co-factor production. The screening of a small compound library using this system identified the angular psoralen angelicin and the insect growth regulator fenoxycarb as activators of the Ultraspiracle (USP) ligand-binding domain. These results demonstrate the utility of this system for the functional dissection of nuclear receptor pathways and for the development of new receptor agonists and antagonists that can be used to modulate metabolism and disease and to develop more effective means of insect control.

  1. Functional requirements driving the gene duplication in 12 Drosophila species.

    Science.gov (United States)

    Zhong, Yan; Jia, Yanxiao; Gao, Yang; Tian, Dacheng; Yang, Sihai; Zhang, Xiaohui

    2013-08-15

    Gene duplication supplies the raw materials for novel gene functions and many gene families arisen from duplication experience adaptive evolution. Most studies of young duplicates have focused on mammals, especially humans, whereas reports describing their genome-wide evolutionary patterns across the closely related Drosophila species are rare. The sequenced 12 Drosophila genomes provide the opportunity to address this issue. In our study, 3,647 young duplicate gene families were identified across the 12 Drosophila species and three types of expansions, species-specific, lineage-specific and complex expansions, were detected in these gene families. Our data showed that the species-specific young duplicate genes predominated (86.6%) over the other two types. Interestingly, many independent species-specific expansions in the same gene family have been observed in many species, even including 11 or 12 Drosophila species. Our data also showed that the functional bias observed in these young duplicate genes was mainly related to responses to environmental stimuli and biotic stresses. This study reveals the evolutionary patterns of young duplicates across 12 Drosophila species on a genomic scale. Our results suggest that convergent evolution acts on young duplicate genes after the species differentiation and adaptive evolution may play an important role in duplicate genes for adaption to ecological factors and environmental changes in Drosophila.

  2. Tissue-specific regulation of BMP signaling by Drosophila N-glycanase 1.

    Science.gov (United States)

    Galeone, Antonio; Han, Seung Yeop; Huang, Chengcheng; Hosomi, Akira; Suzuki, Tadashi; Jafar-Nejad, Hamed

    2017-08-04

    Mutations in the human N- glycanase 1 ( NGLY1 ) cause a rare, multisystem congenital disorder with global developmental delay. However, the mechanisms by which NGLY1 and its homologs regulate embryonic development are not known. Here we show that Drosophila Pngl encodes an N -glycanase and exhibits a high degree of functional conservation with human NGLY1. Loss of Pngl results in developmental midgut defects reminiscent of midgut-specific loss of BMP signaling. Pngl mutant larvae also exhibit a severe midgut clearance defect, which cannot be fully explained by impaired BMP signaling. Genetic experiments indicate that Pngl is primarily required in the mesoderm during Drosophila development. Loss of Pngl results in a severe decrease in the level of Dpp homodimers and abolishes BMP autoregulation in the visceral mesoderm mediated by Dpp and Tkv homodimers. Thus, our studies uncover a novel mechanism for the tissue-specific regulation of an evolutionarily conserved signaling pathway by an N -glycanase enzyme.

  3. Regulation of lifespan, metabolism, and stress responses by the Drosophila SH2B protein, Lnk.

    Directory of Open Access Journals (Sweden)

    Cathy Slack

    2010-03-01

    Full Text Available Drosophila Lnk is the single ancestral orthologue of a highly conserved family of structurally-related intracellular adaptor proteins, the SH2B proteins. As adaptors, they lack catalytic activity but contain several protein-protein interaction domains, thus playing a critical role in signal transduction from receptor tyrosine kinases to form protein networks. Physiological studies of SH2B function in mammals have produced conflicting data. However, a recent study in Drosophila has shown that Lnk is an important regulator of the insulin/insulin-like growth factor (IGF-1 signaling (IIS pathway during growth, functioning in parallel to the insulin receptor substrate, Chico. As this pathway also has an evolutionary conserved role in the determination of organism lifespan, we investigated whether Lnk is required for normal lifespan in Drosophila. Phenotypic analysis of mutants for Lnk revealed that loss of Lnk function results in increased lifespan and improved survival under conditions of oxidative stress and starvation. Starvation resistance was found to be associated with increased metabolic stores of carbohydrates and lipids indicative of impaired metabolism. Biochemical and genetic data suggest that Lnk functions in both the IIS and Ras/Mitogen activated protein Kinase (MapK signaling pathways. Microarray studies support this model, showing transcriptional feedback onto genes in both pathways as well as indicating global changes in both lipid and carbohydrate metabolism. Finally, our data also suggest that Lnk itself may be a direct target of the IIS responsive transcription factor, dFoxo, and that dFoxo may repress Lnk expression. We therefore describe novel functions for a member of the SH2B protein family and provide the first evidence for potential mechanisms of SH2B regulation. Our findings suggest that IIS signaling in Drosophila may require the activity of a second intracellular adaptor, thereby yielding fundamental new insights into the

  4. dRYBP contributes to the negative regulation of the Drosophila Imd pathway.

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    Ricardo Aparicio

    Full Text Available The Drosophila humoral innate immune response fights infection by producing antimicrobial peptides (AMPs through the microbe-specific activation of the Toll or the Imd signaling pathway. Upon systemic infection, the production of AMPs is both positively and negatively regulated to reach a balanced immune response required for survival. Here, we report the function of the dRYBP (drosophila Ring and YY1 Binding Protein protein, which contains a ubiquitin-binding domain, in the Imd pathway. We have found that dRYBP contributes to the negative regulation of AMP production: upon systemic infection with Gram-negative bacteria, Diptericin expression is up-regulated in the absence of dRYBP and down-regulated in the presence of high levels of dRYBP. Epistatic analyses using gain and loss of function alleles of imd, Relish, or skpA and dRYBP suggest that dRYBP functions upstream or together with SKPA, a member of the SCF-E3-ubiquitin ligase complex, to repress the Imd signaling cascade. We propose that the role of dRYBP in the regulation of the Imd signaling pathway is to function as a ubiquitin adaptor protein together with SKPA to promote SCF-dependent proteasomal degradation of Relish. Beyond the identification of dRYBP as a novel component of Imd pathway regulation, our results also suggest that the evolutionarily conserved RYBP protein may be involved in the human innate immune response.

  5. Identification of chromatin-associated regulators of MSL complex targeting in Drosophila dosage compensation.

    Directory of Open Access Journals (Sweden)

    Erica Larschan

    Full Text Available Sex chromosome dosage compensation in Drosophila provides a model for understanding how chromatin organization can modulate coordinate gene regulation. Male Drosophila increase the transcript levels of genes on the single male X approximately two-fold to equal the gene expression in females, which have two X-chromosomes. Dosage compensation is mediated by the Male-Specific Lethal (MSL histone acetyltransferase complex. Five core components of the MSL complex were identified by genetic screens for genes that are specifically required for male viability and are dispensable for females. However, because dosage compensation must interface with the general transcriptional machinery, it is likely that identifying additional regulators that are not strictly male-specific will be key to understanding the process at a mechanistic level. Such regulators would not have been recovered from previous male-specific lethal screening strategies. Therefore, we have performed a cell culture-based, genome-wide RNAi screen to search for factors required for MSL targeting or function. Here we focus on the discovery of proteins that function to promote MSL complex recruitment to "chromatin entry sites," which are proposed to be the initial sites of MSL targeting. We find that components of the NSL (Non-specific lethal complex, and a previously unstudied zinc-finger protein, facilitate MSL targeting and display a striking enrichment at MSL entry sites. Identification of these factors provides new insight into how MSL complex establishes the specialized hyperactive chromatin required for dosage compensation in Drosophila.

  6. Dietary glucose regulates yeast consumption in adult Drosophila males

    Directory of Open Access Journals (Sweden)

    Sebastien eLebreton

    2014-12-01

    Full Text Available The adjustment of feeding behavior in response to hunger and satiety contributes to homeostatic regulation in animals. The fruit fly Drosophila melanogaster feeds on yeasts growing on overripe fruit, providing nutrients required for adult survival, reproduction and larval growth. Here, we present data on how the nutritional value of food affects subsequent yeast consumption in Drosophila adult males. After a period of starvation, flies showed intensive yeast consumption. In comparison, flies stopped feeding after having access to a nutritive cornmeal diet. Interestingly, dietary glucose was equally efficient as the complex cornmeal diet. In contrast, flies fed with sucralose, a non-metabolizable sweetener, behaved as if they were starved. The adipokinetic hormone and insulin-like peptides regulate metabolic processes in insects. We did not find any effect of the adipokinetic hormone pathway on this modulation. Instead, the insulin pathway was involved in these changes. Flies lacking the insulin receptor did not respond to nutrient deprivation by increasing yeast consumption. Together these results show the importance of insulin in the regulation of yeast consumption in response to starvation in adult D. melanogaster males.

  7. Dietary glucose regulates yeast consumption in adult Drosophila males.

    Science.gov (United States)

    Lebreton, Sébastien; Witzgall, Peter; Olsson, Marie; Becher, Paul G

    2014-01-01

    The adjustment of feeding behavior in response to hunger and satiety contributes to homeostatic regulation in animals. The fruit fly Drosophila melanogaster feeds on yeasts growing on overripe fruit, providing nutrients required for adult survival, reproduction and larval growth. Here, we present data on how the nutritional value of food affects subsequent yeast consumption in Drosophila adult males. After a period of starvation, flies showed intensive yeast consumption. In comparison, flies stopped feeding after having access to a nutritive cornmeal diet. Interestingly, dietary glucose was equally efficient as the complex cornmeal diet. In contrast, flies fed with sucralose, a non-metabolizable sweetener, behaved as if they were starved. The adipokinetic hormone and insulin-like peptides regulate metabolic processes in insects. We did not find any effect of the adipokinetic hormone pathway on this modulation. Instead, the insulin pathway was involved in these changes. Flies lacking the insulin receptor (InR) did not respond to nutrient deprivation by increasing yeast consumption. Together these results show the importance of insulin in the regulation of yeast consumption in response to starvation in adult D. melanogaster males.

  8. Mushroom bodies regulate habit formation in Drosophila.

    Science.gov (United States)

    Brembs, Björn

    2009-08-25

    To make good decisions, we evaluate past choices to guide later decisions. In most situations, we have the opportunity to simultaneously learn about both the consequences of our choice (i.e., operantly) and the stimuli associated with correct or incorrect choices (i.e., classically). Interestingly, in many species, including humans, these learning processes occasionally lead to irrational decisions. An extreme case is the habitual drug user consistently administering the drug despite the negative consequences, but we all have experience with our own, less severe habits. The standard animal model employs a combination of operant and classical learning components to bring about habit formation in rodents. After extended training, these animals will press a lever even if the outcome associated with lever-pressing is no longer desired. In this study, experiments with wild-type and transgenic flies revealed that a prominent insect neuropil, the mushroom bodies (MBs), regulates habit formation in flies by inhibiting the operant learning system when a predictive stimulus is present. This inhibition enables generalization of the classical memory and prevents premature habit formation. Extended training in wild-type flies produced a phenocopy of MB-impaired flies, such that generalization was abolished and goal-directed actions were transformed into habitual responses.

  9. Fan-Shaped Body Neurons Are Involved in "Period"-Dependent Regulation of Long-Term Courtship Memory in "Drosophila"

    Science.gov (United States)

    Sakai, Takaomi; Inami, Show; Sato, Shoma; Kitamoto, Toshihiro

    2012-01-01

    In addition to its established function in the regulation of circadian rhythms, the "Drosophila" gene "period" ("per") also plays an important role in processing long-term memory (LTM). Here, we used courtship conditioning as a learning paradigm and revealed that (1) overexpression and knocking down of "per" in subsets of brain neurons enhance and…

  10. The Drosophila PNG kinase complex regulates the translation of cyclin B.

    Science.gov (United States)

    Vardy, Leah; Orr-Weaver, Terry L

    2007-01-01

    The Drosophila PAN GU (PNG) kinase complex regulates the developmental translation of cyclin B. cyclin B mRNA becomes unmasked during oogenesis independent of PNG activity, but PNG is required for translation from egg activation. We find that although polyadenylation of cyclin B augments translation, it is not essential, and a fully elongated poly(A) is not required for translation to proceed. In fact, changes in poly(A) tail length are not sufficient to account for PNG-mediated control of cyclin B translation and of the early embryonic cell cycles. We present evidence that PNG functions instead as an antagonist of PUMILIO-dependent translational repression. Our data argue that changes in poly(A) tail length are not a universal mechanism governing embryonic cell cycles, and that PNG-mediated derepression of translation is an important alternative mechanism in Drosophila.

  11. Developmental regulation of nucleolus size during Drosophila eye differentiation.

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    Nicholas E Baker

    Full Text Available When cell cycle withdrawal accompanies terminal differentiation, biosynthesis and cellular growth are likely to change also. In this study, nucleolus size was monitored during cell fate specification in the Drosophila eye imaginal disc using fibrillarin antibody labeling. Nucleolus size is an indicator of ribosome biogenesis and can correlate with cellular growth rate. Nucleolar size was reduced significantly during cell fate specification and differentiation, predominantly as eye disc cells entered a cell cycle arrest that preceded cell fate specification. This reduction in nucleolus size required Dpp and Hh signaling. A transient enlargement of the nucleolus accompanied cell division in the Second Mitotic Wave. Nucleoli continued to diminish in postmitotic cells following fate specification. These results suggest that cellular growth is regulated early in the transition from proliferating progenitor cells to terminal cell fate specification, contemporary with regulation of the cell cycle, and requiring the same extracellular signals.

  12. Developmental regulation of nucleolus size during Drosophila eye differentiation.

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    Baker, Nicholas E

    2013-01-01

    When cell cycle withdrawal accompanies terminal differentiation, biosynthesis and cellular growth are likely to change also. In this study, nucleolus size was monitored during cell fate specification in the Drosophila eye imaginal disc using fibrillarin antibody labeling. Nucleolus size is an indicator of ribosome biogenesis and can correlate with cellular growth rate. Nucleolar size was reduced significantly during cell fate specification and differentiation, predominantly as eye disc cells entered a cell cycle arrest that preceded cell fate specification. This reduction in nucleolus size required Dpp and Hh signaling. A transient enlargement of the nucleolus accompanied cell division in the Second Mitotic Wave. Nucleoli continued to diminish in postmitotic cells following fate specification. These results suggest that cellular growth is regulated early in the transition from proliferating progenitor cells to terminal cell fate specification, contemporary with regulation of the cell cycle, and requiring the same extracellular signals.

  13. Resources for Functional Genomics Studies in Drosophila melanogaster

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    Mohr, Stephanie E.; Hu, Yanhui; Kim, Kevin; Housden, Benjamin E.; Perrimon, Norbert

    2014-01-01

    Drosophila melanogaster has become a system of choice for functional genomic studies. Many resources, including online databases and software tools, are now available to support design or identification of relevant fly stocks and reagents or analysis and mining of existing functional genomic, transcriptomic, proteomic, etc. datasets. These include large community collections of fly stocks and plasmid clones, “meta” information sites like FlyBase and FlyMine, and an increasing number of more specialized reagents, databases, and online tools. Here, we introduce key resources useful to plan large-scale functional genomics studies in Drosophila and to analyze, integrate, and mine the results of those studies in ways that facilitate identification of highest-confidence results and generation of new hypotheses. We also discuss ways in which existing resources can be used and might be improved and suggest a few areas of future development that would further support large- and small-scale studies in Drosophila and facilitate use of Drosophila information by the research community more generally. PMID:24653003

  14. Functional variation in the gut microbiome of wild Drosophila populations.

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    Bost, Alyssa; Martinson, Vincent G; Franzenburg, Soeren; Adair, Karen L; Albasi, Alice; Wells, Martin T; Douglas, Angela E

    2018-05-26

    Most of the evidence that the gut microbiome of animals is functionally variable, with consequences for the health and fitness of the animal host, is based on laboratory studies, often using inbred animals under tightly controlled conditions. It is largely unknown whether these microbiome effects would be evident in outbred animal populations under natural conditions. In this study, we quantified the functional traits of the gut microbiota (metagenome) and host (gut transcriptome) and the taxonomic composition of the gut microorganisms (16S rRNA gene sequence) in natural populations of three mycophagous Drosophila species. Variation in microbiome function and composition was driven principally by the period of sample collection, while host function varied mostly with Drosophila species, indicating that variation in microbiome traits is determined largely by environmental factors, and not host taxonomy. Despite this, significant correlations between microbiome and host functional traits were obtained. In particular, microbiome functions dominated by metabolism were positively associated with host functions relating to gut epithelial turnover. Much of the functional variation in the microbiome could be attributed to variation in abundance of Bacteroidetes, rather than the two other abundant groups, the γ-Proteobacteria or Lactobacillales. We conclude that functional variation in the interactions between animals and their gut microbiome can be detectable in natural populations and, in mycophagous Drosophila, this variation relates primarily to metabolism and homeostasis of the gut epithelium. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

  15. Regulation of sleep by neuropeptide Y-like system in Drosophila melanogaster.

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    Chunxia He

    Full Text Available Sleep is important for maintenance of normal physiology in animals. In mammals, neuropeptide Y (NPY, a homolog of Drosophila neuropeptide F (NPF, is involved in sleep regulation, with different effects in human and rat. However, the function of NPF on sleep in Drosophila melanogaster has not yet been described. In this study, we investigated the effects of NPF and its receptor-neuropeptide F receptor (NPFR1 on Drosophila sleep. Male flies over-expressing NPF or NPFR1 exhibited increased sleep during the nighttime. Further analysis demonstrated that sleep episode duration during nighttime was greatly increased and sleep latency was significantly reduced, indicating that NPF and NPFR1 promote sleep quality, and their action on sleep is not because of an impact of the NPF signal system on development. Moreover, the homeostatic regulation of flies after sleep deprivation was disrupted by altered NPF signaling, since sleep deprivation decreased transcription of NPF in control flies, and there were less sleep loss during sleep deprivation and less sleep gain after sleep deprivation in flies overexpressing NPF and NPFR1 than in control flies, suggesting that NPF system auto-regulation plays an important role in sleep homeostasis. However, these effects did not occur in females, suggesting a sex-dependent regulatory function in sleep for NPF and NPFR1. NPF in D1 brain neurons showed male-specific expression, providing the cellular locus for male-specific regulation of sleep by NPF and NPFR1. This study brings a new understanding into sleep studies of a sexually dimorphic regulatory mode in female and male flies.

  16. Specification of Drosophila corpora cardiaca neuroendocrine cells from mesoderm is regulated by Notch signaling.

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    Sangbin Park

    2011-08-01

    Full Text Available Drosophila neuroendocrine cells comprising the corpora cardiaca (CC are essential for systemic glucose regulation and represent functional orthologues of vertebrate pancreatic α-cells. Although Drosophila CC cells have been regarded as developmental orthologues of pituitary gland, the genetic regulation of CC development is poorly understood. From a genetic screen, we identified multiple novel regulators of CC development, including Notch signaling factors. Our studies demonstrate that the disruption of Notch signaling can lead to the expansion of CC cells. Live imaging demonstrates localized emergence of extra precursor cells as the basis of CC expansion in Notch mutants. Contrary to a recent report, we unexpectedly found that CC cells originate from head mesoderm. We show that Tinman expression in head mesoderm is regulated by Notch signaling and that the combination of Daughterless and Tinman is sufficient for ectopic CC specification in mesoderm. Understanding the cellular, genetic, signaling, and transcriptional basis of CC cell specification and expansion should accelerate discovery of molecular mechanisms regulating ontogeny of organs that control metabolism.

  17. Negative regulation of MAP kinase signaling in Drosophila by Ptp61F/PTP1B.

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    Tchankouo-Nguetcheu, Stéphane; Udinotti, Mario; Durand, Marjorie; Meng, Tzu-Ching; Taouis, Mohammed; Rabinow, Leonard

    2014-10-01

    PTP1B is an important negative regulator of insulin and other signaling pathways in mammals. However, the role of PTP1B in the regulation of RAS-MAPK signaling remains open to deliberation, due to conflicting evidence from different experimental systems. The Drosophila orthologue of mammalian PTP1B, PTP61F, has until recently remained largely uncharacterized. To establish the potential role of PTP61F in the regulation of signaling pathways in Drosophila and particularly to help resolve its fundamental function in RAS-MAPK signaling, we generated a new allele of Ptp61F as well as employed both RNA interference and overexpression alleles. Our results validate recent data showing that the activity of insulin and Abl kinase signaling is increased in Ptp61F mutants and RNA interference lines. Importantly, we establish negative regulation of the RAS/MAPK pathway by Ptp61F activity in whole animals. Of particular interest, our results document the modulation of hyperactive MAP kinase activity by Ptp61F alleles, showing that the phosphatase intervenes to directly or indirectly regulate MAP kinase itself.

  18. Functional Gustatory Role of Chemoreceptors in Drosophila Wings.

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    Raad, Hussein; Ferveur, Jean-François; Ledger, Neil; Capovilla, Maria; Robichon, Alain

    2016-05-17

    Neuroanatomical evidence argues for the presence of taste sensilla in Drosophila wings; however, the taste physiology of insect wings remains hypothetical, and a comprehensive link to mechanical functions, such as flight, wing flapping, and grooming, is lacking. Our data show that the sensilla of the Drosophila anterior wing margin respond to both sweet and bitter molecules through an increase in cytosolic Ca(2+) levels. Conversely, genetically modified flies presenting a wing-specific reduction in chemosensory cells show severe defects in both wing taste signaling and the exploratory guidance associated with chemodetection. In Drosophila, the chemodetection machinery includes mechanical grooming, which facilitates the contact between tastants and wing chemoreceptors, and the vibrations of flapping wings that nebulize volatile molecules as carboxylic acids. Together, these data demonstrate that the Drosophila wing chemosensory sensilla are a functional taste organ and that they may have a role in the exploration of ecological niches. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

  19. Novel Functional Properties of Drosophila CNS Glutamate Receptors

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    Li, Yan; Dharkar, Poorva; Han, Tae-Hee; Serpe, Mihaela; Lee, Chi-Hon; Mayer, Mark L.

    2016-12-01

    Phylogenetic analysis reveals AMPA, kainate, and NMDA receptor families in insect genomes, suggesting conserved functional properties corresponding to their vertebrate counterparts. However, heterologous expression of the Drosophila kainate receptor DKaiR1D and the AMPA receptor DGluR1A revealed novel ligand selectivity at odds with the classification used for vertebrate glutamate receptor ion channels (iGluRs). DKaiR1D forms a rapidly activating and desensitizing receptor that is inhibited by both NMDA and the NMDA receptor antagonist AP5; crystallization of the KaiR1D ligand-binding domain reveals that these ligands stabilize open cleft conformations, explaining their action as antagonists. Surprisingly, the AMPA receptor DGluR1A shows weak activation by its namesake agonist AMPA and also by quisqualate. Crystallization of the DGluR1A ligand-binding domain reveals amino acid exchanges that interfere with binding of these ligands. The unexpected ligand-binding profiles of insect iGluRs allows classical tools to be used in novel approaches for the study of synaptic regulation.

  20. dyschronic, a Drosophila homolog of a deaf-blindness gene, regulates circadian output and Slowpoke channels.

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    James E C Jepson

    Full Text Available Many aspects of behavior and physiology are under circadian control. In Drosophila, the molecular clock that regulates rhythmic patterns of behavior has been extensively characterized. In contrast, genetic loci involved in linking the clock to alterations in motor activity have remained elusive. In a forward-genetic screen, we uncovered a new component of the circadian output pathway, which we have termed dyschronic (dysc. dysc mutants exhibit arrhythmic locomotor behavior, yet their eclosion rhythms are normal and clock protein cycling remains intact. Intriguingly, dysc is the closest Drosophila homolog of whirlin, a gene linked to type II Usher syndrome, the leading cause of deaf-blindness in humans. Whirlin and other Usher proteins are expressed in the mammalian central nervous system, yet their function in the CNS has not been investigated. We show that DYSC is expressed in major neuronal tracts and regulates expression of the calcium-activated potassium channel SLOWPOKE (SLO, an ion channel also required in the circadian output pathway. SLO and DYSC are co-localized in the brain and control each other's expression post-transcriptionally. Co-immunoprecipitation experiments demonstrate they form a complex, suggesting they regulate each other through protein-protein interaction. Furthermore, electrophysiological recordings of neurons in the adult brain show that SLO-dependent currents are greatly reduced in dysc mutants. Our work identifies a Drosophila homolog of a deaf-blindness gene as a new component of the circadian output pathway and an important regulator of ion channel expression, and suggests novel roles for Usher proteins in the mammalian nervous system.

  1. The PP2C Alphabet is a negative regulator of stress-activated protein kinase signaling in Drosophila.

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    Baril, Caroline; Sahmi, Malha; Ashton-Beaucage, Dariel; Stronach, Beth; Therrien, Marc

    2009-02-01

    The Jun N-terminal kinase and p38 pathways, also known as stress-activated protein kinase (SAPK) pathways, are signaling conduits reiteratively used throughout the development and adult life of metazoans where they play central roles in the control of apoptosis, immune function, and environmental stress responses. We recently identified a Drosophila Ser/Thr phosphatase of the PP2C family, named Alphabet (Alph), which acts as a negative regulator of the Ras/ERK pathway. Here we show that Alph also plays an inhibitory role with respect to Drosophila SAPK signaling during development as well as under stress conditions such as oxidative or genotoxic stresses. Epistasis experiments suggest that Alph acts at a step upstream of the MAPKKs Hep and Lic. Consistent with this interpretation, biochemical experiments identify the upstream MAPKKKs Slpr, Tak1, and Wnd as putative substrates. Together with previous findings, this work identifies Alph as a general attenuator of MAPK signaling in Drosophila.

  2. Regulation of Drosophila Brain Wiring by Neuropil Interactions via a Slit-Robo-RPTP Signaling Complex.

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    Oliva, Carlos; Soldano, Alessia; Mora, Natalia; De Geest, Natalie; Claeys, Annelies; Erfurth, Maria-Luise; Sierralta, Jimena; Ramaekers, Ariane; Dascenco, Dan; Ejsmont, Radoslaw K; Schmucker, Dietmar; Sanchez-Soriano, Natalia; Hassan, Bassem A

    2016-10-24

    The axonal wiring molecule Slit and its Round-About (Robo) receptors are conserved regulators of nerve cord patterning. Robo receptors also contribute to wiring brain circuits. Whether molecular mechanisms regulating these signals are modified to fit more complex brain wiring processes is unclear. We investigated the role of Slit and Robo receptors in wiring Drosophila higher-order brain circuits and identified differences in the cellular and molecular mechanisms of Robo/Slit function. First, we find that signaling by Robo receptors in the brain is regulated by the Receptor Protein Tyrosine Phosphatase RPTP69d. RPTP69d increases membrane availability of Robo3 without affecting its phosphorylation state. Second, we detect no midline localization of Slit during brain development. Instead, Slit is enriched in the mushroom body, a neuronal structure covering large areas of the brain. Thus, a divergent molecular mechanism regulates neuronal circuit wiring in the Drosophila brain, partly in response to signals from the mushroom body. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  3. Genome-wide RNAi Screen Identifies Networks Involved in Intestinal Stem Cell Regulation in Drosophila

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    Xiankun Zeng

    2015-02-01

    Full Text Available The intestinal epithelium is the most rapidly self-renewing tissue in adult animals and maintained by intestinal stem cells (ISCs in both Drosophila and mammals. To comprehensively identify genes and pathways that regulate ISC fates, we performed a genome-wide transgenic RNAi screen in adult Drosophila intestine and identified 405 genes that regulate ISC maintenance and lineage-specific differentiation. By integrating these genes into publicly available interaction databases, we further developed functional networks that regulate ISC self-renewal, ISC proliferation, ISC maintenance of diploid status, ISC survival, ISC-to-enterocyte (EC lineage differentiation, and ISC-to-enteroendocrine (EE lineage differentiation. By comparing regulators among ISCs, female germline stem cells, and neural stem cells, we found that factors related to basic stem cell cellular processes are commonly required in all stem cells, and stem-cell-specific, niche-related signals are required only in the unique stem cell type. Our findings provide valuable insights into stem cell maintenance and lineage-specific differentiation.

  4. Negative regulation of EGFR/MAPK pathway by Pumilio in Drosophila melanogaster.

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    Sung Yun Kim

    Full Text Available In Drosophila melanogaster, specification of wing vein cells and sensory organ precursor (SOP cells, which later give rise to a bristle, requires EGFR signaling. Here, we show that Pumilio (Pum, an RNA-binding translational repressor, negatively regulates EGFR signaling in wing vein and bristle development. We observed that loss of Pum function yielded extra wing veins and additional bristles. Conversely, overexpression of Pum eliminated wing veins and bristles. Heterozygotes for Pum produced no phenotype on their own, but greatly enhanced phenotypes caused by the enhancement of EGFR signaling. Conversely, over-expression of Pum suppressed the effects of ectopic EGFR signaling. Components of the EGFR signaling pathway are encoded by mRNAs that have Nanos Response Element (NRE-like sequences in their 3'UTRs; NREs are known to bind Pum to confer regulation in other mRNAs. We show that these NRE-like sequences bind Pum and confer repression on a luciferase reporter in heterologous cells. Taken together, our evidence suggests that Pum functions as a negative regulator of EGFR signaling by directly targeting components of the pathway in Drosophila.

  5. A Genetic Toolkit for Dissecting Dopamine Circuit Function in Drosophila

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    Tingting Xie

    2018-04-01

    Full Text Available Summary: The neuromodulator dopamine (DA plays a key role in motor control, motivated behaviors, and higher-order cognitive processes. Dissecting how these DA neural networks tune the activity of local neural circuits to regulate behavior requires tools for manipulating small groups of DA neurons. To address this need, we assembled a genetic toolkit that allows for an exquisite level of control over the DA neural network in Drosophila. To further refine targeting of specific DA neurons, we also created reagents that allow for the conversion of any existing GAL4 line into Split GAL4 or GAL80 lines. We demonstrated how this toolkit can be used with recently developed computational methods to rapidly generate additional reagents for manipulating small subsets or individual DA neurons. Finally, we used the toolkit to reveal a dynamic interaction between a small subset of DA neurons and rearing conditions in a social space behavioral assay. : The rapid analysis of how dopaminergic circuits regulate behavior is limited by the genetic tools available to target and manipulate small numbers of these neurons. Xie et al. present genetic tools in Drosophila that allow rational targeting of sparse dopaminergic neuronal subsets and selective knockdown of dopamine signaling. Keywords: dopamine, genetics, behavior, neural circuits, neuromodulation, Drosophila

  6. Functions of the nonsense-mediated mRNA decay pathway in Drosophila development.

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    Mark M Metzstein

    2006-12-01

    Full Text Available Nonsense-mediated mRNA decay (NMD is a cellular surveillance mechanism that degrades transcripts containing premature translation termination codons, and it also influences expression of certain wild-type transcripts. Although the biochemical mechanisms of NMD have been studied intensively, its developmental functions and importance are less clear. Here, we describe the isolation and characterization of Drosophila "photoshop" mutations, which increase expression of green fluorescent protein and other transgenes. Mapping and molecular analyses show that photoshop mutations are loss-of-function mutations in the Drosophila homologs of NMD genes Upf1, Upf2, and Smg1. We find that Upf1 and Upf2 are broadly active during development, and they are required for NMD as well as for proper expression of dozens of wild-type genes during development and for larval viability. Genetic mosaic analysis shows that Upf1 and Upf2 are required for growth and/or survival of imaginal cell clones, but this defect can be overcome if surrounding wild-type cells are eliminated. By contrast, we find that the PI3K-related kinase Smg1 potentiates but is not required for NMD or for viability, implying that the Upf1 phosphorylation cycle that is required for mammalian and Caenorhabditis elegans NMD has a more limited role during Drosophila development. Finally, we show that the SV40 3' UTR, present in many Drosophila transgenes, targets the transgenes for regulation by the NMD pathway. The results establish that the Drosophila NMD pathway is broadly active and essential for development, and one critical function of the pathway is to endow proliferating imaginal cells with a competitive growth advantage that prevents them from being overtaken by other proliferating cells.

  7. Anatomy and behavioral function of serotonin receptors in Drosophila melanogaster larvae.

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    Annina Huser

    Full Text Available The biogenic amine serotonin (5-HT is an important neuroactive molecule in the central nervous system of the majority of animal phyla. 5-HT binds to specific G protein-coupled and ligand-gated ion receptors to regulate particular aspects of animal behavior. In Drosophila, as in many other insects this includes the regulation of locomotion and feeding. Due to its genetic amenability and neuronal simplicity the Drosophila larva has turned into a useful model for studying the anatomical and molecular basis of chemosensory behaviors. This is particularly true for the olfactory system, which is mostly described down to the synaptic level over the first three orders of neuronal information processing. Here we focus on the 5-HT receptor system of the Drosophila larva. In a bipartite approach consisting of anatomical and behavioral experiments we describe the distribution and the implications of individual 5-HT receptors on naïve and acquired chemosensory behaviors. Our data suggest that 5-HT1A, 5-HT1B, and 5-HT7 are dispensable for larval naïve olfactory and gustatory choice behaviors as well as for appetitive and aversive associative olfactory learning and memory. In contrast, we show that 5-HT/5-HT2A signaling throughout development, but not as an acute neuronal function, affects associative olfactory learning and memory using high salt concentration as a negative unconditioned stimulus. These findings describe for the first time an involvement of 5-HT signaling in learning and memory in Drosophila larvae. In the longer run these results may uncover developmental, 5-HT dependent principles related to reinforcement processing possibly shared with adult Drosophila and other insects.

  8. Drosophila pink1 is required for mitochondrial function and interacts genetically with parkin.

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    Clark, Ira E; Dodson, Mark W; Jiang, Changan; Cao, Joseph H; Huh, Jun R; Seol, Jae Hong; Yoo, Soon Ji; Hay, Bruce A; Guo, Ming

    2006-06-29

    Parkinson's disease is the second most common neurodegenerative disorder and is characterized by the degeneration of dopaminergic neurons in the substantia nigra. Mitochondrial dysfunction has been implicated as an important trigger for Parkinson's disease-like pathogenesis because exposure to environmental mitochondrial toxins leads to Parkinson's disease-like pathology. Recently, multiple genes mediating familial forms of Parkinson's disease have been identified, including PTEN-induced kinase 1 (PINK1; PARK6) and parkin (PARK2), which are also associated with sporadic forms of Parkinson's disease. PINK1 encodes a putative serine/threonine kinase with a mitochondrial targeting sequence. So far, no in vivo studies have been reported for pink1 in any model system. Here we show that removal of Drosophila PINK1 homologue (CG4523; hereafter called pink1) function results in male sterility, apoptotic muscle degeneration, defects in mitochondrial morphology and increased sensitivity to multiple stresses including oxidative stress. Pink1 localizes to mitochondria, and mitochondrial cristae are fragmented in pink1 mutants. Expression of human PINK1 in the Drosophila testes restores male fertility and normal mitochondrial morphology in a portion of pink1 mutants, demonstrating functional conservation between human and Drosophila Pink1. Loss of Drosophila parkin shows phenotypes similar to loss of pink1 function. Notably, overexpression of parkin rescues the male sterility and mitochondrial morphology defects of pink1 mutants, whereas double mutants removing both pink1 and parkin function show muscle phenotypes identical to those observed in either mutant alone. These observations suggest that pink1 and parkin function, at least in part, in the same pathway, with pink1 functioning upstream of parkin. The role of the pink1-parkin pathway in regulating mitochondrial function underscores the importance of mitochondrial dysfunction as a central mechanism of Parkinson's disease

  9. The Drosophila MAPK p38c regulates oxidative stress and lipid homeostasis in the intestine.

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    Sveta Chakrabarti

    2014-09-01

    Full Text Available The p38 mitogen-activated protein (MAP kinase signaling cassette has been implicated in stress and immunity in evolutionarily diverse species. In response to a wide variety of physical, chemical and biological stresses p38 kinases phosphorylate various substrates, transcription factors of the ATF family and other protein kinases, regulating cellular adaptation to stress. The Drosophila genome encodes three p38 kinases named p38a, p38b and p38c. In this study, we have analyzed the role of p38c in the Drosophila intestine. The p38c gene is expressed in the midgut and upregulated upon intestinal infection. We showed that p38c mutant flies are more resistant to infection with the lethal pathogen Pseudomonas entomophila but are more susceptible to the non-pathogenic bacterium Erwinia carotovora 15. This phenotype was linked to a lower production of Reactive Oxygen Species (ROS in the gut of p38c mutants, whereby the transcription of the ROS-producing enzyme Duox is reduced in p38c mutant flies. Our genetic analysis shows that p38c functions in a pathway with Mekk1 and Mkk3 to induce the phosphorylation of Atf-2, a transcription factor that controls Duox expression. Interestingly, p38c deficient flies accumulate lipids in the intestine while expressing higher levels of antimicrobial peptide and metabolic genes. The role of p38c in lipid metabolism is mediated by the Atf3 transcription factor. This observation suggests that p38c and Atf3 function in a common pathway in the intestine to regulate lipid metabolism and immune homeostasis. Collectively, our study demonstrates that p38c plays a central role in the intestine of Drosophila. It also reveals that many roles initially attributed to p38a are in fact mediated by p38c.

  10. The Drosophila MAPK p38c regulates oxidative stress and lipid homeostasis in the intestine.

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    Chakrabarti, Sveta; Poidevin, Mickaël; Lemaitre, Bruno

    2014-09-01

    The p38 mitogen-activated protein (MAP) kinase signaling cassette has been implicated in stress and immunity in evolutionarily diverse species. In response to a wide variety of physical, chemical and biological stresses p38 kinases phosphorylate various substrates, transcription factors of the ATF family and other protein kinases, regulating cellular adaptation to stress. The Drosophila genome encodes three p38 kinases named p38a, p38b and p38c. In this study, we have analyzed the role of p38c in the Drosophila intestine. The p38c gene is expressed in the midgut and upregulated upon intestinal infection. We showed that p38c mutant flies are more resistant to infection with the lethal pathogen Pseudomonas entomophila but are more susceptible to the non-pathogenic bacterium Erwinia carotovora 15. This phenotype was linked to a lower production of Reactive Oxygen Species (ROS) in the gut of p38c mutants, whereby the transcription of the ROS-producing enzyme Duox is reduced in p38c mutant flies. Our genetic analysis shows that p38c functions in a pathway with Mekk1 and Mkk3 to induce the phosphorylation of Atf-2, a transcription factor that controls Duox expression. Interestingly, p38c deficient flies accumulate lipids in the intestine while expressing higher levels of antimicrobial peptide and metabolic genes. The role of p38c in lipid metabolism is mediated by the Atf3 transcription factor. This observation suggests that p38c and Atf3 function in a common pathway in the intestine to regulate lipid metabolism and immune homeostasis. Collectively, our study demonstrates that p38c plays a central role in the intestine of Drosophila. It also reveals that many roles initially attributed to p38a are in fact mediated by p38c.

  11. The Drosophila Perlecan gene trol regulates multiple signaling pathways in different developmental contexts

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    Perry Trinity L

    2007-11-01

    Full Text Available Abstract Background Heparan sulfate proteoglycans modulate signaling by a variety of growth factors. The mammalian proteoglycan Perlecan binds and regulates signaling by Sonic Hedgehog, Fibroblast Growth Factors (FGFs, Vascular Endothelial Growth Factor (VEGF and Platelet Derived Growth Factor (PDGF, among others, in contexts ranging from angiogenesis and cardiovascular development to cancer progression. The Drosophila Perlecan homolog trol has been shown to regulate the activity of Hedgehog and Branchless (an FGF homolog to control the onset of stem cell proliferation in the developing brain during first instar. Here we extend analysis of trol mutant phenotypes to show that trol is required for a variety of developmental events and modulates signaling by multiple growth factors in different situations. Results Different mutations in trol allow developmental progression to varying extents, suggesting that trol is involved in multiple cell-fate and patterning decisions. Analysis of the initiation of neuroblast proliferation at second instar demonstrated that trol regulates this event by modulating signaling by Hedgehog and Branchless, as it does during first instar. Trol protein is distributed over the surface of the larval brain, near the regulated neuroblasts that reside on the cortical surface. Mutations in trol also decrease the number of circulating plasmatocytes. This is likely to be due to decreased expression of pointed, the response gene for VEGF/PDGF signaling that is required for plasmatocyte proliferation. Trol is found on plasmatocytes, where it could regulate VEGF/PDGF signaling. Finally, we show that in second instar brains but not third instar brain lobes and eye discs, mutations in trol affect signaling by Decapentaplegic (a Transforming Growth Factor family member, Wingless (a Wnt growth factor and Hedgehog. Conclusion These studies extend the known functions of the Drosophila Perlecan homolog trol in both developmental and

  12. Drosophila Cbp53E Regulates Axon Growth at the Neuromuscular Junction.

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    Kimberly R Hagel

    Full Text Available Calcium is a primary second messenger in all cells that functions in processes ranging from cellular proliferation to synaptic transmission. Proper regulation of calcium is achieved through numerous mechanisms involving channels, sensors, and buffers notably containing one or more EF-hand calcium binding domains. The Drosophila genome encodes only a single 6 EF-hand domain containing protein, Cbp53E, which is likely the prototypic member of a small family of related mammalian proteins that act as calcium buffers and calcium sensors. Like the mammalian homologs, Cbp53E is broadly though discretely expressed throughout the nervous system. Despite the importance of calcium in neuronal function and growth, nothing is known about Cbp53E's function in neuronal development. To address this deficiency, we generated novel null alleles of Drosophila Cbp53E and examined neuronal development at the well-characterized larval neuromuscular junction. Loss of Cbp53E resulted in increases in axonal branching at both peptidergic and glutamatergic neuronal terminals. This overgrowth could be completely rescued by expression of exogenous Cbp53E. Overexpression of Cbp53E, however, only affected the growth of peptidergic neuronal processes. These findings indicate that Cbp53E plays a significant role in neuronal growth and suggest that it may function in both local synaptic and global cellular mechanisms.

  13. Expressionof Drosophila FOXO regulates growth and can phenocopy starvation

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    Lockyer Joseph M

    2003-07-01

    Full Text Available Abstract Background Components of theinsulin signaling pathway are important regulators of growth. TheFOXO (forkhead box, sub-group "O" transcriptionfactors regulate cellular processes under conditions of low levelsof insulin signaling. Studies in mammalian cell culture show thatactivation of FOXO transcription factors causes cell death or cellcycle arrest. The Caenorhabiditis elegans homologue ofFOXO, Daf-16, is required for the formation of dauer larvae in responseto nutritional stress. In addition, FOXO factors have been implicatedin stress resistance and longevity. Results We have identifiedthe Drosophila melanogaster homologue of FOXO (dFOXO,which is conserved in amino acid sequence compared with the mammalianFOXO homologues and Daf-16. Expression of dFOXO during early larvaldevelopment causes inhibition of larval growth and alterations infeeding behavior. Inhibition of larval growth is reversible upondiscontinuation of dFOXO expression. Expression of dFOXO duringthe third larval instar or at low levels during development leadsto the generation of adults that are reduced in size. Analysis ofthe wings and eyes of these small flies indicates that the reductionin size is due to decreases in cell size and cell number. Overexpressionof dFOXO in the developing eye leads to a characteristic phenotypewith reductions in cell size and cell number. This phenotype canbe rescued by co-expression of upstream insulin signaling components,dPI3K and dAkt, however, this rescue is not seen when FOXO is mutatedto a constitutively active form. Conclusions dFOXO is conservedin both sequence and regulatory mechanisms when compared with otherFOXO homologues. The establishment of Drosophila as a model forthe study of FOXO transcription factors should prove beneficialto determining the biological role of these signaling molecules.The alterations in larval development seen upon overexpression ofdFOXO closely mimic the phenotypic effects of starvation, suggestinga

  14. Circadian Rhythm Neuropeptides in Drosophila: Signals for Normal Circadian Function and Circadian Neurodegenerative Disease.

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    He, Qiankun; Wu, Binbin; Price, Jeffrey L; Zhao, Zhangwu

    2017-04-21

    Circadian rhythm is a ubiquitous phenomenon in many organisms ranging from prokaryotes to eukaryotes. During more than four decades, the intrinsic and exogenous regulations of circadian rhythm have been studied. This review summarizes the core endogenous oscillation in Drosophila and then focuses on the neuropeptides, neurotransmitters and hormones that mediate its outputs and integration in Drosophila and the links between several of these (pigment dispersing factor (PDF) and insulin-like peptides) and neurodegenerative disease. These signaling molecules convey important network connectivity and signaling information for normal circadian function, but PDF and insulin-like peptides can also convey signals that lead to apoptosis, enhanced neurodegeneration and cognitive decline in flies carrying circadian mutations or in a senescent state.

  15. Sensory integration regulating male courtship behavior in Drosophila.

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    Dimitrije Krstic

    Full Text Available The courtship behavior of Drosophila melanogaster serves as an excellent model system to study how complex innate behaviors are controlled by the nervous system. To understand how the underlying neural network controls this behavior, it is not sufficient to unravel its architecture, but also crucial to decipher its logic. By systematic analysis of how variations in sensory inputs alter the courtship behavior of a naïve male in the single-choice courtship paradigm, we derive a model describing the logic of the network that integrates the various sensory stimuli and elicits this complex innate behavior. This approach and the model derived from it distinguish (i between initiation and maintenance of courtship, (ii between courtship in daylight and in the dark, where the male uses a scanning strategy to retrieve the decamping female, and (iii between courtship towards receptive virgin females and mature males. The last distinction demonstrates that sexual orientation of the courting male, in the absence of discriminatory visual cues, depends on the integration of gustatory and behavioral feedback inputs, but not on olfactory signals from the courted animal. The model will complement studies on the connectivity and intrinsic properties of the neurons forming the circuitry that regulates male courtship behavior.

  16. Drosophila TRF2 and TAF9 regulate lipid droplet size and phospholipid fatty acid composition.

    Science.gov (United States)

    Fan, Wei; Lam, Sin Man; Xin, Jingxue; Yang, Xiao; Liu, Zhonghua; Liu, Yuan; Wang, Yong; Shui, Guanghou; Huang, Xun

    2017-03-01

    The general transcription factor TBP (TATA-box binding protein) and its associated factors (TAFs) together form the TFIID complex, which directs transcription initiation. Through RNAi and mutant analysis, we identified a specific TBP family protein, TRF2, and a set of TAFs that regulate lipid droplet (LD) size in the Drosophila larval fat body. Among the three Drosophila TBP genes, trf2, tbp and trf1, only loss of function of trf2 results in increased LD size. Moreover, TRF2 and TAF9 regulate fatty acid composition of several classes of phospholipids. Through RNA profiling, we found that TRF2 and TAF9 affects the transcription of a common set of genes, including peroxisomal fatty acid β-oxidation-related genes that affect phospholipid fatty acid composition. We also found that knockdown of several TRF2 and TAF9 target genes results in large LDs, a phenotype which is similar to that of trf2 mutants. Together, these findings provide new insights into the specific role of the general transcription machinery in lipid homeostasis.

  17. Regulation of Energy Stores and Feeding by Neuronal and Peripheral CREB Activity in Drosophila

    Science.gov (United States)

    Iijima, Koichi; Zhao, LiJuan; Shenton, Christopher; Iijima-Ando, Kanae

    2009-01-01

    The cAMP-responsive transcription factor CREB functions in adipose tissue and liver to regulate glycogen and lipid metabolism in mammals. While Drosophila has a homolog of mammalian CREB, dCREB2, its role in energy metabolism is not fully understood. Using tissue-specific expression of a dominant-negative form of CREB (DN-CREB), we have examined the effect of blocking CREB activity in neurons and in the fat body, the primary energy storage depot with functions of adipose tissue and the liver in flies, on energy balance, stress resistance and feeding behavior. We found that disruption of CREB function in neurons reduced glycogen and lipid stores and increased sensitivity to starvation. Expression of DN-CREB in the fat body also reduced glycogen levels, while it did not affect starvation sensitivity, presumably due to increased lipid levels in these flies. Interestingly, blocking CREB activity in the fat body increased food intake. These flies did not show a significant change in overall body size, suggesting that disruption of CREB activity in the fat body caused an obese-like phenotype. Using a transgenic CRE-luciferase reporter, we further demonstrated that disruption of the adipokinetic hormone receptor, which is functionally related to mammalian glucagon and β-adrenergic signaling, in the fat body reduced CRE-mediated transcription in flies. This study demonstrates that CREB activity in either neuronal or peripheral tissues regulates energy balance in Drosophila, and that the key signaling pathway regulating CREB activity in peripheral tissue is evolutionarily conserved. PMID:20041126

  18. Molecular evolution and functional characterization of Drosophila insulin-like peptides.

    Directory of Open Access Journals (Sweden)

    Sebastian Grönke

    2010-02-01

    Full Text Available Multicellular animals match costly activities, such as growth and reproduction, to the environment through nutrient-sensing pathways. The insulin/IGF signaling (IIS pathway plays key roles in growth, metabolism, stress resistance, reproduction, and longevity in diverse organisms including mammals. Invertebrate genomes often contain multiple genes encoding insulin-like ligands, including seven Drosophila insulin-like peptides (DILPs. We investigated the evolution, diversification, redundancy, and functions of the DILPs, combining evolutionary analysis, based on the completed genome sequences of 12 Drosophila species, and functional analysis, based on newly-generated knock-out mutations for all 7 dilp genes in D. melanogaster. Diversification of the 7 DILPs preceded diversification of Drosophila species, with stable gene diversification and family membership, suggesting stabilising selection for gene function. Gene knock-outs demonstrated both synergy and compensation of expression between different DILPs, notably with DILP3 required for normal expression of DILPs 2 and 5 in brain neurosecretory cells and expression of DILP6 in the fat body compensating for loss of brain DILPs. Loss of DILP2 increased lifespan and loss of DILP6 reduced growth, while loss of DILP7 did not affect fertility, contrary to its proposed role as a Drosophila relaxin. Importantly, loss of DILPs produced in the brain greatly extended lifespan but only in the presence of the endosymbiontic bacterium Wolbachia, demonstrating a specific interaction between IIS and Wolbachia in lifespan regulation. Furthermore, loss of brain DILPs blocked the responses of lifespan and fecundity to dietary restriction (DR and the DR response of these mutants suggests that IIS extends lifespan through mechanisms that both overlap with those of DR and through additional mechanisms that are independent of those at work in DR. Evolutionary conservation has thus been accompanied by synergy

  19. NF-1 Dependent Gene Regulation in Drosophila Melanogaster

    National Research Council Canada - National Science Library

    Zhong, Yi

    2004-01-01

    .... We have used an Affymetrix whole genome chip, containing all 13,500 genes of the fruit fly Drosophila, to identify 93 genes with altered expression patterns in flies that have no NF1 protein compared...

  20. The Brakeless co-regulator can directly activate and repress transcription in early Drosophila embryos.

    Science.gov (United States)

    Crona, Filip; Holmqvist, Per-Henrik; Tang, Min; Singla, Bhumica; Vakifahmetoglu-Norberg, Helin; Fantur, Katrin; Mannervik, Mattias

    2015-11-01

    The Brakeless protein performs many important functions during Drosophila development, but how it controls gene expression is poorly understood. We previously showed that Brakeless can function as a transcriptional co-repressor. In this work, we perform transcriptional profiling of brakeless mutant embryos. Unexpectedly, the majority of affected genes are down-regulated in brakeless mutants. We demonstrate that genomic regions in close proximity to some of these genes are occupied by Brakeless, that over-expression of Brakeless causes a reciprocal effect on expression of these genes, and that Brakeless remains an activator of the genes upon fusion to an activation domain. Together, our results show that Brakeless can both repress and activate gene expression. A yeast two-hybrid screen identified the Mediator complex subunit Med19 as interacting with an evolutionarily conserved part of Brakeless. Both down- and up-regulated Brakeless target genes are also affected in Med19-depleted embryos, but only down-regulated targets are influenced in embryos depleted of both Brakeless and Med19. Our data provide support for a Brakeless activator function that regulates transcription by interacting with Med19. We conclude that the transcriptional co-regulator Brakeless can either activate or repress transcription depending on context. Copyright © 2015 Elsevier Inc. All rights reserved.

  1. DIP1 modulates stem cell homeostasis in Drosophila through regulation of sisR-1.

    Science.gov (United States)

    Wong, Jing Ting; Akhbar, Farzanah; Ng, Amanda Yunn Ee; Tay, Mandy Li-Ian; Loi, Gladys Jing En; Pek, Jun Wei

    2017-10-02

    Stable intronic sequence RNAs (sisRNAs) are by-products of splicing and regulate gene expression. How sisRNAs are regulated is unclear. Here we report that a double-stranded RNA binding protein, Disco-interacting protein 1 (DIP1) regulates sisRNAs in Drosophila. DIP1 negatively regulates the abundance of sisR-1 and INE-1 sisRNAs. Fine-tuning of sisR-1 by DIP1 is important to maintain female germline stem cell homeostasis by modulating germline stem cell differentiation and niche adhesion. Drosophila DIP1 localizes to a nuclear body (satellite body) and associates with the fourth chromosome, which contains a very high density of INE-1 transposable element sequences that are processed into sisRNAs. DIP1 presumably acts outside the satellite bodies to regulate sisR-1, which is not on the fourth chromosome. Thus, our study identifies DIP1 as a sisRNA regulatory protein that controls germline stem cell self-renewal in Drosophila.Stable intronic sequence RNAs (sisRNAs) are by-products of splicing from introns with roles in embryonic development in Drosophila. Here, the authors show that the RNA binding protein DIP1 regulates sisRNAs in Drosophila, which is necessary for germline stem cell homeostasis.

  2. Methyl Farnesoate Plays a Dual Role in Regulating Drosophila Metamorphosis

    Science.gov (United States)

    Wen, Di; Rivera-Perez, Crisalejandra; Abdou, Mohamed; Jia, Qiangqiang; He, Qianyu; Liu, Xi; Zyaan, Ola; Xu, Jingjing; Bendena, William G.; Tobe, Stephen S.; Noriega, Fernando G.; Palli, Subba R.; Wang, Jian; Li, Sheng

    2015-01-01

    Corpus allatum (CA) ablation results in juvenile hormone (JH) deficiency and pupal lethality in Drosophila. The fly CA produces and releases three sesquiterpenoid hormones: JH III bisepoxide (JHB3), JH III, and methyl farnesoate (MF). In the whole body extracts, MF is the most abundant sesquiterpenoid, followed by JHB3 and JH III. Knockout of JH acid methyl transferase (jhamt) did not result in lethality; it decreased biosynthesis of JHB3, but MF biosynthesis was not affected. RNAi-mediated reduction of 3-hydroxy-3-methylglutaryl CoA reductase (hmgcr) expression in the CA decreased biosynthesis and titers of the three sesquiterpenoids, resulting in partial lethality. Reducing hmgcr expression in the CA of the jhamt mutant further decreased MF titer to a very low level, and caused complete lethality. JH III, JHB3, and MF function through Met and Gce, the two JH receptors, and induce expression of Kr-h1, a JH primary-response gene. As well, a portion of MF is converted to JHB3 in the hemolymph or peripheral tissues. Topical application of JHB3, JH III, or MF precluded lethality in JH-deficient animals, but not in the Met gce double mutant. Taken together, these experiments show that MF is produced by the larval CA and released into the hemolymph, from where it exerts its anti-metamorphic effects indirectly after conversion to JHB3, as well as acting as a hormone itself through the two JH receptors, Met and Gce. PMID:25774983

  3. Drosophila DH31 Neuropeptide and PDF Receptor Regulate Night-Onset Temperature Preference.

    Science.gov (United States)

    Goda, Tadahiro; Tang, Xin; Umezaki, Yujiro; Chu, Michelle L; Hamada, Fumika N

    2016-11-16

    Body temperature exhibits rhythmic fluctuations over a 24 h period (Refinetti and Menaker, 1992) and decreases during the night, which is associated with sleep initiation (Gilbert et al., 2004; Kräuchi, 2007a,b). However, the underlying mechanism of this temperature decrease is largely unknown. We have previously shown that Drosophila exhibit a daily temperature preference rhythm (TPR), in which their preferred temperatures increase during the daytime and then decrease at the transition from day to night (night-onset) (Kaneko et al., 2012). Because Drosophila are small ectotherms, their body temperature is very close to that of the ambient temperature (Stevenson, 1985), suggesting that their TPR generates their body temperature rhythm. Here, we demonstrate that the neuropeptide diuretic hormone 31 (DH31) and pigment-dispersing factor receptor (PDFR) contribute to regulate the preferred temperature decrease at night-onset. We show that PDFR and tethered-DH31 expression in dorsal neurons 2 (DN2s) restore the preferred temperature decrease at night-onset, suggesting that DH31 acts on PDFR in DN2s. Notably, we previously showed that the molecular clock in DN2s is important for TPR. Although PDF (another ligand of PDFR) is a critical factor for locomotor activity rhythms, Pdf mutants exhibit normal preferred temperature decreases at night-onset. This suggests that DH31-PDFR signaling specifically regulates a preferred temperature decrease at night-onset. Thus, we propose that night-onset TPR and locomotor activity rhythms are differentially controlled not only by clock neurons but also by neuropeptide signaling in the brain. Body temperature rhythm (BTR) is fundamental for the maintenance of functions essential for homeostasis, such as generating metabolic energy and sleep. One major unsolved question is how body temperature decreases dramatically during the night. Previously, we demonstrated that a BTR-like mechanism, referred to as temperature preference rhythm (TPR

  4. Molecular Regulation of Alternative Polyadenylation (APA) within the Drosophila Nervous System.

    Science.gov (United States)

    Vallejos Baier, Raul; Picao-Osorio, Joao; Alonso, Claudio R

    2017-10-27

    Alternative polyadenylation (APA) is a widespread gene regulatory mechanism that generates mRNAs with different 3'-ends, allowing them to interact with different sets of RNA regulators such as microRNAs and RNA-binding proteins. Recent studies have shown that during development, neural tissues produce mRNAs with particularly long 3'UTRs, suggesting that such extensions might be important for neural development and function. Despite this, the mechanisms underlying neural APA are not well understood. Here, we investigate this problem within the Drosophila nervous system, focusing on the roles played by general cleavage and polyadenylation factors (CPA factors). In particular, we examine the model that modulations in CPA factor concentration may affect APA during development. For this, we first analyse the expression of the Drosophila orthologues of all mammalian CPA factors and note that their expression decreases during embryogenesis. In contrast to this global developmental decrease in CPA factor expression, we see that cleavage factor I (CFI) expression is actually elevated in the late embryonic central nervous system, suggesting that CFI might play a special role in neural tissues. To test this, we use the UAS/Gal4 system to deplete CFI proteins from neural tissue and observe that in this condition, multiple genes switch their APA patterns, demonstrating a role of CFI in APA control during Drosophila neural development. Furthermore, analysis of genes with 3'UTR extensions of different length leads us to suggest a novel relation between 3'UTR length and sensitivity to CPA factor expression. Our work thus contributes to the understanding of the mechanisms of APA control within the developing central nervous system. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

  5. Mutational Analysis of Drosophila Basigin Function in the Visual System

    Science.gov (United States)

    Munro, Michelle; Akkam, Yazan; Curtin, Kathryn D.

    2009-01-01

    Drosophila basigin is a cell-surface glycoprotein of the Ig superfamily and a member of a protein family that includes mammalian EMMPRIN/CD147/basigin, neuroplastin, and embigin. Our previous work on Drosophila basigin has shown that it is required for normal photoreceptor cell structure and normal neuron-glia interaction in the fly visual system. Specifically, the photoreceptor neurons of mosaic animals that are mutant in the eye for basigin show altered cell structure with nuclei, mitochondria and rER misplaced and variable axon diameter compared to wild-type. In addition, glia cells in the optic lamina that contact photoreceptor axons are misplaced and show altered structure. All these defects are rescued by expression of either transgenic fly basigin or transgenic mouse basigin in the photoreceptors demonstrating that mouse basigin can functionally replace fly basigin. To determine what regions of the basigin protein are required for each of these functions, we have created mutant basigin transgenes coding for proteins that are altered in conserved residues, introduced these into the fly genome, and tested them for their ability to rescue both photoreceptor cell structure defects and neuron-glia interaction defects of basigin. The results suggest that the highly conserved transmembrane domain and the extracellular domains are crucial for basigin function in the visual system while the short intracellular tail may not play a role in these functions. PMID:19782733

  6. A Screening of UNF Targets Identifies Rnb, a Novel Regulator of Drosophila Circadian Rhythms.

    Science.gov (United States)

    Kozlov, Anatoly; Jaumouillé, Edouard; Machado Almeida, Pedro; Koch, Rafael; Rodriguez, Joseph; Abruzzi, Katharine C; Nagoshi, Emi

    2017-07-12

    Behavioral circadian rhythms are controlled by multioscillator networks comprising functionally different subgroups of clock neurons. Studies have demonstrated that molecular clocks in the fruit fly Drosophila melanogaster are regulated differently in clock neuron subclasses to support their specific functions (Lee et al., 2016; Top et al., 2016). The nuclear receptor unfulfilled ( unf ) represents a regulatory node that provides the small ventral lateral neurons (s-LNvs) unique characteristics as the master pacemaker (Beuchle et al., 2012). We previously showed that UNF interacts with the s-LNv molecular clocks by regulating transcription of the core clock gene period ( per ) (Jaumouillé et al., 2015). To gain more insight into the mechanisms by which UNF contributes to the functioning of the circadian master pacemaker, we identified UNF target genes using chromatin immunoprecipitation. Our data demonstrate that a previously uncharacterized gene CG7837 , which we termed R and B ( Rnb ), acts downstream of UNF to regulate the function of the s-LNvs as the master circadian pacemaker. Mutations and LNv-targeted adult-restricted knockdown of Rnb impair locomotor rhythms. RNB localizes to the nucleus, and its loss-of-function blunts the molecular rhythms and output rhythms of the s-LNvs, particularly the circadian rhythms in PDF accumulation and axonal arbor remodeling. These results establish a second pathway by which UNF interacts with the molecular clocks in the s-LNvs and highlight the mechanistic differences in the molecular clockwork within the pacemaker circuit. SIGNIFICANCE STATEMENT Circadian behavior is generated by a pacemaker circuit comprising diverse classes of pacemaker neurons, each of which contains a molecular clock. In addition to the anatomical and functional diversity, recent studies have shown the mechanistic differences in the molecular clockwork among the pacemaker neurons in Drosophila Here, we identified the molecular characteristics

  7. Functional dissection of the Hox protein Abdominal-B in Drosophila cell culture

    Energy Technology Data Exchange (ETDEWEB)

    Zhai, Zongzhao [Key Laboratory of the Zoological Systematics and Evolution, Institute of Zoology, Chinese Academy of Sciences, Beichen West Road, Chaoyang, Beijing 100101 (China); CellNetworks - Cluster of Excellence, Centre for Organismal Studies (COS) Heidelberg, University of Heidelberg, D-69120 Heidelberg (Germany); Graduate School of Chinese Academy of Sciences, Beijing 100039 (China); Yang, Xingke, E-mail: yangxk@ioz.ac.cn [Key Laboratory of the Zoological Systematics and Evolution, Institute of Zoology, Chinese Academy of Sciences, Beichen West Road, Chaoyang, Beijing 100101 (China); Lohmann, Ingrid, E-mail: ilohmann@flydev.org [CellNetworks - Cluster of Excellence, Centre for Organismal Studies (COS) Heidelberg, University of Heidelberg, D-69120 Heidelberg (Germany)

    2011-11-04

    Highlights: Black-Right-Pointing-Pointer ct340 CRM was identified to be the posterior spiracle enhancer of gene cut. Black-Right-Pointing-Pointer ct340 is under the direct transcriptional control of Hox protein Abd-B. Black-Right-Pointing-Pointer An efficient cloning system was developed to assay protein-DNA interaction. Black-Right-Pointing-Pointer New features of Abd-B dependent target gene regulation were detected. -- Abstract: Hox transcription factors regulate the morphogenesis along the anterior-posterior (A/P) body axis through the interaction with small cis-regulatory modules (CRMs) of their target gene, however so far very few Hox CRMs are known and have been analyzed in detail. In this study we have identified a new Hox CRM, ct340, which guides the expression of the cell type specification gene cut (ct) in the posterior spiracle under the direct control of the Hox protein Abdominal-B (Abd-B). Using the ct340 enhancer activity as readout, an efficient cloning system to generate VP16 activation domain fusion protein was developed to unambiguously test protein-DNA interaction in Drosophila cell culture. By functionally dissecting the Abd-B protein, new features of Abd-B dependent target gene regulation were detected. Due to its easy adaptability, this system can be generally used to map functional domains within sequence-specific transcriptional factors in Drosophila cell culture, and thus provide preliminary knowledge of the protein functional domain structure for further in vivo analysis.

  8. Functional dissection of the Hox protein Abdominal-B in Drosophila cell culture

    International Nuclear Information System (INIS)

    Zhai, Zongzhao; Yang, Xingke; Lohmann, Ingrid

    2011-01-01

    Highlights: ► ct340 CRM was identified to be the posterior spiracle enhancer of gene cut. ► ct340 is under the direct transcriptional control of Hox protein Abd-B. ► An efficient cloning system was developed to assay protein–DNA interaction. ► New features of Abd-B dependent target gene regulation were detected. -- Abstract: Hox transcription factors regulate the morphogenesis along the anterior–posterior (A/P) body axis through the interaction with small cis-regulatory modules (CRMs) of their target gene, however so far very few Hox CRMs are known and have been analyzed in detail. In this study we have identified a new Hox CRM, ct340, which guides the expression of the cell type specification gene cut (ct) in the posterior spiracle under the direct control of the Hox protein Abdominal-B (Abd-B). Using the ct340 enhancer activity as readout, an efficient cloning system to generate VP16 activation domain fusion protein was developed to unambiguously test protein–DNA interaction in Drosophila cell culture. By functionally dissecting the Abd-B protein, new features of Abd-B dependent target gene regulation were detected. Due to its easy adaptability, this system can be generally used to map functional domains within sequence-specific transcriptional factors in Drosophila cell culture, and thus provide preliminary knowledge of the protein functional domain structure for further in vivo analysis.

  9. Fragile X mental retardation protein regulates trans-synaptic signaling in Drosophila

    Directory of Open Access Journals (Sweden)

    Samuel H. Friedman

    2013-11-01

    Fragile X syndrome (FXS, the most common inherited determinant of intellectual disability and autism spectrum disorders, is caused by loss of the fragile X mental retardation 1 (FMR1 gene product (FMRP, an mRNA-binding translational repressor. A number of conserved FMRP targets have been identified in the well-characterized Drosophila FXS disease model, but FMRP is highly pleiotropic in function and the full spectrum of FMRP targets has yet to be revealed. In this study, screens for upregulated neural proteins in Drosophila fmr1 (dfmr1 null mutants reveal strong elevation of two synaptic heparan sulfate proteoglycans (HSPGs: GPI-anchored glypican Dally-like protein (Dlp and transmembrane Syndecan (Sdc. Our recent work has shown that Dlp and Sdc act as co-receptors regulating extracellular ligands upstream of intracellular signal transduction in multiple trans-synaptic pathways that drive synaptogenesis. Consistently, dfmr1 null synapses exhibit altered WNT signaling, with changes in both Wingless (Wg ligand abundance and downstream Frizzled-2 (Fz2 receptor C-terminal nuclear import. Similarly, a parallel anterograde signaling ligand, Jelly belly (Jeb, and downstream ERK phosphorylation (dpERK are depressed at dfmr1 null synapses. In contrast, the retrograde BMP ligand Glass bottom boat (Gbb and downstream signaling via phosphorylation of the transcription factor MAD (pMAD seem not to be affected. To determine whether HSPG upregulation is causative for synaptogenic defects, HSPGs were genetically reduced to control levels in the dfmr1 null background. HSPG correction restored both (1 Wg and Jeb trans-synaptic signaling, and (2 synaptic architecture and transmission strength back to wild-type levels. Taken together, these data suggest that FMRP negatively regulates HSPG co-receptors controlling trans-synaptic signaling during synaptogenesis, and that loss of this regulation causes synaptic structure and function defects characterizing the FXS disease state.

  10. Mondo/ChREBP-Mlx-Regulated Transcriptional Network Is Essential for Dietary Sugar Tolerance in Drosophila

    Science.gov (United States)

    Havula, Essi; Teesalu, Mari; Hyötyläinen, Tuulia; Seppälä, Heini; Hasygar, Kiran; Auvinen, Petri; Orešič, Matej; Sandmann, Thomas; Hietakangas, Ville

    2013-01-01

    Sugars are important nutrients for many animals, but are also proposed to contribute to overnutrition-derived metabolic diseases in humans. Understanding the genetic factors governing dietary sugar tolerance therefore has profound biological and medical significance. Paralogous Mondo transcription factors ChREBP and MondoA, with their common binding partner Mlx, are key sensors of intracellular glucose flux in mammals. Here we report analysis of the in vivo function of Drosophila melanogaster Mlx and its binding partner Mondo (ChREBP) in respect to tolerance to dietary sugars. Larvae lacking mlx or having reduced mondo expression show strikingly reduced survival on a diet with moderate or high levels of sucrose, glucose, and fructose. mlx null mutants display widespread changes in lipid and phospholipid profiles, signs of amino acid catabolism, as well as strongly elevated circulating glucose levels. Systematic loss-of-function analysis of Mlx target genes reveals that circulating glucose levels and dietary sugar tolerance can be genetically uncoupled: Krüppel-like transcription factor Cabut and carbonyl detoxifying enzyme Aldehyde dehydrogenase type III are essential for dietary sugar tolerance, but display no influence on circulating glucose levels. On the other hand, Phosphofructokinase 2, a regulator of the glycolysis pathway, is needed for both dietary sugar tolerance and maintenance of circulating glucose homeostasis. Furthermore, we show evidence that fatty acid synthesis, which is a highly conserved Mondo-Mlx-regulated process, does not promote dietary sugar tolerance. In contrast, survival of larvae with reduced fatty acid synthase expression is sugar-dependent. Our data demonstrate that the transcriptional network regulated by Mondo-Mlx is a critical determinant of the healthful dietary spectrum allowing Drosophila to exploit sugar-rich nutrient sources. PMID:23593032

  11. Structure and novel functional mechanism of Drosophila SNF in sex-lethal splicing.

    Directory of Open Access Journals (Sweden)

    Jicheng Hu

    Full Text Available Sans-fille (SNF is the Drosophila homologue of mammalian general splicing factors U1A and U2B'', and it is essential in Drosophila sex determination. We found that, besides its ability to bind U1 snRNA, SNF can also bind polyuridine RNA tracts flanking the male-specific exon of the master switch gene Sex-lethal (Sxl pre-mRNA specifically, similar to Sex-lethal protein (SXL. The polyuridine RNA binding enables SNF directly inhibit Sxl exon 3 splicing, as the dominant negative mutant SNF(1621 binds U1 snRNA but not polyuridine RNA. Unlike U1A, both RNA recognition motifs (RRMs of SNF can recognize polyuridine RNA tracts independently, even though SNF and U1A share very high sequence identity and overall structure similarity. As SNF RRM1 tends to self-associate on the opposite side of the RNA binding surface, it is possible for SNF to bridge the formation of super-complexes between two introns flanking Sxl exon 3 or between a intron and U1 snRNP, which serves the molecular basis for SNF to directly regulate Sxl splicing. Taken together, a new functional model for SNF in Drosophila sex determination is proposed. The key of the new model is that SXL and SNF function similarly in promoting Sxl male-specific exon skipping with SNF being an auxiliary or backup to SXL, and it is the combined dose of SXL and SNF governs Drosophila sex determination.

  12. EGFR/Ras Signaling Controls Drosophila Intestinal Stem Cell Proliferation via Capicua-Regulated Genes.

    Directory of Open Access Journals (Sweden)

    Yinhua Jin

    2015-12-01

    Full Text Available Epithelial renewal in the Drosophila intestine is orchestrated by Intestinal Stem Cells (ISCs. Following damage or stress the intestinal epithelium produces ligands that activate the epidermal growth factor receptor (EGFR in ISCs. This promotes their growth and division and, thereby, epithelial regeneration. Here we demonstrate that the HMG-box transcriptional repressor, Capicua (Cic, mediates these functions of EGFR signaling. Depleting Cic in ISCs activated them for division, whereas overexpressed Cic inhibited ISC proliferation and midgut regeneration. Epistasis tests showed that Cic acted as an essential downstream effector of EGFR/Ras signaling, and immunofluorescence showed that Cic's nuclear localization was regulated by EGFR signaling. ISC-specific mRNA expression profiling and DNA binding mapping using DamID indicated that Cic represses cell proliferation via direct targets including string (Cdc25, Cyclin E, and the ETS domain transcription factors Ets21C and Pointed (pnt. pnt was required for ISC over-proliferation following Cic depletion, and ectopic pnt restored ISC proliferation even in the presence of overexpressed dominant-active Cic. These studies identify Cic, Pnt, and Ets21C as critical downstream effectors of EGFR signaling in Drosophila ISCs.

  13. Regulation of the formin Cappuccino is critical for polarity of Drosophila oocytes

    Science.gov (United States)

    Bor, Batbileg; Bois, Justin S.; Quinlan, Margot E.

    2014-01-01

    The Drosophila formin Cappuccino (Capu) creates an actin mesh-like structure that traverses the oocyte during mid-oogenesis. This mesh is thought to prevent premature onset of fast cytoplasmic streaming which normally happens during late-oogenesis. Proper cytoskeletal organization and cytoplasmic streaming are crucial for localization of polarity determinants such as osk, grk, bcd and nanos mRNAs. Capu mutants disrupt these events, leading to female sterility. Capu is regulated by another nucleator, Spire, as well as by autoinhibition in vitro. Studies in vivo confirm that Spire modulates Capu’s function in oocytes; however, how autoinhibition contributes is still unclear. To study the role of autoinhibition in flies, we expressed a Capu construct that is missing the Capu Inhibitory Domain, CapuΔN. Consistent with a gain of activity due to loss of autoinhibition, the actin mesh was denser in CapuΔN oocytes. Further, cytoplasmic streaming was delayed and fertility levels decreased. Localization of osk mRNA in early stages, and bcd and nanos in late stages, were disrupted in CapuΔN-expressing oocytes. Finally, evidence that these phenotypes were due to a loss of autoinhibition comes from co-expression of the N-terminal half of Capu with CapuΔN, which suppressed the defects in actin, cytoplasmic streaming and fertility. From these results, we conclude that Capu can be autoinhibited during Drosophila oocyte development. PMID:25557988

  14. Functional characterization of the Drosophila MRP (mitochondrial RNA processing) RNA gene.

    Science.gov (United States)

    Schneider, Mary D; Bains, Anupinder K; Rajendra, T K; Dominski, Zbigniew; Matera, A Gregory; Simmonds, Andrew J

    2010-11-01

    MRP RNA is a noncoding RNA component of RNase mitochondrial RNA processing (MRP), a multi-protein eukaryotic endoribonuclease reported to function in multiple cellular processes, including ribosomal RNA processing, mitochondrial DNA replication, and cell cycle regulation. A recent study predicted a potential Drosophila ortholog of MRP RNA (CR33682) by computer-based genome analysis. We have confirmed the expression of this gene and characterized the phenotype associated with this locus. Flies with mutations that specifically affect MRP RNA show defects in growth and development that begin in the early larval period and end in larval death during the second instar stage. We present several lines of evidence demonstrating a role for Drosophila MRP RNA in rRNA processing. The nuclear fraction of Drosophila MRP RNA localizes to the nucleolus. Further, a mutant strain shows defects in rRNA processing that include a defect in 5.8S rRNA processing, typical of MRP RNA mutants in other species, as well as defects in early stages of rRNA processing.

  15. Identification of functional elements and regulatory circuits by Drosophila modENCODE

    Energy Technology Data Exchange (ETDEWEB)

    Roy, Sushmita; Ernst, Jason; Kharchenko, Peter V.; Kheradpour, Pouya; Negre, Nicolas; Eaton, Matthew L.; Landolin, Jane M.; Bristow, Christopher A.; Ma, Lijia; Lin, Michael F.; Washietl, Stefan; Arshinoff, Bradley I.; Ay, Ferhat; Meyer, Patrick E.; Robine, Nicolas; Washington, Nicole L.; Stefano, Luisa Di; Berezikov, Eugene; Brown, Christopher D.; Candeias, Rogerio; Carlson, Joseph W.; Carr, Adrian; Jungreis, Irwin; Marbach, Daniel; Sealfon, Rachel; Tolstorukov, Michael Y.; Will, Sebastian; Alekseyenko, Artyom A.; Artieri, Carlo; Booth, Benjamin W.; Brooks, Angela N.; Dai, Qi; Davis, Carrie A.; Duff, Michael O.; Feng, Xin; Gorchakov, Andrey A.; Gu, Tingting; Henikoff, Jorja G.; Kapranov, Philipp; Li, Renhua; MacAlpine, Heather K.; Malone, John; Minoda, Aki; Nordman, Jared; Okamura, Katsutomo; Perry, Marc; Powell, Sara K.; Riddle, Nicole C.; Sakai, Akiko; Samsonova, Anastasia; Sandler, Jeremy E.; Schwartz, Yuri B.; Sher, Noa; Spokony, Rebecca; Sturgill, David; van Baren, Marijke; Wan, Kenneth H.; Yang, Li; Yu, Charles; Feingold, Elise; Good, Peter; Guyer, Mark; Lowdon, Rebecca; Ahmad, Kami; Andrews, Justen; Berger, Bonnie; Brenner, Steven E.; Brent, Michael R.; Cherbas, Lucy; Elgin, Sarah C. R.; Gingeras, Thomas R.; Grossman, Robert; Hoskins, Roger A.; Kaufman, Thomas C.; Kent, William; Kuroda, Mitzi I.; Orr-Weaver, Terry; Perrimon, Norbert; Pirrotta, Vincenzo; Posakony, James W.; Ren, Bing; Russell, Steven; Cherbas, Peter; Graveley, Brenton R.; Lewis, Suzanna; Micklem, Gos; Oliver, Brian; Park, Peter J.; Celniker, Susan E.; Henikoff, Steven; Karpen, Gary H.; Lai, Eric C.; MacAlpine, David M.; Stein, Lincoln D.; White, Kevin P.; Kellis, Manolis

    2010-12-22

    To gain insight into how genomic information is translated into cellular and developmental programs, the Drosophila model organism Encyclopedia of DNA Elements (modENCODE) project is comprehensively mapping transcripts, histone modifications, chromosomal proteins, transcription factors, replication proteins and intermediates, and nucleosome properties across a developmental time course and in multiple cell lines. We have generated more than 700 data sets and discovered protein-coding, noncoding, RNA regulatory, replication, and chromatin elements, more than tripling the annotated portion of the Drosophila genome. Correlated activity patterns of these elements reveal a functional regulatory network, which predicts putative new functions for genes, reveals stage- and tissue-specific regulators, and enables gene-expression prediction. Our results provide a foundation for directed experimental and computational studies in Drosophila and related species and also a model for systematic data integration toward comprehensive genomic and functional annotation. Several years after the complete genetic sequencing of many species, it is still unclear how to translate genomic information into a functional map of cellular and developmental programs. The Encyclopedia of DNA Elements (ENCODE) (1) and model organism ENCODE (modENCODE) (2) projects use diverse genomic assays to comprehensively annotate the Homo sapiens (human), Drosophila melanogaster (fruit fly), and Caenorhabditis elegans (worm) genomes, through systematic generation and computational integration of functional genomic data sets. Previous genomic studies in flies have made seminal contributions to our understanding of basic biological mechanisms and genome functions, facilitated by genetic, experimental, computational, and manual annotation of the euchromatic and heterochromatic genome (3), small genome size, short life cycle, and a deep knowledge of development, gene function, and chromosome biology. The functions

  16. Drosophila UTX coordinates with p53 to regulate ku80 expression in response to DNA damage.

    Directory of Open Access Journals (Sweden)

    Chengwan Zhang

    Full Text Available UTX is known as a general factor that activates gene transcription during development. Here, we demonstrate an additional essential role of UTX in the DNA damage response, in which it upregulates the expression of ku80 in Drosophila, both in cultured cells and in third instar larvae. We further showed that UTX mediates the expression of ku80 by the demethylation of H3K27me3 at the ku80 promoter upon exposure to ionizing radiation (IR in a p53-dependent manner. UTX interacts physically with p53, and both UTX and p53 are recruited to the ku80 promoter following IR exposure in an interdependent manner. In contrast, the loss of utx has little impact on the expression of ku70, mre11, hid and reaper, suggesting the specific regulation of ku80 expression by UTX. Thus, our findings further elucidate the molecular function of UTX.

  17. Delivery of circulating lipoproteins to specific neurons in the Drosophila brain regulates systemic insulin signaling.

    Science.gov (United States)

    Brankatschk, Marko; Dunst, Sebastian; Nemetschke, Linda; Eaton, Suzanne

    2014-10-02

    The Insulin signaling pathway couples growth, development and lifespan to nutritional conditions. Here, we demonstrate a function for the Drosophila lipoprotein LTP in conveying information about dietary lipid composition to the brain to regulate Insulin signaling. When yeast lipids are present in the diet, free calcium levels rise in Blood Brain Barrier glial cells. This induces transport of LTP across the Blood Brain Barrier by two LDL receptor-related proteins: LRP1 and Megalin. LTP accumulates on specific neurons that connect to cells that produce Insulin-like peptides, and induces their release into the circulation. This increases systemic Insulin signaling and the rate of larval development on yeast-containing food compared with a plant-based food of similar nutritional content.

  18. A genome-wide gene function prediction resource for Drosophila melanogaster.

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    Han Yan

    2010-08-01

    Full Text Available Predicting gene functions by integrating large-scale biological data remains a challenge for systems biology. Here we present a resource for Drosophila melanogaster gene function predictions. We trained function-specific classifiers to optimize the influence of different biological datasets for each functional category. Our model predicted GO terms and KEGG pathway memberships for Drosophila melanogaster genes with high accuracy, as affirmed by cross-validation, supporting literature evidence, and large-scale RNAi screens. The resulting resource of prioritized associations between Drosophila genes and their potential functions offers a guide for experimental investigations.

  19. Drosophila Cancer Models Identify Functional Differences between Ret Fusions.

    Science.gov (United States)

    Levinson, Sarah; Cagan, Ross L

    2016-09-13

    We generated and compared Drosophila models of RET fusions CCDC6-RET and NCOA4-RET. Both RET fusions directed cells to migrate, delaminate, and undergo EMT, and both resulted in lethality when broadly expressed. In all phenotypes examined, NCOA4-RET was more severe than CCDC6-RET, mirroring their effects on patients. A functional screen against the Drosophila kinome and a library of cancer drugs found that CCDC6-RET and NCOA4-RET acted through different signaling networks and displayed distinct drug sensitivities. Combining data from the kinome and drug screens identified the WEE1 inhibitor AZD1775 plus the multi-kinase inhibitor sorafenib as a synergistic drug combination that is specific for NCOA4-RET. Our work emphasizes the importance of identifying and tailoring a patient's treatment to their specific RET fusion isoform and identifies a multi-targeted therapy that may prove effective against tumors containing the NCOA4-RET fusion. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  20. Glycosphingolipids with extended sugar chain have specialized functions in development and behavior of Drosophila

    DEFF Research Database (Denmark)

    Chen, Ya-Wen; Pedersen, Johannes W; Wandall, Hans H

    2007-01-01

    Glycosphingolipids (GSL) are glycosylated polar lipids in cell membranes essential for development of vertebrates as well as Drosophila. Mutants that impair enzymes involved in biosynthesis of GSL sugar chains provide a means to assess the functions of the sugar chains in vivo. The Drosophila gly...

  1. Lipids in the cell: organisation regulates function.

    Science.gov (United States)

    Santos, Ana L; Preta, Giulio

    2018-06-01

    Lipids are fundamental building blocks of all cells and play important roles in the pathogenesis of different diseases, including inflammation, autoimmune disease, cancer, and neurodegeneration. The lipid composition of different organelles can vary substantially from cell to cell, but increasing evidence demonstrates that lipids become organised specifically in each compartment, and this organisation is essential for regulating cell function. For example, lipid microdomains in the plasma membrane, known as lipid rafts, are platforms for concentrating protein receptors and can influence intra-cellular signalling. Lipid organisation is tightly regulated and can be observed across different model organisms, including bacteria, yeast, Drosophila, and Caenorhabditis elegans, suggesting that lipid organisation is evolutionarily conserved. In this review, we summarise the importance and function of specific lipid domains in main cellular organelles and discuss recent advances that investigate how these specific and highly regulated structures contribute to diverse biological processes.

  2. Drosophila DJ-1 decreases neural sensitivity to stress by negatively regulating Daxx-like protein through dFOXO.

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    Soojin Hwang

    2013-04-01

    Full Text Available DJ-1, a Parkinson's disease (PD-associated gene, has been shown to protect against oxidative stress in Drosophila. However, the molecular mechanism underlying oxidative stress-induced phenotypes, including apoptosis, locomotive defects, and lethality, in DJ-1-deficient flies is not fully understood. Here we showed that Daxx-like protein (DLP, a Drosophila homologue of the mammalian Death domain-associated protein (Daxx, was upregulated under oxidative stress conditions in the loss-of-function mutants of Drosophila DJ-1β, a Drosophila homologue of DJ-1. DLP overexpression induced apoptosis via the c-Jun N-terminal kinase (JNK/Drosophila forkhead box subgroup O (dFOXO pathway, whereas loss of DLP increased resistance to oxidative stress and UV irradiation. Moreover, the oxidative stress-induced phenotypes of DJ-1β mutants were dramatically rescued by DLP deficiency, suggesting that enhanced expression of DLP contributes to the DJ-1β mutant phenotypes. Interestingly, we found that dFOXO was required for the increase in DLP expression in DJ-1β mutants and that dFOXO activity was increased in the heads of DJ-1β mutants. In addition, subcellular localization of DLP appeared to be influenced by DJ-1 expression so that cytosolic DLP was increased in DJ-1β mutants. Similarly, in mammalian cells, Daxx translocation from the nucleus to the cytosol was suppressed by overexpressed DJ-1β under oxidative stress conditions; and, furthermore, targeted expression of DJ-1β to mitochondria efficiently inhibited the Daxx translocation. Taken together, our findings demonstrate that DJ-1β protects flies against oxidative stress- and UV-induced apoptosis by regulating the subcellular localization and gene expression of DLP, thus implying that Daxx-induced apoptosis is involved in the pathogenesis of DJ-1-associated PD.

  3. A genome-wide RNAi screen identifies regulators of cholesterol-modified hedgehog secretion in Drosophila.

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    Reid Aikin

    Full Text Available Hedgehog (Hh proteins are secreted molecules that function as organizers in animal development. In addition to being palmitoylated, Hh is the only metazoan protein known to possess a covalently-linked cholesterol moiety. The absence of either modification severely disrupts the organization of numerous tissues during development. It is currently not known how lipid-modified Hh is secreted and released from producing cells. We have performed a genome-wide RNAi screen in Drosophila melanogaster cells to identify regulators of Hh secretion. We found that cholesterol-modified Hh secretion is strongly dependent on coat protein complex I (COPI but not COPII vesicles, suggesting that cholesterol modification alters the movement of Hh through the early secretory pathway. We provide evidence that both proteolysis and cholesterol modification are necessary for the efficient trafficking of Hh through the ER and Golgi. Finally, we identified several putative regulators of protein secretion and demonstrate a role for some of these genes in Hh and Wingless (Wg morphogen secretion in vivo. These data open new perspectives for studying how morphogen secretion is regulated, as well as provide insight into regulation of lipid-modified protein secretion.

  4. The histone deacetylase HDAC1 positively regulates Notch signaling during Drosophila wing development

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    Zehua Wang

    2018-02-01

    Full Text Available The Notch signaling pathway is highly conserved across different animal species and plays crucial roles in development and physiology. Regulation of Notch signaling occurs at multiple levels in different tissues and cell types. Here, we show that the histone deacetylase HDAC1 acts as a positive regulator of Notch signaling during Drosophila wing development. Depletion of HDAC1 causes wing notches on the margin of adult wing. Consistently, the expression of Notch target genes is reduced in the absence of HDAC1 during wing margin formation. We further provide evidence that HDAC1 acts upstream of Notch activation. Mechanistically, we show that HDAC1 regulates Notch protein levels by promoting Notch transcription. Consistent with this, the HDAC1-associated transcriptional co-repressor Atrophin (Atro is also required for transcriptional activation of Notch in the wing disc. In summary, our results demonstrate that HDAC1 positively regulates Notch signaling and reveal a previously unidentified function of HDAC1 in Notch signaling.

  5. Loss of PTB or negative regulation of Notch mRNA reveals distinct zones of Notch and actin protein accumulation in Drosophila embryo.

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    Cedric S Wesley

    Full Text Available Polypyrimidine Tract Binding (PTB protein is a regulator of mRNA processing and translation. Genetic screens and studies of wing and bristle development during the post-embryonic stages of Drosophila suggest that it is a negative regulator of the Notch pathway. How PTB regulates the Notch pathway is unknown. Our studies of Drosophila embryogenesis indicate that (1 the Notch mRNA is a potential target of PTB, (2 PTB and Notch functions in the dorso-lateral regions of the Drosophila embryo are linked to actin regulation but not their functions in the ventral region, and (3 the actin-related Notch activity in the dorso-lateral regions might require a Notch activity at or near the cell surface that is different from the nuclear Notch activity involved in cell fate specification in the ventral region. These data raise the possibility that the Drosophila embryo is divided into zones of different PTB and Notch activities based on whether or not they are linked to actin regulation. They also provide clues to the almost forgotten role of Notch in cell adhesion and reveal a role for the Notch pathway in cell fusions.

  6. Loss of PTB or Negative Regulation of Notch mRNA Reveals Distinct Zones of Notch and Actin Protein Accumulation in Drosophila Embryo

    Science.gov (United States)

    Wesley, Cedric S.; Guo, Heng; Chaudhry, Kanita A.; Thali, Markus J.; Yin, Jerry C.; Clason, Todd; Wesley, Umadevi V.

    2011-01-01

    Polypyrimidine Tract Binding (PTB) protein is a regulator of mRNA processing and translation. Genetic screens and studies of wing and bristle development during the post-embryonic stages of Drosophila suggest that it is a negative regulator of the Notch pathway. How PTB regulates the Notch pathway is unknown. Our studies of Drosophila embryogenesis indicate that (1) the Notch mRNA is a potential target of PTB, (2) PTB and Notch functions in the dorso-lateral regions of the Drosophila embryo are linked to actin regulation but not their functions in the ventral region, and (3) the actin-related Notch activity in the dorso-lateral regions might require a Notch activity at or near the cell surface that is different from the nuclear Notch activity involved in cell fate specification in the ventral region. These data raise the possibility that the Drosophila embryo is divided into zones of different PTB and Notch activities based on whether or not they are linked to actin regulation. They also provide clues to the almost forgotten role of Notch in cell adhesion and reveal a role for the Notch pathway in cell fusions. PMID:21750738

  7. The silent information regulator 1 (Sirt1) is a positive regulator of the Notch pathway in Drosophila

    Czech Academy of Sciences Publication Activity Database

    Horváth, Matěj; Mihajlović, Zorana; Slaninová, Věra; Perez-Gomez, Raquel; Moshkin, Y.; Krejčí, Alena

    2016-01-01

    Roč. 473, č. 22 (2016), s. 4129-4143 ISSN 0264-6021 R&D Projects: GA ČR(CZ) GA14-08583S Institutional support: RVO:60077344 Keywords : Drosophila * silent information regulator 1 * Notch pathway Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.797, year: 2016

  8. Regulation of axonal development by the nuclear protein hindsight (pebbled) in the Drosophila visual system.

    Science.gov (United States)

    Oliva, Carlos; Sierralta, Jimena

    2010-08-15

    The molecules and networks involved in the process of acquisition and maintenance of the form of a mature neuron are not completely known. Using a misexpression screen we identified the gene hindsight as a gene involved in the process of acquisition of the neuronal morphogenesis in the Drosophila adult nervous system. hindsight encodes a transcription factor known for its role in early developmental processes such as embryonic germ band retraction and dorsal closure, as well as in the establishment of cell morphology, planar cell polarity, and epithelial integrity during retinal development. We describe here a novel function for HNT by showing that both loss and gain of function of HNT affects the pathfinding of the photoreceptors axons. By manipulating the timing and level of HNT expression, together with the number of cells manipulated we show here that the function of HNT in axonal guidance is independent of the HNT functions previously reported in retinal cells. Based on genetic interaction experiments we show that part of HNT function in axonal development is exerted through the regulation of genes involved in the dynamics of the actin cytoskeleton. Copyright 2010 Elsevier Inc. All rights reserved.

  9. Inwardly rectifying potassium channels influence Drosophila wing morphogenesis by regulating Dpp release.

    Science.gov (United States)

    Dahal, Giri Raj; Pradhan, Sarala Joshi; Bates, Emily Anne

    2017-08-01

    Loss of embryonic ion channel function leads to morphological defects, but the underlying reason for these defects remains elusive. Here, we show that inwardly rectifying potassium (Irk) channels regulate release of the Drosophila bone morphogenetic protein Dpp in the developing fly wing and that this is necessary for developmental signaling. Inhibition of Irk channels decreases the incidence of distinct Dpp-GFP release events above baseline fluorescence while leading to a broader distribution of Dpp-GFP. Work by others in different cell types has shown that Irk channels regulate peptide release by modulating membrane potential and calcium levels. We found calcium transients in the developing wing, and inhibition of Irk channels reduces the duration and amplitude of calcium transients. Depolarization with high extracellular potassium evokes Dpp release. Taken together, our data implicate Irk channels as a requirement for regulated release of Dpp, highlighting the importance of the temporal pattern of Dpp presentation for morphogenesis of the wing. © 2017. Published by The Company of Biologists Ltd.

  10. Yorkie regulates epidermal wound healing in Drosophila larvae independently of cell proliferation and apoptosis.

    Science.gov (United States)

    Tsai, Chang-Ru; Anderson, Aimee E; Burra, Sirisha; Jo, Juyeon; Galko, Michael J

    2017-07-01

    Yorkie (Yki), the transcriptional co-activator of the Hippo signaling pathway, has well-characterized roles in balancing apoptosis and cell division during organ growth control. Yki is also required in diverse tissue regenerative contexts. In most cases this requirement reflects its well-characterized roles in balancing apoptosis and cell division. Whether Yki has repair functions outside of the control of cell proliferation, death, and growth is not clear. Here we show that Yki and Scalloped (Sd) are required for epidermal wound closure in the Drosophila larval epidermis. Using a GFP-tagged Yki transgene we show that Yki transiently translocates to some epidermal nuclei upon wounding. Genetic analysis strongly suggests that Yki interacts with the known wound healing pathway, Jun N-terminal kinase (JNK), but not with Platelet Derived Growth Factor/Vascular-Endothelial Growth Factor receptor (Pvr). Yki likely acts downstream of or parallel to JNK signaling and does not appear to regulate either proliferation or apoptosis in the larval epidermis during wound repair. Analysis of actin structures after wounding suggests that Yki and Sd promote wound closure through actin regulation. In sum, we found that Yki regulates an epithelial tissue repair process independently of its previously documented roles in balancing proliferation and apoptosis. Copyright © 2017 Elsevier Inc. All rights reserved.

  11. Drosophila Big bang regulates the apical cytocortex and wing growth through junctional tension.

    Science.gov (United States)

    Tsoumpekos, Giorgos; Nemetschke, Linda; Knust, Elisabeth

    2018-03-05

    Growth of epithelial tissues is regulated by a plethora of components, including signaling and scaffolding proteins, but also by junctional tension, mediated by the actomyosin cytoskeleton. However, how these players are spatially organized and functionally coordinated is not well understood. Here, we identify the Drosophila melanogaster scaffolding protein Big bang as a novel regulator of growth in epithelial cells of the wing disc by ensuring proper junctional tension. Loss of big bang results in the reduction of the regulatory light chain of nonmuscle myosin, Spaghetti squash. This is associated with an increased apical cell surface, decreased junctional tension, and smaller wings. Strikingly, these phenotypic traits of big bang mutant discs can be rescued by expressing constitutively active Spaghetti squash. Big bang colocalizes with Spaghetti squash in the apical cytocortex and is found in the same protein complex. These results suggest that in epithelial cells of developing wings, the scaffolding protein Big bang controls apical cytocortex organization, which is important for regulating cell shape and tissue growth. © 2018 Tsoumpekos et al.

  12. Pink1 and Parkin regulate Drosophila intestinal stem cell proliferation during stress and aging.

    Science.gov (United States)

    Koehler, Christopher L; Perkins, Guy A; Ellisman, Mark H; Jones, D Leanne

    2017-08-07

    Intestinal stem cells (ISCs) maintain the midgut epithelium in Drosophila melanogaster Proper cellular turnover and tissue function rely on tightly regulated rates of ISC division and appropriate differentiation of daughter cells. However, aging and epithelial injury cause elevated ISC proliferation and decreased capacity for terminal differentiation of daughter enteroblasts (EBs). The mechanisms causing functional decline of stem cells with age remain elusive; however, recent findings suggest that stem cell metabolism plays an important role in the regulation of stem cell activity. Here, we investigate how alterations in mitochondrial homeostasis modulate stem cell behavior in vivo via RNA interference-mediated knockdown of factors involved in mitochondrial dynamics. ISC/EB-specific knockdown of the mitophagy-related genes Pink1 or Parkin suppresses the age-related loss of tissue homeostasis, despite dramatic changes in mitochondrial ultrastructure and mitochondrial damage in ISCs/EBs. Maintenance of tissue homeostasis upon reduction of Pink1 or Parkin appears to result from reduction of age- and stress-induced ISC proliferation, in part, through induction of ISC senescence. Our results indicate an uncoupling of cellular, tissue, and organismal aging through inhibition of ISC proliferation and provide insight into strategies used by stem cells to maintain tissue homeostasis despite severe damage to organelles. © 2017 Koehler et al.

  13. DISCO Interacting Protein 2 regulates axonal bifurcation and guidance of Drosophila mushroom body neurons.

    Science.gov (United States)

    Nitta, Yohei; Yamazaki, Daisuke; Sugie, Atsushi; Hiroi, Makoto; Tabata, Tetsuya

    2017-01-15

    Axonal branching is one of the key processes within the enormous complexity of the nervous system to enable a single neuron to send information to multiple targets. However, the molecular mechanisms that control branch formation are poorly understood. In particular, previous studies have rarely addressed the mechanisms underlying axonal bifurcation, in which axons form new branches via splitting of the growth cone. We demonstrate that DISCO Interacting Protein 2 (DIP2) is required for precise axonal bifurcation in Drosophila mushroom body (MB) neurons by suppressing ectopic bifurcation and regulating the guidance of sister axons. We also found that DIP2 localize to the plasma membrane. Domain function analysis revealed that the AMP-synthetase domains of DIP2 are essential for its function, which may involve exerting a catalytic activity that modifies fatty acids. Genetic analysis and subsequent biochemical analysis suggested that DIP2 is involved in the fatty acid metabolization of acyl-CoA. Taken together, our results reveal a function of DIP2 in the developing nervous system and provide a potential functional relationship between fatty acid metabolism and axon morphogenesis. Copyright © 2016 Elsevier Inc. All rights reserved.

  14. Regulated functions and integrability

    Directory of Open Access Journals (Sweden)

    Ján Gunčaga

    2009-04-01

    Full Text Available Properties of functions defined on a bounded closed interval, weaker than continuity, have been considered by many mathematicians. Functions having both sides limits at each point are called regulated and were considered by J. Dieudonné [2], D. Fraňková [3] and others (see for example S. Banach [1], S. Saks [8]. The main class of functions we deal with consists of piece-wise constant ones. These functions play a fundamental role in the integration theory which had been developed by Igor Kluvanek (see Š. Tkacik [9]. We present an outline of this theory.

  15. Drosophila Kismet regulates histone H3 lysine 27 methylation and early elongation by RNA polymerase II.

    Directory of Open Access Journals (Sweden)

    Shrividhya Srinivasan

    2008-10-01

    Full Text Available Polycomb and trithorax group proteins regulate cellular pluripotency and differentiation by maintaining hereditable states of transcription. Many Polycomb and trithorax group proteins have been implicated in the covalent modification or remodeling of chromatin, but how they interact with each other and the general transcription machinery to regulate transcription is not well understood. The trithorax group protein Kismet-L (KIS-L is a member of the CHD subfamily of chromatin-remodeling factors that plays a global role in transcription by RNA polymerase II (Pol II. Mutations in CHD7, the human counterpart of kis, are associated with CHARGE syndrome, a developmental disorder affecting multiple tissues and organs. To clarify how KIS-L activates gene expression and counteracts Polycomb group silencing, we characterized defects resulting from the loss of KIS-L function in Drosophila. These studies revealed that KIS-L acts downstream of P-TEFb recruitment to stimulate elongation by Pol II. The presence of two chromodomains in KIS-L suggested that its recruitment or function might be regulated by the methylation of histone H3 lysine 4 by the trithorax group proteins ASH1 and TRX. Although we observed significant overlap between the distributions of KIS-L, ASH1, and TRX on polytene chromosomes, KIS-L did not bind methylated histone tails in vitro, and loss of TRX or ASH1 function did not alter the association of KIS-L with chromatin. By contrast, loss of kis function led to a dramatic reduction in the levels of TRX and ASH1 associated with chromatin and was accompanied by increased histone H3 lysine 27 methylation-a modification required for Polycomb group repression. A similar increase in H3 lysine 27 methylation was observed in ash1 and trx mutant larvae. Our findings suggest that KIS-L promotes early elongation and counteracts Polycomb group repression by recruiting the ASH1 and TRX histone methyltransferases to chromatin.

  16. Clueless, a protein required for mitochondrial function, interacts with the PINK1-Parkin complex in Drosophila

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    Aditya Sen

    2015-06-01

    Full Text Available Loss of mitochondrial function often leads to neurodegeneration and is thought to be one of the underlying causes of neurodegenerative diseases such as Parkinson's disease (PD. However, the precise events linking mitochondrial dysfunction to neuronal death remain elusive. PTEN-induced putative kinase 1 (PINK1 and Parkin (Park, either of which, when mutated, are responsible for early-onset PD, mark individual mitochondria for destruction at the mitochondrial outer membrane. The specific molecular pathways that regulate signaling between the nucleus and mitochondria to sense mitochondrial dysfunction under normal physiological conditions are not well understood. Here, we show that Drosophila Clueless (Clu, a highly conserved protein required for normal mitochondrial function, can associate with Translocase of the outer membrane (TOM 20, Porin and PINK1, and is thus located at the mitochondrial outer membrane. Previously, we found that clu genetically interacts with park in Drosophila female germ cells. Here, we show that clu also genetically interacts with PINK1, and our epistasis analysis places clu downstream of PINK1 and upstream of park. In addition, Clu forms a complex with PINK1 and Park, further supporting that Clu links mitochondrial function with the PINK1-Park pathway. Lack of Clu causes PINK1 and Park to interact with each other, and clu mutants have decreased mitochondrial protein levels, suggesting that Clu can act as a negative regulator of the PINK1-Park pathway. Taken together, these results suggest that Clu directly modulates mitochondrial function, and that Clu's function contributes to the PINK1-Park pathway of mitochondrial quality control.

  17. Kibra and aPKC regulate starvation-induced autophagy in Drosophila

    Energy Technology Data Exchange (ETDEWEB)

    Jin, Ahrum [Department of Biological Sciences, Korea Advanced Institute of Science and Technology, 291 Daehak-ro, Yuseong-gu, Daejeon 34141 (Korea, Republic of); Neufeld, Thomas P. [Department of Genetics, Cell Biology and Development, University of Minnesota, Minneapolis, MN 55455 (United States); Choe, Joonho, E-mail: jchoe@kaist.ac.kr [Department of Biological Sciences, Korea Advanced Institute of Science and Technology, 291 Daehak-ro, Yuseong-gu, Daejeon 34141 (Korea, Republic of)

    2015-12-04

    Autophagy is a bulk degradation system that functions in response to cellular stresses such as metabolic stress, endoplasmic reticulum stress, oxidative stress, and developmental processes. During autophagy, cytoplasmic components are captured in double-membrane vesicles called autophagosomes. The autophagosome fuses with the lysosome, producing a vacuole known as an autolysosome. The cellular components are degraded by lysosomal proteases and recycled. Autophagy is important for maintaining cellular homeostasis, and the process is evolutionarily conserved. Kibra is an upstream regulator of the hippo signaling pathway, which controls organ size by affecting cell growth, proliferation, and apoptosis. Kibra is mainly localized in the apical membrane domain of epithelial cells and acts as a scaffold protein. We found that Kibra is required for autophagy to function properly. The absence of Kibra caused defects in the formation of autophagic vesicles and autophagic degradation. We also found that the well-known cell polarity protein aPKC interacts with Kibra, and its activity affects autophagy upstream of Kibra. Constitutively active aPKC decreased autophagic vesicle formation and autophagic degradation. We confirmed the interaction between aPKC and Kibra in S2 cells and Drosophila larva. Taken together, our data suggest that Kibra and aPKC are essential for regulating starvation-induced autophagy. - Highlights: • Loss of Kibra causes defects in autophagosome formation and autophagic degradation. • Constitutively-active aPKCs negatively regulate autophagy. • Kibra interacts with aPKC in vitro and in vivo. • Kibra regulates autophagy downstream of aPKC.

  18. Drosophila MOF controls Checkpoint protein2 and regulates genomic stability during early embryogenesis.

    Science.gov (United States)

    Pushpavalli, Sreerangam N C V L; Sarkar, Arpita; Ramaiah, M Janaki; Chowdhury, Debabani Roy; Bhadra, Utpal; Pal-Bhadra, Manika

    2013-01-24

    In Drosophila embryos, checkpoints maintain genome stability by delaying cell cycle progression that allows time for damage repair or to complete DNA synthesis. Drosophila MOF, a member of MYST histone acetyl transferase is an essential component of male X hyperactivation process. Until recently its involvement in G2/M cell cycle arrest and defects in ionizing radiation induced DNA damage pathways was not well established. Drosophila MOF is highly expressed during early embryogenesis. In the present study we show that haplo-insufficiency of maternal MOF leads to spontaneous mitotic defects like mitotic asynchrony, mitotic catastrophe and chromatid bridges in the syncytial embryos. Such abnormal nuclei are eliminated and digested in the yolk tissues by nuclear fall out mechanism. MOF negatively regulates Drosophila checkpoint kinase 2 tumor suppressor homologue. In response to DNA damage the checkpoint gene Chk2 (Drosophila mnk) is activated in the mof mutants, there by causing centrosomal inactivation suggesting its role in response to genotoxic stress. A drastic decrease in the fall out nuclei in the syncytial embryos derived from mof¹/+; mnkp⁶/+ females further confirms the role of DNA damage response gene Chk2 to ensure the removal of abnormal nuclei from the embryonic precursor pool and maintain genome stability. The fact that mof mutants undergo DNA damage has been further elucidated by the increased number of single and double stranded DNA breaks. mof mutants exhibited genomic instability as evidenced by the occurance of frequent mitotic bridges in anaphase, asynchronous nuclear divisions, disruption of cytoskeleton, inactivation of centrosomes finally leading to DNA damage. Our findings are consistent to what has been reported earlier in mammals that; reduced levels of MOF resulted in increased genomic instability while total loss resulted in lethality. The study can be further extended using Drosophila as model system and carry out the interaction of MOF

  19. Drosophila MOF controls Checkpoint protein2 and regulates genomic stability during early embryogenesis

    Directory of Open Access Journals (Sweden)

    Pushpavalli Sreerangam NCVL

    2013-01-01

    Full Text Available Abstract Background In Drosophila embryos, checkpoints maintain genome stability by delaying cell cycle progression that allows time for damage repair or to complete DNA synthesis. Drosophila MOF, a member of MYST histone acetyl transferase is an essential component of male X hyperactivation process. Until recently its involvement in G2/M cell cycle arrest and defects in ionizing radiation induced DNA damage pathways was not well established. Results Drosophila MOF is highly expressed during early embryogenesis. In the present study we show that haplo-insufficiency of maternal MOF leads to spontaneous mitotic defects like mitotic asynchrony, mitotic catastrophe and chromatid bridges in the syncytial embryos. Such abnormal nuclei are eliminated and digested in the yolk tissues by nuclear fall out mechanism. MOF negatively regulates Drosophila checkpoint kinase 2 tumor suppressor homologue. In response to DNA damage the checkpoint gene Chk2 (Drosophila mnk is activated in the mof mutants, there by causing centrosomal inactivation suggesting its role in response to genotoxic stress. A drastic decrease in the fall out nuclei in the syncytial embryos derived from mof1/+; mnkp6/+ females further confirms the role of DNA damage response gene Chk2 to ensure the removal of abnormal nuclei from the embryonic precursor pool and maintain genome stability. The fact that mof mutants undergo DNA damage has been further elucidated by the increased number of single and double stranded DNA breaks. Conclusion mof mutants exhibited genomic instability as evidenced by the occurance of frequent mitotic bridges in anaphase, asynchronous nuclear divisions, disruption of cytoskeleton, inactivation of centrosomes finally leading to DNA damage. Our findings are consistent to what has been reported earlier in mammals that; reduced levels of MOF resulted in increased genomic instability while total loss resulted in lethality. The study can be further extended using

  20. The cell-mediated immunity of Drosophila melanogaster: hemocyte lineages, immune compartments, microanatomy and regulation.

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    Honti, Viktor; Csordás, Gábor; Kurucz, Éva; Márkus, Róbert; Andó, István

    2014-01-01

    In the animal kingdom, innate immunity is the first line of defense against invading pathogens. The dangers of microbial and parasitic attacks are countered by similar mechanisms, involving the prototypes of the cell-mediated immune responses, the phagocytosis and encapsulation. Work on Drosophila has played an important role in promoting an understanding of the basic mechanisms of phylogenetically conserved modules of innate immunity. The aim of this review is to survey the developments in the identification and functional definition of immune cell types and the immunological compartments of Drosophila melanogaster. We focus on the molecular and developmental aspects of the blood cell types and compartments, as well as the dynamics of blood cell development and the immune response. Further advances in the characterization of the innate immune mechanisms in Drosophila will provide basic clues to the understanding of the importance of the evolutionary conserved mechanisms of innate immune defenses in the animal kingdom. Copyright © 2013 Elsevier Ltd. All rights reserved.

  1. Activin signaling targeted by insulin/dFOXO regulates aging and muscle proteostasis in Drosophila.

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    Hua Bai

    2013-11-01

    Full Text Available Reduced insulin/IGF signaling increases lifespan in many animals. To understand how insulin/IGF mediates lifespan in Drosophila, we performed chromatin immunoprecipitation-sequencing analysis with the insulin/IGF regulated transcription factor dFOXO in long-lived insulin/IGF signaling genotypes. Dawdle, an Activin ligand, is bound and repressed by dFOXO when reduced insulin/IGF extends lifespan. Reduced Activin signaling improves performance and protein homeostasis in muscles of aged flies. Activin signaling through the Smad binding element inhibits the transcription of Autophagy-specific gene 8a (Atg8a within muscle, a factor controlling the rate of autophagy. Expression of Atg8a within muscle is sufficient to increase lifespan. These data reveal how insulin signaling can regulate aging through control of Activin signaling that in turn controls autophagy, representing a potentially conserved molecular basis for longevity assurance. While reduced Activin within muscle autonomously retards functional aging of this tissue, these effects in muscle also reduce secretion of insulin-like peptides at a distance from the brain. Reduced insulin secretion from the brain may subsequently reinforce longevity assurance through decreased systemic insulin/IGF signaling.

  2. Conservation patterns in different functional sequence categoriesof divergent Drosophila species

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    Papatsenko, Dmitri; Kislyuk, Andrey; Levine, Michael; Dubchak, Inna

    2005-10-01

    We have explored the distributions of fully conservedungapped blocks in genome-wide pairwise alignments of recently completedspecies of Drosophila: D.yakuba, D.ananassae, D.pseudoobscura, D.virilisand D.mojavensis. Based on these distributions we have found that nearlyevery functional sequence category possesses its own distinctiveconservation pattern, sometimes independent of the overall sequenceconservation level. In the coding and regulatory regions, the ungappedblocks were longer than in introns, UTRs and non-functional sequences. Atthe same time, the blocks in the coding regions carried 3N+2 signaturecharacteristic to synonymic substitutions in the 3rd codon positions.Larger block sizes in transcription regulatory regions can be explainedby the presence of conserved arrays of binding sites for transcriptionfactors. We also have shown that the longest ungapped blocks, or'ultraconserved' sequences, are associated with specific gene groups,including those encoding ion channels and components of the cytoskeleton.We discussed how restrained conservation patterns may help in mappingfunctional sequence categories and improving genomeannotation.

  3. Small RNA-Seq analysis reveals microRNA-regulation of the Imd pathway during Escherichia coli infection in Drosophila.

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    Li, Shengjie; Shen, Li; Sun, Lianjie; Xu, Jiao; Jin, Ping; Chen, Liming; Ma, Fei

    2017-05-01

    Drosophila have served as a model for research on innate immunity for decades. However, knowledge of the post-transcriptional regulation of immune gene expression by microRNAs (miRNAs) remains rudimentary. In the present study, using small RNA-seq and bioinformatics analysis, we identified 67 differentially expressed miRNAs in Drosophila infected with Escherichia coli compared to injured flies at three time-points. Furthermore, we found that 21 of these miRNAs were potentially involved in the regulation of Imd pathway-related genes. Strikingly, based on UAS-miRNAs line screening and Dual-luciferase assay, we identified that miR-9a and miR-981 could both negatively regulate Drosophila antibacterial defenses and decrease the level of the antibacterial peptide, Diptericin. Taken together, these data support the involvement of miRNAs in the regulation of the Drosophila Imd pathway. Copyright © 2017 Elsevier Ltd. All rights reserved.

  4. Experimental evidence for nutrition regulated stress resistance in Drosophila ananassae.

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    Seema Sisodia

    Full Text Available The amount and quality of nutrients consumed by organisms have a strong impact on stress resistance, life-history traits and reproduction. The balance between energy acquisition and expenditure is crucial to the survival and reproductive success of animals. The ability of organisms to adjust their development, physiology or behavior in response to environmental conditions, called phenotypic plasticity, is a defining property of life. One of the most familiar and important examples of phenotypic plasticity is the response of stress tolerance and reproduction to changes in developmental nutrition. Larval nutrition may affect a range of different life-history traits as well as responses to environmental stress in adult.Here we investigate the effect of larval nutrition on desiccation, starvation, chill-coma recovery, heat resistance as well as egg to adult viability, egg production and ovariole number in Drosophila ananassae. We raised larvae on either protein rich diet or carbohydrate rich diet. We found that flies consuming protein rich diet have higher desiccation and heat shock resistance whereas flies developed on carbohydrate rich diet have higher starvation and cold resistance. Egg production was higher in females developed on protein rich diet and we also found trade-off between egg production and Egg to adult viability of the flies. Viability was higher in carbohydrate rich diet. However, sex specific viability was found in different nutritional regimes. Higher Egg production might be due to higher ovariole number in females of protein rich diet.Thus, Drosophila ananassae adapts different stress tolerance and life-history strategies according to the quality of the available diet, which are correlated with phenotypic adjustment at anatomical and physiological levels.

  5. Experimental evidence for nutrition regulated stress resistance in Drosophila ananassae.

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    Sisodia, Seema; Singh, Bashisth N

    2012-01-01

    The amount and quality of nutrients consumed by organisms have a strong impact on stress resistance, life-history traits and reproduction. The balance between energy acquisition and expenditure is crucial to the survival and reproductive success of animals. The ability of organisms to adjust their development, physiology or behavior in response to environmental conditions, called phenotypic plasticity, is a defining property of life. One of the most familiar and important examples of phenotypic plasticity is the response of stress tolerance and reproduction to changes in developmental nutrition. Larval nutrition may affect a range of different life-history traits as well as responses to environmental stress in adult. Here we investigate the effect of larval nutrition on desiccation, starvation, chill-coma recovery, heat resistance as well as egg to adult viability, egg production and ovariole number in Drosophila ananassae. We raised larvae on either protein rich diet or carbohydrate rich diet. We found that flies consuming protein rich diet have higher desiccation and heat shock resistance whereas flies developed on carbohydrate rich diet have higher starvation and cold resistance. Egg production was higher in females developed on protein rich diet and we also found trade-off between egg production and Egg to adult viability of the flies. Viability was higher in carbohydrate rich diet. However, sex specific viability was found in different nutritional regimes. Higher Egg production might be due to higher ovariole number in females of protein rich diet. Thus, Drosophila ananassae adapts different stress tolerance and life-history strategies according to the quality of the available diet, which are correlated with phenotypic adjustment at anatomical and physiological levels.

  6. Dystroglycan and mitochondrial ribosomal protein L34 regulate differentiation in the Drosophila eye.

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    Yougen Zhan

    2010-05-01

    Full Text Available Mutations that diminish the function of the extracellular matrix receptor Dystroglycan (DG result in muscular dystrophies, with associated neuronal migration defects in the brain and mental retardation e.g. Muscle Eye Brain Disease. To gain insight into the function of DG in the nervous system we initiated a study to examine its contribution to development of the eye of Drosophila melanogaster. Immuno-histochemistry showed that DG is concentrated on the apical surface of photoreceptors (R cells during specification of cell-fate in the third instar larva and is maintained at this location through early pupal stages. In point mutations that are null for DG we see abortive R cell elongation during differentiation that first appears in the pupa and results in stunted R cells in the adult. Overexpression of DG in R cells results in a small but significant increase in their size. R cell differentiation defects appear at the same stage in a deficiency line Df(2RDg(248 that affects Dg and the neighboring mitochondrial ribosomal gene, mRpL34. In the adult, these flies have severely disrupted R cells as well as defects in the lens and ommatidia. Expression of an mRpL34 transgene rescues much of this phenotype. We conclude that DG does not affect neuronal commitment but functions R cell autonomously to regulate neuronal elongation during differentiation in the pupa. We discuss these findings in view of recent work implicating DG as a regulator of cell metabolism and its genetic interaction with mRpL34, a member of a class of mitochondrial genes essential for normal metabolic function.

  7. The Hrs/Stam complex acts as a positive and negative regulator of RTK signaling during Drosophila development.

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    Hélène Chanut-Delalande

    Full Text Available BACKGROUND: Endocytosis is a key regulatory step of diverse signalling pathways, including receptor tyrosine kinase (RTK signalling. Hrs and Stam constitute the ESCRT-0 complex that controls the initial selection of ubiquitinated proteins, which will subsequently be degraded in lysosomes. It has been well established ex vivo and during Drosophila embryogenesis that Hrs promotes EGFR down regulation. We have recently isolated the first mutations of stam in flies and shown that Stam is required for air sac morphogenesis, a larval respiratory structure whose formation critically depends on finely tuned levels of FGFR activity. This suggest that Stam, putatively within the ESCRT-0 complex, modulates FGF signalling, a possibility that has not been examined in Drosophila yet. PRINCIPAL FINDINGS: Here, we assessed the role of the Hrs/Stam complex in the regulation of signalling activity during Drosophila development. We show that stam and hrs are required for efficient FGFR signalling in the tracheal system, both during cell migration in the air sac primordium and during the formation of fine cytoplasmic extensions in terminal cells. We find that stam and hrs mutant cells display altered FGFR/Btl localisation, likely contributing to impaired signalling levels. Electron microscopy analyses indicate that endosome maturation is impaired at distinct steps by hrs and stam mutations. These somewhat unexpected results prompted us to further explore the function of stam and hrs in EGFR signalling. We show that while stam and hrs together downregulate EGFR signalling in the embryo, they are required for full activation of EGFR signalling during wing development. CONCLUSIONS/SIGNIFICANCE: Our study shows that the ESCRT-0 complex differentially regulates RTK signalling, either positively or negatively depending on tissues and developmental stages, further highlighting the importance of endocytosis in modulating signalling pathways during development.

  8. DISCO interacting protein 2 determines direction of axon projection under the regulation of c-Jun N-terminal kinase in the Drosophila mushroom body

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    Nitta, Yohei; Sugie, Atsushi

    2017-01-01

    Precisely controlled axon guidance for complex neuronal wiring is essential for appropriate neuronal function. c-Jun N-terminal kinase (JNK) was found to play a role in axon guidance recently as well as in cell proliferation, protection and apoptosis. In spite of many genetic and molecular studies on these biological processes regulated by JNK, how JNK regulates axon guidance accurately has not been fully explained thus far. To address this question, we use the Drosophila mushroom body (MB) as a model since the α/β axons project in two distinct directions. Here we show that DISCO interacting protein 2 (DIP2) is required for the accurate direction of axonal guidance. DIP2 expression is under the regulation of Basket (Bsk), the Drosophila homologue of JNK. We additionally found that the Bsk/DIP2 pathway is independent from the AP-1 transcriptional factor complex pathway, which is directly activated by Bsk. In conclusion, our findings revealed DIP2 as a novel effector downstream of Bsk modulating the direction of axon projection. - Highlights: • DIP2 is required for accurate direction of axon guidance in Drosophila mushroom body. • DIP2 is a downstream of JNK in the axon guidance of Drosophila mushroom body neuron. • JNK/DIP2 pathway is independent from JNK/AP-1 transcriptional factor complex pathway.

  9. Wash functions downstream of Rho1 GTPase in a subset of Drosophila immune cell developmental migrations

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    Verboon, Jeffrey M.; Rahe, Travis K.; Rodriguez-Mesa, Evelyn; Parkhurst, Susan M.

    2015-01-01

    Drosophila immune cells, the hemocytes, undergo four stereotypical developmental migrations to populate the embryo, where they provide immune reconnoitering, as well as a number of non–immune-related functions necessary for proper embryogenesis. Here, we describe a role for Rho1 in one of these developmental migrations in which posteriorly located hemocytes migrate toward the head. This migration requires the interaction of Rho1 with its downstream effector Wash, a Wiskott–Aldrich syndrome family protein. Both Wash knockdown and a Rho1 transgene harboring a mutation that prevents Wash binding exhibit the same developmental migratory defect as Rho1 knockdown. Wash activates the Arp2/3 complex, whose activity is needed for this migration, whereas members of the WASH regulatory complex (SWIP, Strumpellin, and CCDC53) are not. Our results suggest a WASH complex–independent signaling pathway to regulate the cytoskeleton during a subset of hemocyte developmental migrations. PMID:25739458

  10. Mood stabilizing drugs regulate transcription of immune, neuronal and metabolic pathway genes in Drosophila.

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    Herteleer, L; Zwarts, L; Hens, K; Forero, D; Del-Favero, J; Callaerts, P

    2016-05-01

    Lithium and valproate (VPA) are drugs used in the management of bipolar disorder. Even though they reportedly act on various pathways, the transcriptional targets relevant for disease mechanism and therapeutic effect remain unclear. Furthermore, multiple studies used lymphoblasts of bipolar patients as a cellular proxy, but it remains unclear whether peripheral cells provide a good readout for the effects of these drugs in the brain. We used Drosophila culture cells and adult flies to analyze the transcriptional effects of lithium and VPA and define mechanistic pathways. Transcriptional profiles were determined for Drosophila S2-cells and adult fly heads following lithium or VPA treatment. Gene ontology categories were identified using the DAVID functional annotation tool with a cut-off of p neuronal development, neuronal function, and metabolism. (i) Transcriptional effects of lithium and VPA in Drosophila S2 cells and heads show significant overlap. (ii) The overlap between transcriptional alterations in peripheral versus neuronal cells at the single gene level is negligible, but at the gene ontology and pathway level considerable overlap can be found. (iii) Lithium and VPA act on evolutionarily conserved pathways in Drosophila and mammalian models.

  11. Dietary composition regulates Drosophila mobility and cardiac physiology

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    Bazzell, Brian; Ginzberg, Sara; Healy, Lindsey; Wessells, R. J.

    2013-01-01

    SUMMARY The impact of dietary composition on exercise capacity is a subject of intense study in both humans and model organisms. Interactions between diet and genetics are a crucial component of optimized dietary design. However, the genetic factors governing exercise response are still not well understood. The recent development of invertebrate models for endurance exercise is likely to facilitate study designs examining the conserved interactions between diet, exercise and genetics. As a first step, we used the Drosophila model to describe the effects of varying dietary composition on several physiological indices, including fatigue tolerance and climbing speed, cardiac performance, lipid storage and autophagy. We found that flies of two divergent genetic backgrounds optimize endurance and cardiac performance on relatively balanced low calorie diets. When flies are provided with unbalanced diets, diets higher in sugar than in yeast facilitate greater endurance at the expense of cardiac performance. Importantly, we found that dietary composition has a profound effect on various physiological indices, whereas total caloric intake per se has very little predictive value for performance. We also found that the effects of diet on endurance are completely reversible within 48 h if flies are switched to a different diet. PMID:23155082

  12. The histone deacetylase HDAC4 regulates long-term memory in Drosophila.

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    Helen L Fitzsimons

    Full Text Available A growing body of research indicates that pharmacological inhibition of histone deacetylases (HDACs correlates with enhancement of long-term memory and current research is concentrated on determining the roles that individual HDACs play in cognitive function. Here, we investigate the role of HDAC4 in long-term memory formation in Drosophila. We show that overexpression of HDAC4 in the adult mushroom body, an important structure for memory formation, resulted in a specific impairment in long-term courtship memory, but had no affect on short-term memory. Overexpression of an HDAC4 catalytic mutant also abolished LTM, suggesting a mode of action independent of catalytic activity. We found that overexpression of HDAC4 resulted in a redistribution of the transcription factor MEF2 from a relatively uniform distribution through the nucleus into punctate nuclear bodies, where it colocalized with HDAC4. As MEF2 has also been implicated in regulation of long-term memory, these data suggest that the repressive effects of HDAC4 on long-term memory may be through interaction with MEF2. In the same genetic background, we also found that RNAi-mediated knockdown of HDAC4 impairs long-term memory, therefore we demonstrate that HDAC4 is not only a repressor of long-term memory, but also modulates normal memory formation.

  13. The histone deacetylase HDAC4 regulates long-term memory in Drosophila.

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    Fitzsimons, Helen L; Schwartz, Silvia; Given, Fiona M; Scott, Maxwell J

    2013-01-01

    A growing body of research indicates that pharmacological inhibition of histone deacetylases (HDACs) correlates with enhancement of long-term memory and current research is concentrated on determining the roles that individual HDACs play in cognitive function. Here, we investigate the role of HDAC4 in long-term memory formation in Drosophila. We show that overexpression of HDAC4 in the adult mushroom body, an important structure for memory formation, resulted in a specific impairment in long-term courtship memory, but had no affect on short-term memory. Overexpression of an HDAC4 catalytic mutant also abolished LTM, suggesting a mode of action independent of catalytic activity. We found that overexpression of HDAC4 resulted in a redistribution of the transcription factor MEF2 from a relatively uniform distribution through the nucleus into punctate nuclear bodies, where it colocalized with HDAC4. As MEF2 has also been implicated in regulation of long-term memory, these data suggest that the repressive effects of HDAC4 on long-term memory may be through interaction with MEF2. In the same genetic background, we also found that RNAi-mediated knockdown of HDAC4 impairs long-term memory, therefore we demonstrate that HDAC4 is not only a repressor of long-term memory, but also modulates normal memory formation.

  14. Control of G1 in the developing Drosophila eye: rca1 regulates Cyclin A.

    Science.gov (United States)

    Dong, X; Zavitz, K H; Thomas, B J; Lin, M; Campbell, S; Zipursky, S L

    1997-01-01

    In the developing eye of Drosophila melanogaster, cells become synchronized in the G1 phase of the cell cycle just prior to the onset of cellular differentiation and morphogenesis. In roughex (rux) mutants, cells enter S phase precociously because of ectopic activation of a Cyclin A/Cdk complex in early G1. This leads to defects in cell fate and pattern formation, and results in abnormalities in the morphology of the adult eye. A screen for dominant suppressors of the rux eye phenotype led to the identification of mutations in cyclin A, string (cdc25), and new cell cycle genes. One of these genes, regulator of cyclin A (rca1), encodes a novel protein required for both mitotic and meiotic cell cycle progression. rca1 mutants arrest in G2 of embryonic cell cycle 16 with a phenotype very similar to cyclin A loss of function mutants. Expression of rca1 transgenes in G1 or in postmitotic neurons promotes Cyclin A protein accumulation and drives cells into S phase in a Cyclin A-dependent fashion.

  15. Compartmentalized Regulation of Parkin-Mediated Mitochondrial Quality Control in the Drosophila Nervous System In Vivo

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    Sung, Hyun; Tandarich, Lauren C.; Nguyen, Kenny

    2016-01-01

    In neurons, the normal distribution and selective removal of mitochondria are considered essential for maintaining the functions of the large asymmetric cell and its diverse compartments. Parkin, a E3 ubiquitin ligase associated with familial Parkinson's disease, has been implicated in mitochondrial dynamics and removal in cells including neurons. However, it is not clear how Parkin functions in mitochondrial turnover in vivo, or whether Parkin-dependent events of the mitochondrial life cycle occur in all neuronal compartments. Here, using the live Drosophila nervous system, we investigated the involvement of Parkin in mitochondrial dynamics, distribution, morphology, and removal. Contrary to our expectations, we found that Parkin-deficient animals do not accumulate senescent mitochondria in their motor axons or neuromuscular junctions; instead, they contain far fewer axonal mitochondria, and these displayed normal motility behavior, morphology, and metabolic state. However, the loss of Parkin did produce abnormal tubular and reticular mitochondria restricted to the motor cell bodies. In addition, in contrast to drug-treated, immortalized cells in vitro, mature motor neurons rarely displayed Parkin-dependent mitophagy. These data indicate that the cell body is the focus of Parkin-dependent mitochondrial quality control in neurons, and argue that a selection process allows only healthy mitochondria to pass from cell bodies to axons, perhaps to limit the impact of mitochondrial dysfunction. SIGNIFICANCE STATEMENT Parkin has been proposed to police mitochondrial fidelity by binding to dysfunctional mitochondria via PTEN (phosphatase and tensin homolog)-induced putative kinase 1 (PINK1) and targeting them for autophagic degradation. However, it is unknown whether and how the PINK1/Parkin pathway regulates the mitochondrial life cycle in neurons in vivo. Using Drosophila motor neurons, we show that parkin disruption generates an abnormal mitochondrial network in cell

  16. Dendritic Cytoskeletal Architecture Is Modulated by Combinatorial Transcriptional Regulation in Drosophila melanogaster.

    Science.gov (United States)

    Das, Ravi; Bhattacharjee, Shatabdi; Patel, Atit A; Harris, Jenna M; Bhattacharya, Surajit; Letcher, Jamin M; Clark, Sarah G; Nanda, Sumit; Iyer, Eswar Prasad R; Ascoli, Giorgio A; Cox, Daniel N

    2017-12-01

    Transcription factors (TFs) have emerged as essential cell autonomous mediators of subtype specific dendritogenesis; however, the downstream effectors of these TFs remain largely unknown, as are the cellular events that TFs control to direct morphological change. As dendritic morphology is largely dictated by the organization of the actin and microtubule (MT) cytoskeletons, elucidating TF-mediated cytoskeletal regulatory programs is key to understanding molecular control of diverse dendritic morphologies. Previous studies in Drosophila melanogaster have demonstrated that the conserved TFs Cut and Knot exert combinatorial control over aspects of dendritic cytoskeleton development, promoting actin and MT-based arbor morphology, respectively. To investigate transcriptional targets of Cut and/or Knot regulation, we conducted systematic neurogenomic studies, coupled with in vivo genetic screens utilizing multi-fluor cytoskeletal and membrane marker reporters. These analyses identified a host of putative Cut and/or Knot effector molecules, and a subset of these putative TF targets converge on modulating dendritic cytoskeletal architecture, which are grouped into three major phenotypic categories, based upon neuromorphometric analyses: complexity enhancer, complexity shifter, and complexity suppressor. Complexity enhancer genes normally function to promote higher order dendritic growth and branching with variable effects on MT stabilization and F-actin organization, whereas complexity shifter and complexity suppressor genes normally function in regulating proximal-distal branching distribution or in restricting higher order branching complexity, respectively, with spatially restricted impacts on the dendritic cytoskeleton. Collectively, we implicate novel genes and cellular programs by which TFs distinctly and combinatorially govern dendritogenesis via cytoskeletal modulation. Copyright © 2017 by the Genetics Society of America.

  17. Toll-8/Tollo negatively regulates antimicrobial response in the Drosophila respiratory epithelium.

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    Idir Akhouayri

    2011-10-01

    Full Text Available Barrier epithelia that are persistently exposed to microbes have evolved potent immune tools to eliminate such pathogens. If mechanisms that control Drosophila systemic responses are well-characterized, the epithelial immune responses remain poorly understood. Here, we performed a genetic dissection of the cascades activated during the immune response of the Drosophila airway epithelium i.e. trachea. We present evidence that bacteria induced-antimicrobial peptide (AMP production in the trachea is controlled by two signalling cascades. AMP gene transcription is activated by the inducible IMD pathway that acts non-cell autonomously in trachea. This IMD-dependent AMP activation is antagonized by a constitutively active signalling module involving the receptor Toll-8/Tollo, the ligand Spätzle2/DNT1 and Ect-4, the Drosophila ortholog of the human Sterile alpha and HEAT/ARMadillo motif (SARM. Our data show that, in addition to Toll-1 whose function is essential during the systemic immune response, Drosophila relies on another Toll family member to control the immune response in the respiratory epithelium.

  18. Sex Differences in Drosophila Somatic Gene Expression: Variation and Regulation by doublesex

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    Michelle N. Arbeitman

    2016-07-01

    Full Text Available Sex differences in gene expression have been widely studied in Drosophila melanogaster. Sex differences vary across strains, but many molecular studies focus on only a single strain, or on genes that show sexually dimorphic expression in many strains. How extensive variability is and whether this variability occurs among genes regulated by sex determination hierarchy terminal transcription factors is unknown. To address these questions, we examine differences in sexually dimorphic gene expression between two strains in Drosophila adult head tissues. We also examine gene expression in doublesex (dsx mutant strains to determine which sex-differentially expressed genes are regulated by DSX, and the mode by which DSX regulates expression. We find substantial variation in sex-differential expression. The sets of genes with sexually dimorphic expression in each strain show little overlap. The prevalence of different DSX regulatory modes also varies between the two strains. Neither the patterns of DSX DNA occupancy, nor mode of DSX regulation explain why some genes show consistent sex-differential expression across strains. We find that the genes identified as regulated by DSX in this study are enriched with known sites of DSX DNA occupancy. Finally, we find that sex-differentially expressed genes and genes regulated by DSX are highly enriched on the fourth chromosome. These results provide insights into a more complete pool of potential DSX targets, as well as revealing the molecular flexibility of DSX regulation.

  19. The Drosophila T-box transcription factor Midline functions within the Notch–Delta signaling pathway to specify sensory organ precursor cell fates and regulates cell survival within the eye imaginal disc

    Science.gov (United States)

    Das, Sudeshna; Chen, Q. Brent; Saucier, Joseph D.; Drescher, Brandon; Zong, Yan; Morgan, Sarah; Forstall, John; Meriwether, Andrew; Toranzo, Randy; Leal, Sandra M.

    2014-01-01

    We report that the T-box transcription factor Midline (Mid), an evolutionary conserved homolog of the vertebrate Tbx20 protein, functions within the Notch–Delta signaling pathway essential for specifying the fates of sensory organ precursor cells. This complements an established history of research showing that Mid regulates the cell-fate specification of diverse cell types within the developing heart, epidermis and central nervous system. Tbx20 has been detected in diverse neuronal and epithelial cells of embryonic eye tissues in both mice and humans. However, the mechanisms by which either Mid or Tbx20 function to regulate cell-fate specification or other critical aspects of eye development including cell survival have not yet been elucidated. We have also gathered preliminary evidence suggesting that Mid may play an indirect, but vital role in selecting SOP cells within the third-instar larval eye disc by regulating the expression of the proneural gene atonal. During subsequent pupal stages, Mid specifies SOP cell fates as a member of the Notch–Delta signaling hierarchy and is essential for maintaining cell viability within by inhibiting apoptotic pathways. We present several new hypotheses that seek to understand the role of Mid in regulating developmental processes downstream of the Notch receptor that are critical for specifying unique cell fates, patterning the adult eye and maintaining cellular homeostasis during eye disc morphogenesis. PMID:23962751

  20. The Drosophila T-box transcription factor Midline functions within the Notch-Delta signaling pathway to specify sensory organ precursor cell fates and regulates cell survival within the eye imaginal disc.

    Science.gov (United States)

    Das, Sudeshna; Chen, Q Brent; Saucier, Joseph D; Drescher, Brandon; Zong, Yan; Morgan, Sarah; Forstall, John; Meriwether, Andrew; Toranzo, Randy; Leal, Sandra M

    2013-01-01

    We report that the T-box transcription factor Midline (Mid), an evolutionary conserved homolog of the vertebrate Tbx20 protein, functions within the Notch-Delta signaling pathway essential for specifying the fates of sensory organ precursor (SOP) cells. These findings complement an established history of research showing that Mid regulates the cell-fate specification of diverse cell types within the developing heart, epidermis and central nervous system. Tbx20 has been detected in unique neuronal and epithelial cells of embryonic eye tissues in both mice and humans. However, the mechanisms by which either Mid or Tbx20 function to regulate cell-fate specification or other critical aspects of eye development including cell survival have not yet been elucidated. We have also gathered preliminary evidence suggesting that Mid may play an indirect, but vital role in selecting SOP cells within the third-instar larval eye disc by regulating the expression of the proneural gene atonal. During subsequent pupal stages, Mid specifies SOP cell fates as a member of the Notch-Delta signaling hierarchy and is essential for maintaining cell viability by inhibiting apoptotic pathways. We present several new hypotheses that seek to understand the role of Mid in regulating developmental processes downstream of the Notch receptor that are critical for specifying unique cell fates, patterning the adult eye and maintaining cellular homeostasis during eye disc morphogenesis. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  1. miR-11 regulates pupal size of Drosophila melanogaster via directly targeting Ras85D.

    Science.gov (United States)

    Li, Yao; Li, Shengjie; Jin, Ping; Chen, Liming; Ma, Fei

    2017-01-01

    MicroRNAs play diverse roles in various physiological processes during Drosophila development. In the present study, we reported that miR-11 regulates pupal size during Drosophila metamorphosis via targeting Ras85D with the following evidences: pupal size was increased in the miR-11 deletion mutant; restoration of miR-11 in the miR-11 deletion mutant rescued the increased pupal size phenotype observed in the miR-11 deletion mutant; ectopic expression of miR-11 in brain insulin-producing cells (IPCs) and whole body shows consistent alteration of pupal size; Dilps and Ras85D expressions were negatively regulated by miR-11 in vivo; miR-11 targets Ras85D through directly binding to Ras85D 3'-untranslated region in vitro; removal of one copy of Ras85D in the miR-11 deletion mutant rescued the increased pupal size phenotype observed in the miR-11 deletion mutant. Thus, our current work provides a novel mechanism of pupal size determination by microRNAs during Drosophila melanogaster metamorphosis. Copyright © 2017 the American Physiological Society.

  2. Drosophila Insulin receptor regulates the persistence of injury-induced nociceptive sensitization

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    Patel, Atit A.

    2018-01-01

    ABSTRACT Diabetes-associated nociceptive hypersensitivity affects diabetic patients with hard-to-treat chronic pain. Because multiple tissues are affected by systemic alterations in insulin signaling, the functional locus of insulin signaling in diabetes-associated hypersensitivity remains obscure. Here, we used Drosophila nociception/nociceptive sensitization assays to investigate the role of Insulin receptor (Insulin-like receptor, InR) in nociceptive hypersensitivity. InR mutant larvae exhibited mostly normal baseline thermal nociception (absence of injury) and normal acute thermal hypersensitivity following UV-induced injury. However, their acute thermal hypersensitivity persists and fails to return to baseline, unlike in controls. Remarkably, injury-induced persistent hypersensitivity is also observed in larvae that exhibit either type 1 or type 2 diabetes. Cell type-specific genetic analysis indicates that InR function is required in multidendritic sensory neurons including nociceptive class IV neurons. In these same nociceptive sensory neurons, only modest changes in dendritic morphology were observed in the InRRNAi-expressing and diabetic larvae. At the cellular level, InR-deficient nociceptive sensory neurons show elevated calcium responses after injury. Sensory neuron-specific expression of InR rescues the persistent thermal hypersensitivity of InR mutants and constitutive activation of InR in sensory neurons ameliorates the hypersensitivity observed with a type 2-like diabetic state. Our results suggest that a sensory neuron-specific function of InR regulates the persistence of injury-associated hypersensitivity. It is likely that this new system will be an informative genetically tractable model of diabetes-associated hypersensitivity. PMID:29752280

  3. The serotonergic central nervous system of the Drosophila larva: anatomy and behavioral function.

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    Annina Huser

    Full Text Available The Drosophila larva has turned into a particularly simple model system for studying the neuronal basis of innate behaviors and higher brain functions. Neuronal networks involved in olfaction, gustation, vision and learning and memory have been described during the last decade, often up to the single-cell level. Thus, most of these sensory networks are substantially defined, from the sensory level up to third-order neurons. This is especially true for the olfactory system of the larva. Given the wealth of genetic tools in Drosophila it is now possible to address the question how modulatory systems interfere with sensory systems and affect learning and memory. Here we focus on the serotonergic system that was shown to be involved in mammalian and insect sensory perception as well as learning and memory. Larval studies suggested that the serotonergic system is involved in the modulation of olfaction, feeding, vision and heart rate regulation. In a dual anatomical and behavioral approach we describe the basic anatomy of the larval serotonergic system, down to the single-cell level. In parallel, by expressing apoptosis-inducing genes during embryonic and larval development, we ablate most of the serotonergic neurons within the larval central nervous system. When testing these animals for naïve odor, sugar, salt and light perception, no profound phenotype was detectable; even appetitive and aversive learning was normal. Our results provide the first comprehensive description of the neuronal network of the larval serotonergic system. Moreover, they suggest that serotonin per se is not necessary for any of the behaviors tested. However, our data do not exclude that this system may modulate or fine-tune a wide set of behaviors, similar to its reported function in other insect species or in mammals. Based on our observations and the availability of a wide variety of genetic tools, this issue can now be addressed.

  4. Reading the maps: Organization and function of chromatin types in Drosophila

    NARCIS (Netherlands)

    Braunschweig, U.

    2010-01-01

    The work presented in this thesis shows that the Drosophila genome is organized in chromatin domains with many implications for gene regulation, nuclear organization, and evolution. Furthermore it provides examples of how maps of chromatin protein binding, combined with computational approaches, can

  5. Ligand binding pocket function of drosophila USP is necessary for metamorphosis

    Science.gov (United States)

    The widely accepted paradigm that epoxidized methyl farnesoates (“juvenile hormones,” JHs) are the principle sesquiterpenoid hormones regulating insect metamorphosis was assessed in Drosophila melanogaster. GC-MS analysis showed that methyl farnesoate, rather than methyl epoxyfarnesoate (= JH III), ...

  6. Astrocyte-specific regulation of hMeCP2 expression in Drosophila

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    David L. Hess-Homeier

    2014-10-01

    Full Text Available Alterations in the expression of Methyl-CpG-binding protein 2 (MeCP2 either by mutations or gene duplication leads to a wide spectrum of neurodevelopmental disorders including Rett Syndrome and MeCP2 duplication disorder. Common features of Rett Syndrome (RTT, MeCP2 duplication disorder, and neuropsychiatric disorders indicate that even moderate changes in MeCP2 protein levels result in functional and structural cell abnormalities. In this study, we investigated two areas of MeCP2 pathophysiology using Drosophila as a model system: the effects of MeCP2 glial gain-of-function activity on circuits controlling sleep behavior, and the cell-type specific regulation of MeCP2 expression. In this study, we first examined the effects of elevated MeCP2 levels on microcircuits by expressing human MeCP2 (hMeCP2 in astrocytes and distinct subsets of amine neurons including dopamine and octopamine (OA neurons. Depending on the cell-type, hMeCP2 expression reduced sleep levels, altered daytime/nighttime sleep patterns, and generated sleep maintenance deficits. Second, we identified a 498 base pair region of the MeCP2e2 isoform that is targeted for regulation in distinct subsets of astrocytes. Levels of the full-length hMeCP2e2 and mutant RTT R106W protein decreased in astrocytes in a temporally and spatially regulated manner. In contrast, expression of the deletion Δ166 hMeCP2 protein was not altered in the entire astrocyte population. qPCR experiments revealed a reduction in full-length hMeCP2e2 transcript levels suggesting transgenic hMeCP2 expression is regulated at the transcriptional level. Given the phenotypic complexities that are caused by alterations in MeCP2 levels, our results provide insight into distinct cellular mechanisms that control MeCP2 expression and link microcircuit abnormalities with defined behavioral deficits.

  7. Functional Interaction between HEXIM and Hedgehog Signaling during Drosophila Wing Development.

    Science.gov (United States)

    Nguyen, Duy; Fayol, Olivier; Buisine, Nicolas; Lecorre, Pierrette; Uguen, Patricia

    2016-01-01

    Studying the dynamic of gene regulatory networks is essential in order to understand the specific signals and factors that govern cell proliferation and differentiation during development. This also has direct implication in human health and cancer biology. The general transcriptional elongation regulator P-TEFb regulates the transcriptional status of many developmental genes. Its biological activity is controlled by an inhibitory complex composed of HEXIM and the 7SK snRNA. Here, we examine the function of HEXIM during Drosophila development. Our key finding is that HEXIM affects the Hedgehog signaling pathway. HEXIM knockdown flies display strong phenotypes and organ failures. In the wing imaginal disc, HEXIM knockdown initially induces ectopic expression of Hedgehog (Hh) and its transcriptional effector Cubitus interuptus (Ci). In turn, deregulated Hedgehog signaling provokes apoptosis, which is continuously compensated by apoptosis-induced cell proliferation. Thus, the HEXIM knockdown mutant phenotype does not result from the apoptotic ablation of imaginal disc; but rather from the failure of dividing cells to commit to a proper developmental program due to Hedgehog signaling defects. Furthermore, we show that ci is a genetic suppressor of hexim. Thus, HEXIM ensures the integrity of Hedgehog signaling in wing imaginal disc, by a yet unknown mechanism. To our knowledge, this is the first time that the physiological function of HEXIM has been addressed in such details in vivo.

  8. Functional Interaction between HEXIM and Hedgehog Signaling during Drosophila Wing Development.

    Directory of Open Access Journals (Sweden)

    Duy Nguyen

    Full Text Available Studying the dynamic of gene regulatory networks is essential in order to understand the specific signals and factors that govern cell proliferation and differentiation during development. This also has direct implication in human health and cancer biology. The general transcriptional elongation regulator P-TEFb regulates the transcriptional status of many developmental genes. Its biological activity is controlled by an inhibitory complex composed of HEXIM and the 7SK snRNA. Here, we examine the function of HEXIM during Drosophila development. Our key finding is that HEXIM affects the Hedgehog signaling pathway. HEXIM knockdown flies display strong phenotypes and organ failures. In the wing imaginal disc, HEXIM knockdown initially induces ectopic expression of Hedgehog (Hh and its transcriptional effector Cubitus interuptus (Ci. In turn, deregulated Hedgehog signaling provokes apoptosis, which is continuously compensated by apoptosis-induced cell proliferation. Thus, the HEXIM knockdown mutant phenotype does not result from the apoptotic ablation of imaginal disc; but rather from the failure of dividing cells to commit to a proper developmental program due to Hedgehog signaling defects. Furthermore, we show that ci is a genetic suppressor of hexim. Thus, HEXIM ensures the integrity of Hedgehog signaling in wing imaginal disc, by a yet unknown mechanism. To our knowledge, this is the first time that the physiological function of HEXIM has been addressed in such details in vivo.

  9. BMPs regulate msx gene expression in the dorsal neuroectoderm of Drosophila and vertebrates by distinct mechanisms.

    Science.gov (United States)

    Esteves, Francisco F; Springhorn, Alexander; Kague, Erika; Taylor, Erika; Pyrowolakis, George; Fisher, Shannon; Bier, Ethan

    2014-09-01

    In a broad variety of bilaterian species the trunk central nervous system (CNS) derives from three primary rows of neuroblasts. The fates of these neural progenitor cells are determined in part by three conserved transcription factors: vnd/nkx2.2, ind/gsh and msh/msx in Drosophila melanogaster/vertebrates, which are expressed in corresponding non-overlapping patterns along the dorsal-ventral axis. While this conserved suite of "neural identity" gene expression strongly suggests a common ancestral origin for the patterning systems, it is unclear whether the original regulatory mechanisms establishing these patterns have been similarly conserved during evolution. In Drosophila, genetic evidence suggests that Bone Morphogenetic Proteins (BMPs) act in a dosage-dependent fashion to repress expression of neural identity genes. BMPs also play a dose-dependent role in patterning the dorsal and lateral regions of the vertebrate CNS, however, the mechanism by which they achieve such patterning has not yet been clearly established. In this report, we examine the mechanisms by which BMPs act on cis-regulatory modules (CRMs) that control localized expression of the Drosophila msh and zebrafish (Danio rerio) msxB in the dorsal central nervous system (CNS). Our analysis suggests that BMPs act differently in these organisms to regulate similar patterns of gene expression in the neuroectoderm: repressing msh expression in Drosophila, while activating msxB expression in the zebrafish. These findings suggest that the mechanisms by which the BMP gradient patterns the dorsal neuroectoderm have reversed since the divergence of these two ancient lineages.

  10. BMPs regulate msx gene expression in the dorsal neuroectoderm of Drosophila and vertebrates by distinct mechanisms.

    Directory of Open Access Journals (Sweden)

    Francisco F Esteves

    2014-09-01

    Full Text Available In a broad variety of bilaterian species the trunk central nervous system (CNS derives from three primary rows of neuroblasts. The fates of these neural progenitor cells are determined in part by three conserved transcription factors: vnd/nkx2.2, ind/gsh and msh/msx in Drosophila melanogaster/vertebrates, which are expressed in corresponding non-overlapping patterns along the dorsal-ventral axis. While this conserved suite of "neural identity" gene expression strongly suggests a common ancestral origin for the patterning systems, it is unclear whether the original regulatory mechanisms establishing these patterns have been similarly conserved during evolution. In Drosophila, genetic evidence suggests that Bone Morphogenetic Proteins (BMPs act in a dosage-dependent fashion to repress expression of neural identity genes. BMPs also play a dose-dependent role in patterning the dorsal and lateral regions of the vertebrate CNS, however, the mechanism by which they achieve such patterning has not yet been clearly established. In this report, we examine the mechanisms by which BMPs act on cis-regulatory modules (CRMs that control localized expression of the Drosophila msh and zebrafish (Danio rerio msxB in the dorsal central nervous system (CNS. Our analysis suggests that BMPs act differently in these organisms to regulate similar patterns of gene expression in the neuroectoderm: repressing msh expression in Drosophila, while activating msxB expression in the zebrafish. These findings suggest that the mechanisms by which the BMP gradient patterns the dorsal neuroectoderm have reversed since the divergence of these two ancient lineages.

  11. ERK phosphorylation regulates sleep and plasticity in Drosophila.

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    William M Vanderheyden

    Full Text Available Given the relationship between sleep and plasticity, we examined the role of Extracellular signal-regulated kinase (ERK in regulating baseline sleep, and modulating the response to waking experience. Both sleep deprivation and social enrichment increase ERK phosphorylation in wild-type flies. The effects of both sleep deprivation and social enrichment on structural plasticity in the LNvs can be recapitulated by expressing an active version of ERK (UAS-ERK(SEM pan-neuronally in the adult fly using GeneSwitch (Gsw Gsw-elav-GAL4. Conversely, disrupting ERK reduces sleep and prevents both the behavioral and structural plasticity normally induced by social enrichment. Finally, using transgenic flies carrying a cAMP response Element (CRE-luciferase reporter we show that activating ERK enhances CRE-Luc activity while disrupting ERK reduces it. These data suggest that ERK phosphorylation is an important mediator in transducing waking experience into sleep.

  12. De novo loss-of-function mutations in WAC cause a recognizable intellectual disability syndrome and learning deficits in Drosophila.

    Science.gov (United States)

    Lugtenberg, Dorien; Reijnders, Margot R F; Fenckova, Michaela; Bijlsma, Emilia K; Bernier, Raphael; van Bon, Bregje W M; Smeets, Eric; Vulto-van Silfhout, Anneke T; Bosch, Danielle; Eichler, Evan E; Mefford, Heather C; Carvill, Gemma L; Bongers, Ernie M H F; Schuurs-Hoeijmakers, Janneke Hm; Ruivenkamp, Claudia A; Santen, Gijs W E; van den Maagdenberg, Arn M J M; Peeters-Scholte, Cacha M P C D; Kuenen, Sabine; Verstreken, Patrik; Pfundt, Rolph; Yntema, Helger G; de Vries, Petra F; Veltman, Joris A; Hoischen, Alexander; Gilissen, Christian; de Vries, Bert B A; Schenck, Annette; Kleefstra, Tjitske; Vissers, Lisenka E L M

    2016-08-01

    Recently WAC was reported as a candidate gene for intellectual disability (ID) based on the identification of a de novo mutation in an individual with severe ID. WAC regulates transcription-coupled histone H2B ubiquitination and has previously been implicated in the 10p12p11 contiguous gene deletion syndrome. In this study, we report on 10 individuals with de novo WAC mutations which we identified through routine (diagnostic) exome sequencing and targeted resequencing of WAC in 2326 individuals with unexplained ID. All but one mutation was expected to lead to a loss-of-function of WAC. Clinical evaluation of all individuals revealed phenotypic overlap for mild ID, hypotonia, behavioral problems and distinctive facial dysmorphisms, including a square-shaped face, deep set eyes, long palpebral fissures, and a broad mouth and chin. These clinical features were also previously reported in individuals with 10p12p11 microdeletion syndrome. To investigate the role of WAC in ID, we studied the importance of the Drosophila WAC orthologue (CG8949) in habituation, a non-associative learning paradigm. Neuronal knockdown of Drosophila CG8949 resulted in impaired learning, suggesting that WAC is required in neurons for normal cognitive performance. In conclusion, we defined a clinically recognizable ID syndrome, caused by de novo loss-of-function mutations in WAC. Independent functional evidence in Drosophila further supported the role of WAC in ID. On the basis of our data WAC can be added to the list of ID genes with a role in transcription regulation through histone modification.

  13. Functional characterization of dopamine transporter in vivo using Drosophila melanogaster behavioral analysis.

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    Taro eUeno

    2014-09-01

    Full Text Available Dopamine mediates diverse functions such as motivation, reward, attention, learning/memory and sleep/arousal. Recent studies using model organisms including the fruit fly, have elucidated various physiological functions of dopamine, and identified specific neural circuits for these functions. Flies with mutations in the Drosophila dopamine transporter (dDAT gene show enhanced dopamine signaling, and short sleep and memory impairment phenotypes. However, understanding the mechanism by which dopamine signaling causes these phenotypes requires an understanding of the dynamics of dopamine release. Here we report the effects of dDAT expression on behavioral traits. We show that dDAT expression in a subset of dopaminergic neurons is sufficient for normal sleep. dDAT expression in other cell types such as Kenyon cells and glial cells can also rescue the short sleep phenotype of dDAT mutants. dDAT mutants also show a down-regulation of the D1-like dopamine receptor dDA1, and this phenotype is rescued when dDAT is expressed in the same cell types in which it rescues sleep. On the other hand, dDAT overexpression in mushroom bodies, which are the target of memory forming dopamine neurons, abolishes olfactory aversive memory. Our data demonstrate that expression of extrasynaptic dopamine transporters can rescue some aspects of dopamine signaling in dopamine transporter mutants. These results provide novel insights into regulatory systems that modulate dopamine signaling.

  14. Necessity of angiotensin-converting enzyme-related gene for cardiac functions and longevity of Drosophila melanogaster assessed by optical coherence tomography

    Science.gov (United States)

    Liao, Fang-Tsu; Chang, Cheng-Yi; Su, Ming-Tsan; Kuo, Wen-Chuan

    2014-01-01

    Prior studies have established the necessity of an angiotensin-converting enzyme-related (ACER) gene for heart morphogenesis of Drosophila. Nevertheless, the physiology of ACER has yet to be comprehensively understood. Herein, we employed RNA interference to down-regulate the expression of ACER in Drosophila's heart and swept source optical coherence tomography to assess whether ACER is required for cardiac functions in living adult flies. Several contractile parameters of Drosophila heart, including the heart rate (HR), end-diastolic diameter (EDD), end-systolic diameter (ESD), percent fractional shortening (%FS), and stress-induced cardiac performance, are shown, which are age dependent. These age-dependent cardiac functions declined significantly when ACER was down-regulated. Moreover, the lifespans of ACER knock-down flies were significantly shorter than those of wild-type control flies. Thus, we posit that ACER, the Drosophila ortholog of mammalian angiotensin-converting enzyme 2 (ACE2), is essential for both heart physiology and longevity of animals. Since mammalian ACE2 controls many cardiovascular physiological features and is implicated in cardiomyopathies, our findings that ACER plays conserved roles in genetically tractable animals will pave the way for uncovering the genetic pathway that controls the renin-angiotensin system.

  15. Analysis of synaptic growth and function in Drosophila with an extended larval stage.

    Science.gov (United States)

    Miller, Daniel L; Ballard, Shannon L; Ganetzky, Barry

    2012-10-03

    The Drosophila larval neuromuscular junction (NMJ) is a powerful system for the genetic and molecular analysis of neuronal excitability, synaptic transmission, and synaptic development. However, its use for studying age-dependent processes, such as maintenance of neuronal viability and synaptic stability, are temporally limited by the onset of pupariation and metamorphosis. Here we characterize larval NMJ growth, growth regulation, structure, and function in a developmental variant with an extended third instar (ETI). RNAi-knockdown of the prothoracicotropic hormone receptor, torso, in the ring gland of developing larvae leaves the timing of first and second instar molts largely unchanged, but triples duration of the third instar from 3 to 9.5 d (McBrayer et al., 2007; Rewitz et al., 2009). During this ETI period, NMJs undergo additional growth (adding >50 boutons/NMJ), and this growth remains under the control of the canonical regulators Highwire and the TGFβ/BMP pathway. NMJ growth during the ETI period occurs via addition of new branches, satellite boutons, and interstitial boutons, and continues even after muscle growth levels off. Throughout the ETI, organization of synapses and active zones remains normal, and synaptic transmission is unchanged. These results establish the ETI larval system as a viable model for studying motor neuron diseases and for investigating time-dependent effects of perturbations that impair mechanisms of neuroprotection, synaptic maintenance, and response to neural injury.

  16. Molecular cloning, functional expression, and gene silencing of two Drosophila receptors for the Drosophila neuropeptide pyrokinin-2

    DEFF Research Database (Denmark)

    Rosenkilde, Carina; Cazzamali, Giuseppe; Williamson, Michael

    2003-01-01

    The database of the Drosophila Genome Project contains the sequences of two genes, CG8784 and CG8795, predicted to code for two structurally related G protein-coupled receptors. We have cloned these genes and expressed their coding parts in Chinese hamster ovary cells. We found that both receptors...... can be activated by low concentrations of the Drosophila neuropeptide pyrokinin-2 (CG8784, EC(50) for pyrokinin-2, 1x10(-9)M; CG8795, EC(50) for pyrokinin-2, 5 x 10(-10)M). The precise role of Drosophila pyrokinin-2 (SVPFKPRLamide) in Drosophila is unknown, but in other insects, pyrokinins have...... embryos and first instar larvae. In addition to the two Drosophila receptors, we also identified two probable pyrokinin receptors in the genomic database from the malaria mosquito Anopheles gambiae. The two Drosophila pyrokinin receptors are, to our knowledge, the first invertebrate pyrokinin receptors...

  17. The Atg1-Tor pathway regulates yolk catabolism in Drosophila embryos.

    Science.gov (United States)

    Kuhn, Hallie; Sopko, Richelle; Coughlin, Margaret; Perrimon, Norbert; Mitchison, Tim

    2015-11-15

    Yolk provides an important source of nutrients during the early development of oviparous organisms. It is composed mainly of vitellogenin proteins packed into membrane-bound compartments called yolk platelets. Catabolism of yolk is initiated by acidification of the yolk platelet, leading to the activation of Cathepsin-like proteinases, but it is unknown how this process is triggered. Yolk catabolism initiates at cellularization in Drosophila melanogaster embryos. Using maternal shRNA technology we found that yolk catabolism depends on the Tor pathway and on the autophagy-initiating kinase Atg1. Whereas Atg1 was required for a burst of spatially regulated autophagy during late cellularization, autophagy was not required for initiating yolk catabolism. We propose that the conserved Tor metabolic sensing pathway regulates yolk catabolism, similar to Tor-dependent metabolic regulation on the lysosome. © 2015. Published by The Company of Biologists Ltd.

  18. The Drosophila cell adhesion molecule Neuroglian regulates Lissencephaly-1 localisation in circulating immunosurveillance cells

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    Williams Michael J

    2009-03-01

    Full Text Available Abstract Background When the parasitoid wasp Leptopilina boulardi lays its eggs in Drosophila larvae phagocytic cells called plasmatocytes and specialized cells known as lamellocytes encapsulate the egg. This requires these circulating immunosurveillance cells (haemocytes to change from a non-adhesive to an adhesive state enabling them to bind to the invader. Interestingly, attachment of leukocytes, platelets, and insect haemocytes requires the same adhesion complexes as epithelial and neuronal cells. Results Here evidence is presented showing that the Drosophila L1-type cell adhesion molecule Neuroglian (Nrg is required for haemocytes to encapsulate L. boulardi wasp eggs. The amino acid sequence FIGQY containing a conserved phosphorylated tyrosine is found in the intracellular domain of all L1-type cell adhesion molecules. This conserved tyrosine is phosphorylated at the cell periphery of plasmatocytes and lamellocytes prior to parasitisation, but dephosphorylated after immune activation. Intriguingly, another pool of Nrg located near the nucleus of plasmatocytes remains phosphorylated after parasitisation. In mammalian neuronal cells phosphorylated neurofascin, another L1-type cell adhesion molecule interacts with a nucleokinesis complex containing the microtubule binding protein lissencephaly-1 (Lis1 1. Interestingly in plasmatocytes from Nrg mutants the nucleokinesis regulating protein Lissencephaly-1 (Lis1 fails to localise properly around the nucleus and is instead found diffuse throughout the cytoplasm and at unidentified perinuclear structures. After attaching to the wasp egg control plasmatocytes extend filopodia laterally from their cell periphery; as well as extending lateral filopodia plasmatocytes from Nrg mutants also extend many filopodia from their apical surface. Conclusion The Drosophila cellular adhesion molecule Neuroglian is expressed in haemocytes and its activity is required for the encapsulation of L. boularli eggs. At

  19. Phospholipid Homeostasis Regulates Dendrite Morphogenesis in Drosophila Sensory Neurons

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    Shan Meltzer

    2017-10-01

    Full Text Available Disruptions in lipid homeostasis have been observed in many neurodevelopmental disorders that are associated with dendrite morphogenesis defects. However, the molecular mechanisms of how lipid homeostasis affects dendrite morphogenesis are unclear. We find that easily shocked (eas, which encodes a kinase with a critical role in phospholipid phosphatidylethanolamine (PE synthesis, and two other enzymes in this synthesis pathway are required cell autonomously in sensory neurons for dendrite growth and stability. Furthermore, we show that the level of Sterol Regulatory Element-Binding Protein (SREBP activity is important for dendrite development. SREBP activity increases in eas mutants, and decreasing the level of SREBP and its transcriptional targets in eas mutants largely suppresses the dendrite growth defects. Furthermore, reducing Ca2+ influx in neurons of eas mutants ameliorates the dendrite morphogenesis defects. Our study uncovers a role for EAS kinase and reveals the in vivo function of phospholipid homeostasis in dendrite morphogenesis.

  20. Class IIa histone deacetylases are conserved regulators of circadian function.

    Science.gov (United States)

    Fogg, Paul C M; O'Neill, John S; Dobrzycki, Tomasz; Calvert, Shaun; Lord, Emma C; McIntosh, Rebecca L L; Elliott, Christopher J H; Sweeney, Sean T; Hastings, Michael H; Chawla, Sangeeta

    2014-12-05

    Class IIa histone deacetylases (HDACs) regulate the activity of many transcription factors to influence liver gluconeogenesis and the development of specialized cells, including muscle, neurons, and lymphocytes. Here, we describe a conserved role for class IIa HDACs in sustaining robust circadian behavioral rhythms in Drosophila and cellular rhythms in mammalian cells. In mouse fibroblasts, overexpression of HDAC5 severely disrupts transcriptional rhythms of core clock genes. HDAC5 overexpression decreases BMAL1 acetylation on Lys-537 and pharmacological inhibition of class IIa HDACs increases BMAL1 acetylation. Furthermore, we observe cyclical nucleocytoplasmic shuttling of HDAC5 in mouse fibroblasts that is characteristically circadian. Mutation of the Drosophila homolog HDAC4 impairs locomotor activity rhythms of flies and decreases period mRNA levels. RNAi-mediated knockdown of HDAC4 in Drosophila clock cells also dampens circadian function. Given that the localization of class IIa HDACs is signal-regulated and influenced by Ca(2+) and cAMP signals, our findings offer a mechanism by which extracellular stimuli that generate these signals can feed into the molecular clock machinery. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  1. Functional evolution of cis-regulatory modules at a homeotic gene in Drosophila.

    Directory of Open Access Journals (Sweden)

    Margaret C W Ho

    2009-11-01

    Full Text Available It is a long-held belief in evolutionary biology that the rate of molecular evolution for a given DNA sequence is inversely related to the level of functional constraint. This belief holds true for the protein-coding homeotic (Hox genes originally discovered in Drosophila melanogaster. Expression of the Hox genes in Drosophila embryos is essential for body patterning and is controlled by an extensive array of cis-regulatory modules (CRMs. How the regulatory modules functionally evolve in different species is not clear. A comparison of the CRMs for the Abdominal-B gene from different Drosophila species reveals relatively low levels of overall sequence conservation. However, embryonic enhancer CRMs from other Drosophila species direct transgenic reporter gene expression in the same spatial and temporal patterns during development as their D. melanogaster orthologs. Bioinformatic analysis reveals the presence of short conserved sequences within defined CRMs, representing gap and pair-rule transcription factor binding sites. One predicted binding site for the gap transcription factor KRUPPEL in the IAB5 CRM was found to be altered in Superabdominal (Sab mutations. In Sab mutant flies, the third abdominal segment is transformed into a copy of the fifth abdominal segment. A model for KRUPPEL-mediated repression at this binding site is presented. These findings challenge our current understanding of the relationship between sequence evolution at the molecular level and functional activity of a CRM. While the overall sequence conservation at Drosophila CRMs is not distinctive from neighboring genomic regions, functionally critical transcription factor binding sites within embryonic enhancer CRMs are highly conserved. These results have implications for understanding mechanisms of gene expression during embryonic development, enhancer function, and the molecular evolution of eukaryotic regulatory modules.

  2. Disruption of Lysosome Function Promotes Tumor Growth and Metastasis in Drosophila *

    OpenAIRE

    Chi, Congwu; Zhu, Huanhu; Han, Min; Zhuang, Yuan; Wu, Xiaohui; Xu, Tian

    2010-01-01

    Lysosome function is essential to many physiological processes. It has been suggested that deregulation of lysosome function could contribute to cancer. Through a genetic screen in Drosophila, we have discovered that mutations disrupting lysosomal degradation pathway components contribute to tumor development and progression. Loss-of-function mutations in the Class C vacuolar protein sorting (VPS) gene, deep orange (dor), dramatically promote tumor overgrowth and invasion of the RasV12 cells....

  3. Neurofibromin Loss of Function Drives Excessive Grooming in Drosophila

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    Lanikea B. King

    2016-04-01

    Full Text Available 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, and 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 provides 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, we combined traditional activity monitoring 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, we found that 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.

  4. Neurofibromin Loss of Function Drives Excessive Grooming in Drosophila.

    Science.gov (United States)

    King, Lanikea B; Koch, Marta; Murphy, Keith R; Velazquez, Yoheilly; Ja, William W; Tomchik, Seth M

    2016-04-07

    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, and 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 provides 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, we combined traditional activity monitoring 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, we found that 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. Copyright © 2016 King et al.

  5. A novel Drosophila model of TDP-43 proteinopathies: N-terminal sequences combined with the Q/N domain induce protein functional loss and locomotion defects

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    Simona Langellotti

    2016-06-01

    Full Text Available Transactive response DNA-binding protein 43 kDa (TDP-43, also known as TBPH in Drosophila melanogaster and TARDBP in mammals is the main protein component of the pathological inclusions observed in neurons of patients affected by different neurodegenerative disorders, including amyotrophic lateral sclerosis (ALS and fronto-temporal lobar degeneration (FTLD. The number of studies investigating the molecular mechanisms underlying neurodegeneration is constantly growing; however, the role played by TDP-43 in disease onset and progression is still unclear. A fundamental shortcoming that hampers progress is the lack of animal models showing aggregation of TDP-43 without overexpression. In this manuscript, we have extended our cellular model of aggregation to a transgenic Drosophila line. Our fly model is not based on the overexpression of a wild-type TDP-43 transgene. By contrast, we engineered a construct that includes only the specific TDP-43 amino acid sequences necessary to trigger aggregate formation and capable of trapping endogenous Drosophila TDP-43 into a non-functional insoluble form. Importantly, the resulting recombinant product lacks functional RNA recognition motifs (RRMs and, thus, does not have specific TDP-43-physiological functions (i.e. splicing regulation ability that might affect the animal phenotype per se. This novel Drosophila model exhibits an evident degenerative phenotype with reduced lifespan and early locomotion defects. Additionally, we show that important proteins involved in neuromuscular junction function, such as syntaxin (SYX, decrease their levels as a consequence of TDP-43 loss of function implying that the degenerative phenotype is a consequence of TDP-43 sequestration into the aggregates. Our data lend further support to the role of TDP-43 loss-of-function in the pathogenesis of neurodegenerative disorders. The novel transgenic Drosophila model presented in this study will help to gain further insight into the

  6. Functional conservation of the Drosophila gooseberry gene and its evolutionary alleles.

    Directory of Open Access Journals (Sweden)

    Wei Liu

    Full Text Available The Drosophila Pax gene gooseberry (gsb is required for development of the larval cuticle and CNS, survival to adulthood, and male fertility. These functions can be rescued in gsb mutants by two gsb evolutionary alleles, gsb-Prd and gsb-Pax3, which express the Drosophila Paired and mouse Pax3 proteins under the control of gooseberry cis-regulatory region. Therefore, both Paired and Pax3 proteins have conserved all the Gsb functions that are required for survival of embryos to fertile adults, despite the divergent primary sequences in their C-terminal halves. As gsb-Prd and gsb-Pax3 uncover a gsb function involved in male fertility, construction of evolutionary alleles may provide a powerful strategy to dissect hitherto unknown gene functions. Our results provide further evidence for the essential role of cis-regulatory regions in the functional diversification of duplicated genes during evolution.

  7. Sex chromosome-specific regulation in the Drosophila male germline but little evidence for chromosomal dosage compensation or meiotic inactivation.

    Directory of Open Access Journals (Sweden)

    Colin D Meiklejohn

    2011-08-01

    Full Text Available The evolution of heteromorphic sex chromosomes (e.g., XY in males or ZW in females has repeatedly elicited the evolution of two kinds of chromosome-specific regulation: dosage compensation--the equalization of X chromosome gene expression in males and females--and meiotic sex chromosome inactivation (MSCI--the transcriptional silencing and heterochromatinization of the X during meiosis in the male (or Z in the female germline. How the X chromosome is regulated in the Drosophila melanogaster male germline is unclear. Here we report three new findings concerning gene expression from the X in Drosophila testes. First, X chromosome-wide dosage compensation appears to be absent from most of the Drosophila male germline. Second, microarray analysis provides no evidence for X chromosome-specific inactivation during meiosis. Third, we confirm the previous discovery that the expression of transgene reporters driven by autosomal spermatogenesis-specific promoters is strongly reduced when inserted on the X chromosome versus the autosomes; but we show that this chromosomal difference in expression is established in premeiotic cells and persists in meiotic cells. The magnitude of the X-autosome difference in transgene expression cannot be explained by the absence of dosage compensation, suggesting that a previously unrecognized mechanism limits expression from the X during spermatogenesis in Drosophila. These findings help to resolve several previously conflicting reports and have implications for patterns of genome evolution and speciation in Drosophila.

  8. The function of the frizzled pathway in the Drosophila wing is dependent on inturned and fuzzy.

    OpenAIRE

    Lee, Haeryun; Adler, Paul N

    2002-01-01

    The Drosophila epidermis is characterized by a dramatic planar or tissue polarity. The frizzled pathway has been shown to be a key regulator of planar polarity for hairs on the wing, ommatidia in the eye, and sensory bristles on the notum. We have investigated the genetic relationships between putative frizzled pathway downstream genes inturned, fuzzy, and multiple wing hairs (inturned-like genes) and upstream genes such as frizzled, prickle, and starry night (frizzled-like genes). Previous d...

  9. Cooperation of the BTB-Zinc finger protein, Abrupt, with cytoskeletal regulators in Drosophila epithelial tumorigenesis

    Directory of Open Access Journals (Sweden)

    Nezaket Turkel

    2015-08-01

    Full Text Available The deregulation of cell polarity or cytoskeletal regulators is a common occurrence in human epithelial cancers. Moreover, there is accumulating evidence in human epithelial cancer that BTB-ZF genes, such as Bcl6 and ZBTB7A, are oncogenic. From our previous studies in the vinegar fly, Drosophila melanogaster, we have identified a cooperative interaction between a mutation in the apico-basal cell polarity regulator Scribble (Scrib and overexpression of the BTB-ZF protein Abrupt (Ab. Herein, we show that co-expression of ab with actin cytoskeletal regulators, RhoGEF2 or Src64B, in the developing eye-antennal epithelial tissue results in the formation of overgrown amorphous tumours, whereas ab and DRac1 co-expression leads to non-cell autonomous overgrowth. Together with ab, these genes affect the expression of differentiation genes, resulting in tumours locked in a progenitor cell fate. Finally, we show that the expression of two mammalian genes related to ab, Bcl6 and ZBTB7A, which are oncogenes in mammalian epithelial cancers, significantly correlate with the upregulation of cytoskeletal genes or downregulation of apico-basal cell polarity neoplastic tumour suppressor genes in colorectal, lung and other human epithelial cancers. Altogether, this analysis has revealed that upregulation of cytoskeletal regulators cooperate with Abrupt in Drosophila epithelial tumorigenesis, and that high expression of human BTB-ZF genes, Bcl6 and ZBTB7A, shows significant correlations with cytoskeletal and cell polarity gene expression in specific epithelial tumour types. This highlights the need for further investigation of the cooperation between these genes in mammalian systems.

  10. Interorgan Communication Pathways in Physiology: Focus on Drosophila

    OpenAIRE

    Droujinine, Ilia A.; Perrimon, Norbert

    2016-01-01

    Studies in mammals and Drosophila have demonstrated the existence and significance of secreted factors involved in communication between distal organs. In this review, primarily focusing on Drosophila, we examine the known interorgan communication factors and their functions, physiological inducers, and integration in regulating physiology. Moreover, we describe how organ-sensing screens in Drosophila can systematically identify novel conserved interorgan communication factors. Finally, we di...

  11. Activated Cdc42 kinase regulates Dock localization in male germ cells during Drosophila spermatogenesis.

    Science.gov (United States)

    Abdallah, Abbas M; Zhou, Xin; Kim, Christine; Shah, Kushani K; Hogden, Christopher; Schoenherr, Jessica A; Clemens, James C; Chang, Henry C

    2013-06-15

    Deregulation of the non-receptor tyrosine kinase ACK1 (Activated Cdc42-associated kinase) correlates with poor prognosis in cancers and has been implicated in promoting metastasis. To further understand its in vivo function, we have characterized the developmental defects of a null mutation in Drosophila Ack, which bears a high degree of sequence similarity to mammalian ACK1 but lacks a CRIB domain. We show that Ack, while not essential for viability, is critical for sperm formation. This function depends on Ack tyrosine kinase activity and is required cell autonomously in differentiating male germ cells at or after the spermatocyte stage. Ack associates predominantly with endocytic clathrin sites in spermatocytes, but disruption of Ack function has no apparent effect on clathrin localization and receptor-mediated internalization of Boss (Bride of sevenless) protein in eye discs. Instead, Ack is required for the subcellular distribution of Dock (dreadlocks), the Drosophila homolog of the SH2- and SH3-containing adaptor protein Nck. Moreover, Dock forms a complex with Ack, and the localization of Dock in male germ cells depends on its SH2 domain. Together, our results suggest that Ack-dependent tyrosine phosphorylation recruits Dock to promote sperm differentiation. Copyright © 2013 Elsevier Inc. All rights reserved.

  12. Isoform-specific functions of Mud/NuMA mediate binucleation of Drosophila male accessory gland cells.

    Science.gov (United States)

    Taniguchi, Kiichiro; Kokuryo, Akihiko; Imano, Takao; Minami, Ryunosuke; Nakagoshi, Hideki; Adachi-Yamada, Takashi

    2014-12-20

    In standard cell division, the cells undergo karyokinesis and then cytokinesis. Some cells, however, such as cardiomyocytes and hepatocytes, can produce binucleate cells by going through mitosis without cytokinesis. This cytokinesis skipping is thought to be due to the inhibition of cytokinesis machinery such as the central spindle or the contractile ring, but the mechanisms regulating it are unclear. We investigated them by characterizing the binucleation event during development of the Drosophila male accessory gland, in which all cells are binucleate. The accessory gland cells arrested the cell cycle at 50 hours after puparium formation (APF) and in the middle of the pupal stage stopped proliferating for 5 hours. They then restarted the cell cycle and at 55 hours APF entered the M-phase synchronously. At this stage, accessory gland cells binucleated by mitosis without cytokinesis. Binucleating cells displayed the standard karyokinesis progression but also showed unusual features such as a non-round shape, spindle orientation along the apico-basal axis, and poor assembly of the central spindle. Mud, a Drosophila homolog of NuMA, regulated the processes responsible for these three features, the classical isoform Mud(PBD) and the two newly characterized isoforms Mud(L) and Mud(S) regulated them differently: Mud(L) repressed cell rounding, Mud(PBD) and Mud(S) oriented the spindle along the apico-basal axis, and Mud(S) and Mud(L) repressed central spindle assembly. Importantly, overexpression of Mud(S) induced binucleation even in standard proliferating cells such as those in imaginal discs. We characterized the binucleation in the Drosophila male accessory gland and examined mechanisms that regulated unusual morphologies of binucleating cells. We demonstrated that Mud, a microtubule binding protein regulating spindle orientation, was involved in this binucleation. We suggest that atypical functions exerted by three structurally different isoforms of Mud regulate

  13. Discovery of functional elements in 12 Drosophila genomes using evolutionary signatures

    DEFF Research Database (Denmark)

    Stark, Alexander; Lin, Michael F; Kheradpour, Pouya

    2007-01-01

    Sequencing of multiple related species followed by comparative genomics analysis constitutes a powerful approach for the systematic understanding of any genome. Here, we use the genomes of 12 Drosophila species for the de novo discovery of functional elements in the fly. Each type of functional e...... individual motif instances with high confidence. We also study how discovery power scales with the divergence and number of species compared, and we provide general guidelines for comparative studies....

  14. Somatic insulin signaling regulates a germline starvation response in Drosophila egg chambers

    Science.gov (United States)

    Burn, K. Mahala; Shimada, Yuko; Ayers, Kathleen; Lu, Feiyue; Hudson, Andrew M.; Cooley, Lynn

    2014-01-01

    Egg chambers from starved Drosophila females contain large aggregates of processing (P) bodies and cortically enriched microtubules. As this response to starvation is rapidly reversed upon re-feeding females or culturing egg chambers with exogenous bovine insulin, we examined the role of endogenous insulin signaling in mediating the starvation response. We found that systemic Drosophila insulin-like peptides (dILPs) activate the insulin pathway in follicle cells, which then regulate both microtubule and P body organization in the underlying germline cells. This organization is modulated by the motor proteins Dynein and Kinesin. Dynein activity is required for microtubule and P body organization during starvation, while Kinesin activity is required during nutrient-rich conditions. Blocking the ability of egg chambers to form P body aggregates in response to starvation correlated with reduced progeny survival. These data suggest a potential mechanism to maximize fecundity even during periods of poor nutrient availability, by mounting a protective response in immature egg chambers. PMID:25481758

  15. The Sex Determination Gene transformer Regulates Male-Female Differences in Drosophila Body Size.

    Science.gov (United States)

    Rideout, Elizabeth J; Narsaiya, Marcus S; Grewal, Savraj S

    2015-12-01

    Almost all animals show sex differences in body size. For example, in Drosophila, females are larger than males. Although Drosophila is widely used as a model to study growth, the mechanisms underlying this male-female difference in size remain unclear. Here, we describe a novel role for the sex determination gene transformer (tra) in promoting female body growth. Normally, Tra is expressed only in females. We find that loss of Tra in female larvae decreases body size, while ectopic Tra expression in males increases body size. Although we find that Tra exerts autonomous effects on cell size, we also discovered that Tra expression in the fat body augments female body size in a non cell-autonomous manner. These effects of Tra do not require its only known targets doublesex and fruitless. Instead, Tra expression in the female fat body promotes growth by stimulating the secretion of insulin-like peptides from insulin producing cells in the brain. Our data suggest a model of sex-specific growth in which body size is regulated by a previously unrecognized branch of the sex determination pathway, and identify Tra as a novel link between sex and the conserved insulin signaling pathway.

  16. Altered gene regulation and synaptic morphology in Drosophila learning and memory mutants

    Science.gov (United States)

    Guan, Zhuo; Buhl, Lauren K.; Quinn, William G.; Littleton, J. Troy

    2011-01-01

    Genetic studies in Drosophila have revealed two separable long-term memory pathways defined as anesthesia-resistant memory (ARM) and long-lasting long-term memory (LLTM). ARM is disrupted in radish (rsh) mutants, whereas LLTM requires CREB-dependent protein synthesis. Although the downstream effectors of ARM and LLTM are distinct, pathways leading to these forms of memory may share the cAMP cascade critical for associative learning. Dunce, which encodes a cAMP-specific phosphodiesterase, and rutabaga, which encodes an adenylyl cyclase, both disrupt short-term memory. Amnesiac encodes a pituitary adenylyl cyclase-activating peptide homolog and is required for middle-term memory. Here, we demonstrate that the Radish protein localizes to the cytoplasm and nucleus and is a PKA phosphorylation target in vitro. To characterize how these plasticity pathways may manifest at the synaptic level, we assayed synaptic connectivity and performed an expression analysis to detect altered transcriptional networks in rutabaga, dunce, amnesiac, and radish mutants. All four mutants disrupt specific aspects of synaptic connectivity at larval neuromuscular junctions (NMJs). Genome-wide DNA microarray analysis revealed ∼375 transcripts that are altered in these mutants, suggesting defects in multiple neuronal signaling pathways. In particular, the transcriptional target Lapsyn, which encodes a leucine-rich repeat cell adhesion protein, localizes to synapses and regulates synaptic growth. This analysis provides insights into the Radish-dependent ARM pathway and novel transcriptional targets that may contribute to memory processing in Drosophila. PMID:21422168

  17. Regulation of the activity of the tumor suppressor PTEN by thioredoxin in Drosophila melanogaster

    International Nuclear Information System (INIS)

    Song, Zuohe; Saghafi, Negin; Gokhale, Vijay; Brabant, Marc; Meuillet, Emmanuelle J.

    2007-01-01

    Human Thioredoxin-1 (hTrx-1) is a small redox protein with a molecular weight of 12 kDa that contains two cysteine residues found in its catalytic site. HTrx-1 plays an important role in cell growth, apoptosis, and cancer patient prognosis. Recently, we have demonstrated that hTrx-1 binds to the C2 domain of the human tumor suppressor, PTEN, in a redox dependent manner. This binding leads to the inhibition of PTEN lipid phosphatase activity in mammalian tissue culture systems. In this study, we show that over-expression of hTrx-1 in Drosophila melanogaster promotes cell growth and proliferation during eye development as measured by eye size and ommatidia size. Furthermore, hTrx-1 rescues the small eye phenotype induced by the over-expression of PTEN. We demonstrate that this rescue of the PTEN-induced eye size phenotype requires cysteine-218 in the C2 domain of PTEN. We also show that hTrx-1 over-expression results in increased Akt phosphorylation in fly head extracts supporting our observations that the hTrx-1-induced eye size increase results from the inhibition of PTEN activity. Our study confirms the redox regulation of PTEN through disulfide bond formation with the hTrx-1 in Drosophila and suggests conserved mechanisms for thioredoxins and their interactions with the phosphatidylinositol-3-kinase signaling pathway in humans and fruit flies

  18. The Sex Determination Gene transformer Regulates Male-Female Differences in Drosophila Body Size.

    Directory of Open Access Journals (Sweden)

    Elizabeth J Rideout

    2015-12-01

    Full Text Available Almost all animals show sex differences in body size. For example, in Drosophila, females are larger than males. Although Drosophila is widely used as a model to study growth, the mechanisms underlying this male-female difference in size remain unclear. Here, we describe a novel role for the sex determination gene transformer (tra in promoting female body growth. Normally, Tra is expressed only in females. We find that loss of Tra in female larvae decreases body size, while ectopic Tra expression in males increases body size. Although we find that Tra exerts autonomous effects on cell size, we also discovered that Tra expression in the fat body augments female body size in a non cell-autonomous manner. These effects of Tra do not require its only known targets doublesex and fruitless. Instead, Tra expression in the female fat body promotes growth by stimulating the secretion of insulin-like peptides from insulin producing cells in the brain. Our data suggest a model of sex-specific growth in which body size is regulated by a previously unrecognized branch of the sex determination pathway, and identify Tra as a novel link between sex and the conserved insulin signaling pathway.

  19. Astrocytic glutamate transport regulates a Drosophila CNS synapse that lacks astrocyte ensheathment.

    Science.gov (United States)

    MacNamee, Sarah E; Liu, Kendra E; Gerhard, Stephan; Tran, Cathy T; Fetter, Richard D; Cardona, Albert; Tolbert, Leslie P; Oland, Lynne A

    2016-07-01

    Anatomical, molecular, and physiological interactions between astrocytes and neuronal synapses regulate information processing in the brain. The fruit fly Drosophila melanogaster has become a valuable experimental system for genetic manipulation of the nervous system and has enormous potential for elucidating mechanisms that mediate neuron-glia interactions. Here, we show the first electrophysiological recordings from Drosophila astrocytes and characterize their spatial and physiological relationship with particular synapses. Astrocyte intrinsic properties were found to be strongly analogous to those of vertebrate astrocytes, including a passive current-voltage relationship, low membrane resistance, high capacitance, and dye-coupling to local astrocytes. Responses to optogenetic stimulation of glutamatergic premotor neurons were correlated directly with anatomy using serial electron microscopy reconstructions of homologous identified neurons and surrounding astrocytic processes. Robust bidirectional communication was present: neuronal activation triggered astrocytic glutamate transport via excitatory amino acid transporter 1 (Eaat1), and blocking Eaat1 extended glutamatergic interneuron-evoked inhibitory postsynaptic currents in motor neurons. The neuronal synapses were always located within 1 μm of an astrocytic process, but none were ensheathed by those processes. Thus, fly astrocytes can modulate fast synaptic transmission via neurotransmitter transport within these anatomical parameters. J. Comp. Neurol. 524:1979-1998, 2016. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  20. Genetic and systems level analysis of Drosophila sticky/citron kinase and dFmr1 mutants reveals common regulation of genetic networks

    Directory of Open Access Journals (Sweden)

    Zarnescu Daniela C

    2008-11-01

    Full Text Available Abstract Background In Drosophila, the genes sticky and dFmr1 have both been shown to regulate cytoskeletal dynamics and chromatin structure. These genes also genetically interact with Argonaute family microRNA regulators. Furthermore, in mammalian systems, both genes have been implicated in neuronal development. Given these genetic and functional similarities, we tested Drosophila sticky and dFmr1 for a genetic interaction and measured whole genome expression in both mutants to assess similarities in gene regulation. Results We found that sticky mutations can dominantly suppress a dFmr1 gain-of-function phenotype in the developing eye, while phenotypes produced by RNAi knock-down of sticky were enhanced by dFmr1 RNAi and a dFmr1 loss-of-function mutation. We also identified a large number of transcripts that were misexpressed in both mutants suggesting that sticky and dFmr1 gene products similarly regulate gene expression. By integrating gene expression data with a protein-protein interaction network, we found that mutations in sticky and dFmr1 resulted in misexpression of common gene networks, and consequently predicted additional specific phenotypes previously not known to be associated with either gene. Further phenotypic analyses validated these predictions. Conclusion These findings establish a functional link between two previously unrelated genes. Microarray analysis indicates that sticky and dFmr1 are both required for regulation of many developmental genes in a variety of cell types. The diversity of transcripts regulated by these two genes suggests a clear cause of the pleiotropy that sticky and dFmr1 mutants display and provides many novel, testable hypotheses about the functions of these genes. As both of these genes are implicated in the development and function of the mammalian brain, these results have relevance to human health as well as to understanding more general biological processes.

  1. Affecting Rhomboid-3 function causes a dilated heart in adult Drosophila.

    Directory of Open Access Journals (Sweden)

    Lin Yu

    2010-05-01

    Full Text Available Drosophila is a well recognized model of several human diseases, and recent investigations have demonstrated that Drosophila can be used as a model of human heart failure. Previously, we described that optical coherence tomography (OCT can be used to rapidly examine the cardiac function in adult, awake flies. This technique provides images that are similar to echocardiography in humans, and therefore we postulated that this approach could be combined with the vast resources that are available in the fly community to identify new mutants that have abnormal heart function, a hallmark of certain cardiovascular diseases. Using OCT to examine the cardiac function in adult Drosophila from a set of molecularly-defined genomic deficiencies from the DrosDel and Exelixis collections, we identified an abnormally enlarged cardiac chamber in a series of deficiency mutants spanning the rhomboid 3 locus. Rhomboid 3 is a member of a highly conserved family of intramembrane serine proteases and processes Spitz, an epidermal growth factor (EGF-like ligand. Using multiple approaches based on the examination of deficiency stocks, a series of mutants in the rhomboid-Spitz-EGF receptor pathway, and cardiac-specific transgenic rescue or dominant-negative repression of EGFR, we demonstrate that rhomboid 3 mediated activation of the EGF receptor pathway is necessary for proper adult cardiac function. The importance of EGF receptor signaling in the adult Drosophila heart underscores the concept that evolutionarily conserved signaling mechanisms are required to maintain normal myocardial function. Interestingly, prior work showing the inhibition of ErbB2, a member of the EGF receptor family, in transgenic knock-out mice or individuals that received herceptin chemotherapy is associated with the development of dilated cardiomyopathy. Our results, in conjunction with the demonstration that altered ErbB2 signaling underlies certain forms of mammalian cardiomyopathy, suggest

  2. Myostatin-like proteins regulate synaptic function and neuronal morphology.

    Science.gov (United States)

    Augustin, Hrvoje; McGourty, Kieran; Steinert, Joern R; Cochemé, Helena M; Adcott, Jennifer; Cabecinha, Melissa; Vincent, Alec; Halff, Els F; Kittler, Josef T; Boucrot, Emmanuel; Partridge, Linda

    2017-07-01

    Growth factors of the TGFβ superfamily play key roles in regulating neuronal and muscle function. Myostatin (or GDF8) and GDF11 are potent negative regulators of skeletal muscle mass. However, expression of myostatin and its cognate receptors in other tissues, including brain and peripheral nerves, suggests a potential wider biological role. Here, we show that Myoglianin (MYO), the Drosophila homolog of myostatin and GDF11, regulates not only body weight and muscle size, but also inhibits neuromuscular synapse strength and composition in a Smad2-dependent manner. Both myostatin and GDF11 affected synapse formation in isolated rat cortical neuron cultures, suggesting an effect on synaptogenesis beyond neuromuscular junctions. We also show that MYO acts in vivo to inhibit synaptic transmission between neurons in the escape response neural circuit of adult flies. Thus, these anti-myogenic proteins act as important inhibitors of synapse function and neuronal growth. © 2017. Published by The Company of Biologists Ltd.

  3. A Circadian Clock Gene, Cry, Affects Heart Morphogenesis and Function in Drosophila as Revealed by Optical Coherence Microscopy.

    Directory of Open Access Journals (Sweden)

    Aneesh Alex

    Full Text Available Circadian rhythms are endogenous, entrainable oscillations of physical, mental and behavioural processes in response to local environmental cues such as daylight, which are present in the living beings, including humans. Circadian rhythms have been related to cardiovascular function and pathology. However, the role that circadian clock genes play in heart development and function in a whole animal in vivo are poorly understood. The Drosophila cryptochrome (dCry is a circadian clock gene that encodes a major component of the circadian clock negative feedback loop. Compared to the embryonic stage, the relative expression levels of dCry showed a significant increase (>100-fold in Drosophila during the pupa and adult stages. In this study, we utilized an ultrahigh resolution optical coherence microscopy (OCM system to perform non-invasive and longitudinal analysis of functional and morphological changes in the Drosophila heart throughout its post-embryonic lifecycle for the first time. The Drosophila heart exhibited major morphological and functional alterations during its development. Notably, heart rate (HR and cardiac activity period (CAP of Drosophila showed significant variations during the pupa stage, when heart remodeling took place. From the M-mode (2D + time OCM images, cardiac structural and functional parameters of Drosophila at different developmental stages were quantitatively determined. In order to study the functional role of dCry on Drosophila heart development, we silenced dCry by RNAi in the Drosophila heart and mesoderm, and quantitatively measured heart morphology and function in those flies throughout its development. Silencing of dCry resulted in slower HR, reduced CAP, smaller heart chamber size, pupal lethality and disrupted posterior segmentation that was related to increased expression of a posterior compartment protein, wingless. Collectively, our studies provided novel evidence that the circadian clock gene, dCry, plays

  4. A Circadian Clock Gene, Cry, Affects Heart Morphogenesis and Function in Drosophila as Revealed by Optical Coherence Microscopy

    Science.gov (United States)

    Zeng, Xianxu; Tate, Rebecca E.; McKee, Mary L.; Capen, Diane E.; Zhang, Zhan; Tanzi, Rudolph E.; Zhou, Chao

    2015-01-01

    Circadian rhythms are endogenous, entrainable oscillations of physical, mental and behavioural processes in response to local environmental cues such as daylight, which are present in the living beings, including humans. Circadian rhythms have been related to cardiovascular function and pathology. However, the role that circadian clock genes play in heart development and function in a whole animal in vivo are poorly understood. The Drosophila cryptochrome (dCry) is a circadian clock gene that encodes a major component of the circadian clock negative feedback loop. Compared to the embryonic stage, the relative expression levels of dCry showed a significant increase (>100-fold) in Drosophila during the pupa and adult stages. In this study, we utilized an ultrahigh resolution optical coherence microscopy (OCM) system to perform non-invasive and longitudinal analysis of functional and morphological changes in the Drosophila heart throughout its post-embryonic lifecycle for the first time. The Drosophila heart exhibited major morphological and functional alterations during its development. Notably, heart rate (HR) and cardiac activity period (CAP) of Drosophila showed significant variations during the pupa stage, when heart remodeling took place. From the M-mode (2D + time) OCM images, cardiac structural and functional parameters of Drosophila at different developmental stages were quantitatively determined. In order to study the functional role of dCry on Drosophila heart development, we silenced dCry by RNAi in the Drosophila heart and mesoderm, and quantitatively measured heart morphology and function in those flies throughout its development. Silencing of dCry resulted in slower HR, reduced CAP, smaller heart chamber size, pupal lethality and disrupted posterior segmentation that was related to increased expression of a posterior compartment protein, wingless. Collectively, our studies provided novel evidence that the circadian clock gene, dCry, plays an essential

  5. Analysis of functional importance of binding sites in the Drosophila gap gene network model.

    Science.gov (United States)

    Kozlov, Konstantin; Gursky, Vitaly V; Kulakovskiy, Ivan V; Dymova, Arina; Samsonova, Maria

    2015-01-01

    The statistical thermodynamics based approach provides a promising framework for construction of the genotype-phenotype map in many biological systems. Among important aspects of a good model connecting the DNA sequence information with that of a molecular phenotype (gene expression) is the selection of regulatory interactions and relevant transcription factor bindings sites. As the model may predict different levels of the functional importance of specific binding sites in different genomic and regulatory contexts, it is essential to formulate and study such models under different modeling assumptions. We elaborate a two-layer model for the Drosophila gap gene network and include in the model a combined set of transcription factor binding sites and concentration dependent regulatory interaction between gap genes hunchback and Kruppel. We show that the new variants of the model are more consistent in terms of gene expression predictions for various genetic constructs in comparison to previous work. We quantify the functional importance of binding sites by calculating their impact on gene expression in the model and calculate how these impacts correlate across all sites under different modeling assumptions. The assumption about the dual interaction between hb and Kr leads to the most consistent modeling results, but, on the other hand, may obscure existence of indirect interactions between binding sites in regulatory regions of distinct genes. The analysis confirms the previously formulated regulation concept of many weak binding sites working in concert. The model predicts a more or less uniform distribution of functionally important binding sites over the sets of experimentally characterized regulatory modules and other open chromatin domains.

  6. A Drosophila Genome-Wide Screen Identifies Regulators of Steroid Hormone Production and Developmental Timing

    DEFF Research Database (Denmark)

    Thomas Danielsen, E.; E. Møller, Morten; Yamanaka, Naoki

    2016-01-01

    Steroid hormones control important developmental processes and are linked to many diseases. To systematically identify genes and pathways required for steroid production, we performed a Drosophila genome-wide in vivo RNAi screen and identified 1,906 genes with potential roles in steroidogenesis...... and developmental timing. Here, we use our screen as a resource to identify mechanisms regulating intracellular levels of cholesterol, a substrate for steroidogenesis. We identify a conserved fatty acid elongase that underlies a mechanism that adjusts cholesterol trafficking and steroidogenesis with nutrition...... and developmental programs. In addition, we demonstrate the existence of an autophagosomal cholesterol mobilization mechanism and show that activation of this system rescues Niemann-Pick type C1 deficiency that causes a disorder characterized by cholesterol accumulation. These cholesterol-trafficking mechanisms...

  7. Macronutrients mediate the functional relationship between Drosophila and Wolbachia

    Science.gov (United States)

    Ponton, Fleur; Wilson, Kenneth; Holmes, Andrew; Raubenheimer, David; Robinson, Katie L.; Simpson, Stephen J.

    2015-01-01

    Wolbachia are maternally inherited bacterial endosymbionts that naturally infect a diverse array of arthropods. They are primarily known for their manipulation of host reproductive biology, and recently, infections with Wolbachia have been proposed as a new strategy for controlling insect vectors and subsequent human-transmissible diseases. Yet, Wolbachia abundance has been shown to vary greatly between individuals and the magnitude of the effects of infection on host life-history traits and protection against infection is correlated to within-host Wolbachia abundance. It is therefore essential to better understand the factors that modulate Wolbachia abundance and effects on host fitness. Nutrition is known to be one of the most important mediators of host–symbiont interactions. Here, we used nutritional geometry to quantify the role of macronutrients on insect–Wolbachia relationships in Drosophila melanogaster. Our results show fundamental interactions between diet composition, host diet selection, Wolbachia abundance and effects on host lifespan and fecundity. The results and methods described here open a new avenue in the study of insect–Wolbachia relationships and are of general interest to numerous research disciplines, ranging from nutrition and life-history theory to public health. PMID:25520356

  8. Mislocalization of the Drosophila centromere-specific histone CIDpromotes formation of functional ectopic kinetochores

    Energy Technology Data Exchange (ETDEWEB)

    Heun, Patrick; Erhardt, Sylvia; Blower, Michael D.; Weiss,Samara; Skora, Andrew D.; Karpen, Gary H.

    2006-01-30

    The centromere-specific histone variant CENP-A (CID in Drosophila) is a structural and functional foundation for kinetochore formation and chromosome segregation. Here, we show that overexpressed CID is mislocalized into normally non-centromeric regions in Drosophila tissue culture cells and animals. Analysis of mitoses in living and fixed cells reveals that mitotic delays, anaphase bridges, chromosome fragmentation, and cell and organismal lethality are all direct consequences of CID mislocalization. In addition, proteins that are normally restricted to endogenous kinetochores assemble at a subset of ectopic CID incorporation regions. The presence of microtubule motors and binding proteins, spindle attachments, and aberrant chromosome morphologies demonstrate that these ectopic kinetochores are functional. We conclude that CID mislocalization promotes formation of ectopic centromeres and multicentric chromosomes, which causes chromosome missegregation, aneuploidy, and growth defects. Thus, CENP-A mislocalization is one possible mechanism for genome instability during cancer progression, as well as centromere plasticity during evolution.

  9. Astrocyte-like glial cells physiologically regulate olfactory processing through the modification of ORN-PN synaptic strength in Drosophila.

    Science.gov (United States)

    Liu, He; Zhou, Bangyu; Yan, Wenjun; Lei, Zhengchang; Zhao, Xiaoliang; Zhang, Ke; Guo, Aike

    2014-09-01

    Astrocyte-like glial cells are abundant in the central nervous system of adult Drosophila and exhibit morphology similar to astrocytes of mammals. Previous evidence has shown that astrocyte-like glial cells are strongly associated with synapses in the antennal lobe (AL), the first relay of the olfactory system, where olfactory receptor neurons (ORNs) transmit information into projection neurons (PNs). However, the function of astrocyte-like glia in the AL remains obscure. In this study, using in vivo calcium imaging, we found that astrocyte-like glial cells exhibited spontaneous microdomain calcium elevations. Using simultaneous manipulation of glial activity and monitoring of neuronal function, we found that the astrocyte-like glial activation, but not ensheathing glial activation, could inhibit odor-evoked responses of PNs. Ensheathing glial cells are another subtype of glia, and are of functional importance in the AL. Electrophysiological experiments indicated that astrocyte-like glial activation decreased the amplitude and slope of excitatory postsynaptic potentials evoked through electrical stimulation of the antennal nerve. These results suggest that astrocyte-like glial cells may regulate olfactory processing through negative regulation of ORN-PN synaptic strength. Beyond the antennal lobe we observed astrocyte-like glial spontaneous calcium activities in the ventromedial protocerebrum, indicating that astrocyte-like glial spontaneous calcium elevations might be general in the adult fly brain. Overall, our study demonstrates a new function for astrocyte-like glial cells in the physiological modulation of olfactory information transmission, possibly through regulating ORN-PN synapse strength. © 2014 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.

  10. Rapid male-specific regulatory divergence and down regulation of spermatogenesis genes in Drosophila species hybrids.

    Directory of Open Access Journals (Sweden)

    Jennifer Ferguson

    Full Text Available In most crosses between closely related species of Drosophila, the male hybrids are sterile and show postmeiotic abnormalities. A series of gene expression studies using genomic approaches have found significant down regulation of postmeiotic spermatogenesis genes in sterile male hybrids. These results have led some to suggest a direct relationship between down regulation in gene expression and hybrid sterility. An alternative explanation to a cause-and-effect relationship between misregulation of gene expression and male sterility is rapid divergence of male sex regulatory elements leading to incompatible interactions in an interspecies hybrid genome. To test the effect of regulatory divergence in spermatogenesis gene expression, we isolated 35 fertile D. simulans strains with D. mauritiana introgressions in either the X, second or third chromosome. We analyzed gene expression in these fertile hybrid strains for a subset of spermatogenesis genes previously reported as significantly under expressed in sterile hybrids relative to D. simulans. We found that fertile autosomal introgressions can cause levels of gene down regulation similar to that of sterile hybrids. We also found that X chromosome heterospecific introgressions cause significantly less gene down regulation than autosomal introgressions. Our results provide evidence that rapid male sex gene regulatory divergence can explain misexpression of spermatogenesis genes in hybrids.

  11. Lipid phosphate phosphatase activity regulates dispersal and bilateral sorting of embryonic germ cells in Drosophila

    Science.gov (United States)

    Renault, Andrew D.; Kunwar, Prabhat S.; Lehmann, Ruth

    2010-01-01

    In Drosophila, germ cell survival and directionality of migration are controlled by two lipid phosphate phosphatases (LPP), wunen (wun) and wunen-2 (wun2). wun wun2 double mutant analysis reveals that the two genes, hereafter collectively called wunens, act redundantly in primordial germ cells. We find that wunens mediate germ cell-germ cell repulsion and that this repulsion is necessary for germ cell dispersal and proper transepithelial migration at the onset of migration and for the equal sorting of the germ cells between the two embryonic gonads during their migration. We propose that this dispersal function optimizes adult fecundity by assuring maximal germ cell occupancy of both gonads. Furthermore, we find that the requirement for wunens in germ cell survival can be eliminated by blocking germ cell migration. We suggest that this essential function of Wunen is needed to maintain cell integrity in actively migrating germ cells. PMID:20431117

  12. RNAi Screen in Drosophila melanogastor Identifies Regulators of Steroidogenesis and Developmental Maturation

    DEFF Research Database (Denmark)

    Danielsen, Erik Thomas

    and duration required for juvenile-adult transition. This PhD project demonstrates the power of Drosophila genetics by taking an in vivo genome-wide RNAi screening approach to uncover genes required for the function of steroid producing tissue and developmental maturation. In total, 1909 genes were found...... to be required for the prothoracic gland function and affected the developmental timing for the juvenile-adult transition. Among the screen hits, we focused on an uncharacterized gene, sit (CG5278), which is highly expressed in the gland and is required for ecdysone production. Sit is a homolog of mammalian very...... flux of cholesterol uptake in the gland cells and affected the endosomal trafficking. Therefore this gene was suggested to be named stuck in traffic (sit). Sit’s role in cholesterol uptake was also supported by the observation that the developmental delayed phenotype from loss of sit expression...

  13. Diverse Roles of Axonemal Dyneins in Drosophila Auditory Neuron Function and Mechanical Amplification in Hearing.

    Science.gov (United States)

    Karak, Somdatta; Jacobs, Julie S; Kittelmann, Maike; Spalthoff, Christian; Katana, Radoslaw; Sivan-Loukianova, Elena; Schon, Michael A; Kernan, Maurice J; Eberl, Daniel F; Göpfert, Martin C

    2015-11-26

    Much like vertebrate hair cells, the chordotonal sensory neurons that mediate hearing in Drosophila are motile and amplify the mechanical input of the ear. Because the neurons bear mechanosensory primary cilia whose microtubule axonemes display dynein arms, we hypothesized that their motility is powered by dyneins. Here, we describe two axonemal dynein proteins that are required for Drosophila auditory neuron function, localize to their primary cilia, and differently contribute to mechanical amplification in hearing. Promoter fusions revealed that the two axonemal dynein genes Dmdnah3 (=CG17150) and Dmdnai2 (=CG6053) are expressed in chordotonal neurons, including the auditory ones in the fly's ear. Null alleles of both dyneins equally abolished electrical auditory neuron responses, yet whereas mutations in Dmdnah3 facilitated mechanical amplification, amplification was abolished by mutations in Dmdnai2. Epistasis analysis revealed that Dmdnah3 acts downstream of Nan-Iav channels in controlling the amplificatory gain. Dmdnai2, in addition to being required for amplification, was essential for outer dynein arms in auditory neuron cilia. This establishes diverse roles of axonemal dyneins in Drosophila auditory neuron function and links auditory neuron motility to primary cilia and axonemal dyneins. Mutant defects in sperm competition suggest that both dyneins also function in sperm motility.

  14. Drosophila Pumilio protein contains multiple autonomous repression domains that regulate mRNAs independently of Nanos and brain tumor.

    Science.gov (United States)

    Weidmann, Chase A; Goldstrohm, Aaron C

    2012-01-01

    Drosophila melanogaster Pumilio is an RNA-binding protein that potently represses specific mRNAs. In developing embryos, Pumilio regulates a key morphogen, Hunchback, in collaboration with the cofactor Nanos. To investigate repression by Pumilio and Nanos, we created cell-based assays and found that Pumilio inhibits translation and enhances mRNA decay independent of Nanos. Nanos robustly stimulates repression through interactions with the Pumilio RNA-binding domain. We programmed Pumilio to recognize a new binding site, which garners repression of new target mRNAs. We show that cofactors Brain Tumor and eIF4E Homologous Protein are not obligatory for Pumilio and Nanos activity. The conserved RNA-binding domain of Pumilio was thought to be sufficient for its function. Instead, we demonstrate that three unique domains in the N terminus of Pumilio possess the major repressive activity and can function autonomously. The N termini of insect and vertebrate Pumilio and Fem-3 binding factors (PUFs) are related, and we show that corresponding regions of human PUM1 and PUM2 have repressive activity. Other PUF proteins lack these repression domains. Our findings suggest that PUF proteins have evolved new regulatory functions through protein sequences appended to their conserved PUF repeat RNA-binding domains.

  15. Functional Maps of Mechanosensory Features in the Drosophila Brain.

    Science.gov (United States)

    Patella, Paola; Wilson, Rachel I

    2018-04-09

    Johnston's organ is the largest mechanosensory organ in Drosophila. It contributes to hearing, touch, vestibular sensing, proprioception, and wind sensing. In this study, we used in vivo 2-photon calcium imaging and unsupervised image segmentation to map the tuning properties of Johnston's organ neurons (JONs) at the site where their axons enter the brain. We then applied the same methodology to study two key brain regions that process signals from JONs: the antennal mechanosensory and motor center (AMMC) and the wedge, which is downstream of the AMMC. First, we identified a diversity of JON response types that tile frequency space and form a rough tonotopic map. Some JON response types are direction selective; others are specialized to encode amplitude modulations over a specific range (dynamic range fractionation). Next, we discovered that both the AMMC and the wedge contain a tonotopic map, with a significant increase in tonotopy-and a narrowing of frequency tuning-at the level of the wedge. Whereas the AMMC tonotopic map is unilateral, the wedge tonotopic map is bilateral. Finally, we identified a subregion of the AMMC/wedge that responds preferentially to the coherent rotation of the two mechanical organs in the same angular direction, indicative of oriented steady air flow (directional wind). Together, these maps reveal the broad organization of the primary and secondary mechanosensory regions of the brain. They provide a framework for future efforts to identify the specific cell types and mechanisms that underlie the hierarchical re-mapping of mechanosensory information in this system. Copyright © 2018 Elsevier Ltd. All rights reserved.

  16. Forkhead, a new cross regulator of metabolism and innate immunity downstream of TOR in Drosophila.

    Science.gov (United States)

    Varma, Disha; Bülow, Margret H; Pesch, Yanina-Yasmin; Loch, Gerrit; Hoch, Michael

    2014-10-01

    Antimicrobial peptides (AMPs) are conserved cationic peptides which act both as defense molecules of the host immune system and as regulators of the commensal microbiome. Expression of AMPs is induced in response to infection by the Toll and Imd pathway. Under non-infected conditions, the transcription factor dFOXO directly regulates a set of AMP expression at low levels when nutrients are limited. Here we have analyzed whether target of rapamycin (TOR), another major regulator of growth and metabolism, also modulates AMP responses in Drosophila. We found that downregulation of TOR by feeding the drug rapamycin or by overexpressing the negative TOR regulators TSC1/TSC2, resulted in a specific induction of the AMPs Diptericin (Dpt) and Metchnikowin (Mtk). In contrast, overexpression of Rheb, which positively regulates TOR led to a repression of the two AMPs. Genetic and pharmacological experiments indicate that Dpt and Mtk activation is controlled by the transcription factor Forkhead (FKH), the founding member of the FoxO family. Shuttling of FKH from the cytoplasm to the nucleus is induced in the fat body and in the posterior midgut in response to TOR downregulation. The FKH-dependent induction of Dpt and Mtk can be triggered in dFOXO null mutants and in immune-compromised Toll and IMD pathway mutants indicating that FKH acts in parallel to these regulators. Together, we have discovered that FKH is the second conserved member of the FoxO family cross-regulating metabolism and innate immunity. dFOXO and FKH, which are activated upon downregulation of insulin or TOR activities, respectively, act in parallel to induce different sets of AMPs, thereby modulating the immune status of metabolic tissues such as the fat body or the gut in response to the oscillating energy status of the organism. Copyright © 2014 Elsevier Ltd. All rights reserved.

  17. Functional Differences between Global Pre- and Postsynaptic Inhibition in the Drosophila Olfactory Circuit.

    Science.gov (United States)

    Oizumi, Masafumi; Satoh, Ryota; Kazama, Hokto; Okada, Masato

    2012-01-01

    The Drosophila antennal lobe is subdivided into multiple glomeruli, each of which represents a unique olfactory information processing channel. In each glomerulus, feedforward input from olfactory receptor neurons (ORNs) is transformed into activity of projection neurons (PNs), which represent the output. Recent investigations have indicated that lateral presynaptic inhibitory input from other glomeruli controls the gain of this transformation. Here, we address why this gain control acts "pre"-synaptically rather than "post"-synaptically. Postsynaptic inhibition could work similarly to presynaptic inhibition with regard to regulating the firing rates of PNs depending on the stimulus intensity. We investigate the differences between pre- and postsynaptic gain control in terms of odor discriminability by simulating a network model of the Drosophila antennal lobe with experimental data. We first demonstrate that only presynaptic inhibition can reproduce the type of gain control observed in experiments. We next show that presynaptic inhibition decorrelates PN responses whereas postsynaptic inhibition does not. Due to this effect, presynaptic gain control enhances the accuracy of odor discrimination by a linear decoder while its postsynaptic counterpart only diminishes it. Our results provide the reason gain control operates "pre"-synaptically but not "post"-synaptically in the Drosophila antennal lobe.

  18. Functional differences between global pre- and postsynaptic inhibition in the Drosophila olfactory circuit

    Directory of Open Access Journals (Sweden)

    Masafumi eOizumi

    2012-03-01

    Full Text Available The Drosophila antennal lobe is subdivided into multiple glomeruli, each of which represents a unique olfactory information processing channel. In each glomerulus, feedforward input from olfactory receptor neurons (ORNs is transformed into activity of projection neurons (PNs, which represent the output. Recent investigations have indicated that lateral pre-synaptic inhibitory input from other glomeruli controls the gain of this transformation. Here, we address why this gain control acts `pre'-synaptically rather than `post'-synaptically. Postsynaptic inhibition could work similarly to presynaptic inhibition with regard to regulating the firing rates of PNs depending on the stimulus intensity. We investigate the differences between pre- and postsynaptic gain control in terms of odor discriminability by simulating a network model of the Drosophila antennal lobe with experimental data. We first demonstrate that only presynaptic inhibition can reproduce the type of gain control observed in experiments. We next show that presynaptic inhibition decorrelates PN responses whereas postsynaptic inhibition does not. Due to this effect, presynaptic gain control enhances the accuracy of odor discrimination by a linear decoder while its postsynaptic counterpart only diminishes it. Our results provide the reason gain control operates `pre'-synaptically but not `post'-synaptically in the Drosophila antennal lobe.

  19. Phenotypic Plasticity through Transcriptional Regulation of the Evolutionary Hotspot Gene tan in Drosophila melanogaster.

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    Jean-Michel Gibert

    2016-08-01

    Full Text Available Phenotypic plasticity is the ability of a given genotype to produce different phenotypes in response to distinct environmental conditions. Phenotypic plasticity can be adaptive. Furthermore, it is thought to facilitate evolution. Although phenotypic plasticity is a widespread phenomenon, its molecular mechanisms are only beginning to be unravelled. Environmental conditions can affect gene expression through modification of chromatin structure, mainly via histone modifications, nucleosome remodelling or DNA methylation, suggesting that phenotypic plasticity might partly be due to chromatin plasticity. As a model of phenotypic plasticity, we study abdominal pigmentation of Drosophila melanogaster females, which is temperature sensitive. Abdominal pigmentation is indeed darker in females grown at 18°C than at 29°C. This phenomenon is thought to be adaptive as the dark pigmentation produced at lower temperature increases body temperature. We show here that temperature modulates the expression of tan (t, a pigmentation gene involved in melanin production. t is expressed 7 times more at 18°C than at 29°C in female abdominal epidermis. Genetic experiments show that modulation of t expression by temperature is essential for female abdominal pigmentation plasticity. Temperature modulates the activity of an enhancer of t without modifying compaction of its chromatin or level of the active histone mark H3K27ac. By contrast, the active mark H3K4me3 on the t promoter is strongly modulated by temperature. The H3K4 methyl-transferase involved in this process is likely Trithorax, as we show that it regulates t expression and the H3K4me3 level on the t promoter and also participates in female pigmentation and its plasticity. Interestingly, t was previously shown to be involved in inter-individual variation of female abdominal pigmentation in Drosophila melanogaster, and in abdominal pigmentation divergence between Drosophila species. Sensitivity of t

  20. Functional Characterization of CCHamide and Muscarinic Acetylcholine Receptor Signalling in Drosophila melanogaster

    DEFF Research Database (Denmark)

    Ren, Guilin Robin

    G-protein coupled receptors (GPCRs) constitute a large and ancient superfamily of membraneproteins responsible for the transduction of extracellular signals to the inside of the cells. In thisPh.D. thesis, Drosophila melanogaster (Dm) was used as a model organism to investigate a numberof topics...... is a newly discovered insect peptide hormone. The function of this novel peptide hasnot been well characterised. In this Ph.D. thesis, I identified CCHamide-2 peptides in endocrinecells of the gut and neurones of the brain of larvae and endocrine cells of the gut of adultDrosophila. Behavioural assays...... little is known about muscarinic acetylcholine receptorsignalling in insects. In this study, I found that two types of mAChRs occur in D. melanogaster, onecoupling to Gq (A-type) and the other to Gi (B-type). Both A- and B-type Dm-mAChRs can beactivated by acetylcholine (ACh), but the classical...

  1. Painting of fourth and chromosome-wide regulation of the 4th chromosome in Drosophila melanogaster.

    Science.gov (United States)

    Johansson, Anna-Mia; Stenberg, Per; Bernhardsson, Carolina; Larsson, Jan

    2007-05-02

    Drosophila melanogaster exhibits two expression-regulating systems that target whole, specific chromosomes: the dosage compensation system whereby the male-specific lethal complex doubles transcription of genes on the male X-chromosome and the chromosome 4-specific protein Painting of fourth, POF. POF is the first example of an autosome-specific protein and its presence raises the question of the universality of chromosome-specific regulation. Here we show that POF and heterochromatin protein 1 (HP1) are involved in the global regulation of the 4th chromosome. Contrary to previous conclusions, Pof is not essential for survival of diplo-4th karyotype flies. However, Pof is essential for survival of haplo-4th individuals and expression of chromosome 4 genes in diplo-4th individuals is decreased in the absence of Pof. Mapping of POF using chromatin immunoprecipitation suggested that it binds within genes. Furthermore, we show that POF binding is dependent on heterochromatin and that POF and HP1 bind interdependently to the 4th chromosome. We propose a balancing mechanism involving POF and HP1 that provides a feedback system for fine-tuning expression status of genes on the 4th chromosome.

  2. Relationship between organization and function of ribosomal genes in Drosophila melanogaster

    International Nuclear Information System (INIS)

    Karpen, G.H.

    1987-01-01

    In most eukaryotic organisms, the genes that encode the 18S and 28S ribosomal RNAs (rDNA genes) are tandemly repeated, and are located in constitutive heterochromatin and/or centromeric or telomeric regions. P-element mediated transformation was used to investigate the relationship between rDNA organization and function in Drosophila melanogaster. Tritiated-uridine incorporation under heat shock conditions and in situ hybridization to rRNA were used to demonstrate that a single rDNA gene inserted into euchromatin can be transcribed at a high rate, in polytene nuclei. P-element-mediated transformation of a single Drosophila rDNA gene was also utilized to investigate the ability of ribosomal DNA to organize a nucleolus. Cytological approaches demonstrated that structures resembling the endogenous nucleoli were preferentially associated with four different sites of rDNA insertion, in polytene nuclei. These mini-nucleoli also contained components specific to the nucleolus, as shown by in situ hybridization to rRNA and indirect immunofluorescence with an antibody that binds to Drosophila nucleoli. The transformed genes were able to partially rescue mutant phenotypes due to a deficiency of rDNA, indicating that the mini-nucleoli were functional

  3. Quantitative Assessment of Eye Phenotypes for Functional Genetic Studies Using Drosophila melanogaster

    Directory of Open Access Journals (Sweden)

    Janani Iyer

    2016-05-01

    Full Text Available About two-thirds of the vital genes in the Drosophila genome are involved in eye development, making the fly eye an excellent genetic system to study cellular function and development, neurodevelopment/degeneration, and complex diseases such as cancer and diabetes. We developed a novel computational method, implemented as Flynotyper software (http://flynotyper.sourceforge.net, to quantitatively assess the morphological defects in the Drosophila eye resulting from genetic alterations affecting basic cellular and developmental processes. Flynotyper utilizes a series of image processing operations to automatically detect the fly eye and the individual ommatidium, and calculates a phenotypic score as a measure of the disorderliness of ommatidial arrangement in the fly eye. As a proof of principle, we tested our method by analyzing the defects due to eye-specific knockdown of Drosophila orthologs of 12 neurodevelopmental genes to accurately document differential sensitivities of these genes to dosage alteration. We also evaluated eye images from six independent studies assessing the effect of overexpression of repeats, candidates from peptide library screens, and modifiers of neurotoxicity and developmental processes on eye morphology, and show strong concordance with the original assessment. We further demonstrate the utility of this method by analyzing 16 modifiers of sine oculis obtained from two genome-wide deficiency screens of Drosophila and accurately quantifying the effect of its enhancers and suppressors during eye development. Our method will complement existing assays for eye phenotypes, and increase the accuracy of studies that use fly eyes for functional evaluation of genes and genetic interactions.

  4. Neuronal Cbl Controls Biosynthesis of Insulin-Like Peptides in Drosophila melanogaster

    OpenAIRE

    Yu, Yue; Sun, Ying; He, Shengqi; Yan, Cheng; Rui, Liangyou; Li, Wenjun; Liu, Yong

    2012-01-01

    The Cbl family proteins function as both E3 ubiquitin ligases and adaptor proteins to regulate various cellular signaling events, including the insulin/insulin-like growth factor 1 (IGF1) and epidermal growth factor (EGF) pathways. These pathways play essential roles in growth, development, metabolism, and survival. Here we show that in Drosophila melanogaster, Drosophila Cbl (dCbl) regulates longevity and carbohydrate metabolism through downregulating the production of Drosophila insulin-lik...

  5. A functional screen reveals an extensive layer of transcriptional and splicing control underlying RAS/MAPK signaling in Drosophila.

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    Dariel Ashton-Beaucage

    2014-03-01

    Full Text Available The small GTPase RAS is among the most prevalent oncogenes. The evolutionarily conserved RAF-MEK-MAPK module that lies downstream of RAS is one of the main conduits through which RAS transmits proliferative signals in normal and cancer cells. Genetic and biochemical studies conducted over the last two decades uncovered a small set of factors regulating RAS/MAPK signaling. Interestingly, most of these were found to control RAF activation, thus suggesting a central regulatory role for this event. Whether additional factors are required at this level or further downstream remains an open question. To obtain a comprehensive view of the elements functionally linked to the RAS/MAPK cascade, we used a quantitative assay in Drosophila S2 cells to conduct a genome-wide RNAi screen for factors impacting RAS-mediated MAPK activation. The screen led to the identification of 101 validated hits, including most of the previously known factors associated to this pathway. Epistasis experiments were then carried out on individual candidates to determine their position relative to core pathway components. While this revealed several new factors acting at different steps along the pathway--including a new protein complex modulating RAF activation--we found that most hits unexpectedly work downstream of MEK and specifically influence MAPK expression. These hits mainly consist of constitutive splicing factors and thereby suggest that splicing plays a specific role in establishing MAPK levels. We further characterized two representative members of this group and surprisingly found that they act by regulating mapk alternative splicing. This study provides an unprecedented assessment of the factors modulating RAS/MAPK signaling in Drosophila. In addition, it suggests that pathway output does not solely rely on classical signaling events, such as those controlling RAF activation, but also on the regulation of MAPK levels. Finally, it indicates that core splicing

  6. Bar represses dPax2 and decapentaplegic to regulate cell fate and morphogenetic cell death in Drosophila eye.

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    Jongkyun Kang

    Full Text Available The coordinated regulation of cell fate and cell survival is crucial for normal pattern formation in developing organisms. In Drosophila compound eye development, crystalline arrays of hexagonal ommatidia are established by precise assembly of diverse cell types, including the photoreceptor cells, cone cells and interommatidial (IOM pigment cells. The molecular basis for controlling the number of cone and IOM pigment cells during ommatidial pattern formation is not well understood. Here we present evidence that BarH1 and BarH2 homeobox genes are essential for eye patterning by inhibiting excess cone cell differentiation and promoting programmed death of IOM cells. Specifically, we show that loss of Bar from the undifferentiated retinal precursor cells leads to ectopic expression of Prospero and dPax2, two transcription factors essential for cone cell specification, resulting in excess cone cell differentiation. We also show that loss of Bar causes ectopic expression of the TGFβ homolog Decapentaplegic (Dpp posterior to the morphogenetic furrow in the larval eye imaginal disc. The ectopic Dpp expression is not responsible for the formation of excess cone cells in Bar loss-of-function mutant eyes. Instead, it causes reduction in IOM cell death in the pupal stage by antagonizing the function of pro-apoptotic gene reaper. Taken together, this study suggests a novel regulatory mechanism in the control of developmental cell death in which the repression of Dpp by Bar in larval eye disc is essential for IOM cell death in pupal retina.

  7. A dual function for Deep orange in programmed autophagy in the Drosophila melanogaster fat body

    International Nuclear Information System (INIS)

    Lindmo, Karine; Simonsen, Anne; Brech, Andreas; Finley, Kim; Rusten, Tor Erik; Stenmark, Harald

    2006-01-01

    Lysosomal degradation of cytoplasm by way of autophagy is essential for cellular amino acid homeostasis and for tissue remodeling. In insects such as Drosophila, autophagy is developmentally upregulated in the larval fat body prior to metamorphosis. Here, autophagy is induced by the hormone ecdysone through down-regulation of the autophagy-suppressive phosphoinositide 3-kinase (PI3K) signaling pathway. In yeast, Vps18 and other members of the HOPS complex have been found essential for autophagic degradation. In Drosophila, the Vps18 homologue Deep orange (Dor) has previously been shown to mediate fusion of multivesicular endosomes with lysosomes. A requirement of Dor for ecdysone-mediated chromosome puffing has also been reported. In the present report, we have tested the hypothesis that Dor may control programmed autophagy at the level of ecdysone signaling as well as by mediating autophagosome-to-lysosome fusion. We show that dor mutants are defective in programmed autophagy and provide evidence that autophagy is blocked at two levels. First, PI3K activity was not down-regulated correctly in dor larvae, which correlated with a decrease in ecdysone reporter activity. The down-regulation of PI3K activity was restored by feeding ecdysone to the mutant larvae. Second, neither exogenous ecdysone nor overexpression of PTEN, a silencer of PI3K signaling, restored fusion of autophagosomes with lysosomes in the fat body of dor mutants. These results indicate that Dor controls autophagy indirectly, via ecdysone signaling, as well as directly, via autolysosomal fusion

  8. The Drosophila melanogaster methuselah gene: a novel gene with ancient functions.

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    Ana Rita Araújo

    Full Text Available The Drosophila melanogaster G protein-coupled receptor gene, methuselah (mth, has been described as a novel gene that is less than 10 million years old. Nevertheless, it shows a highly specific expression pattern in embryos, larvae, and adults, and has been implicated in larval development, stress resistance, and in the setting of adult lifespan, among others. Although mth belongs to a gene subfamily with 16 members in D. melanogaster, there is no evidence for functional redundancy in this subfamily. Therefore, it is surprising that a novel gene influences so many traits. Here, we explore the alternative hypothesis that mth is an old gene. Under this hypothesis, in species distantly related to D. melanogaster, there should be a gene with features similar to those of mth. By performing detailed phylogenetic, synteny, protein structure, and gene expression analyses we show that the D. virilis GJ12490 gene is the orthologous of mth in species distantly related to D. melanogaster. We also show that, in D. americana (a species of the virilis group of Drosophila, a common amino acid polymorphism at the GJ12490 orthologous gene is significantly associated with developmental time, size, and lifespan differences. Our results imply that GJ12490 orthologous genes are candidates for developmental time and lifespan differences in Drosophila in general.

  9. p53- and ERK7-dependent ribosome surveillance response regulates Drosophila insulin-like peptide secretion.

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    Kiran Hasygar

    2014-11-01

    Full Text Available Insulin-like signalling is a conserved mechanism that coordinates animal growth and metabolism with nutrient status. In Drosophila, insulin-producing median neurosecretory cells (IPCs regulate larval growth by secreting insulin-like peptides (dILPs in a diet-dependent manner. Previous studies have shown that nutrition affects dILP secretion through humoral signals derived from the fat body. Here we uncover a novel mechanism that operates cell autonomously in the IPCs to regulate dILP secretion. We observed that impairment of ribosome biogenesis specifically in the IPCs strongly inhibits dILP secretion, which consequently leads to reduced body size and a delay in larval development. This response is dependent on p53, a known surveillance factor for ribosome biogenesis. A downstream effector of this growth inhibitory response is an atypical MAP kinase ERK7 (ERK8/MAPK15, which is upregulated in the IPCs following impaired ribosome biogenesis as well as starvation. We show that ERK7 is sufficient and essential to inhibit dILP secretion upon impaired ribosome biogenesis, and it acts epistatically to p53. Moreover, we provide evidence that p53 and ERK7 contribute to the inhibition of dILP secretion upon starvation. Thus, we conclude that a cell autonomous ribosome surveillance response, which leads to upregulation of ERK7, inhibits dILP secretion to impede tissue growth under limiting dietary conditions.

  10. Integrins Regulate Apical Constriction via Microtubule Stabilization in the Drosophila Eye Disc Epithelium

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    Vilaiwan M. Fernandes

    2014-12-01

    Full Text Available During morphogenesis, extracellular signals trigger actomyosin contractility in subpopulations of cells to coordinate changes in cell shape. To illuminate the link between signaling-mediated tissue patterning and cytoskeletal remodeling, we study the progression of the morphogenetic furrow (MF, the wave of apical constriction that traverses the Drosophila eye imaginal disc preceding photoreceptor neurogenesis. Apical constriction depends on actomyosin contractility downstream of the Hedgehog (Hh and bone morphogenetic protein (BMP pathways. We identify a role for integrin adhesion receptors in MF progression. We show that Hh and BMP regulate integrin expression, the loss of which disrupts apical constriction and slows furrow progression; conversely, elevated integrins accelerate furrow progression. We present evidence that integrins regulate MF progression by promoting microtubule stabilization, since reducing microtubule stability rescues integrin-mediated furrow acceleration. Thus, integrins act as a genetic link between tissue-level signaling events and morphological change at the cellular level, leading to morphogenesis and neurogenesis in the eye.

  11. Drosophila KDM2 is a H3K4me3 demethylase regulating nucleolar organization

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    Birchler James A

    2009-10-01

    Full Text Available Abstract Background CG11033 (dKDM2 is the Drosophila homolog of the gene KDM2B. dKDM2 has been known to possess histone lysine demethylase activity towards H3K36me2 in cell lines and it regulates H2A ubiquitination. The human homolog of the gene has dual activity towards H3K36me2 as well as H3K4me3, and plays an important role in cellular senescence. Findings We have used transgenic flies bearing an RNAi construct for the dKDM2 gene. The knockdown of dKDM2 gene was performed by crossing UAS-RNAi-dKDM2 flies with actin-Gal4 flies. Western blots of acid extracted histones and immunofluoresence analysis of polytene chromosome showed the activity of the enzyme dKDM2 to be specific for H3K4me3 in adult flies. Immunofluoresence analysis of polytene chromosome also revealed the presence of multiple nucleoli in RNAi knockdown mutants of dKDM2 and decreased H3-acetylation marks associated with active transcription. Conclusion Our findings indicate that dKDM2 is a histone lysine demethylase with specificity for H3K4me3 and regulates nucleolar organization.

  12. A pair of pharyngeal gustatory receptor neurons regulates caffeine-dependent ingestion in Drosophila larvae

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    Jaekyun Choi

    2016-07-01

    Full Text Available The sense of taste is an essential chemosensory modality that enables animals to identify appropriate food sources and control feeding behavior. In particular, the recognition of bitter taste prevents animals from feeding on harmful substances. Feeding is a complex behavior comprised of multiple steps, and food quality is continuously assessed. We here examined the role of pharyngeal gustatory organs in ingestion behavior. As a first step, we constructed a gustatory receptor-to-neuron map of the larval pharyngeal sense organs, and examined corresponding gustatory receptor neuron projections in the larval brain. Out of 22 candidate bitter compounds, we found 14 bitter compounds that elicit inhibition of ingestion in a dose-dependent manner. We provide evidence that certain pharyngeal gustatory receptor neurons are necessary and sufficient for the ingestion response of larvae to caffeine. Additionally, we show that a specific pair of pharyngeal gustatory receptor neurons, DP1, responds to caffeine by calcium imaging. In this study we show that a specific pair of gustatory receptor neurons in the pharyngeal sense organs coordinates caffeine sensing with regulation of behavioral responses such as ingestion. Our results indicate that in Drosophila larvae, the pharyngeal gustatory receptor neurons have a major role in sensing food palatability to regulate ingestion behavior. The pharyngeal sense organs are prime candidates to influence ingestion due to their position in the pharynx, and they may act as first level sensors of ingested food.

  13. Regulation of spindle orientation and neural stem cell fate in the Drosophila optic lobe

    Directory of Open Access Journals (Sweden)

    Brand Andrea H

    2007-01-01

    Full Text Available Abstract Background The choice of a stem cell to divide symmetrically or asymmetrically has profound consequences for development and disease. Unregulated symmetric division promotes tumor formation, whereas inappropriate asymmetric division affects organ morphogenesis. Despite its importance, little is known about how spindle positioning is regulated. In some tissues cell fate appears to dictate the type of cell division, whereas in other tissues it is thought that stochastic variation in spindle position dictates subsequent sibling cell fate. Results Here we investigate the relationship between neural progenitor identity and spindle positioning in the Drosophila optic lobe. We use molecular markers and live imaging to show that there are two populations of progenitors in the optic lobe: symmetrically dividing neuroepithelial cells and asymmetrically dividing neuroblasts. We use genetically marked single cell clones to show that neuroepithelial cells give rise to neuroblasts. To determine if a change in spindle orientation can trigger a neuroepithelial to neuroblast transition, we force neuroepithelial cells to divide along their apical/basal axis by misexpressing Inscuteable. We find that this does not induce neuroblasts, nor does it promote premature neuronal differentiation. Conclusion We show that symmetrically dividing neuroepithelial cells give rise to asymmetrically dividing neuroblasts in the optic lobe, and that regulation of spindle orientation and division symmetry is a consequence of cell type specification, rather than a mechanism for generating cell type diversity.

  14. Reph, a regulator of Eph receptor expression in the Drosophila melanogaster optic lobe.

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    Richard E Dearborn

    Full Text Available Receptors of the Eph family of tyrosine kinases and their Ephrin ligands are involved in developmental processes as diverse as angiogenesis, axon guidance and cell migration. However, our understanding of the Eph signaling pathway is incomplete, and could benefit from an analysis by genetic methods. To this end, we performed a genetic modifier screen for mutations that affect Eph signaling in Drosophila melanogaster. Several dozen loci were identified on the basis of their suppression or enhancement of an eye defect induced by the ectopic expression of Ephrin during development; many of these mutant loci were found to disrupt visual system development. One modifier locus, reph (regulator of eph expression, was characterized in molecular detail and found to encode a putative nuclear protein that interacts genetically with Eph signaling pathway mutations. Reph is an autonomous regulator of Eph receptor expression, required for the graded expression of Eph protein and the establishment of an optic lobe axonal topographic map. These results reveal a novel component of the regulatory pathway controlling expression of eph and identify reph as a novel factor in the developing visual system.

  15. Drosophila SLC5A11 Mediates Hunger by Regulating K(+) Channel Activity.

    Science.gov (United States)

    Park, Jin-Yong; Dus, Monica; Kim, Seonil; Abu, Farhan; Kanai, Makoto I; Rudy, Bernardo; Suh, Greg S B

    2016-08-08

    Hunger is a powerful drive that stimulates food intake. Yet, the mechanism that determines how the energy deficits that result in hunger are represented in the brain and promote feeding is not well understood. We previously described SLC5A11-a sodium/solute co-transporter-like-(or cupcake) in Drosophila melanogaster, which is required for the fly to select a nutritive sugar over a sweeter nonnutritive sugar after periods of food deprivation. SLC5A11 acts on approximately 12 pairs of ellipsoid body (EB) R4 neurons to trigger the selection of nutritive sugars, but the underlying mechanism is not understood. Here, we report that the excitability of SLC5A11-expressing EB R4 neurons increases dramatically during starvation and that this increase is abolished in the SLC5A11 mutation. Artificial activation of SLC5A11-expresssing neurons is sufficient to promote feeding and hunger-driven behaviors; silencing these neurons has the opposite effect. Notably, SLC5A11 transcript levels in the brain increase significantly when flies are starved and decrease shortly after starved flies are refed. Furthermore, expression of SLC5A11 is sufficient for promoting hunger-driven behaviors and enhancing the excitability of SLC5A11-expressing neurons. SLC5A11 inhibits the function of the Drosophila KCNQ potassium channel in a heterologous expression system. Accordingly, a knockdown of dKCNQ expression in SLC5A11-expressing neurons produces hunger-driven behaviors even in fed flies, mimicking the overexpression of SLC5A11. We propose that starvation increases SLC5A11 expression, which enhances the excitability of SLC5A11-expressing neurons by suppressing dKCNQ channels, thereby conferring the hunger state. Copyright © 2016 Elsevier Ltd. All rights reserved.

  16. Male sex interspecies divergence and down regulation of expression of spermatogenesis genes in Drosophila sterile hybrids.

    Science.gov (United States)

    Sundararajan, Vignesh; Civetta, Alberto

    2011-01-01

    Male sex genes have shown a pattern of rapid interspecies divergence at both the coding and gene expression level. A common outcome from crosses between closely-related species is hybrid male sterility. Phenotypic and genetic studies in Drosophila sterile hybrid males have shown that spermatogenesis arrest is postmeiotic with few exceptions, and that most misregulated genes are involved in late stages of spermatogenesis. Comparative studies of gene regulation in sterile hybrids and parental species have mainly used microarrays providing a whole genome representation of regulatory problems in sterile hybrids. Real-time PCR studies can reject or reveal differences not observed in microarray assays. Moreover, differences in gene expression between samples can be dependant on the source of RNA (e.g., whole body vs. tissue). Here we survey expression in D. simulans, D. mauritiana and both intra and interspecies hybrids using a real-time PCR approach for eight genes expressed at the four main stages of sperm development. We find that all genes show a trend toward under expression in the testes of sterile hybrids relative to parental species with only the two proliferation genes (bam and bgcn) and the two meiotic class genes (can and sa) showing significant down regulation. The observed pattern of down regulation for the genes tested can not fully explain hybrid male sterility. We discuss the down regulation of spermatogenesis genes in hybrids between closely-related species within the contest of rapid divergence experienced by the male genome, hybrid sterility and possible allometric changes due to subtle testes-specific developmental abnormalities.

  17. Protein expression profiling of the drosophila fragile X mutant brain reveals up-regulation of monoamine synthesis.

    Science.gov (United States)

    Zhang, Yong Q; Friedman, David B; Wang, Zhe; Woodruff, Elvin; Pan, Luyuan; O'donnell, Janis; Broadie, Kendal

    2005-03-01

    Fragile X syndrome is the most common form of inherited mental retardation, associated with both cognitive and behavioral anomalies. The disease is caused by silencing of the fragile X mental retardation 1 (fmr1) gene, which encodes the mRNA-binding, translational regulator FMRP. Previously we established a disease model through mutation of Drosophila fmr1 (dfmr1) and showed that loss of dFMRP causes defects in neuronal structure, function, and behavioral output similar to the human disease state. To uncover molecular targets of dFMRP in the brain, we use here a proteomic approach involving two-dimensional difference gel electrophoresis analyses followed by mass spectrometry identification of proteins with significantly altered expression in dfmr1 null mutants. We then focus on two misregulated enzymes, phenylalanine hydroxylase (Henna) and GTP cyclohydrolase (Punch), both of which mediate in concert the synthetic pathways of two key monoamine neuromodulators, dopamine and serotonin. Brain enzymatic assays show a nearly 2-fold elevation of Punch activity in dfmr1 null mutants. Consistently brain neurochemical assays show that both dopamine and serotonin are significantly increased in dfmr1 null mutants. At a cellular level, dfmr1 null mutant neurons display a highly significant elevation of the dense core vesicles that package these monoamine neuromodulators for secretion. Taken together, these data indicate that dFMRP normally down-regulates the monoamine pathway, which is consequently up-regulated in the mutant condition. Elevated brain levels of dopamine and serotonin provide a plausible mechanistic explanation for aspects of cognitive and behavioral deficits in human patients.

  18. Sex-lethal enables germline stem cell differentiation by down-regulating Nanos protein levels during Drosophila oogenesis.

    Science.gov (United States)

    Chau, Johnnie; Kulnane, Laura Shapiro; Salz, Helen K

    2012-06-12

    Drosophila ovarian germ cells require Sex-lethal (Sxl) to exit from the stem cell state and to enter the differentiation pathway. Sxl encodes a female-specific RNA binding protein and in somatic cells serves as the developmental switch gene for somatic sex determination and X-chromosome dosage compensation. None of the known Sxl target genes are required for germline differentiation, leaving open the question of how Sxl promotes the transition from stem cell to committed daughter cell. We address the mechanism by which Sxl regulates this transition through the identification of nanos as one of its target genes. Previous studies have shown that Nanos protein is necessary for GSC self-renewal and is rapidly down-regulated in the daughter cells fated to differentiate in the adult ovary. We find that this dynamic expression pattern is limited to female germ cells and is under Sxl control. In the absence of Sxl, or in male germ cells, Nanos protein is continuously expressed. Furthermore, this female-specific expression pattern is dependent on the presence of canonical Sxl binding sites located in the nanos 3' untranslated region. These results, combined with the observation that nanos RNA associates with the Sxl protein in ovarian extracts and loss and gain of function studies, suggest that Sxl enables the switch from germline stem cell to committed daughter cell by posttranscriptional down-regulation of nanos expression. These findings connect sexual identity to the stem cell self-renewal/differentiation decision and highlight the importance of posttranscriptional gene regulatory networks in controlling stem cell behavior.

  19. The Drosophila Netrin receptor frazzled/DCC functions as an invasive tumor suppressor

    Directory of Open Access Journals (Sweden)

    Duman-Scheel Molly

    2011-06-01

    Full Text Available Abstract Background Loss of heterozygosity at 18q, which includes the Deleted in Colorectal Cancer (DCC gene, has been linked to many human cancers. However, it is unclear if loss of DCC is the specific underlying cause of these cancers. The Drosophila imaginal discs are excellent systems in which to study DCC function, as it is possible to model human tumors through the generation of somatic clones of cells bearing multiple genetic lesions. Here, these attributes of the fly system were utilized to investigate the potential tumor suppressing functions of the Drosophila DCC homologue frazzled (fra during eye-antennal disc development. Results Most fra loss of function clones are eliminated during development. However, when mutant clone cells generated in the developing eye were rescued from death, partially differentiated eye cells were found outside of the normal eye field, and in extreme cases distant sites of the body. Characterization of these cells during development indicates that fra mutant cells display characteristics of invasive tumor cells, including increased levels of phospho-ERK, phospho-JNK, and Mmp-1, changes in cadherin expression, remodeling of the actin cytoskeleton, and loss of polarity. Mutation of fra promotes basement membrane degradation and invasion which are repressed by inhibition of Rho1 signaling. Although inhibition of JNK signaling blocks invasive phenotypes in some metastatic cancer models in flies, blocking JNK signaling inhibits fra mutant cell death, thereby enhancing the fra mutant phenotype. Conclusions The results of this investigation provide the first direct link between point mutations in fra/DCC and metastatic phenotypes in an animal model and suggest that Fra functions as an invasive tumor suppressor during Drosophila development.

  20. Drosophila Studies on Autism Spectrum Disorders

    Institute of Scientific and Technical Information of China (English)

    Yao Tian; Zi Chao Zhang; Junhai Han

    2017-01-01

    In the past decade,numerous genes associated with autism spectrum disorders (ASDs) have been identified.These genes encode key regulators of synaptogenesis,synaptic function,and synaptic plasticity.Drosophila is a prominent model system for ASD studies to define novel genes linked to ASDs and decipher their molecular roles in synaptogenesis,synaptic function,synaptic plasticity,and neural circuit assembly and consolidation.Here,we review Drosophila studies on ASD genes that regulate synaptogenesis,synaptic function,and synaptic plasticity through modulating chromatin remodeling,transcription,protein synthesis and degradation,cytoskeleton dynamics,and synaptic scaffolding.

  1. Overlapping functions of argonaute proteins in patterning and morphogenesis of Drosophila embryos.

    Directory of Open Access Journals (Sweden)

    Wibke J Meyer

    2006-08-01

    Full Text Available Argonaute proteins are essential components of the molecular machinery that drives RNA silencing. In Drosophila, different members of the Argonaute family of proteins have been assigned to distinct RNA silencing pathways. While Ago1 is required for microRNA function, Ago2 is a crucial component of the RNA-induced silencing complex in siRNA-triggered RNA interference. Drosophila Ago2 contains an unusual amino-terminus with two types of imperfect glutamine-rich repeats (GRRs of unknown function. Here we show that the GRRs of Ago2 are essential for the normal function of the protein. Alleles with reduced numbers of GRRs cause specific disruptions in two morphogenetic processes associated with the midblastula transition: membrane growth and microtubule-based organelle transport. These defects do not appear to result from disruption of siRNA-dependent processes but rather suggest an interference of the mutant Ago2 proteins in an Ago1-dependent pathway. Using loss-of-function alleles, we further demonstrate that Ago1 and Ago2 act in a partially redundant manner to control the expression of the segment-polarity gene wingless in the early embryo. Our findings argue against a strict separation of Ago1 and Ago2 functions and suggest that these proteins act in concert to control key steps of the midblastula transition and of segmental patterning.

  2. Drosophila MOF regulates DIAP1 and induces apoptosis in a JNK dependent pathway.

    Science.gov (United States)

    Pushpavalli, Sreerangam N C V L; Sarkar, Arpita; Ramaiah, M Janaki; Koteswara Rao, G; Bag, Indira; Bhadra, Utpal; Pal-Bhadra, Manika

    2016-03-01

    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 (1) 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 we propose that apoptosis triggered by mof (RNAi) proceeds through a caspase-dependent and JNK mediated pathway.

  3. Tailless and Atrophin control Drosophila aggression by regulating neuropeptide signalling in the pars intercerebralis

    Science.gov (United States)

    Davis, Shaun M.; Thomas, Amanda L.; Nomie, Krystle J.; Huang, Longwen; Dierick, Herman A.

    2014-02-01

    Aggressive behaviour is widespread throughout the animal kingdom. However, its mechanisms are poorly understood, and the degree of molecular conservation between distantly related species is unknown. Here we show that knockdown of tailless (tll) increases aggression in Drosophila, similar to the effect of its mouse orthologue Nr2e1. Tll localizes to the adult pars intercerebralis (PI), which shows similarity to the mammalian hypothalamus. Knockdown of tll in the PI is sufficient to increase aggression and is rescued by co-expressing human NR2E1. Knockdown of Atrophin, a Tll co-repressor, also increases aggression, and both proteins physically interact in the PI. tll knockdown-induced aggression is fully suppressed by blocking neuropeptide processing or release from the PI. In addition, genetically activating PI neurons increases aggression, mimicking the aggression-inducing effect of hypothalamic stimulation. Together, our results suggest that a transcriptional control module regulates neuropeptide signalling from the neurosecretory cells of the brain to control aggressive behaviour.

  4. A fat-derived metabolite regulates a peptidergic feeding circuit in Drosophila.

    Directory of Open Access Journals (Sweden)

    Do-Hyoung Kim

    2017-03-01

    Full Text Available Here, we show that the enzymatic cofactor tetrahydrobiopterin (BH4 inhibits feeding in Drosophila. BH4 biosynthesis requires the sequential action of the conserved enzymes Punch, Purple, and Sepiapterin Reductase (Sptr. Although we observe increased feeding upon loss of Punch and Purple in the adult fat body, loss of Sptr must occur in the brain. We found Sptr expression is required in four adult neurons that express neuropeptide F (NPF, the fly homologue of the vertebrate appetite regulator neuropeptide Y (NPY. As expected, feeding flies BH4 rescues the loss of Punch and Purple in the fat body and the loss of Sptr in NPF neurons. Mechanistically, we found BH4 deficiency reduces NPF staining, likely by promoting its release, while excess BH4 increases NPF accumulation without altering its expression. We thus show that, because of its physically distributed biosynthesis, BH4 acts as a fat-derived signal that induces satiety by inhibiting the activity of the NPF neurons.

  5. Unfolded Protein Response-regulated Drosophila Fic (dFic) Protein Reversibly AMPylates BiP Chaperone during Endoplasmic Reticulum Homeostasis*

    Science.gov (United States)

    Ham, Hyeilin; Woolery, Andrew R.; Tracy, Charles; Stenesen, Drew; Krämer, Helmut; Orth, Kim

    2014-01-01

    Drosophila Fic (dFic) mediates AMPylation, a covalent attachment of adenosine monophosphate (AMP) from ATP to hydroxyl side chains of protein substrates. Here, we identified the endoplasmic reticulum (ER) chaperone BiP as a substrate for dFic and mapped the modification site to Thr-366 within the ATPase domain. The level of AMPylated BiP in Drosophila S2 cells is high during homeostasis, whereas the level of AMPylated BiP decreases upon the accumulation of misfolded proteins in the ER. Both dFic and BiP are transcriptionally activated upon ER stress, supporting the role of dFic in the unfolded protein response pathway. The inactive conformation of BiP is the preferred substrate for dFic, thus endorsing a model whereby AMPylation regulates the function of BiP as a chaperone, allowing acute activation of BiP by deAMPylation during an ER stress response. These findings not only present the first substrate of eukaryotic AMPylator but also provide a target for regulating the unfolded protein response, an emerging avenue for cancer therapy. PMID:25395623

  6. Functional analysis of the glycogen binding subunit CG9238/Gbs-70E of protein phosphatase 1 in Drosophila melanogaster.

    Science.gov (United States)

    Kerekes, Éva; Kókai, Endre; Páldy, Ferenc Sándor; Dombrádi, Viktor

    2014-06-01

    The product of the CG9238 gene that we termed glycogen binding subunit 70E (Gbs-70E) was characterized by biochemical and molecular genetics methods. The interaction between Gbs-70E and all catalytic subunits of protein phosphatase 1 (Pp1-87B, Pp1-9C, Pp1-96A and Pp1-13C) of Drosophila melanogaster was confirmed by pairwise yeast two-hybrid tests, co-immunoprecipitation and pull down experiments. The binding of Gbs-70E to glycogen was demonstrated by sedimentation analysis. With RT-PCR we found that the mRNAs coding for the longer Gbs-70E PB/PC protein were expressed in all developmental stages of the fruit flies while the mRNA for the shorter Gbs-70E PA was restricted to the eggs and the ovaries of the adult females. The development specific expression of the shorter splice variant was not conserved in different Drosophila species. The expression level of the gene was manipulated by P-element insertions and gene deletion to analyze the functions of the gene product. A small or moderate reduction in the gene expression resulted in no significant changes, however, a deletion mutant expressing very low level of the transcript lived shorter and exhibited reduced glycogen content in the imagos. In addition, the gene deletion decreased the fertility of the fruit flies. Our results prove that Gbs-70E functions as the glycogen binding subunit of protein phosphatase 1 that regulates glycogen content and plays a role in the development of eggs in D. melanogaster. Copyright © 2014 Elsevier Ltd. All rights reserved.

  7. Metabolic and functional characterization of effects of developmental temperature in Drosophila melanogaster

    DEFF Research Database (Denmark)

    Schou, Mads Fristrup; Kristensen, Torsten Nygaard; Pedersen, Anders

    2017-01-01

    , and in particular, how physiological stress at extreme temperatures may counteract beneficial acclimation responses at benign temperatures. We exposed Drosophila melanogaster to ten developmental temperatures covering their entire permissible temperature range. We obtained metabolic profiles and reaction norms...... for several functional traits: egg-to-adult viability, developmental time, and heat and cold tolerance. Females were more heat tolerant than males, whereas no sexual dimorphism was found in cold tolerance. A group of metabolites, mainly free amino acids, had linear reaction norms. Several energy carrying...

  8. Olfactory memory traces in Drosophila

    OpenAIRE

    Berry, Jacob; Krause, William C.; Davis, Ronald L.

    2008-01-01

    In Drosophila the fruit fly, coincident exposure to an odor and an aversive electric shock can produce robust behavioral memory. This behavioral memory is thought to be regulated by cellular memory traces within the central nervous system of the fly. These molecular, physiological or structural changes in neurons, induced by pairing odor and shock, regulate behavior by altering the neurons’ response to the learned environment. Recently, novel in vivo functional imaging techniques have allowed...

  9. The neuropeptide allatostatin A regulates metabolism and feeding decisions in Drosophila

    DEFF Research Database (Denmark)

    Hentze, Julie Lilith; Carlsson, Mikael A.; Kondo, Shu

    2015-01-01

    Coordinating metabolism and feeding is important to avoid obesity and metabolic diseases, yet the underlying mechanisms, balancing nutrient intake and metabolic expenditure, are poorly understood. Several mechanisms controlling these processes are conserved in Drosophila, where homeostasis and en...

  10. Distinct functions for the Drosophila piRNA pathway in genome maintenance and telomere protection.

    Directory of Open Access Journals (Sweden)

    Jaspreet S Khurana

    2010-12-01

    Full Text Available Transposons and other selfish DNA elements can be found in all phyla, and mobilization of these elements can compromise genome integrity. The piRNA (PIWI-interacting RNA pathway silences transposons in the germline, but it is unclear if this pathway has additional functions during development. Here we show that mutations in the Drosophila piRNA pathway genes, armi, aub, ago3, and rhi, lead to extensive fragmentation of the zygotic genome during the cleavage stage of embryonic divisions. Additionally, aub and armi show defects in telomere resolution during meiosis and the cleavage divisions; and mutations in lig-IV, which disrupt non-homologous end joining, suppress these fusions. By contrast, lig-IV mutations enhance chromosome fragmentation. Chromatin immunoprecipitation studies show that aub and armi mutations disrupt telomere binding of HOAP, which is a component of the telomere protection complex, and reduce expression of a subpopulation of 19- to 22-nt telomere-specific piRNAs. Mutations in rhi and ago3, by contrast, do not block HOAP binding or production of these piRNAs. These findings uncover genetically separable functions for the Drosophila piRNA pathway. The aub, armi, rhi, and ago3 genes silence transposons and maintain chromosome integrity during cleavage-stage embryonic divisions. However, the aub and armi genes have an additional function in assembly of the telomere protection complex.

  11. The Drosophila IKK-related kinase (Ik2 and Spindle-F proteins are part of a complex that regulates cytoskeleton organization during oogenesis

    Directory of Open Access Journals (Sweden)

    Shaanan Boaz

    2008-09-01

    Full Text Available Abstract Background IkappaB kinases (IKKs regulate the activity of Rel/NF-kappaB transcription factors by targeting their inhibitory partner proteins, IkappaBs, for degradation. The Drosophila genome encodes two members of the IKK family. Whereas the first is a kinase essential for activation of the NF-kappaB pathway, the latter does not act as IkappaB kinase. Instead, recent findings indicate that Ik2 regulates F-actin assembly by mediating the function of nonapoptotic caspases via degradation of DIAP1. Also, it has been suggested that ik2 regulates interactions between the minus ends of the microtubules and the actin-rich cortex in the oocyte. Since spn-F mutants display oocyte defects similar to those of ik2 mutant, we decided to investigate whether Spn-F could be a direct regulatory target of Ik2. Results We found that Ik2 binds physically to Spn-F, biomolecular interaction analysis of Spn-F and Ik2 demonstrating that both proteins bind directly and form a complex. We showed that Ik2 phosphorylates Spn-F and demonstrated that this phosphorylation does not lead to Spn-F degradation. Ik2 is localized to the anterior ring of the oocyte and to punctate structures in the nurse cells together with Spn-F protein, and both proteins are mutually required for their localization. Conclusion We conclude that Ik2 and Spn-F form a complex, which regulates cytoskeleton organization during Drosophila oogenesis and in which Spn-F is the direct regulatory target for Ik2. Interestingly, Ik2 in this complex does not function as a typical IKK in that it does not direct SpnF for degradation following phosphorylation.

  12. Drosophila Regulate Yeast Density and Increase Yeast Community Similarity in a Natural Substrate

    OpenAIRE

    Stamps, Judy A.; Yang, Louie H.; Morales, Vanessa M.; Boundy-Mills, Kyria L.

    2012-01-01

    Drosophila melanogaster adults and larvae, but especially larvae, had profound effects on the densities and community structure of yeasts that developed in banana fruits. Pieces of fruit exposed to adult female flies previously fed fly-conditioned bananas developed higher yeast densities than pieces of the same fruits that were not exposed to flies, supporting previous suggestions that adult Drosophila vector yeasts to new substrates. However, larvae alone had dramatic effects on yeast densit...

  13. Genome-wide analysis reveals novel regulators of growth in Drosophila melanogaster

    OpenAIRE

    Vonesch, Sibylle; Mackay, Trudy; Lamparter, David; Hafen, Ernst; Bergmann, Sven

    2015-01-01

    Organismal size depends on the interplay between genetic and environmental factors. Genome-wide association (GWA) analyses in humans have implied many genes in the control of height but suffer from the inability to control the environment. Genetic analyses in Drosophila have identified conserved signaling pathways controlling size; however, how these pathways control phenotypic diversity is unclear. We performed GWA of size traits using the Drosophila Genetic Reference Panel of inbred, sequen...

  14. The Drosophila FoxA ortholog Fork head regulates growth and gene expression downstream of Target of rapamycin.

    Directory of Open Access Journals (Sweden)

    Margret H Bülow

    2010-12-01

    Full Text Available Forkhead transcription factors of the FoxO subfamily regulate gene expression programs downstream of the insulin signaling network. It is less clear which proteins mediate transcriptional control exerted by Target of rapamycin (TOR signaling, but recent studies in nematodes suggest a role for FoxA transcription factors downstream of TOR. In this study we present evidence that outlines a similar connection in Drosophila, in which the FoxA protein Fork head (FKH regulates cellular and organismal size downstream of TOR. We find that ectopic expression and targeted knockdown of FKH in larval tissues elicits different size phenotypes depending on nutrient state and TOR signaling levels. FKH overexpression has a negative effect on growth under fed conditions, and this phenotype is not further exacerbated by inhibition of TOR via rapamycin feeding. Under conditions of starvation or low TOR signaling levels, knockdown of FKH attenuates the size reduction associated with these conditions. Subcellular localization of endogenous FKH protein is shifted from predominantly cytoplasmic on a high-protein diet to a pronounced nuclear accumulation in animals with reduced levels of TOR or fed with rapamycin. Two putative FKH target genes, CG6770 and cabut, are transcriptionally induced by rapamycin or FKH expression, and silenced by FKH knockdown. Induction of both target genes in heterozygous TOR mutant animals is suppressed by mutations in fkh. Furthermore, TOR signaling levels and FKH impact on transcription of the dFOXO target gene d4E-BP, implying a point of crosstalk with the insulin pathway. In summary, our observations show that an alteration of FKH levels has an effect on cellular and organismal size, and that FKH function is required for the growth inhibition and target gene induction caused by low TOR signaling levels.

  15. bantam miRNA is important for Drosophila blood cell homeostasis and a regulator of proliferation in the hematopoietic progenitor niche

    Energy Technology Data Exchange (ETDEWEB)

    Lam, Victoria; Tokusumi, Tsuyoshi; Tokusumi, Yumiko; Schulz, Robert A., E-mail: rschulz@nd.edu

    2014-10-24

    Highlights: • bantam miRNA is endogenously expressed in the hematopoietic progenitor niche. • bantam is necessary and sufficient to induce cellular proliferation in the PSC. • bantam is upstream of the Insulin Receptor signaling pathway. • A model for positive regulation of hematopoietic niche growth is proposed. - Abstract: The Drosophila hematopoietic system is utilized in this study to gain novel insights into the process of growth control of the hematopoietic progenitor niche in blood development. The niche microenvironment is an essential component controlling the balance between progenitor populations and differentiated, mature blood cells and has been shown to lead to hematopoietic malignancies in humans when misregulated. MicroRNAs are one class of regulators associated with blood malignancies; however, there remains a relative paucity of information about the role of miRNAs in the niche. Here we demonstrate that bantam miRNA is endogenously active in the Drosophila hematopoietic progenitor niche, the posterior signaling center (PSC), and functions in the primary hematopoietic organ, the lymph gland, as a positive regulator of growth. Loss of bantam leads to a significant reduction in the PSC and overall lymph gland size, as well as a loss of the progenitor population and correlative premature differentiation of mature hemocytes. Interestingly, in addition to being essential for proper lymph gland development, we have determined bantam to be a novel upstream component of the insulin signaling cascade in the PSC and have unveiled dMyc as one factor central to bantam activity. These important findings identify bantam as a new hematopoietic regulator, place it in an evolutionarily conserved signaling pathway, present one way in which it is regulated, and provide a mechanism through which it facilitates cellular proliferation in the hematopoietic niche.

  16. Hindsight regulates photoreceptor axon targeting through transcriptional control of jitterbug/Filamin and multiple genes involved in axon guidance in Drosophila.

    Science.gov (United States)

    Oliva, Carlos; Molina-Fernandez, Claudia; Maureira, Miguel; Candia, Noemi; López, Estefanía; Hassan, Bassem; Aerts, Stein; Cánovas, José; Olguín, Patricio; Sierralta, Jimena

    2015-09-01

    During axon targeting, a stereotyped pattern of connectivity is achieved by the integration of intrinsic genetic programs and the response to extrinsic long and short-range directional cues. How this coordination occurs is the subject of intense study. Transcription factors play a central role due to their ability to regulate the expression of multiple genes required to sense and respond to these cues during development. Here we show that the transcription factor HNT regulates layer-specific photoreceptor axon targeting in Drosophila through transcriptional control of jbug/Filamin and multiple genes involved in axon guidance and cytoskeleton organization.Using a microarray analysis we identified 235 genes whose expression levels were changed by HNT overexpression in the eye primordia. We analyzed nine candidate genes involved in cytoskeleton regulation and axon guidance, six of which displayed significantly altered gene expression levels in hnt mutant retinas. Functional analysis confirmed the role of OTK/PTK7 in photoreceptor axon targeting and uncovered Tiggrin, an integrin ligand, and Jbug/Filamin, a conserved actin- binding protein, as new factors that participate of photoreceptor axon targeting. Moreover, we provided in silico and molecular evidence that supports jbug/Filamin as a direct transcriptional target of HNT and that HNT acts partially through Jbug/Filamin in vivo to regulate axon guidance. Our work broadens the understanding of how HNT regulates the coordinated expression of a group of genes to achieve the correct connectivity pattern in the Drosophila visual system. © 2015 Wiley Periodicals, Inc. Develop Neurobiol 75: 1018-1032, 2015. © 2015 Wiley Periodicals, Inc.

  17. bantam miRNA is important for Drosophila blood cell homeostasis and a regulator of proliferation in the hematopoietic progenitor niche

    International Nuclear Information System (INIS)

    Lam, Victoria; Tokusumi, Tsuyoshi; Tokusumi, Yumiko; Schulz, Robert A.

    2014-01-01

    Highlights: • bantam miRNA is endogenously expressed in the hematopoietic progenitor niche. • bantam is necessary and sufficient to induce cellular proliferation in the PSC. • bantam is upstream of the Insulin Receptor signaling pathway. • A model for positive regulation of hematopoietic niche growth is proposed. - Abstract: The Drosophila hematopoietic system is utilized in this study to gain novel insights into the process of growth control of the hematopoietic progenitor niche in blood development. The niche microenvironment is an essential component controlling the balance between progenitor populations and differentiated, mature blood cells and has been shown to lead to hematopoietic malignancies in humans when misregulated. MicroRNAs are one class of regulators associated with blood malignancies; however, there remains a relative paucity of information about the role of miRNAs in the niche. Here we demonstrate that bantam miRNA is endogenously active in the Drosophila hematopoietic progenitor niche, the posterior signaling center (PSC), and functions in the primary hematopoietic organ, the lymph gland, as a positive regulator of growth. Loss of bantam leads to a significant reduction in the PSC and overall lymph gland size, as well as a loss of the progenitor population and correlative premature differentiation of mature hemocytes. Interestingly, in addition to being essential for proper lymph gland development, we have determined bantam to be a novel upstream component of the insulin signaling cascade in the PSC and have unveiled dMyc as one factor central to bantam activity. These important findings identify bantam as a new hematopoietic regulator, place it in an evolutionarily conserved signaling pathway, present one way in which it is regulated, and provide a mechanism through which it facilitates cellular proliferation in the hematopoietic niche

  18. Changes of mitochondrial ultrastructure and function during ageing in mice and Drosophila.

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    Brandt, Tobias; Mourier, Arnaud; Tain, Luke S; Partridge, Linda; Larsson, Nils-Göran; Kühlbrandt, Werner

    2017-07-12

    Ageing is a progressive decline of intrinsic physiological functions. We examined the impact of ageing on the ultrastructure and function of mitochondria in mouse and fruit flies ( Drosophila melanogaster ) by electron cryo-tomography and respirometry. We discovered distinct age-related changes in both model organisms. Mitochondrial function and ultrastructure are maintained in mouse heart, whereas subpopulations of mitochondria from mouse liver show age-related changes in membrane morphology. Subpopulations of mitochondria from young and old mouse kidney resemble those described for apoptosis. In aged flies, respiratory activity is compromised and the production of peroxide radicals is increased. In about 50% of mitochondria from old flies, the inner membrane organization breaks down. This establishes a clear link between inner membrane architecture and functional decline. Mitochondria were affected by ageing to very different extents, depending on the organism and possibly on the degree to which tissues within the same organism are protected against mitochondrial damage.

  19. Global regulation of mRNA translation and stability in the early Drosophila embryo by the Smaug RNA-binding protein.

    Science.gov (United States)

    Chen, Linan; Dumelie, Jason G; Li, Xiao; Cheng, Matthew Hk; Yang, Zhiyong; Laver, John D; Siddiqui, Najeeb U; Westwood, J Timothy; Morris, Quaid; Lipshitz, Howard D; Smibert, Craig A

    2014-01-07

    Smaug is an RNA-binding protein that induces the degradation and represses the translation of mRNAs in the early Drosophila embryo. Smaug has two identified direct target mRNAs that it differentially regulates: nanos and Hsp83. Smaug represses the translation of nanos mRNA but has only a modest effect on its stability, whereas it destabilizes Hsp83 mRNA but has no detectable effect on Hsp83 translation. Smaug is required to destabilize more than one thousand mRNAs in the early embryo, but whether these transcripts represent direct targets of Smaug is unclear and the extent of Smaug-mediated translational repression is unknown. To gain a panoramic view of Smaug function in the early embryo, we identified mRNAs that are bound to Smaug using RNA co-immunoprecipitation followed by hybridization to DNA microarrays. We also identified mRNAs that are translationally repressed by Smaug using polysome gradients and microarrays. Comparison of the bound mRNAs to those that are translationally repressed by Smaug and those that require Smaug for their degradation suggests that a large fraction of Smaug's target mRNAs are both translationally repressed and degraded by Smaug. Smaug directly regulates components of the TRiC/CCT chaperonin, the proteasome regulatory particle and lipid droplets, as well as many metabolic enzymes, including several glycolytic enzymes. Smaug plays a direct and global role in regulating the translation and stability of a large fraction of the mRNAs in the early Drosophila embryo, and has unanticipated functions in control of protein folding and degradation, lipid droplet function and metabolism.

  20. Lasp-1 regulates podosome function.

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    Miriam Stölting

    Full Text Available Eukaryotic cells form a variety of adhesive structures to connect with their environment and to regulate cell motility. In contrast to classical focal adhesions, podosomes, highly dynamic structures of different cell types, are actively engaged in matrix remodelling and degradation. Podosomes are composed of an actin-rich core region surrounded by a ring-like structure containing signalling molecules, motor proteins as well as cytoskeleton-associated proteins. Lasp-1 is a ubiquitously expressed, actin-binding protein that is known to regulate cytoskeleton architecture and cell migration. This multidomain protein is predominantely present at focal adhesions, however, a second pool of Lasp-1 molecules is also found at lamellipodia and vesicle-like microdomains in the cytosol.In this report, we show that Lasp-1 is a novel component and regulator of podosomes. Immunofluorescence studies reveal a localization of Lasp-1 in the podosome ring structure, where it colocalizes with zyxin and vinculin. Life cell imaging experiments demonstrate that Lasp-1 is recruited in early steps of podosome assembly. A siRNA-mediated Lasp-1 knockdown in human macrophages affects podosome dynamics as well as their matrix degradation capacity. In summary, our data indicate that Lasp-1 is a novel component of podosomes and is involved in the regulation of podosomal function.

  1. Redox Regulation of Mitochondrial Function

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    Handy, Diane E.

    2012-01-01

    Abstract Redox-dependent processes influence most cellular functions, such as differentiation, proliferation, and apoptosis. Mitochondria are at the center of these processes, as mitochondria both generate reactive oxygen species (ROS) that drive redox-sensitive events and respond to ROS-mediated changes in the cellular redox state. In this review, we examine the regulation of cellular ROS, their modes of production and removal, and the redox-sensitive targets that are modified by their flux. In particular, we focus on the actions of redox-sensitive targets that alter mitochondrial function and the role of these redox modifications on metabolism, mitochondrial biogenesis, receptor-mediated signaling, and apoptotic pathways. We also consider the role of mitochondria in modulating these pathways, and discuss how redox-dependent events may contribute to pathobiology by altering mitochondrial function. Antioxid. Redox Signal. 16, 1323–1367. PMID:22146081

  2. Precise temporal regulation of roughest is required for correct salivary gland autophagic cell death in Drosophila.

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    Simon, Claudio R; Moda, Livia M R; Octacilio-Silva, Shirlei; Anhezini, Lucas; Machado-Gitai, Luciana C H; Ramos, Ricardo Guelerman P

    2009-07-01

    The Drosophila roughest (rst) locus encodes an immunoglobulin superfamily transmembrane glycoprotein implicated in a variety of embryonic and postembryonic developmental processes. Here we demonstrate a previously unnoticed role for this gene in the autophagic elimination of larval salivary glands during early pupal stages by showing that overexpression of the Rst protein ectodomain in early pupa leads to persistence of salivary glands up to at least 12 hours after head eversion, although with variable penetrance. The same phenotype is observed in individuals carrying the dominant regulatory allele rst(D), but not in loss of function alleles. Analysis of persistent glands at the ultrastructural level showed that programmed cell death starts at the right time but is arrested at an early stage of the process. Finally we describe the expression pattern and intracellular distribution of Rst in wild type and rst(D) mutants, showing that its downregulation in salivary glands at the beginning of pupal stage is an important factor in the correct implementation of the autophagic program of this tissue in space and time. 2009 Wiley-Liss, Inc.

  3. The Drosophila blood-brain barrier: Development and function of a glial endothelium

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    Stefanie eLimmer

    2014-11-01

    Full Text Available The efficacy of neuronal function requires a well-balanced extracellular ion homeostasis and a steady supply with nutrients and metabolites. Therefore, all organisms equipped with a complex nervous system developed a so-called blood-brain barrier, protecting it from an uncontrolled entry of solutes, metabolites or pathogens. In higher vertebrates, this diffusion barrier is established by polarized endothelial cells that form extensive tight junctions, whereas in lower vertebrates and invertebrates the blood-brain barrier is exclusively formed by glial cells. Here, we review the development and function of the glial blood-brain barrier of Drosophila melanogaster. In the Drosophila nervous system, at least seven morphologically distinct glial cell classes can be distinguished. Two of these glial classes form the blood-brain barrier. Perineurial glial cells participate in nutrient uptake and establish a first diffusion barrier. The subperineurial glial cells form septate junctions, which block paracellular diffusion and thus seal the nervous system from the hemolymph. We summarize the molecular basis of septate junction formation and address the different transport systems expressed by the blood-brain barrier forming glial cells.

  4. The Drosophila blood-brain barrier: development and function of a glial endothelium.

    Science.gov (United States)

    Limmer, Stefanie; Weiler, Astrid; Volkenhoff, Anne; Babatz, Felix; Klämbt, Christian

    2014-01-01

    The efficacy of neuronal function requires a well-balanced extracellular ion homeostasis and a steady supply with nutrients and metabolites. Therefore, all organisms equipped with a complex nervous system developed a so-called blood-brain barrier, protecting it from an uncontrolled entry of solutes, metabolites or pathogens. In higher vertebrates, this diffusion barrier is established by polarized endothelial cells that form extensive tight junctions, whereas in lower vertebrates and invertebrates the blood-brain barrier is exclusively formed by glial cells. Here, we review the development and function of the glial blood-brain barrier of Drosophila melanogaster. In the Drosophila nervous system, at least seven morphologically distinct glial cell classes can be distinguished. Two of these glial classes form the blood-brain barrier. Perineurial glial cells participate in nutrient uptake and establish a first diffusion barrier. The subperineurial glial (SPG) cells form septate junctions, which block paracellular diffusion and thus seal the nervous system from the hemolymph. We summarize the molecular basis of septate junction formation and address the different transport systems expressed by the blood-brain barrier forming glial cells.

  5. Polytene Chromosomes - A Portrait of Functional Organization of the Drosophila Genome.

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    Zykova, Tatyana Yu; Levitsky, Victor G; Belyaeva, Elena S; Zhimulev, Igor F

    2018-04-01

    This mini-review is devoted to the problem genetic meaning of main polytene chromosome structures - bands and interbands. Generally, densely packed chromatin forms black bands, moderately condensed regions form grey loose bands, whereas decondensed regions of the genome appear as interbands. Recent progress in the annotation of the Drosophila genome and epigenome has made it possible to compare the banding pattern and the structural organization of genes, as well as their activity. This was greatly aided by our ability to establish the borders of bands and interbands on the physical map, which allowed to perform comprehensive side-by-side comparisons of cytology, genetic and epigenetic maps and to uncover the association between the morphological structures and the functional domains of the genome. These studies largely conclude that interbands 5'-ends of housekeeping genes that are active across all cell types. Interbands are enriched with proteins involved in transcription and nucleosome remodeling, as well as with active histone modifications. Notably, most of the replication origins map to interband regions. As for grey loose bands adjacent to interbands, they typically host the bodies of house-keeping genes. Thus, the bipartite structure composed of an interband and an adjacent grey band functions as a standalone genetic unit. Finally, black bands harbor tissue-specific genes with narrow temporal and tissue expression profiles. Thus, the uniform and permanent activity of interbands combined with the inactivity of genes in bands forms the basis of the universal banding pattern observed in various Drosophila tissues.

  6. Mechanism of Notch Pathway Activation and Its Role in the Regulation of Olfactory Plasticity in Drosophila melanogaster.

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    Simon Kidd

    Full Text Available The neural plasticity of sensory systems is being increasingly recognized as playing a role in learning and memory. We have previously shown that Notch, part of an evolutionarily conserved intercellular signaling pathway, is required in adult Drosophila melanogaster olfactory receptor neurons (ORNs for the structural and functional plasticity of olfactory glomeruli that is induced by chronic odor exposure. In this paper we address how long-term exposure to odor activates Notch and how Notch in conjunction with chronic odor mediates olfactory plasticity. We show that upon chronic odor exposure a non-canonical Notch pathway mediates an increase in the volume of glomeruli by a mechanism that is autonomous to ORNs. In addition to activating a pathway that is autonomous to ORNs, chronic odor exposure also activates the Notch ligand Delta in second order projection neurons (PNs, but this does not appear to require acetylcholine receptor activation in PNs. Delta on PNs then feeds back to activate canonical Notch signaling in ORNs, which restricts the extent of the odor induced increase in glomerular volume. Surprisingly, even though the pathway that mediates the increase in glomerular volume is autonomous to ORNs, nonproductive transsynaptic Delta/Notch interactions that do not activate the canonical pathway can block the increase in volume. In conjunction with chronic odor, the canonical Notch pathway also enhances cholinergic activation of PNs. We present evidence suggesting that this is due to increased acetylcholine release from ORNs. In regulating physiological plasticity, Notch functions solely by the canonical pathway, suggesting that there is no direct connection between morphological and physiological plasticity.

  7. The Gemin associates of survival motor neuron are required for motor function in Drosophila.

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    Borg, Rebecca; Cauchi, Ruben J

    2013-01-01

    Membership of the survival motor neuron (SMN) complex extends to nine factors, including the SMN protein, the product of the spinal muscular atrophy (SMA) disease gene, Gemins 2-8 and Unrip. The best-characterised function of this macromolecular machine is the assembly of the Sm-class of uridine-rich small nuclear ribonucleoprotein (snRNP) particles and each SMN complex member has a key role during this process. So far, however, only little is known about the function of the individual Gemin components in vivo. Here, we make use of the Drosophila model organism to uncover loss-of-function phenotypes of Gemin2, Gemin3 and Gemin5, which together with SMN form the minimalistic fly SMN complex. We show that ectopic overexpression of the dead helicase Gem3(ΔN) mutant or knockdown of Gemin3 result in similar motor phenotypes, when restricted to muscle, and in combination cause lethality, hence suggesting that Gem3(ΔN) overexpression mimics a loss-of-function. Based on the localisation pattern of Gem3(ΔN), we predict that the nucleus is the primary site of the antimorphic or dominant-negative mechanism of Gem3(ΔN)-mediated interference. Interestingly, phenotypes induced by human SMN overexpression in Drosophila exhibit similarities to those induced by overexpression of Gem3(ΔN). Through enhanced knockdown we also uncover a requirement of Gemin2, Gemin3 and Gemin5 for viability and motor behaviour, including locomotion as well as flight, in muscle. Notably, in the case of Gemin3 and Gemin5, such function also depends on adequate levels of the respective protein in neurons. Overall, these findings lead us to speculate that absence of any one member is sufficient to arrest the SMN-Gemins complex function in a nucleocentric pathway, which is critical for motor function in vivo.

  8. Inverse regulation of two classic Hippo pathway target genes in Drosophila by the dimerization hub protein Ctp.

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    Barron, Daniel A; Moberg, Kenneth

    2016-03-14

    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, we find 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.

  9. Cap-n-Collar Promotes Tissue Regeneration by Regulating ROS and JNK Signaling in the Drosophila melanogaster Wing Imaginal Disc.

    Science.gov (United States)

    Brock, Amanda R; Seto, Mabel; Smith-Bolton, Rachel K

    2017-07-01

    Regeneration is a complex process that requires an organism to recognize and repair tissue damage, as well as grow and pattern new tissue. Here, we describe a genetic screen to identify novel regulators of regeneration. We ablated the Drosophila melanogaster larval wing primordium by inducing apoptosis in a spatially and temporally controlled manner and allowed the tissue to regenerate and repattern. To identify genes that regulate regeneration, we carried out a dominant-modifier screen by assessing the amount and quality of regeneration in adult wings heterozygous for isogenic deficiencies. We have identified 31 regions on the right arm of the third chromosome that modify the regenerative response. Interestingly, we observed several distinct phenotypes: mutants that regenerated poorly, mutants that regenerated faster or better than wild-type, and mutants that regenerated imperfectly and had patterning defects. We mapped one deficiency region to cap-n-collar ( cnc ), the Drosophila Nrf2 ortholog, which is required for regeneration. Cnc regulates reactive oxygen species levels in the regenerating epithelium, and affects c-Jun N-terminal protein kinase (JNK) signaling, growth, debris localization, and pupariation timing. Here, we present the results of our screen and propose a model wherein Cnc regulates regeneration by maintaining an optimal level of reactive oxygen species to promote JNK signaling. Copyright © 2017 by the Genetics Society of America.

  10. Model-Based Analysis for Qualitative Data: An Application in Drosophila Germline Stem Cell Regulation

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    Pargett, Michael; Rundell, Ann E.; Buzzard, Gregery T.; Umulis, David M.

    2014-01-01

    Discovery in developmental biology is often driven by intuition that relies on the integration of multiple types of data such as fluorescent images, phenotypes, and the outcomes of biochemical assays. Mathematical modeling helps elucidate the biological mechanisms at play as the networks become increasingly large and complex. However, the available data is frequently under-utilized due to incompatibility with quantitative model tuning techniques. This is the case for stem cell regulation mechanisms explored in the Drosophila germarium through fluorescent immunohistochemistry. To enable better integration of biological data with modeling in this and similar situations, we have developed a general parameter estimation process to quantitatively optimize models with qualitative data. The process employs a modified version of the Optimal Scaling method from social and behavioral sciences, and multi-objective optimization to evaluate the trade-off between fitting different datasets (e.g. wild type vs. mutant). Using only published imaging data in the germarium, we first evaluated support for a published intracellular regulatory network by considering alternative connections of the same regulatory players. Simply screening networks against wild type data identified hundreds of feasible alternatives. Of these, five parsimonious variants were found and compared by multi-objective analysis including mutant data and dynamic constraints. With these data, the current model is supported over the alternatives, but support for a biochemically observed feedback element is weak (i.e. these data do not measure the feedback effect well). When also comparing new hypothetical models, the available data do not discriminate. To begin addressing the limitations in data, we performed a model-based experiment design and provide recommendations for experiments to refine model parameters and discriminate increasingly complex hypotheses. PMID:24626201

  11. Drosophila sosie functions with βH-Spectrin and actin organizers in cell migration, epithelial morphogenesis and cortical stability

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    Urwyler, Olivier; Cortinas-Elizondo, Fabiola; Suter, Beat

    2012-01-01

    Summary Morphogenesis in multicellular organisms requires the careful coordination of cytoskeletal elements, dynamic regulation of cell adhesion and extensive cell migration. sosie (sie) is a novel gene required in various morphogenesis processes in Drosophila oogenesis. Lack of sie interferes with normal egg chamber packaging, maintenance of epithelial integrity and control of follicle cell migration, indicating that sie is involved in controlling epithelial integrity and cell migration. For these functions sie is required both in the germ line and in the soma. Consistent with this, Sosie localizes to plasma membranes in the germ line and in the somatic follicle cells and is predicted to present an EGF-like domain on the extracellular side. Two positively charged residues, C-terminal to the predicted transmembrane domain (on the cytoplasmic side), are required for normal plasma membrane localization of Sosie. Because sie also contributes to normal cortical localization of βH-Spectrin, it appears that cortical βH-Spectrin mediates some of the functions of sosie. sie also interacts with the genes coding for the actin organizers Filamin and Profilin and, in the absence of sie function, F-actin is less well organized and nurse cells frequently fuse. PMID:23213377

  12. Drosophila sosie functions with β(H)-Spectrin and actin organizers in cell migration, epithelial morphogenesis and cortical stability.

    Science.gov (United States)

    Urwyler, Olivier; Cortinas-Elizondo, Fabiola; Suter, Beat

    2012-10-15

    Morphogenesis in multicellular organisms requires the careful coordination of cytoskeletal elements, dynamic regulation of cell adhesion and extensive cell migration. sosie (sie) is a novel gene required in various morphogenesis processes in Drosophila oogenesis. Lack of sie interferes with normal egg chamber packaging, maintenance of epithelial integrity and control of follicle cell migration, indicating that sie is involved in controlling epithelial integrity and cell migration. For these functions sie is required both in the germ line and in the soma. Consistent with this, Sosie localizes to plasma membranes in the germ line and in the somatic follicle cells and is predicted to present an EGF-like domain on the extracellular side. Two positively charged residues, C-terminal to the predicted transmembrane domain (on the cytoplasmic side), are required for normal plasma membrane localization of Sosie. Because sie also contributes to normal cortical localization of β(H)-Spectrin, it appears that cortical β(H)-Spectrin mediates some of the functions of sosie. sie also interacts with the genes coding for the actin organizers Filamin and Profilin and, in the absence of sie function, F-actin is less well organized and nurse cells frequently fuse.

  13. Roles for the VCP co-factors Npl4 and Ufd1 in neuronal function in Drosophila melanogaster.

    Science.gov (United States)

    Byrne, Dwayne J; Harmon, Mark J; Simpson, Jeremy C; Blackstone, Craig; O'Sullivan, Niamh C

    2017-10-20

    The VCP-Ufd1-Npl4 complex regulates proteasomal processing within cells by delivering ubiquitinated proteins to the proteasome for degradation. Mutations in VCP are associated with two neurodegenerative diseases, amyotrophic lateral sclerosis (ALS) and inclusion body myopathy with Paget's disease of the bone and frontotemporal dementia (IBMPFD), and extensive study has revealed crucial functions of VCP within neurons. By contrast, little is known about the functions of Npl4 or Ufd1 in vivo. Using neuronal-specific knockdown of Npl4 or Ufd1 in Drosophila melanogaster, we infer that Npl4 contributes to microtubule organization within developing motor neurons. Moreover, Npl4 RNAi flies present with neurodegenerative phenotypes including progressive locomotor deficits, reduced lifespan and increased accumulation of TAR DNA-binding protein-43 homolog (TBPH). Knockdown, but not overexpression, of TBPH also exacerbates Npl4 RNAi-associated adult-onset neurodegenerative phenotypes. In contrast, we find that neuronal knockdown of Ufd1 has little effect on neuromuscular junction (NMJ) organization, TBPH accumulation or adult behaviour. These findings suggest the differing neuronal functions of Npl4 and Ufd1 in vivo. Copyright © 2017 The Authors. Published by Elsevier Ltd.. All rights reserved.

  14. A Genome-wide RNAi Screen for Microtubule Bundle Formation and Lysosome Motility Regulation in Drosophila S2 Cells

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    Amber L. Jolly

    2016-01-01

    Full Text Available Long-distance intracellular transport of organelles, mRNA, and proteins (“cargo” occurs along the microtubule cytoskeleton by the action of kinesin and dynein motor proteins, but the vast network of factors involved in regulating intracellular cargo transport are still unknown. We capitalize on the Drosophila melanogaster S2 model cell system to monitor lysosome transport along microtubule bundles, which require enzymatically active kinesin-1 motor protein for their formation. We use an automated tracking program and a naive Bayesian classifier for the multivariate motility data to analyze 15,683 gene phenotypes and find 98 proteins involved in regulating lysosome motility along microtubules and 48 involved in the formation of microtubule filled processes in S2 cells. We identify innate immunity genes, ion channels, and signaling proteins having a role in lysosome motility regulation and find an unexpected relationship between the dynein motor, Rab7a, and lysosome motility regulation.

  15. The apical scaffold big bang binds to spectrins and regulates the growth of Drosophila melanogaster wing discs.

    Science.gov (United States)

    Forest, Elodie; Logeay, Rémi; Géminard, Charles; Kantar, Diala; Frayssinoux, Florence; Heron-Milhavet, Lisa; Djiane, Alexandre

    2018-03-05

    During development, cell numbers are tightly regulated, ensuring that tissues and organs reach their correct size and shape. Recent evidence has highlighted the intricate connections between the cytoskeleton and the regulation of the key growth control Hippo pathway. Looking for apical scaffolds regulating tissue growth, we describe that Drosophila melanogaster big bang (Bbg), a poorly characterized multi-PDZ scaffold, controls epithelial tissue growth without affecting epithelial polarity and architecture. bbg -mutant tissues are smaller, with fewer cells that are less apically constricted than normal. We show that Bbg binds to and colocalizes tightly with the β-heavy-Spectrin/Kst subunit at the apical cortex and promotes Yki activity, F-actin enrichment, and the phosphorylation of the myosin II regulatory light chain Spaghetti squash. We propose a model in which the spectrin cytoskeleton recruits Bbg to the cortex, where Bbg promotes actomyosin contractility to regulate epithelial tissue growth. © 2018 Forest et al.

  16. The influence of Adh function on ethanol preference and tolerance in adult Drosophila melanogaster.

    Science.gov (United States)

    Ogueta, Maite; Cibik, Osman; Eltrop, Rouven; Schneider, Andrea; Scholz, Henrike

    2010-11-01

    Preference determines behavioral choices such as choosing among food sources and mates. One preference-affecting chemical is ethanol, which guides insects to fermenting fruits or leaves. Here, we show that adult Drosophila melanogaster prefer food containing up to 5% ethanol over food without ethanol and avoid food with high levels (23%) of ethanol. Although female and male flies behaved differently at ethanol-containing food sources, there was no sexual dimorphism in the preference for food containing modest ethanol levels. We also investigated whether Drosophila preference, sensitivity and tolerance to ethanol was related to the activity of alcohol dehydrogenase (Adh), the primary ethanol-metabolizing enzyme in D. melanogaster. Impaired Adh function reduced ethanol preference in both D. melanogaster and a related species, D. sechellia. Adh-impaired flies also displayed reduced aversion to high ethanol concentrations, increased sensitivity to the effects of ethanol on postural control, and negative tolerance/sensitization (i.e., a reduction of the increased resistance to ethanol's effects that normally occurs upon repeated exposure). These data strongly indicate a linkage between ethanol-induced behavior and ethanol metabolism in adult fruit flies: Adh deficiency resulted in reduced preference to low ethanol concentrations and reduced aversion to high ones, despite recovery from ethanol being strongly impaired.

  17. Notch is required in adult Drosophila sensory neurons for morphological and functional plasticity of the olfactory circuit.

    Directory of Open Access Journals (Sweden)

    Simon Kidd

    2015-05-01

    Full Text Available Olfactory receptor neurons (ORNs convey odor information to the central brain, but like other sensory neurons were thought to play a passive role in memory formation and storage. Here we show that Notch, part of an evolutionarily conserved intercellular signaling pathway, is required in adult Drosophila ORNs for the structural and functional plasticity of olfactory glomeruli that is induced by chronic odor exposure. Specifically, we show that Notch activity in ORNs is necessary for the odor specific increase in the volume of glomeruli that occurs as a consequence of prolonged odor exposure. Calcium imaging experiments indicate that Notch in ORNs is also required for the chronic odor induced changes in the physiology of ORNs and the ensuing changes in the physiological response of their second order projection neurons (PNs. We further show that Notch in ORNs acts by both canonical cleavage-dependent and non-canonical cleavage-independent pathways. The Notch ligand Delta (Dl in PNs switches the balance between the pathways. These data define a circuit whereby, in conjunction with odor, N activity in the periphery regulates the activity of neurons in the central brain and Dl in the central brain regulates N activity in the periphery. Our work highlights the importance of experience dependent plasticity at the first olfactory synapse.

  18. New genes often acquire male-specific functions but rarely become essential in Drosophila.

    Science.gov (United States)

    Kondo, Shu; Vedanayagam, Jeffrey; Mohammed, Jaaved; Eizadshenass, Sogol; Kan, Lijuan; Pang, Nan; Aradhya, Rajaguru; Siepel, Adam; Steinhauer, Josefa; Lai, Eric C

    2017-09-15

    Relatively little is known about the in vivo functions of newly emerging genes, especially in metazoans. Although prior RNAi studies reported prevalent lethality among young gene knockdowns, our phylogenomic analyses reveal that young Drosophila genes are frequently restricted to the nonessential male reproductive system. We performed large-scale CRISPR/Cas9 mutagenesis of "conserved, essential" and "young, RNAi-lethal" genes and broadly confirmed the lethality of the former but the viability of the latter. Nevertheless, certain young gene mutants exhibit defective spermatogenesis and/or male sterility. Moreover, we detected widespread signatures of positive selection on young male-biased genes. Thus, young genes have a preferential impact on male reproductive system function. © 2017 Kondo et al.; Published by Cold Spring Harbor Laboratory Press.

  19. Functional architecture of reward learning in mushroom body extrinsic neurons of larval Drosophila.

    Science.gov (United States)

    Saumweber, Timo; Rohwedder, Astrid; Schleyer, Michael; Eichler, Katharina; Chen, Yi-Chun; Aso, Yoshinori; Cardona, Albert; Eschbach, Claire; Kobler, Oliver; Voigt, Anne; Durairaja, Archana; Mancini, Nino; Zlatic, Marta; Truman, James W; Thum, Andreas S; Gerber, Bertram

    2018-03-16

    The brain adaptively integrates present sensory input, past experience, and options for future action. The insect mushroom body exemplifies how a central brain structure brings about such integration. Here we use a combination of systematic single-cell labeling, connectomics, transgenic silencing, and activation experiments to study the mushroom body at single-cell resolution, focusing on the behavioral architecture of its input and output neurons (MBINs and MBONs), and of the mushroom body intrinsic APL neuron. Our results reveal the identity and morphology of almost all of these 44 neurons in stage 3 Drosophila larvae. Upon an initial screen, functional analyses focusing on the mushroom body medial lobe uncover sparse and specific functions of its dopaminergic MBINs, its MBONs, and of the GABAergic APL neuron across three behavioral tasks, namely odor preference, taste preference, and associative learning between odor and taste. Our results thus provide a cellular-resolution study case of how brains organize behavior.

  20. The GTP- and Phospholipid-Binding Protein TTD14 Regulates Trafficking of the TRPL Ion Channel in Drosophila Photoreceptor Cells

    Science.gov (United States)

    Cerny, Alexander C.; Altendorfer, André; Schopf, Krystina; Baltner, Karla; Maag, Nathalie; Sehn, Elisabeth; Wolfrum, Uwe; Huber, Armin

    2015-01-01

    Recycling of signaling proteins is a common phenomenon in diverse signaling pathways. In photoreceptors of Drosophila, light absorption by rhodopsin triggers a phospholipase Cβ-mediated opening of the ion channels transient receptor potential (TRP) and TRP-like (TRPL) and generates the visual response. The signaling proteins are located in a plasma membrane compartment called rhabdomere. The major rhodopsin (Rh1) and TRP are predominantly localized in the rhabdomere in light and darkness. In contrast, TRPL translocates between the rhabdomeral plasma membrane in the dark and a storage compartment in the cell body in the light, from where it can be recycled to the plasma membrane upon subsequent dark adaptation. Here, we identified the gene mutated in trpl translocation defective 14 (ttd14), which is required for both TRPL internalization from the rhabdomere in the light and recycling of TRPL back to the rhabdomere in the dark. TTD14 is highly conserved in invertebrates and binds GTP in vitro. The ttd14 mutation alters a conserved proline residue (P75L) in the GTP-binding domain and abolishes binding to GTP. This indicates that GTP binding is essential for TTD14 function. TTD14 is a cytosolic protein and binds to PtdIns(3)P, a lipid enriched in early endosome membranes, and to phosphatidic acid. In contrast to TRPL, rhabdomeral localization of the membrane proteins Rh1 and TRP is not affected in the ttd14 P75L mutant. The ttd14 P75L mutation results in Rh1-independent photoreceptor degeneration and larval lethality suggesting that other processes are also affected by the ttd14 P75L mutation. In conclusion, TTD14 is a novel regulator of TRPL trafficking, involved in internalization and subsequent sorting of TRPL into the recycling pathway that enables this ion channel to return to the plasma membrane. PMID:26509977

  1. The GTP- and Phospholipid-Binding Protein TTD14 Regulates Trafficking of the TRPL Ion Channel in Drosophila Photoreceptor Cells.

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    Alexander C Cerny

    2015-10-01

    Full Text Available Recycling of signaling proteins is a common phenomenon in diverse signaling pathways. In photoreceptors of Drosophila, light absorption by rhodopsin triggers a phospholipase Cβ-mediated opening of the ion channels transient receptor potential (TRP and TRP-like (TRPL and generates the visual response. The signaling proteins are located in a plasma membrane compartment called rhabdomere. The major rhodopsin (Rh1 and TRP are predominantly localized in the rhabdomere in light and darkness. In contrast, TRPL translocates between the rhabdomeral plasma membrane in the dark and a storage compartment in the cell body in the light, from where it can be recycled to the plasma membrane upon subsequent dark adaptation. Here, we identified the gene mutated in trpl translocation defective 14 (ttd14, which is required for both TRPL internalization from the rhabdomere in the light and recycling of TRPL back to the rhabdomere in the dark. TTD14 is highly conserved in invertebrates and binds GTP in vitro. The ttd14 mutation alters a conserved proline residue (P75L in the GTP-binding domain and abolishes binding to GTP. This indicates that GTP binding is essential for TTD14 function. TTD14 is a cytosolic protein and binds to PtdIns(3P, a lipid enriched in early endosome membranes, and to phosphatidic acid. In contrast to TRPL, rhabdomeral localization of the membrane proteins Rh1 and TRP is not affected in the ttd14P75L mutant. The ttd14P75L mutation results in Rh1-independent photoreceptor degeneration and larval lethality suggesting that other processes are also affected by the ttd14P75L mutation. In conclusion, TTD14 is a novel regulator of TRPL trafficking, involved in internalization and subsequent sorting of TRPL into the recycling pathway that enables this ion channel to return to the plasma membrane.

  2. Functional correlates of positional and gender-specific renal asymmetry in Drosophila.

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    Venkateswara R Chintapalli

    Full Text Available In humans and other animals, the internal organs are positioned asymmetrically in the body cavity, and disruption of this body plan can be fatal in humans. The mechanisms by which internal asymmetry are established are presently the subject of intense study; however, the functional significance of internal asymmetry (outside the brain is largely unexplored. Is internal asymmetry functionally significant, or merely an expedient way of packing organs into a cavity?Like humans, Drosophila shows internal asymmetry, with the gut thrown into stereotyped folds. There is also renal asymmetry, with the rightmost pair of renal (Malpighian tubules always ramifying anteriorly, and the leftmost pair always sitting posteriorly in the body cavity. Accordingly, transcriptomes of anterior-directed (right-side and posterior-directed (left-side Malpighian (renal tubules were compared in both adult male and female Drosophila. Although genes encoding the basic functions of the tubules (transport, signalling were uniformly expressed, some functions (like innate immunity showed positional or gender differences in emphasis; others, like calcium handling or the generation of potentially toxic ammonia, were reserved for just the right-side or left-side tubules, respectively. These findings correlated with the distinct locations of each tubule pair within the body cavity. Well known developmental genes (like dorsocross, dachshund and doublesex showed continuing, patterned expression in adult tubules, implying that somatic tissues maintain both left-right and gender identities throughout life. Gender asymmetry was also noted, both in defence and in male-specific expression of receptors for neuropeptide F and sex-peptide: NPF elevated calcium only in male tubules.Accordingly, the physical asymmetry of the tubules in the body cavity is directly adaptive. Now that the detailed machinery underlying internal asymmetry is starting to be delineated, our work invites the

  3. Basic leucine zipper protein Cnc-C is a substrate and transcriptional regulator of the Drosophila 26S proteasome.

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    Grimberg, Kristian Björk; Beskow, Anne; Lundin, Daniel; Davis, Monica M; Young, Patrick

    2011-02-01

    While the 26S proteasome is a key proteolytic complex, little is known about how proteasome levels are maintained in higher eukaryotic cells. Here we describe an RNA interference (RNAi) screen of Drosophila melanogaster that was used to identify transcription factors that may play a role in maintaining levels of the 26S proteasome. We used an RNAi library against 993 Drosophila transcription factor genes to identify genes whose suppression in Schneider 2 cells stabilized a ubiquitin-green fluorescent protein reporter protein. This screen identified Cnc (cap 'n' collar [CNC]; basic region leucine zipper) as a candidate transcriptional regulator of proteasome component expression. In fact, 20S proteasome activity was reduced in cells depleted of cnc. Immunoblot assays against proteasome components revealed a general decline in both 19S regulatory complex and 20S proteasome subunits after RNAi depletion of this transcription factor. Transcript-specific silencing revealed that the longest of the seven transcripts for the cnc gene, cnc-C, was needed for proteasome and p97 ATPase production. Quantitative reverse transcription-PCR confirmed the role of Cnc-C in activation of transcription of genes encoding proteasome components. Expression of a V5-His-tagged form of Cnc-C revealed that the transcription factor is itself a proteasome substrate that is stabilized when the proteasome is inhibited. We propose that this single cnc gene in Drosophila resembles the ancestral gene family of mammalian nuclear factor erythroid-derived 2-related transcription factors, which are essential in regulating oxidative stress and proteolysis.

  4. Altered Gene Regulation and Synaptic Morphology in "Drosophila" Learning and Memory Mutants

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    Guan, Zhuo; Buhl, Lauren K.; Quinn, William G.; Littleton, J. Troy

    2011-01-01

    Genetic studies in "Drosophila" have revealed two separable long-term memory pathways defined as anesthesia-resistant memory (ARM) and long-lasting long-term memory (LLTM). ARM is disrupted in "radish" ("rsh") mutants, whereas LLTM requires CREB-dependent protein synthesis. Although the downstream effectors of ARM and LLTM are distinct, pathways…

  5. Distinct Shifts in Microbiota Composition during Drosophila Aging Impair Intestinal Function and Drive Mortality

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    Rebecca I. Clark

    2015-09-01

    Full Text Available Alterations in the composition of the intestinal microbiota have been correlated with aging and measures of frailty in the elderly. However, the relationships between microbial dynamics, age-related changes in intestinal physiology, and organismal health remain poorly understood. Here, we show that dysbiosis of the intestinal microbiota, characterized by an expansion of the Gammaproteobacteria, is tightly linked to age-onset intestinal barrier dysfunction in Drosophila. Indeed, alterations in the microbiota precede and predict the onset of intestinal barrier dysfunction in aged flies. Changes in microbial composition occurring prior to intestinal barrier dysfunction contribute to changes in excretory function and immune gene activation in the aging intestine. In addition, we show that a distinct shift in microbiota composition follows intestinal barrier dysfunction, leading to systemic immune activation and organismal death. Our results indicate that alterations in microbiota dynamics could contribute to and also predict varying rates of health decline during aging in mammals.

  6. Rampant adaptive evolution in regions of proteins with unknown function in Drosophila simulans.

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    Alisha K Holloway

    2007-10-01

    Full Text Available Adaptive protein evolution is pervasive in Drosophila. Genomic studies, thus far, have analyzed each protein as a single entity. However, the targets of adaptive events may be localized to particular parts of proteins, such as protein domains or regions involved in protein folding. We compared the population genetic mechanisms driving sequence polymorphism and divergence in defined protein domains and non-domain regions. Interestingly, we find that non-domain regions of proteins are more frequent targets of directional selection. Protein domains are also evolving under directional selection, but appear to be under stronger purifying selection than non-domain regions. Non-domain regions of proteins clearly play a major role in adaptive protein evolution on a genomic scale and merit future investigations of their functional properties.

  7. Functions and Mechanisms of Fibroblast Growth Factor (FGF Signalling in Drosophila melanogaster

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    Hans-Arno J. Müller

    2013-03-01

    Full Text Available Intercellular signalling via growth factors plays an important role in controlling cell differentiation and cell movements during the development of multicellular animals. Fibroblast Growth Factor (FGF signalling induces changes in cellular behaviour allowing cells in the embryo to move, to survive, to divide or to differentiate. Several examples argue that FGF signalling is used in multi-step morphogenetic processes to achieve and maintain a transitional state of the cells required for the control of cell fate. In the genetic model Drosophila melanogaster, FGF signalling via the receptor tyrosine kinases Heartless (Htl and Breathless (Btl is particularly well studied. These FGF receptors affect gene expression, cell shape and cell–cell interactions during mesoderm layer formation, caudal visceral muscle (CVM formation, tracheal morphogenesis and glia differentiation. Here, we will address the current knowledge of the biological functions of FGF signalling in the fly on the tissue, at a cellular and molecular level.

  8. The Rho-family GTPase Rac1 regulates integrin localization in Drosophila immunosurveillance cells.

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    Miguel J Xavier

    Full Text Available BACKGROUND: When the parasitoid wasp Leptopilina boulardi lays an egg in a Drosophila larva, phagocytic cells called plasmatocytes and specialized cells known as lamellocytes encapsulate the egg. The Drosophila β-integrin Myospheroid (Mys is necessary for lamellocytes to adhere to the cellular capsule surrounding L. boulardi eggs. Integrins are heterodimeric adhesion receptors consisting of α and β subunits, and similar to other plasma membrane receptors undergo ligand-dependent endocytosis. In mammalian cells it is known that integrin binding to the extracellular matrix induces the activation of Rac GTPases, and we have previously shown that Rac1 and Rac2 are necessary for a proper encapsulation response in Drosophila larvae. We wanted to test the possibility that Myospheroid and Rac GTPases interact during the Drosophila anti-parasitoid immune response. RESULTS: In the current study we demonstrate that Rac1 is required for the proper localization of Myospheroid to the cell periphery of haemocytes after parasitization. Interestingly, the mislocalization of Myospheroid in Rac1 mutants is rescued by hyperthermia, involving the heat shock protein Hsp83. From these results we conclude that Rac1 and Hsp83 are required for the proper localization of Mys after parasitization. SIGNIFICANCE: We show for the first time that the small GTPase Rac1 is required for Mysopheroid localization. Interestingly, the necessity of Rac1 in Mys localization was negated by hyperthermia. This presents a problem, in Drosophila we quite often raise larvae at 29°C when using the GAL4/UAS misexpression system. If hyperthermia rescues receptor endosomal recycling defects, raising larvae in hyperthermic conditions may mask potentially interesting phenotypes.

  9. Unique and Overlapping Functions of Formins Frl and DAAM During Ommatidial Rotation and Neuronal Development in Drosophila

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    Dollar, Gretchen; Gombos, Rita; Barnett, Austen A.; Sanchez Hernandez, David; Maung, Saw M. T.; Mih?ly, Jozsef; Jenny, Andreas

    2016-01-01

    The noncanonical Frizzled/planar cell polarity (PCP) pathway regulates establishment of polarity within the plane of an epithelium to generate diversity of cell fates, asymmetric, but highly aligned structures, or to orchestrate the directional migration of cells during convergent extension during vertebrate gastrulation. In Drosophila, PCP signaling is essential to orient actin wing hairs and to align ommatidia in the eye, in part by coordinating the movement of groups of photoreceptor cells...

  10. Analysis of temporal transcription expression profiles reveal links between protein function and developmental stages of Drosophila melanogaster.

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    Wan, Cen; Lees, Jonathan G; Minneci, Federico; Orengo, Christine A; Jones, David T

    2017-10-01

    Accurate gene or protein function prediction is a key challenge in the post-genome era. Most current methods perform well on molecular function prediction, but struggle to provide useful annotations relating to biological process functions due to the limited power of sequence-based features in that functional domain. In this work, we systematically evaluate the predictive power of temporal transcription expression profiles for protein function prediction in Drosophila melanogaster. Our results show significantly better performance on predicting protein function when transcription expression profile-based features are integrated with sequence-derived features, compared with the sequence-derived features alone. We also observe that the combination of expression-based and sequence-based features leads to further improvement of accuracy on predicting all three domains of gene function. Based on the optimal feature combinations, we then propose a novel multi-classifier-based function prediction method for Drosophila melanogaster proteins, FFPred-fly+. Interpreting our machine learning models also allows us to identify some of the underlying links between biological processes and developmental stages of Drosophila melanogaster.

  11. Analysis of temporal transcription expression profiles reveal links between protein function and developmental stages of Drosophila melanogaster.

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    Cen Wan

    2017-10-01

    Full Text Available Accurate gene or protein function prediction is a key challenge in the post-genome era. Most current methods perform well on molecular function prediction, but struggle to provide useful annotations relating to biological process functions due to the limited power of sequence-based features in that functional domain. In this work, we systematically evaluate the predictive power of temporal transcription expression profiles for protein function prediction in Drosophila melanogaster. Our results show significantly better performance on predicting protein function when transcription expression profile-based features are integrated with sequence-derived features, compared with the sequence-derived features alone. We also observe that the combination of expression-based and sequence-based features leads to further improvement of accuracy on predicting all three domains of gene function. Based on the optimal feature combinations, we then propose a novel multi-classifier-based function prediction method for Drosophila melanogaster proteins, FFPred-fly+. Interpreting our machine learning models also allows us to identify some of the underlying links between biological processes and developmental stages of Drosophila melanogaster.

  12. Conserved microRNA miR-8 in fat body regulates innate immune homeostasis in Drosophila.

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    Choi, In Kyou; Hyun, Seogang

    2012-05-01

    Antimicrobial peptides (AMPs) constitute a major arm of the innate immune system across diverse organisms. In Drosophila, septic injury by microbial pathogens rapidly induces the production of the AMPs in fat body via well elucidated pathways such as Toll and IMD. However, several epithelial tissues were reported to locally express AMPs without septic injury via poorly characterized ways. Here, we report that microRNA miR-8 regulates the levels of AMPs basally expressed in Drosophila. The levels of AMPs such as Drosomycin and Diptericin are significantly increased in miR-8 null animals in non-pathogen stimulated conditions. Analysis of various larval tissues revealed that the increase of Drosomycin is fat body specific. Supporting this observation, re-introduction of miR-8 only in the fat body restored the altered AMP expression in miR-8 null flies. Although loss of miR-8 impedes PI3K in the fat body, inhibition of PI3K does not phenocopy the AMP expression of miR-8 null flies, indicating that miR-8 regulates AMP independently of PI3K. Together, our findings suggest a role of miR-8 in systemic immune homeostasis in generally non-pathogenic conditions in flies. Copyright © 2011 Elsevier Ltd. All rights reserved.

  13. A Quantitative RNAi Screen for JNK Modifiers Identifies Pvr as a Novel Regulator of Drosophila Immune Signaling

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    Bond, David; Foley, Edan

    2009-01-01

    Drosophila melanogaster responds to gram-negative bacterial challenges through the IMD pathway, a signal transduction cassette that is driven by the coordinated activities of JNK, NF-κB and caspase modules. While many modifiers of NF-κB activity were identified in cell culture and in vivo assays, the regulatory apparatus that determines JNK inputs into the IMD pathway is relatively unexplored. In this manuscript, we present the first quantitative screen of the entire genome of Drosophila for novel regulators of JNK activity in the IMD pathway. We identified a large number of gene products that negatively or positively impact on JNK activation in the IMD pathway. In particular, we identified the Pvr receptor tyrosine kinase as a potent inhibitor of JNK activation. In a series of in vivo and cell culture assays, we demonstrated that activation of the IMD pathway drives JNK-dependent expression of the Pvr ligands, Pvf2 and Pvf3, which in turn act through the Pvr/ERK MAP kinase pathway to attenuate the JNK and NF-κB arms of the IMD pathway. Our data illuminate a poorly understood arm of a critical and evolutionarily conserved innate immune response. Furthermore, given the pleiotropic involvement of JNK in eukaryotic cell biology, we believe that many of the novel regulators identified in this screen are of interest beyond immune signaling. PMID:19893628

  14. Phosphorylation regulates SIRT1 function.

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    Tsutomu Sasaki

    Full Text Available BACKGROUND: SIR2 is an NAD(+-dependent deacetylase [1]-[3] implicated in the regulation of lifespan in species as diverse as yeast [4], worms [5], and flies [6]. We previously reported that the level of SIRT1, the mammalian homologue of SIR2 [7], [8], is coupled to the level of mitotic activity in cells both in vitro and in vivo[9]. Cells from long-lived mice maintained SIRT1 levels of young mice in tissues that undergo continuous cell replacement by proliferating stem cells. Changes in SIRT1 protein level were not associated with changes in mRNA level, suggesting that SIRT1 could be regulated post-transcriptionally. However, other than a recent report on sumoylation [10] and identification of SIRT1 as a nuclear phospho-protein by mass spectrometry [11], post-translational modifications of this important protein have not been reported. METHODOLOGY/PRINCIPAL FINDINGS: We identified 13 residues in SIRT1 that are phosphorylated in vivo using mass spectrometry. Dephosphorylation by phosphatases in vitro resulted in decreased NAD(+-dependent deacetylase activity. We identified cyclinB/Cdk1 as a cell cycle-dependent kinase that forms a complex with and phosphorylates SIRT1. Mutation of two residues phosphorylated by Cyclin B/Cdk1 (threonine 530 and serine 540 disturbs normal cell cycle progression and fails to rescue proliferation defects in SIRT1-deficient cells [12], [13]. CONCLUSIONS/SIGNIFICANCE: Pharmacological manipulation of SIRT1 activity is currently being tested as a means of extending lifespan in mammals. Treatment of obese mice with resveratrol, a pharmacological activator of SIRT1, modestly but significantly improved longevity and, perhaps more importantly, offered some protection against the development of type 2 diabetes mellitus and metabolic syndrome [14]-[16]. Understanding the endogenous mechanisms that regulate the level and activity of SIRT1, therefore, has obvious relevance to human health and disease. Our results identify

  15. Iron Sulfur and Molybdenum Cofactor Enzymes Regulate the Drosophila Life Cycle by Controlling Cell Metabolism

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    Marelja, Zvonimir; Leimkühler, Silke; Missirlis, Fanis

    2018-01-01

    Iron sulfur (Fe-S) clusters and the molybdenum cofactor (Moco) are present at enzyme sites, where the active metal facilitates electron transfer. Such enzyme systems are soluble in the mitochondrial matrix, cytosol and nucleus, or embedded in the inner mitochondrial membrane, but virtually absent from the cell secretory pathway. They are of ancient evolutionary origin supporting respiration, DNA replication, transcription, translation, the biosynthesis of steroids, heme, catabolism of purines, hydroxylation of xenobiotics, and cellular sulfur metabolism. Here, Fe-S cluster and Moco biosynthesis in Drosophila melanogaster is reviewed and the multiple biochemical and physiological functions of known Fe-S and Moco enzymes are described. We show that RNA interference of Mocs3 disrupts Moco biosynthesis and the circadian clock. Fe-S-dependent mitochondrial respiration is discussed in the context of germ line and somatic development, stem cell differentiation and aging. The subcellular compartmentalization of the Fe-S and Moco assembly machinery components and their connections to iron sensing mechanisms and intermediary metabolism are emphasized. A biochemically active Fe-S core complex of heterologously expressed fly Nfs1, Isd11, IscU, and human frataxin is presented. Based on the recent demonstration that copper displaces the Fe-S cluster of yeast and human ferredoxin, an explanation for why high dietary copper leads to cytoplasmic iron deficiency in flies is proposed. Another proposal that exosomes contribute to the transport of xanthine dehydrogenase from peripheral tissues to the eye pigment cells is put forward, where the Vps16a subunit of the HOPS complex may have a specialized role in concentrating this enzyme within pigment granules. Finally, we formulate a hypothesis that (i) mitochondrial superoxide mobilizes iron from the Fe-S clusters in aconitase and succinate dehydrogenase; (ii) increased iron transiently displaces manganese on superoxide dismutase, which

  16. Iron Sulfur and Molybdenum Cofactor Enzymes Regulate the Drosophila Life Cycle by Controlling Cell Metabolism

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    Zvonimir Marelja

    2018-02-01

    Full Text Available Iron sulfur (Fe-S clusters and the molybdenum cofactor (Moco are present at enzyme sites, where the active metal facilitates electron transfer. Such enzyme systems are soluble in the mitochondrial matrix, cytosol and nucleus, or embedded in the inner mitochondrial membrane, but virtually absent from the cell secretory pathway. They are of ancient evolutionary origin supporting respiration, DNA replication, transcription, translation, the biosynthesis of steroids, heme, catabolism of purines, hydroxylation of xenobiotics, and cellular sulfur metabolism. Here, Fe-S cluster and Moco biosynthesis in Drosophila melanogaster is reviewed and the multiple biochemical and physiological functions of known Fe-S and Moco enzymes are described. We show that RNA interference of Mocs3 disrupts Moco biosynthesis and the circadian clock. Fe-S-dependent mitochondrial respiration is discussed in the context of germ line and somatic development, stem cell differentiation and aging. The subcellular compartmentalization of the Fe-S and Moco assembly machinery components and their connections to iron sensing mechanisms and intermediary metabolism are emphasized. A biochemically active Fe-S core complex of heterologously expressed fly Nfs1, Isd11, IscU, and human frataxin is presented. Based on the recent demonstration that copper displaces the Fe-S cluster of yeast and human ferredoxin, an explanation for why high dietary copper leads to cytoplasmic iron deficiency in flies is proposed. Another proposal that exosomes contribute to the transport of xanthine dehydrogenase from peripheral tissues to the eye pigment cells is put forward, where the Vps16a subunit of the HOPS complex may have a specialized role in concentrating this enzyme within pigment granules. Finally, we formulate a hypothesis that (i mitochondrial superoxide mobilizes iron from the Fe-S clusters in aconitase and succinate dehydrogenase; (ii increased iron transiently displaces manganese on superoxide

  17. The MAP kinase ERK and its scaffold protein MP1 interact with the chromatin regulator Corto during Drosophila wing tissue development

    Science.gov (United States)

    2011-01-01

    Background Mitogen-activated protein kinase (MAPK) cascades (p38, JNK, ERK pathways) are involved in cell fate acquisition during development. These kinase modules are associated with scaffold proteins that control their activity. In Drosophila, dMP1, that encodes an ERK scaffold protein, regulates ERK signaling during wing development and contributes to intervein and vein cell differentiation. Functional relationships during wing development between a chromatin regulator, the Enhancer of Trithorax and Polycomb Corto, ERK and its scaffold protein dMP1, are examined here. Results Genetic interactions show that corto and dMP1 act together to antagonize rolled (which encodes ERK) in the future intervein cells, thus promoting intervein fate. Although Corto, ERK and dMP1 are present in both cytoplasmic and nucleus compartments, they interact exclusively in nucleus extracts. Furthermore, Corto, ERK and dMP1 co-localize on several sites on polytene chromosomes, suggesting that they regulate gene expression directly on chromatin. Finally, Corto is phosphorylated. Interestingly, its phosphorylation pattern differs between cytoplasm and nucleus and changes upon ERK activation. Conclusions Our data therefore suggest that the Enhancer of Trithorax and Polycomb Corto could participate in regulating vein and intervein genes during wing tissue development in response to ERK signaling. PMID:21401930

  18. The MAP kinase ERK and its scaffold protein MP1 interact with the chromatin regulator Corto during Drosophila wing tissue development.

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    Mouchel-Vielh, Emmanuèle; Rougeot, Julien; Decoville, Martine; Peronnet, Frédérique

    2011-03-14

    Mitogen-activated protein kinase (MAPK) cascades (p38, JNK, ERK pathways) are involved in cell fate acquisition during development. These kinase modules are associated with scaffold proteins that control their activity. In Drosophila, dMP1, that encodes an ERK scaffold protein, regulates ERK signaling during wing development and contributes to intervein and vein cell differentiation. Functional relationships during wing development between a chromatin regulator, the Enhancer of Trithorax and Polycomb Corto, ERK and its scaffold protein dMP1, are examined here. Genetic interactions show that corto and dMP1 act together to antagonize rolled (which encodes ERK) in the future intervein cells, thus promoting intervein fate. Although Corto, ERK and dMP1 are present in both cytoplasmic and nucleus compartments, they interact exclusively in nucleus extracts. Furthermore, Corto, ERK and dMP1 co-localize on several sites on polytene chromosomes, suggesting that they regulate gene expression directly on chromatin. Finally, Corto is phosphorylated. Interestingly, its phosphorylation pattern differs between cytoplasm and nucleus and changes upon ERK activation. Our data therefore suggest that the Enhancer of Trithorax and Polycomb Corto could participate in regulating vein and intervein genes during wing tissue development in response to ERK signaling.

  19. The MAP kinase ERK and its scaffold protein MP1 interact with the chromatin regulator Corto during Drosophila wing tissue development

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    Peronnet Frédérique

    2011-03-01

    Full Text Available Abstract Background Mitogen-activated protein kinase (MAPK cascades (p38, JNK, ERK pathways are involved in cell fate acquisition during development. These kinase modules are associated with scaffold proteins that control their activity. In Drosophila, dMP1, that encodes an ERK scaffold protein, regulates ERK signaling during wing development and contributes to intervein and vein cell differentiation. Functional relationships during wing development between a chromatin regulator, the Enhancer of Trithorax and Polycomb Corto, ERK and its scaffold protein dMP1, are examined here. Results Genetic interactions show that corto and dMP1 act together to antagonize rolled (which encodes ERK in the future intervein cells, thus promoting intervein fate. Although Corto, ERK and dMP1 are present in both cytoplasmic and nucleus compartments, they interact exclusively in nucleus extracts. Furthermore, Corto, ERK and dMP1 co-localize on several sites on polytene chromosomes, suggesting that they regulate gene expression directly on chromatin. Finally, Corto is phosphorylated. Interestingly, its phosphorylation pattern differs between cytoplasm and nucleus and changes upon ERK activation. Conclusions Our data therefore suggest that the Enhancer of Trithorax and Polycomb Corto could participate in regulating vein and intervein genes during wing tissue development in response to ERK signaling.

  20. Mutations in the catalytic loop HRD motif alter the activity and function of Drosophila Src64.

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    Taylor C Strong

    Full Text Available The catalytic loop HRD motif is found in most protein kinases and these amino acids are predicted to perform functions in catalysis, transition to, and stabilization of the active conformation of the kinase domain. We have identified mutations in a Drosophila src gene, src64, that alter the three HRD amino acids. We have analyzed the mutants for both biochemical activity and biological function during development. Mutation of the aspartate to asparagine eliminates biological function in cytoskeletal processes and severely reduces fertility, supporting the amino acid's critical role in enzymatic activity. The arginine to cysteine mutation has little to no effect on kinase activity or cytoskeletal reorganization, suggesting that the HRD arginine may not be critical for coordinating phosphotyrosine in the active conformation. The histidine to leucine mutant retains some kinase activity and biological function, suggesting that this amino acid may have a biochemical function in the active kinase that is independent of its side chain hydrogen bonding interactions in the active site. We also describe the phenotypic effects of other mutations in the SH2 and tyrosine kinase domains of src64, and we compare them to the phenotypic effects of the src64 null allele.

  1. The Drosophila nerfin-1 mRNA requires multiple microRNAs to regulate its spatial and temporal translation dynamics in the developing nervous system.

    Science.gov (United States)

    Kuzin, Alexander; Kundu, Mukta; Brody, Thomas; Odenwald, Ward F

    2007-10-01

    The mRNA encoding the Drosophila Zn-finger transcription factor Nerfin-1, required for CNS axon pathfinding events, is subject to post-transcriptional silencing. Although nerfin-1 mRNA is expressed in many neural precursor cells including all early delaminating CNS neuroblasts, the encoded Nerfin-1 protein is detected only in the nuclei of neural precursors that divide just once to generate neurons and then only transiently in nascent neurons. Using a nerfin-1 promoter-controlled reporter transgene, replacement of the nerfin-1 3' UTR with the viral SV-40 3' UTR releases the neuroblast translational block and prolongs reporter protein expression in neurons. Comparative genomics analysis reveals that the nerfin-1 mRNA 3' UTR contains multiple highly conserved sequence blocks that either harbor and/or overlap 21 predicted binding sites for 18 different microRNAs. To determine the functional significance of these microRNA-binding sites and less conserved microRNA target sites, we have studied their ability to block or limit the expression of reporter protein in nerfin-1-expressing cells during embryonic development. Our results indicate that no single microRNA is sufficient to fully inhibit protein expression but rather multiple microRNAs that target different binding sites are required to block ectopic protein expression in neural precursor cells and temporally restrict expression in neurons. Taken together, these results suggest that multiple microRNAs play a cooperative role in the post-transcriptional regulation of nerfin-1 mRNA, and the high degree of microRNA-binding site evolutionary conservation indicates that all members of the Drosophila genus employ a similar strategy to regulate the onset and extinction dynamics of Nerfin-1 expression.

  2. Interorgan Communication Pathways in Physiology: Focus on Drosophila.

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    Droujinine, Ilia A; Perrimon, Norbert

    2016-11-23

    Studies in mammals and Drosophila have demonstrated the existence and significance of secreted factors involved in communication between distal organs. In this review, primarily focusing on Drosophila, we examine the known interorgan communication factors and their functions, physiological inducers, and integration in regulating physiology. Moreover, we describe how organ-sensing screens in Drosophila can systematically identify novel conserved interorgan communication factors. Finally, we discuss how interorgan communication enabled and evolved as a result of specialization of organs. Together, we anticipate that future studies will establish a model for metazoan interorgan communication network (ICN) and how it is deregulated in disease.

  3. Transcriptomics of aged Drosophila motor neurons reveals a matrix metalloproteinase that impairs motor function.

    Science.gov (United States)

    Azpurua, Jorge; Mahoney, Rebekah E; Eaton, Benjamin A

    2018-04-01

    The neuromuscular junction (NMJ) is responsible for transforming nervous system signals into motor behavior and locomotion. In the fruit fly Drosophila melanogaster, an age-dependent decline in motor function occurs, analogous to the decline experienced in mice, humans, and other mammals. The molecular and cellular underpinnings of this decline are still poorly understood. By specifically profiling the transcriptome of Drosophila motor neurons across age using custom microarrays, we found that the expression of the matrix metalloproteinase 1 (dMMP1) gene reproducibly increased in motor neurons in an age-dependent manner. Modulation of physiological aging also altered the rate of dMMP1 expression, validating dMMP1 expression as a bona fide aging biomarker for motor neurons. Temporally controlled overexpression of dMMP1 specifically in motor neurons was sufficient to induce deficits in climbing behavior and cause a decrease in neurotransmitter release at neuromuscular synapses. These deficits were reversible if the dMMP1 expression was shut off again immediately after the onset of motor dysfunction. Additionally, repression of dMMP1 enzymatic activity via overexpression of a tissue inhibitor of metalloproteinases delayed the onset of age-dependent motor dysfunction. MMPs are required for proper tissue architecture during development. Our results support the idea that matrix metalloproteinase 1 is acting as a downstream effector of antagonistic pleiotropy in motor neurons and is necessary for proper development, but deleterious when reactivated at an advanced age. © 2018 The Authors. Aging Cell published by the Anatomical Society and John Wiley & Sons Ltd.

  4. Computational identification of developmental enhancers:conservation and function of transcription factor binding-site clustersin drosophila melanogaster and drosophila psedoobscura

    Energy Technology Data Exchange (ETDEWEB)

    Berman, Benjamin P.; Pfeiffer, Barret D.; Laverty, Todd R.; Salzberg, Steven L.; Rubin, Gerald M.; Eisen, Michael B.; Celniker, SusanE.

    2004-08-06

    The identification of sequences that control transcription in metazoans is a major goal of genome analysis. In a previous study, we demonstrated that searching for clusters of predicted transcription factor binding sites could discover active regulatory sequences, and identified 37 regions of the Drosophila melanogaster genome with high densities of predicted binding sites for five transcription factors involved in anterior-posterior embryonic patterning. Nine of these clusters overlapped known enhancers. Here, we report the results of in vivo functional analysis of 27 remaining clusters. We generated transgenic flies carrying each cluster attached to a basal promoter and reporter gene, and assayed embryos for reporter gene expression. Six clusters are enhancers of adjacent genes: giant, fushi tarazu, odd-skipped, nubbin, squeeze and pdm2; three drive expression in patterns unrelated to those of neighboring genes; the remaining 18 do not appear to have enhancer activity. We used the Drosophila pseudoobscura genome to compare patterns of evolution in and around the 15 positive and 18 false-positive predictions. Although conservation of primary sequence cannot distinguish true from false positives, conservation of binding-site clustering accurately discriminates functional binding-site clusters from those with no function. We incorporated conservation of binding-site clustering into a new genome-wide enhancer screen, and predict several hundred new regulatory sequences, including 85 adjacent to genes with embryonic patterns. Measuring conservation of sequence features closely linked to function--such as binding-site clustering--makes better use of comparative sequence data than commonly used methods that examine only sequence identity.

  5. Genome-Wide Analysis Reveals Novel Regulators of Growth in Drosophila melanogaster.

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    Sibylle Chantal Vonesch

    2016-01-01

    Full Text Available Organismal size depends on the interplay between genetic and environmental factors. Genome-wide association (GWA analyses in humans have implied many genes in the control of height but suffer from the inability to control the environment. Genetic analyses in Drosophila have identified conserved signaling pathways controlling size; however, how these pathways control phenotypic diversity is unclear. We performed GWA of size traits using the Drosophila Genetic Reference Panel of inbred, sequenced lines. We find that the top associated variants differ between traits and sexes; do not map to canonical growth pathway genes, but can be linked to these by epistasis analysis; and are enriched for genes and putative enhancers. Performing GWA on well-studied developmental traits under controlled conditions expands our understanding of developmental processes underlying phenotypic diversity.

  6. Disruption of lysosome function promotes tumor growth and metastasis in Drosophila.

    Science.gov (United States)

    Chi, Congwu; Zhu, Huanhu; Han, Min; Zhuang, Yuan; Wu, Xiaohui; Xu, Tian

    2010-07-09

    Lysosome function is essential to many physiological processes. It has been suggested that deregulation of lysosome function could contribute to cancer. Through a genetic screen in Drosophila, we have discovered that mutations disrupting lysosomal degradation pathway components contribute to tumor development and progression. Loss-of-function mutations in the Class C vacuolar protein sorting (VPS) gene, deep orange (dor), dramatically promote tumor overgrowth and invasion of the Ras(V12) cells. Knocking down either of the two other components of the Class C VPS complex, carnation (car) and vps16A, also renders Ras(V12) cells capable for uncontrolled growth and metastatic behavior. Finally, chemical disruption of the lysosomal function by feeding animals with antimalarial drugs, chloroquine or monensin, leads to malignant tumor growth of the Ras(V12) cells. Taken together, our data provide evidence for a causative role of lysosome dysfunction in tumor growth and invasion and indicate that members of the Class C VPS complex behave as tumor suppressors.

  7. Temporal regulation of Drosophila salivary gland degeneration by the Broad-Complex transcription factors

    Czech Academy of Sciences Publication Activity Database

    Kuchárová-Mahmood, S.; Raška, Ivan; Mechler, B. M.; Farkaš, R.

    2002-01-01

    Roč. 140, - (2002), s. 67-78 ISSN 1047-8477 R&D Projects: GA ČR GA304/02/0342 Grant - others:GA-(SK) VEGA:2/7194/20 Institutional research plan: CEZ:AV0Z5039906; CEZ:MSM 111100003 Keywords : programmed cell death * BR-C transcription factors * drosophila Subject RIV: EA - Cell Biology Impact factor: 4.194, year: 2002

  8. The cell adhesion molecule Fasciclin2 regulates brush border length and organization in Drosophila renal tubules

    DEFF Research Database (Denmark)

    Halberg, Kenneth Agerlin; Rainey, Stephanie M.; Veland, Iben Rønn

    2016-01-01

    Multicellular organisms rely on cell adhesion molecules to coordinate cell-cell interactions, and to provide navigational cues during tissue formation. In Drosophila, Fasciclin 2 (Fas2) has been intensively studied due to its role in nervous system development and maintenance; yet, Fas2 is most...... role for this well-known cell adhesion molecule, and propose that Fas2-mediated intermicrovillar homophilic adhesion complexes help stabilize the brush border....

  9. Aging and Intermittent Fasting Impact on Transcriptional Regulation and Physiological Responses of Adult Drosophila Neuronal and Muscle Tissues.

    Science.gov (United States)

    Zhang, Sharon; Ratliff, Eric P; Molina, Brandon; El-Mecharrafie, Nadja; Mastroianni, Jessica; Kotzebue, Roxanne W; Achal, Madhulika; Mauntz, Ruth E; Gonzalez, Arysa; Barekat, Ayeh; Bray, William A; Macias, Andrew M; Daugherty, Daniel; Harris, Greg L; Edwards, Robert A; Finley, Kim D

    2018-04-10

    The progressive decline of the nervous system, including protein aggregate formation, reflects the subtle dysregulation of multiple functional pathways. Our previous work has shown intermittent fasting (IF) enhances longevity, maintains adult behaviors and reduces aggregates, in part, by promoting autophagic function in the aging Drosophila brain. To clarify the impact that IF-treatment has upon aging, we used high throughput RNA-sequencing technology to examine the changing transcriptome in adult Drosophila tissues. Principle component analysis (PCA) and other analyses showed ~1200 age-related transcriptional differences in head and muscle tissues, with few genes having matching expression patterns. Pathway components showing age-dependent expression differences were involved with stress response, metabolic, neural and chromatin remodeling functions. Middle-aged tissues also showed a significant increase in transcriptional drift-variance (TD), which in the CNS included multiple proteolytic pathway components. Overall, IF-treatment had a demonstrably positive impact on aged transcriptomes, partly ameliorating both fold and variance changes. Consistent with these findings, aged IF-treated flies displayed more youthful metabolic, behavioral and basal proteolytic profiles that closely correlated with transcriptional alterations to key components. These results indicate that even modest dietary changes can have therapeutic consequences, slowing the progressive decline of multiple cellular systems, including proteostasis in the aging nervous system.

  10. Aging and Intermittent Fasting Impact on Transcriptional Regulation and Physiological Responses of Adult Drosophila Neuronal and Muscle Tissues

    Directory of Open Access Journals (Sweden)

    Sharon Zhang

    2018-04-01

    Full Text Available The progressive decline of the nervous system, including protein aggregate formation, reflects the subtle dysregulation of multiple functional pathways. Our previous work has shown intermittent fasting (IF enhances longevity, maintains adult behaviors and reduces aggregates, in part, by promoting autophagic function in the aging Drosophila brain. To clarify the impact that IF-treatment has upon aging, we used high throughput RNA-sequencing technology to examine the changing transcriptome in adult Drosophila tissues. Principle component analysis (PCA and other analyses showed ~1200 age-related transcriptional differences in head and muscle tissues, with few genes having matching expression patterns. Pathway components showing age-dependent expression differences were involved with stress response, metabolic, neural and chromatin remodeling functions. Middle-aged tissues also showed a significant increase in transcriptional drift-variance (TD, which in the CNS included multiple proteolytic pathway components. Overall, IF-treatment had a demonstrably positive impact on aged transcriptomes, partly ameliorating both fold and variance changes. Consistent with these findings, aged IF-treated flies displayed more youthful metabolic, behavioral and basal proteolytic profiles that closely correlated with transcriptional alterations to key components. These results indicate that even modest dietary changes can have therapeutic consequences, slowing the progressive decline of multiple cellular systems, including proteostasis in the aging nervous system.

  11. Regulation of Centromere Localization of the Drosophila Shugoshin MEI-S332 and Sister-Chromatid Cohesion in Meiosis

    Science.gov (United States)

    Nogueira, Cristina; Kashevsky, Helena; Pinto, Belinda; Clarke, Astrid; Orr-Weaver, Terry L.

    2014-01-01

    The Shugoshin (Sgo) protein family helps to ensure proper chromosome segregation by protecting cohesion at the centromere by preventing cleavage of the cohesin complex. Some Sgo proteins also influence other aspects of kinetochore-microtubule attachments. Although many Sgo members require Aurora B kinase to localize to the centromere, factors controlling delocalization are poorly understood and diverse. Moreover, it is not clear how Sgo function is inactivated and whether this is distinct from delocalization. We investigated these questions in Drosophila melanogaster, an organism with superb chromosome cytology to monitor Sgo localization and quantitative assays to test its function in sister-chromatid segregation in meiosis. Previous research showed that in mitosis in cell culture, phosphorylation of the Drosophila Sgo, MEI-S332, by Aurora B promotes centromere localization, whereas Polo phosphorylation promotes delocalization. These studies also suggested that MEI-S332 can be inactivated independently of delocalization, a conclusion supported here by localization and function studies in meiosis. Phosphoresistant and phosphomimetic mutants for the Aurora B and Polo phosphorylation sites were examined for effects on MEI-S332 localization and chromosome segregation in meiosis. Strikingly, MEI-S332 with a phosphomimetic mutation in the Aurora B phosphorylation site prematurely dissociates from the centromeres in meiosis I. Despite the absence of MEI-S332 on meiosis II centromeres in male meiosis, sister chromatids segregate normally, demonstrating that detectable levels of this Sgo are not essential for chromosome congression, kinetochore biorientation, or spindle assembly. PMID:25081981

  12. Downregulation of dTps1 in Drosophila melanogaster larvae confirms involvement of trehalose in redox regulation following desiccation.

    Science.gov (United States)

    Thorat, Leena; Mani, Krishna-Priya; Thangaraj, Pradeep; Chatterjee, Suvro; Nath, Bimalendu B

    2016-03-01

    As a survival strategy to environmental water deficits, desiccation-tolerant organisms are commonly known for their ability to recruit stress-protective biomolecules such as trehalose. We have previously reported the pivotal role of trehalose in larval desiccation tolerance in Drosophila melanogaster. Trehalose has emerged as a versatile molecule, serving mainly as energy source in insects and also being a stress protectant. While several recent reports have revealed the unconventional role of trehalose in scavenging reactive oxygen species in yeast and plants, this aspect has not received much attention in animals. We examined the status of desiccation-induced generation of reactive oxygen species in D. melanogaster larvae and the possible involvement of trehalose in ameliorating the harmful consequences thereof. Insect trehalose synthesis is governed by the enzyme trehalose 6-phosphate synthase 1 (TPS1). Using the ubiquitous da-GAL4-driven expression of the dTps1-RNAi transgene, we generated dTps1-downregulated Drosophila larvae possessing depleted levels of dTps1 transcripts. This resulted in the inability of the larvae for trehalose synthesis, thereby allowing us to elucidate the significance of trehalose in the regulation of desiccation-responsive redox homeostasis. Furthermore, the results from molecular genetics studies, biochemical assays, electron spin resonance analyses and a simple, non-invasive method of whole larval live imaging suggested that trehalose in collaboration with superoxide dismutase (SOD) is involved in the maintenance of redox state in D. melanogaster.

  13. Drosophila's contribution to stem cell research [version 2; referees: 2 approved

    Directory of Open Access Journals (Sweden)

    Gyanesh Singh

    2016-08-01

    Full Text Available The discovery of Drosophila stem cells with striking similarities to mammalian stem cells has brought new hope for stem cell research. Recent developments in Drosophila stem cell research is bringing wider opportunities for contemporary stem cell biologists. In this regard, Drosophila germ cells are becoming a popular model of stem cell research. In several cases, genes that controlled Drosophila stem cells were later discovered to have functional homologs in mammalian stem cells. Like mammals, Drosophila germline stem cells (GSCs are controlled by both intrinsic as well as external signals. Inside the Drosophila testes, germline and somatic stem cells form a cluster of cells (the hub. Hub cells depend on JAK-STAT signaling, and, in absence of this signal, they do not self-renew. In Drosophila, significant changes occur within the stem cell niche that contributes to a decline in stem cell number over time. In case of aging Drosophila, somatic niche cells show reduced DE-cadherin and unpaired (Upd proteins. Unpaired proteins are known to directly decrease stem cell number within the niches, and, overexpression of upd within niche cells restored GSCs in older males also . Stem cells in the midgut of Drosophila are also very promising. Reduced Notch signaling was found to increase the number of midgut progenitor cells. On the other hand, activation of the Notch pathway decreased proliferation of these cells. Further research in this area should lead to the discovery of additional factors that regulate stem and progenitor cells in Drosophila.

  14. Timed Knickkopf function is essential for wing cuticle formation in Drosophila melanogaster.

    Science.gov (United States)

    Li, Kaixia; Zhang, Xubo; Zuo, Ying; Liu, Weimin; Zhang, Jianzhen; Moussian, Bernard

    2017-10-01

    The insect cuticle is an extracellular matrix that consists of the polysaccharide chitin, proteins, lipids and organic molecules that are arranged in distinct horizontal layers. In Drosophila melanogaster, these layers are not formed sequentially, but, at least partially, at the same time. Timing of the underlying molecular mechanisms is conceivably crucial for cuticle formation. To study this issue, we determined the time period during which the function of Knickkopf (Knk), a key factor of chitin organization, is required for wing cuticle differentiation in D. melanogaster. Although knk is expressed throughout metamorphosis, we demonstrate that its expression 30 h prior and 48 h after pupariation is essential for correct wing cuticle formation. In other words, expression beyond this period is futile. Importantly, manipulation of Knk expression during this time causes wing bending suggesting an effect of Knk amounts on the physical properties of the wing cuticle. Manipulation of Knk expression also interferes with the structure and function of the cuticle surface. First, we show that the shape of surface nano-structures depends on the expression levels of knk. Second, we find that cuticle impermeability is compromised in wings with reduced knk expression. In summary, despite the extended supply of Knk during metamorphosis, controlled amounts of Knk are important for correct wing cuticle differentiation and function in a concise period of time. Copyright © 2017 Elsevier Ltd. All rights reserved.

  15. A transgenic Drosophila model demonstrates that the Helicobacter pylori CagA protein functions as a eukaryotic Gab adaptor.

    Directory of Open Access Journals (Sweden)

    Crystal M Botham

    2008-05-01

    Full Text Available Infection with the human gastric pathogen Helicobacter pylori is associated with a spectrum of diseases including gastritis, peptic ulcers, gastric adenocarcinoma, and gastric mucosa-associated lymphoid tissue lymphoma. The cytotoxin-associated gene A (CagA protein of H. pylori, which is translocated into host cells via a type IV secretion system, is a major risk factor for disease development. Experiments in gastric tissue culture cells have shown that once translocated, CagA activates the phosphatase SHP-2, which is a component of receptor tyrosine kinase (RTK pathways whose over-activation is associated with cancer formation. Based on CagA's ability to activate SHP-2, it has been proposed that CagA functions as a prokaryotic mimic of the eukaryotic Grb2-associated binder (Gab adaptor protein, which normally activates SHP-2. We have developed a transgenic Drosophila model to test this hypothesis by investigating whether CagA can function in a well-characterized Gab-dependent process: the specification of photoreceptors cells in the Drosophila eye. We demonstrate that CagA expression is sufficient to rescue photoreceptor development in the absence of the Drosophila Gab homologue, Daughter of Sevenless (DOS. Furthermore, CagA's ability to promote photoreceptor development requires the SHP-2 phosphatase Corkscrew (CSW. These results provide the first demonstration that CagA functions as a Gab protein within the tissue of an organism and provide insight into CagA's oncogenic potential. Since many translocated bacterial proteins target highly conserved eukaryotic cellular processes, such as the RTK signaling pathway, the transgenic Drosophila model should be of general use for testing the in vivo function of bacterial effector proteins and for identifying the host genes through which they function.

  16. PH Domain-Arf G Protein Interactions Localize the Arf-GEF Steppke for Cleavage Furrow Regulation in Drosophila.

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    Donghoon M Lee

    Full Text Available The recruitment of GDP/GTP exchange factors (GEFs to specific subcellular sites dictates where they activate small G proteins for the regulation of various cellular processes. Cytohesins are a conserved family of plasma membrane GEFs for Arf small G proteins that regulate endocytosis. Analyses of mammalian cytohesins have identified a number of recruitment mechanisms for these multi-domain proteins, but the conservation and developmental roles for these mechanisms are unclear. Here, we report how the pleckstrin homology (PH domain of the Drosophila cytohesin Steppke affects its localization and activity at cleavage furrows of the early embryo. We found that the PH domain is necessary for Steppke furrow localization, and for it to regulate furrow structure. However, the PH domain was not sufficient for the localization. Next, we examined the role of conserved PH domain amino acid residues that are required for mammalian cytohesins to bind PIP3 or GTP-bound Arf G proteins. We confirmed that the Steppke PH domain preferentially binds PIP3 in vitro through a conserved mechanism. However, disruption of residues for PIP3 binding had no apparent effect on GFP-Steppke localization and effects. Rather, residues for binding to GTP-bound Arf G proteins made major contributions to this Steppke localization and activity. By analyzing GFP-tagged Arf and Arf-like small G proteins, we found that Arf1-GFP, Arf6-GFP and Arl4-GFP, but not Arf4-GFP, localized to furrows. However, analyses of embryos depleted of Arf1, Arf6 or Arl4 revealed either earlier defects than occur in embryos depleted of Steppke, or no detectable furrow defects, possibly because of redundancies, and thus it was difficult to assess how individual Arf small G proteins affect Steppke. Nonetheless, our data show that the Steppke PH domain and its conserved residues for binding to GTP-bound Arf G proteins have substantial effects on Steppke localization and activity in early Drosophila embryos.

  17. Hippo, TGF-β, and Src-MAPK pathways regulate transcription of the upd3 cytokine in Drosophila enterocytes upon bacterial infection.

    Science.gov (United States)

    Houtz, Philip; Bonfini, Alessandro; Liu, Xi; Revah, Jonathan; Guillou, Aurélien; Poidevin, Mickael; Hens, Korneel; Huang, Hsin-Yi; Deplancke, Bart; Tsai, Yu-Chen; Buchon, Nicolas

    2017-11-01

    Cytokine signaling is responsible for coordinating conserved epithelial regeneration and immune responses in the digestive tract. In the Drosophila midgut, Upd3 is a major cytokine, which is induced in enterocytes (EC) and enteroblasts (EB) upon oral infection, and initiates intestinal stem cell (ISC) dependent tissue repair. To date, the genetic network directing upd3 transcription remains largely uncharacterized. Here, we have identified the key infection-responsive enhancers of the upd3 gene and show that distinct enhancers respond to various stresses. Furthermore, through functional genetic screening, bioinformatic analyses and yeast one-hybrid screening, we determined that the transcription factors Scalloped (Sd), Mothers against dpp (Mad), and D-Fos are principal regulators of upd3 expression. Our study demonstrates that upd3 transcription in the gut is regulated by the activation of multiple pathways, including the Hippo, TGF-β/Dpp, and Src, as well as p38-dependent MAPK pathways. Thus, these essential pathways, which are known to control ISC proliferation cell-autonomously, are also activated in ECs to promote tissue turnover the regulation of upd3 transcription.

  18. Scalloped a member of the Hippo tumor suppressor pathway controls mushroom body size in Drosophila brain by non-canonical regulation of neuroblast proliferation.

    Science.gov (United States)

    Rohith, Basavanahalli Nanjundaiah; Shyamala, Baragur Venkatanarayanasetty

    2017-12-15

    Cell proliferation, growth and survival are three different basic processes which converge at determining a fundamental property -the size of an organism. Scalloped (Sd) is the first characterised transcriptional partner to Yorkie (Yki), the downstream effector of the Hippo pathway which is a highly potential and evolutionarily conserved regulator of organ size. Here we have studied the hypomorphic effect of sd on the development of Mushroom Bodies (MBs) in Drosophila brain. We show that, sd non-function results in an increase in the size of MBs. We demonstrate that, sd regulation on MB size operates through multiple routes. Sd expressed in the differentiated MB neurons, imposes non-cell autonomous repression on the proliferation of MB precursor cells, and Sd expression in the MB neuroblasts (NB) cell autonomously represses mushroom body neuroblast (MBNB) proliferation. Further Sd in Kenyon cells (KCs) imparts a cell autonomous restriction on their growth. Our findings are distinctive because, while the classical sd loss of function phenotypes in eye, wing and lymph gland are reported as loss of tissue or reduced organ size, the present study shows that, Sd inactivation in the developing MB, promotes precursor cell proliferation and results in an increase in the organ size. Copyright © 2017 Elsevier Inc. All rights reserved.

  19. The E3 ligase Ubr3 regulates Usher syndrome and MYH9 disorder proteins in the auditory organs of Drosophila and mammals.

    Science.gov (United States)

    Li, Tongchao; Giagtzoglou, Nikolaos; Eberl, Daniel F; Jaiswal, Sonal Nagarkar; Cai, Tiantian; Godt, Dorothea; Groves, Andrew K; Bellen, Hugo J

    2016-06-22

    Myosins play essential roles in the development and function of auditory organs and multiple myosin genes are associated with hereditary forms of deafness. Using a forward genetic screen in Drosophila, we identified an E3 ligase, Ubr3, as an essential gene for auditory organ development. Ubr3 negatively regulates the mono-ubiquitination of non-muscle Myosin II, a protein associated with hearing loss in humans. The mono-ubiquitination of Myosin II promotes its physical interaction with Myosin VIIa, a protein responsible for Usher syndrome type IB. We show that ubr3 mutants phenocopy pathogenic variants of Myosin II and that Ubr3 interacts genetically and physically with three Usher syndrome proteins. The interactions between Myosin VIIa and Myosin IIa are conserved in the mammalian cochlea and in human retinal pigment epithelium cells. Our work reveals a novel mechanism that regulates protein complexes affected in two forms of syndromic deafness and suggests a molecular function for Myosin IIa in auditory organs.

  20. Poly(ADP-ribose) Glycohydrolase and Poly(ADP-ribose)-interacting Protein Hrp38 Regulate Pattern Formation during Drosophila Eye Development

    Science.gov (United States)

    Ji, Yingbiao; Jarnik, Michael; Tulin, Alexei V.

    2013-01-01

    Drosophila Hrp38, a homolog of human hnRNP A1, has been shown to regulate splicing, but its function can be modified by poly(ADP-ribosyl)ation. Notwithstanding such findings, our understanding of the roles of poly(ADP-ribosyl)ated Hrp38 on development is limited. Here, we have demonstrated that Hrp38 is essential for fly eye development based on a rough-eye phenotype with disorganized ommatidia observed in adult escapers of the hrp38 mutant. We also observed that Poly(ADP-ribose) Glycohydrolase (Parg) loss-of-function, which caused increased Hrp38 poly(ADP-ribosyl)ation, also resulted in the rough-eye phenotype with disrupted ommatidial lattice and reduced number of photoreceptor cells. In addition, ectopic expression of DE-cadherin, which is required for retinal morphogenesis, fully rescued the rough-eye phenotype of the hrp38 mutant. Similarly, Parg mutant eye clones had decreased expression level of DE-cadherin with orientation defects, which is reminiscent of DE-cadherin mutant eye phenotype. Therefore, our results suggest that Hrp38 poly(ADP-ribosyl)ation controls eye pattern formation via regulation of DE-cadherin expression, a finding which has implications for understanding the pathogenic mechanisms of Hrp38-related Fragile X syndrome and PARP1-related retinal degeneration diseases. PMID:23711619

  1. The E3 ligase Ubr3 regulates Usher syndrome and MYH9 disorder proteins in the auditory organs of Drosophila and mammals

    Science.gov (United States)

    Li, Tongchao; Giagtzoglou, Nikolaos; Eberl, Daniel F; Jaiswal, Sonal Nagarkar; Cai, Tiantian; Godt, Dorothea; Groves, Andrew K; Bellen, Hugo J

    2016-01-01

    Myosins play essential roles in the development and function of auditory organs and multiple myosin genes are associated with hereditary forms of deafness. Using a forward genetic screen in Drosophila, we identified an E3 ligase, Ubr3, as an essential gene for auditory organ development. Ubr3 negatively regulates the mono-ubiquitination of non-muscle Myosin II, a protein associated with hearing loss in humans. The mono-ubiquitination of Myosin II promotes its physical interaction with Myosin VIIa, a protein responsible for Usher syndrome type IB. We show that ubr3 mutants phenocopy pathogenic variants of Myosin II and that Ubr3 interacts genetically and physically with three Usher syndrome proteins. The interactions between Myosin VIIa and Myosin IIa are conserved in the mammalian cochlea and in human retinal pigment epithelium cells. Our work reveals a novel mechanism that regulates protein complexes affected in two forms of syndromic deafness and suggests a molecular function for Myosin IIa in auditory organs. DOI: http://dx.doi.org/10.7554/eLife.15258.001 PMID:27331610

  2. Circadian Activators Are Expressed Days before They Initiate Clock Function in Late Pacemaker Neurons from Drosophila.

    Science.gov (United States)

    Liu, Tianxin; Mahesh, Guruswamy; Houl, Jerry H; Hardin, Paul E

    2015-06-03

    Circadian pacemaker neurons in the Drosophila brain control daily rhythms in locomotor activity. These pacemaker neurons can be subdivided into early or late groups depending on whether rhythms in period (per) and timeless (tim) expression are initiated at the first instar (L1) larval stage or during metamorphosis, respectively. Because CLOCK-CYCLE (CLK-CYC) heterodimers initiate circadian oscillator function by activating per and tim transcription, a Clk-GFP transgene was used to mark when late pacemaker neurons begin to develop. We were surprised to see that CLK-GFP was already expressed in four of five clusters of late pacemaker neurons during the third instar (L3) larval stage. CLK-GFP is only detected in postmitotic neurons from L3 larvae, suggesting that these four late pacemaker neuron clusters are formed before the L3 larval stage. A GFP-cyc transgene was used to show that CYC, like CLK, is also expressed exclusively in pacemaker neurons from L3 larval brains, demonstrating that CLK-CYC is not sufficient to activate per and tim in late pacemaker neurons at the L3 larval stage. These results suggest that most late pacemaker neurons develop days before novel factors activate circadian oscillator function during metamorphosis. Copyright © 2015 the authors 0270-6474/15/358662-10$15.00/0.

  3. The Human dsRNA binding protein PACT is unable to functionally substitute for the Drosophila dsRNA binding protein R2D2 [v1; ref status: indexed, http://f1000r.es/201

    Directory of Open Access Journals (Sweden)

    Benjamin K Dickerman

    2013-10-01

    Full Text Available The primary function of the dsRNA binding protein (dsRBP PACT/RAX is to activate the dsRNA dependent protein kinase PKR in response to stress signals.  Additionally, it has been identified as a component of the small RNA processing pathway.  A role for PACT/RAX in this pathway represents an important interplay between two modes of post-transcriptional gene regulation.  The function of PACT/RAX in this context is poorly understood.  Thus, additional models are required to clarify the mechanism by which PACT/RAX functions.  In this study, Drosophila melanogaster was employed to identify functionally orthologous dsRNA-binding proteins.  Transgenic Drosophila expressing human PACT were generated to determine whether PACT is capable of functionally substituting for the Drosophila dsRBP R2D2, which has a well-defined role in small RNA biogenesis.  Results presented here indicate that PACT is unable to substitute for R2D2 at the whole organism level.

  4. Mammalian enabled (Mena) is a critical regulator of cardiac function.

    Science.gov (United States)

    Aguilar, Frédérick; Belmonte, Stephen L; Ram, Rashmi; Noujaim, Sami F; Dunaevsky, Olga; Protack, Tricia L; Jalife, Jose; Todd Massey, H; Gertler, Frank B; Blaxall, Burns C

    2011-05-01

    Mammalian enabled (Mena) of the Drosophila enabled/vasodilator-stimulated phosphoprotein gene family is a cytoskeletal protein implicated in actin regulation and cell motility. Cardiac Mena expression is enriched in intercalated discs (ICD), the critical intercellular communication nexus between adjacent muscle cells. We previously identified Mena gene expression to be a key predictor of human and murine heart failure (HF). To determine the in vivo function of Mena in the heart, we assessed Mena protein expression in multiple HF models and characterized the effects of genetic Mena deletion on cardiac structure and function. Immunoblot analysis revealed significant upregulation of Mena protein expression in left ventricle tissue from patients with end-stage HF, calsequestrin-overexpressing mice, and isoproterenol-infused mice. Characterization of the baseline cardiac function of adult Mena knockout mice (Mena(-/-)) via echocardiography demonstrated persistent cardiac dysfunction, including a significant reduction in percent fractional shortening compared with wild-type littermates. Electrocardiogram PR and QRS intervals were significantly prolonged in Mena(-/-) mice, manifested by slowed conduction on optical mapping studies. Ultrastructural analysis of Mena(-/-) hearts revealed disrupted organization and widening of ICD structures, mislocalization of the gap junction protein connexin 43 (Cx43) to the lateral borders of cardiomyoycytes, and increased Cx43 expression. Furthermore, the expression of vinculin (an adherens junction protein) was significantly reduced in Mena(-/-) mice. We report for the first time that genetic ablation of Mena results in cardiac dysfunction, highlighted by diminished contractile performance, disrupted ICD structure, and slowed electrical conduction.

  5. Drosophila insulin and target of rapamycin (TOR pathways regulate GSK3 beta activity to control Myc stability and determine Myc expression in vivo

    Directory of Open Access Journals (Sweden)

    Parisi Federica

    2011-09-01

    Full Text Available Abstract Background Genetic studies in Drosophila melanogaster reveal an important role for Myc in controlling growth. Similar studies have also shown how components of the insulin and target of rapamycin (TOR pathways are key regulators of growth. Despite a few suggestions that Myc transcriptional activity lies downstream of these pathways, a molecular mechanism linking these signaling pathways to Myc has not been clearly described. Using biochemical and genetic approaches we tried to identify novel mechanisms that control Myc activity upon activation of insulin and TOR signaling pathways. Results Our biochemical studies show that insulin induces Myc protein accumulation in Drosophila S2 cells, which correlates with a decrease in the activity of glycogen synthase kinase 3-beta (GSK3β a kinase that is responsible for Myc protein degradation. Induction of Myc by insulin is inhibited by the presence of the TOR inhibitor rapamycin, suggesting that insulin-induced Myc protein accumulation depends on the activation of TOR complex 1. Treatment with amino acids that directly activate the TOR pathway results in Myc protein accumulation, which also depends on the ability of S6K kinase to inhibit GSK3β activity. Myc upregulation by insulin and TOR pathways is a mechanism conserved in cells from the wing imaginal disc, where expression of Dp110 and Rheb also induces Myc protein accumulation, while inhibition of insulin and TOR pathways result in the opposite effect. Our functional analysis, aimed at quantifying the relative contribution of Myc to ommatidial growth downstream of insulin and TOR pathways, revealed that Myc activity is necessary to sustain the proliferation of cells from the ommatidia upon Dp110 expression, while its contribution downstream of TOR is significant to control the size of the ommatidia. Conclusions Our study presents novel evidence that Myc activity acts downstream of insulin and TOR pathways to control growth in Drosophila. At

  6. Drosophila insulin and target of rapamycin (TOR) pathways regulate GSK3 beta activity to control Myc stability and determine Myc expression in vivo.

    Science.gov (United States)

    Parisi, Federica; Riccardo, Sara; Daniel, Margaret; Saqcena, Mahesh; Kundu, Nandini; Pession, Annalisa; Grifoni, Daniela; Stocker, Hugo; Tabak, Esteban; Bellosta, Paola

    2011-09-27

    Genetic studies in Drosophila melanogaster reveal an important role for Myc in controlling growth. Similar studies have also shown how components of the insulin and target of rapamycin (TOR) pathways are key regulators of growth. Despite a few suggestions that Myc transcriptional activity lies downstream of these pathways, a molecular mechanism linking these signaling pathways to Myc has not been clearly described. Using biochemical and genetic approaches we tried to identify novel mechanisms that control Myc activity upon activation of insulin and TOR signaling pathways. Our biochemical studies show that insulin induces Myc protein accumulation in Drosophila S2 cells, which correlates with a decrease in the activity of glycogen synthase kinase 3-beta (GSK3β ) a kinase that is responsible for Myc protein degradation. Induction of Myc by insulin is inhibited by the presence of the TOR inhibitor rapamycin, suggesting that insulin-induced Myc protein accumulation depends on the activation of TOR complex 1. Treatment with amino acids that directly activate the TOR pathway results in Myc protein accumulation, which also depends on the ability of S6K kinase to inhibit GSK3β activity. Myc upregulation by insulin and TOR pathways is a mechanism conserved in cells from the wing imaginal disc, where expression of Dp110 and Rheb also induces Myc protein accumulation, while inhibition of insulin and TOR pathways result in the opposite effect. Our functional analysis, aimed at quantifying the relative contribution of Myc to ommatidial growth downstream of insulin and TOR pathways, revealed that Myc activity is necessary to sustain the proliferation of cells from the ommatidia upon Dp110 expression, while its contribution downstream of TOR is significant to control the size of the ommatidia. Our study presents novel evidence that Myc activity acts downstream of insulin and TOR pathways to control growth in Drosophila. At the biochemical level we found that both these pathways

  7. Mutational analysis and genetic cloning of the agnostic locus, which regulates learning ability in Drosophila.

    Science.gov (United States)

    Peresleni, A I; Savvateeva, E V; Peresleni, I V; Sharagina, L M

    1997-01-01

    P-insertion mutations were obtained and localized by in situ methods at the agnostic gene (agn: 1-38.9; 11AB) in Drosophila. All agn mutants showed a wide spectrum of pleiotropic effects: an EMS-induced mutation of the agn-ts398 improved the ability to develop a conditioned defensive response and increased the activity of cAMP metabolic enzymes; spontaneous mutation of agnX1 showed morphological defects of the brain. P-insertion mutations were used to clone the gene; a restriction map of 80 kb in length was determined, and the insertion was localized to a fragment of 9 kb.

  8. Functional Dissection of the Blocking and Bypass Activities of the Fab-8 Boundary in the Drosophila Bithorax Complex.

    Science.gov (United States)

    Kyrchanova, Olga; Mogila, Vladic; Wolle, Daniel; Deshpande, Girish; Parshikov, Alexander; Cléard, Fabienne; Karch, Francois; Schedl, Paul; Georgiev, Pavel

    2016-07-01

    Functionally autonomous regulatory domains direct the parasegment-specific expression of the Drosophila Bithorax complex (BX-C) homeotic genes. Autonomy is conferred by boundary/insulator elements that separate each regulatory domain from its neighbors. For six of the nine parasegment (PS) regulatory domains in the complex, at least one boundary is located between the domain and its target homeotic gene. Consequently, BX-C boundaries must not only block adventitious interactions between neighboring regulatory domains, but also be permissive (bypass) for regulatory interactions between the domains and their gene targets. To elucidate how the BX-C boundaries combine these two contradictory activities, we have used a boundary replacement strategy. We show that a 337 bp fragment spanning the Fab-8 boundary nuclease hypersensitive site and lacking all but 83 bp of the 625 bp Fab-8 PTS (promoter targeting sequence) fully rescues a Fab-7 deletion. It blocks crosstalk between the iab-6 and iab-7 regulatory domains, and has bypass activity that enables the two downstream domains, iab-5 and iab-6, to regulate Abdominal-B (Abd-B) transcription in spite of two intervening boundary elements. Fab-8 has two dCTCF sites and we show that they are necessary both for blocking and bypass activity. However, CTCF sites on their own are not sufficient for bypass. While multimerized dCTCF (or Su(Hw)) sites have blocking activity, they fail to support bypass. Moreover, this bypass defect is not rescued by the full length PTS. Finally, we show that orientation is critical for the proper functioning the Fab-8 replacement. Though the inverted Fab-8 boundary still blocks crosstalk, it disrupts the topology of the Abd-B regulatory domains and does not support bypass. Importantly, altering the orientation of the Fab-8 dCTCF sites is not sufficient to disrupt bypass, indicating that orientation dependence is conferred by other factors.

  9. Developmental shaping of dendritic arbors in Drosophila relies on tightly regulated intra-neuronal activity of protein kinase A (PKA).

    Science.gov (United States)

    Copf, Tijana

    2014-09-15

    Dendrites develop morphologies characterized by multiple levels of complexity that involve neuron type specific dendritic length and particular spatial distribution. How this is developmentally regulated and in particular which signaling molecules are crucial in the process is still not understood. Using Drosophila class IV dendritic arborization (da) neurons we test in vivo the effects of cell-autonomous dose-dependent changes in the activity levels of the cAMP-dependent Protein Kinase A (PKA) on the formation of complex dendritic arbors. We find that genetic manipulations of the PKA activity levels affect profoundly the arbor complexity with strongest impact on distal branches. Both decreasing and increasing PKA activity result in a reduced complexity of the arbors, as reflected in decreased dendritic length and number of branching points, suggesting an inverted U-shape response to PKA. The phenotypes are accompanied by changes in organelle distribution: Golgi outposts and early endosomes in distal dendritic branches are reduced in PKA mutants. By using Rab5 dominant negative we find that PKA interacts genetically with the early endosomal pathway. We test if the possible relationship between PKA and organelles may be the result of phosphorylation of the microtubule motor dynein components or Rab5. We find that Drosophila cytoplasmic dynein components are direct PKA phosphorylation targets in vitro, but not in vivo, thus pointing to a different putative in vivo target. Our data argue that tightly controlled dose-dependent intra-neuronal PKA activity levels are critical in determining the dendritic arbor complexity, one of the possible ways being through the regulation of organelle distribution. Copyright © 2014 Elsevier Inc. All rights reserved.

  10. Epigenetic regulation of macrophage function

    NARCIS (Netherlands)

    Hoeksema, M.A.

    2016-01-01

    Atherosclerosis is a lipid-driven chronic inflammatory disorder with a key role for macrophages in all disease stages. Macrophages are involved as scavengers of lipids, regulate inflammation, attract other immune cells and contribute to the resolution of inflammation, fibrosis and plaque stability.

  11. No significant regulation of bicoid mRNA by Pumilio or Nanos in the early Drosophila embryo.

    Science.gov (United States)

    Wharton, Tammy H; Nomie, Krystle J; Wharton, Robin P

    2018-01-01

    Drosophila Pumilio (Pum) is a founding member of the conserved Puf domain class of RNA-binding translational regulators. Pum binds with high specificity, contacting eight nucleotides, one with each of the repeats in its RNA-binding domain. In general, Pum is thought to block translation in collaboration with Nanos (Nos), which exhibits no binding specificity in isolation but is recruited jointly to regulatory sequences containing a Pum binding site in the 3'-UTRs of target mRNAs. Unlike Pum, which is ubiquitous in the early embryo, Nos is tightly restricted to the posterior, ensuring that repression of its best-characterized target, maternal hunchback (hb) mRNA, takes place exclusively in the posterior. An exceptional case of Nos-independent regulation by Pum has been described-repression of maternal bicoid (bcd) mRNA at the anterior pole of the early embryo, dependent on both Pum and conserved Pum binding sites in the 3'-UTR of the mRNA. We have re-investigated regulation of bcd in the early embryo; our experiments reveal no evidence of a role for Pum or its conserved binding sites in regulation of the perdurance of bcd mRNA or protein. Instead, we find that Pum and Nos control the accumulation of bcd mRNA in testes.

  12. Dissecting the function and assembly of acentriolar microtubule organizing centers in Drosophila cells in vivo.

    Directory of Open Access Journals (Sweden)

    Janina Baumbach

    2015-05-01

    Full Text Available Acentriolar microtubule organizing centers (aMTOCs are formed during meiosis and mitosis in several cell types, but their function and assembly mechanism is unclear. Importantly, aMTOCs can be overactive in cancer cells, enhancing multipolar spindle formation, merotelic kinetochore attachment and aneuploidy. Here we show that aMTOCs can form in acentriolar Drosophila somatic cells in vivo via an assembly pathway that depends on Asl, Cnn and, to a lesser extent, Spd-2--the same proteins that appear to drive mitotic centrosome assembly in flies. This finding enabled us to ablate aMTOC formation in acentriolar cells, and so perform a detailed genetic analysis of the contribution of aMTOCs to acentriolar mitotic spindle formation. Here we show that although aMTOCs can nucleate microtubules, they do not detectably increase the efficiency of acentriolar spindle assembly in somatic fly cells. We find that they are required, however, for robust microtubule array assembly in cells without centrioles that also lack microtubule nucleation from around the chromatin. Importantly, aMTOCs are also essential for dynein-dependent acentriolar spindle pole focusing and for robust cell proliferation in the absence of centrioles and HSET/Ncd (a kinesin essential for acentriolar spindle pole focusing in many systems. We propose an updated model for acentriolar spindle pole coalescence by the molecular motors Ncd/HSET and dynein in conjunction with aMTOCs.

  13. Corazonin neurons function in sexually dimorphic circuitry that shape behavioral responses to stress in Drosophila.

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    Yan Zhao

    Full Text Available All organisms are confronted with dynamic environmental changes that challenge homeostasis, which is the operational definition of stress. Stress produces adaptive behavioral and physiological responses, which, in the Metazoa, are mediated through the actions of various hormones. Based on its associated phenotypes and its expression profiles, a candidate stress hormone in Drosophila is the corazonin neuropeptide. We evaluated the potential roles of corazonin in mediating stress-related changes in target behaviors and physiologies through genetic alteration of corazonin neuronal excitability. Ablation of corazonin neurons confers resistance to metabolic, osmotic, and oxidative stress, as measured by survival. Silencing and activation of corazonin neurons lead to differential lifespan under stress, and these effects showed a strong dependence on sex. Additionally, altered corazonin neuron physiology leads to fundamental differences in locomotor activity, and these effects were also sex-dependent. The dynamics of altered locomotor behavior accompanying stress was likewise altered in flies with altered corazonin neuronal function. We report that corazonin transcript expression is altered under starvation and osmotic stress, and that triglyceride and dopamine levels are equally impacted in corazonin neuronal alterations and these phenotypes similarly show significant sexual dimorphisms. Notably, these sexual dimorphisms map to corazonin neurons. These results underscore the importance of central peptidergic processing within the context of stress and place corazonin signaling as a critical feature of neuroendocrine events that shape stress responses and may underlie the inherent sexual dimorphic differences in stress responses.

  14. Functional divergence caused by ancient positive selection of a Drosophila hybrid incompatibility locus.

    Directory of Open Access Journals (Sweden)

    Daniel A Barbash

    2004-06-01

    Full Text Available Interspecific hybrid lethality and sterility are a consequence of divergent evolution between species and serve to maintain the discrete identities of species. The evolution of hybrid incompatibilities has been described in widely accepted models by Dobzhansky and Muller where lineage-specific functional divergence is the essential characteristic of hybrid incompatibility genes. Experimentally tractable models are required to identify and test candidate hybrid incompatibility genes. Several Drosophila melanogaster genes involved in hybrid incompatibility have been identified but none has yet been shown to have functionally diverged in accordance with the Dobzhansky-Muller model. By introducing transgenic copies of the X-linked Hybrid male rescue (Hmr gene into D. melanogaster from its sibling species D. simulans and D. mauritiana, we demonstrate that Hmr has functionally diverged to cause F1 hybrid incompatibility between these species. Consistent with the Dobzhansky-Muller model, we find that Hmr has diverged extensively in the D. melanogaster lineage, but we also find extensive divergence in the sibling-species lineage. Together, these findings implicate over 13% of the amino acids encoded by Hmr as candidates for causing hybrid incompatibility. The exceptional level of divergence at Hmr cannot be explained by neutral processes because we use phylogenetic methods and population genetic analyses to show that the elevated amino-acid divergence in both lineages is due to positive selection in the distant past-at least one million generations ago. Our findings suggest that multiple substitutions driven by natural selection may be a general phenomenon required to generate hybrid incompatibility alleles.

  15. Receptor Tyrosine Kinases in Drosophila Development

    Science.gov (United States)

    Sopko, Richelle; Perrimon, Norbert

    2013-01-01

    Tyrosine phosphorylation plays a significant role in a wide range of cellular processes. The Drosophila genome encodes more than 20 receptor tyrosine kinases and extensive studies in the past 20 years have illustrated their diverse roles and complex signaling mechanisms. Although some receptor tyrosine kinases have highly specific functions, others strikingly are used in rather ubiquitous manners. Receptor tyrosine kinases regulate a broad expanse of processes, ranging from cell survival and proliferation to differentiation and patterning. Remarkably, different receptor tyrosine kinases share many of the same effectors and their hierarchical organization is retained in disparate biological contexts. In this comprehensive review, we summarize what is known regarding each receptor tyrosine kinase during Drosophila development. Astonishingly, very little is known for approximately half of all Drosophila receptor tyrosine kinases. PMID:23732470

  16. Autocrine regulation of ecdysone synthesis by β3-octopamine receptor in the prothoracic gland is essential for Drosophila metamorphosis.

    Science.gov (United States)

    Ohhara, Yuya; Shimada-Niwa, Yuko; Niwa, Ryusuke; Kayashima, Yasunari; Hayashi, Yoshiki; Akagi, Kazutaka; Ueda, Hitoshi; Yamakawa-Kobayashi, Kimiko; Kobayashi, Satoru

    2015-02-03

    In Drosophila, pulsed production of the steroid hormone ecdysone plays a pivotal role in developmental transitions such as metamorphosis. Ecdysone production is regulated in the prothoracic gland (PG) by prothoracicotropic hormone (PTTH) and insulin-like peptides (Ilps). Here, we show that monoaminergic autocrine regulation of ecdysone biosynthesis in the PG is essential for metamorphosis. PG-specific knockdown of a monoamine G protein-coupled receptor, β3-octopamine receptor (Octβ3R), resulted in arrested metamorphosis due to lack of ecdysone. Knockdown of tyramine biosynthesis genes expressed in the PG caused similar defects in ecdysone production and metamorphosis. Moreover, PTTH and Ilps signaling were impaired by Octβ3R knockdown in the PG, and activation of these signaling pathways rescued the defect in metamorphosis. Thus, monoaminergic autocrine signaling in the PG regulates ecdysone biogenesis in a coordinated fashion on activation by PTTH and Ilps. We propose that monoaminergic autocrine signaling acts downstream of a body size checkpoint that allows metamorphosis to occur when nutrients are sufficiently abundant.

  17. Metal Homeostasis Regulators Suppress FRDA Phenotypes in a Drosophila Model of the Disease.

    Directory of Open Access Journals (Sweden)

    Sirena Soriano

    Full Text Available Friedreich's ataxia (FRDA, the most commonly inherited ataxia in populations of European origin, is a neurodegenerative disorder caused by a decrease in frataxin levels. One of the hallmarks of the disease is the accumulation of iron in several tissues including the brain, and frataxin has been proposed to play a key role in iron homeostasis. We found that the levels of zinc, copper, manganese and aluminum were also increased in a Drosophila model of FRDA, and that copper and zinc chelation improve their impaired motor performance. By means of a candidate genetic screen, we identified that genes implicated in iron, zinc and copper transport and metal detoxification can restore frataxin deficiency-induced phenotypes. Taken together, these results demonstrate that the metal dysregulation in FRDA includes other metals besides iron, therefore providing a new set of potential therapeutic targets.

  18. Evidence for tension-based regulation of Drosophila MAL and SRF during invasive cell migration.

    Science.gov (United States)

    Somogyi, Kálmán; Rørth, Pernille

    2004-07-01

    Cells migrating through a tissue exert force via their cytoskeleton and are themselves subject to tension, but the effects of physical forces on cell behavior in vivo are poorly understood. Border cell migration during Drosophila oogenesis is a useful model for invasive cell movement. We report that this migration requires the activity of the transcriptional factor serum response factor (SRF) and its cofactor MAL-D and present evidence that nuclear accumulation of MAL-D is induced by cell stretching. Border cells that cannot migrate lack nuclear MAL-D but can accumulate it if they are pulled by other migrating cells. Like mammalian MAL, MAL-D also responds to activated Diaphanous, which affects actin dynamics. MAL-D/SRF activity is required to build a robust actin cytoskeleton in the migrating cells; mutant cells break apart when initiating migration. Thus, tension-induced MAL-D activity may provide a feedback mechanism for enhancing cytoskeletal strength during invasive migration.

  19. Drosophila and genome-wide association studies: a review and resource for the functional dissection of human complex traits

    Science.gov (United States)

    Wangler, Michael F.; Hu, Yanhui

    2017-01-01

    ABSTRACT Human genome-wide association studies (GWAS) have successfully identified thousands of susceptibility loci for common diseases with complex genetic etiologies. Although the susceptibility variants identified by GWAS usually have only modest effects on individual disease risk, they contribute to a substantial burden of trait variation in the overall population. GWAS also offer valuable clues to disease mechanisms that have long proven to be elusive. These insights could lead the way to breakthrough treatments; however, several challenges hinder progress, making innovative approaches to accelerate the follow-up of results from GWAS an urgent priority. Here, we discuss the largely untapped potential of the fruit fly, Drosophila melanogaster, for functional investigation of findings from human GWAS. We highlight selected examples where strong genomic conservation with humans along with the rapid and powerful genetic tools available for flies have already facilitated fine mapping of association signals, elucidated gene mechanisms, and revealed novel disease-relevant biology. We emphasize current research opportunities in this rapidly advancing field, and present bioinformatic analyses that systematically explore the applicability of Drosophila for interrogation of susceptibility signals implicated in more than 1000 human traits, based on all GWAS completed to date. Thus, our discussion is targeted at both human geneticists seeking innovative strategies for experimental validation of findings from GWAS, as well as the Drosophila research community, by whom ongoing investigations of the implicated genes will powerfully inform our understanding of human disease. PMID:28151408

  20. Influence of Quercetin in the Temporal Regulation of Redox Homeostasis in Drosophila melanogaster.

    Science.gov (United States)

    Subramanian, Perumal; Kaliyamoorthy, Kanimozhi; Jayapalan, Jaime Jacqueline; Abdul-Rahman, Puteri Shafinaz; Haji Hashim, Onn

    2017-01-01

    Numerous biological processes are governed by the biological clock. Studies using Drosophila melanogaster (L.) are valuable that could be of importance for their effective applications on rodent studies. In this study, the beneficial role of quercetin (a flavonoid) on H2O2 induced stress in D. melanogaster was investigated. D. melanogaster flies were divided into four groups (group I - control, group II - H2O2 (acute exposure), group III - quercetin, and group IV - quercetin + H2O2 treated). Negative geotaxis assay, oxidative stress indicators (protein carbonyls, thiobarbituric reactive substances [TBARS]), and antioxidants (superoxide dismutase [SOD], catalase [CAT], glutathione-S-transferase [GST], glutathione peroxidase, and reduced glutathione [GSH]) were measured at 4 h intervals over 24 h and temporal expression of heat shock protein-70 (Hsp70), Upd1 (homolog of IL-6 in Drosophila), and nitric oxide synthase (Nos) was analyzed by Western blotting. Groups II and IV showed altered biochemical rhythms (compared with controls). Decreased mesor values of negative geotaxis, SOD, CAT, GST, and GSH were noticed in H2O2, increased mesor of oxidative stress indicators (TBARS and protein carbonyl content) and a reversibility of the rhythmic characteristics were conspicuous after quercetin treatment. The expression levels of Hsp70, Upd1, and Nos were noticeably maximum at 04:00. Significant elevation of expression by H2O2 was nearly normalized by quercetin treatment. The possible mechanism by which quercetin modulates oxidant-antioxidant imbalance under oxidative stress could be ascribed to the modulation of the rhythmic properties. Our results will be helpful to understand the molecular interlink between circadian rhythm and oxidative stress mechanism. © The Author 2017. Published by Oxford University Press on behalf of the Entomological Society of America.

  1. Genetic architecture and functional characterization of genes underlying the rapid diversification of male external genitalia between Drosophila simulans and Drosophila mauritiana.

    Science.gov (United States)

    Tanaka, Kentaro M; Hopfen, Corinna; Herbert, Matthew R; Schlötterer, Christian; Stern, David L; Masly, John P; McGregor, Alistair P; Nunes, Maria D S

    2015-05-01

    Male sexual characters are often among the first traits to diverge between closely related species and identifying the genetic basis of such changes can contribute to our understanding of their evolutionary history. However, little is known about the genetic architecture or the specific genes underlying the evolution of male genitalia. The morphology of the claspers, posterior lobes, and anal plates exhibit striking differences between Drosophila mauritiana and D. simulans. Using QTL and introgression-based high-resolution mapping, we identified several small regions on chromosome arms 3L and 3R that contribute to differences in these traits. However, we found that the loci underlying the evolution of clasper differences between these two species are independent from those that contribute to posterior lobe and anal plate divergence. Furthermore, while most of the loci affect each trait in the same direction and act additively, we also found evidence for epistasis between loci for clasper bristle number. In addition, we conducted an RNAi screen in D. melanogaster to investigate if positional and expression candidate genes located on chromosome 3L, are also involved in genital development. We found that six of these genes, including components of Wnt signaling and male-specific lethal 3 (msl3), regulate the development of genital traits consistent with the effects of the introgressed regions where they are located and that thus represent promising candidate genes for the evolution these traits. Copyright © 2015 by the Genetics Society of America.

  2. SUMOylation in Drosophila Development

    Directory of Open Access Journals (Sweden)

    Albert J. Courey

    2012-07-01

    Full Text Available Small ubiquitin-related modifier (SUMO, an ~90 amino acid ubiquitin-like protein, is highly conserved throughout the eukaryotic domain. Like ubiquitin, SUMO is covalently attached to lysine side chains in a large number of target proteins. In contrast to ubiquitin, SUMO does not have a direct role in targeting proteins for proteasomal degradation. However, like ubiquitin, SUMO does modulate protein function in a variety of other ways. This includes effects on protein conformation, subcellular localization, and protein–protein interactions. Significant insight into the in vivo role of SUMOylation has been provided by studies in Drosophila that combine genetic manipulation, proteomic, and biochemical analysis. Such studies have revealed that the SUMO conjugation pathway regulates a wide variety of critical cellular and developmental processes, including chromatin/chromosome function, eggshell patterning, embryonic pattern formation, metamorphosis, larval and pupal development, neurogenesis, development of the innate immune system, and apoptosis. This review discusses our current understanding of the diverse roles for SUMO in Drosophila development.

  3. A conserved serine residue regulates the stability of Drosophila Salvador and human WW domain-containing adaptor 45 through proteasomal degradation

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Di, E-mail: DiWu@mail.nankai.edu.cn; Wu, Shian

    2013-04-19

    Highlights: •Ser-17 is key for the stability of Drosophila Sav. •Ala mutation of Ser-17 promotes the proteasomal degradation of Sav. •Ser-17 residue is not the main target of Hpo-induced Sav stabilization. •Hpo-dependent and -independent mechanisms regulate Sav stability. •This mechanism is conserved in the homologue of Sav, human WW45. -- Abstract: The Hippo (Hpo) pathway is a conserved tumor suppressor pathway that controls organ size through the coordinated regulation of apoptosis and proliferation. Drosophila Salvador (Sav), which limits organ size, is a core component of the Hpo pathway. In this study, Ser-17 was shown to be important for the stability of Sav. Alanine mutation of Ser-17 promoted the proteasomal degradation of Sav. Destabilization and stabilization of the Sav protein mediated by alanine mutation of Ser-17 and by Hpo, respectively, were independent of each other. This implies that the stability of Sav is controlled by two mechanisms, one that is Ser-17-dependent and Hpo-independent, and another that is Ser-17-independent and Hpo-dependent. These dual mechanisms also regulated the human counterpart of Drosophila Sav, WW domain-containing adaptor 45 (WW45). The conservation of this regulation adds to its significance in normal physiology and tumorigenesis.

  4. Genetic modifiers of comatose mutations in Drosophila: insights into neuronal NSF (N-ethylmaleimide-sensitive fusion factor) functions.

    Science.gov (United States)

    Sanyal, Subhabrata; Krishnan, K S

    2012-09-01

    By the middle of the 20th century, development of powerful genetic approaches had ensured that the fruit fly would remain a model organism of choice for genetic and developmental studies. But in the 1970s, a few pioneering groups turned their attention to the prospect of using the fly for neurophysiological experiments. They proposed that in a poikilothermic organism such as Drosophila, temperature-sensitive or "ts" mutations in proteins that controlled nerve function would translate to a "ts" paralytic phenotype. This was by no means an obvious or even a likely assumption. However, following directed screens these groups soon reported dramatic demonstrations of reversible ts paralysis in fly mutants. Resultantly, these "simple" experiments led to the isolation of a number of conditional mutations including shibire, paralytic, and comatose. All have since been cloned and have enabled deep mechanistic insights into synaptic transmission and nerve conduction. comatose (comt) mutations, for example, were found to map to missense changes in dNSF1, a neuron-specific fly homolog of mammalian NSF (N-ethylmaleimide-sensitive fusion factor). Studies on comt were also some of the first to discriminate between nuanced models of NSF function during presynaptic transmitter release that have since been borne out by experiments in multiple preparations. Here, the authors present an overview of NSF function as it is understood today, with an emphasis on contributions from Drosophila beginning with experiments carried out by Obaid Siddiqi in the Benzer laboratory. The authors also outline initial results from a genetic screen for phenotypic modifiers of comt that hold the promise of further elucidating NSF function at the synapse. Over the years, the neuromuscular system of Drosophila has served as a uniquely accessible model to unravel mechanisms underlying synaptic transmission. To this day, ts paralysis remains one of the most emphatic demonstrations of nerve function in an

  5. Epigenetic Regulation of Monocyte and Macrophage Function

    NARCIS (Netherlands)

    Hoeksema, Marten A.; de Winther, Menno P. J.

    2016-01-01

    Monocytes and macrophages are key players in tissue homeostasis and immune responses. Epigenetic processes tightly regulate cellular functioning in health and disease. Recent Advances: Recent technical developments have allowed detailed characterizations of the transcriptional circuitry underlying

  6. Transposable element dynamics and PIWI regulation impacts lncRNA and gene expression diversity in Drosophila ovarian cell cultures.

    Science.gov (United States)

    Sytnikova, Yuliya A; Rahman, Reazur; Chirn, Gung-Wei; Clark, Josef P; Lau, Nelson C

    2014-12-01

    Piwi proteins and Piwi-interacting RNAs (piRNAs) repress transposable elements (TEs) from mobilizing in gonadal cells. To determine the spectrum of piRNA-regulated targets that may extend beyond TEs, we conducted a genome-wide survey for transcripts associated with PIWI and for transcripts affected by PIWI knockdown in Drosophila ovarian somatic sheet (OSS) cells, a follicle cell line expressing the Piwi pathway. Despite the immense sequence diversity among OSS cell piRNAs, our analysis indicates that TE transcripts are the major transcripts associated with and directly regulated by PIWI. However, several coding genes were indirectly regulated by PIWI via an adjacent de novo TE insertion that generated a nascent TE transcript. Interestingly, we noticed that PIWI-regulated genes in OSS cells greatly differed from genes affected in a related follicle cell culture, ovarian somatic cells (OSCs). Therefore, we characterized the distinct genomic TE insertions across four OSS and OSC lines and discovered dynamic TE landscapes in gonadal cultures that were defined by a subset of active TEs. Particular de novo TEs appeared to stimulate the expression of novel candidate long noncoding RNAs (lncRNAs) in a cell lineage-specific manner, and some of these TE-associated lncRNAs were associated with PIWI and overlapped PIWI-regulated genes. Our analyses of OSCs and OSS cells demonstrate that despite having a Piwi pathway to suppress endogenous mobile elements, gonadal cell TE landscapes can still dramatically change and create transcriptome diversity. © 2014 Sytnikova et al.; Published by Cold Spring Harbor Laboratory Press.

  7. Mid-embryo patterning and precision in Drosophila segmentation: Krüppel dual regulation of hunchback.

    Directory of Open Access Journals (Sweden)

    David M Holloway

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

  8. [Drosophila melanogaster as a model for studying the function of animal viral proteins].

    Science.gov (United States)

    Omelianchuk, L V; Iudina, O S

    2011-07-01

    Studies in which Drosophila melanogaster individuals carrying transgenes of animal viruses were used to analyze the action of animal viral proteins on the cell are reviewed. The data presented suggest that host specificity of viruses is determined by their proteins responsible for the penetration of the virus into the cell, while viral proteins responsible for interactions with the host cell are much less host-specific. Due to this, the model of Drosophila with its developed system of searching for genetic interactions can be used to find intracellular targets for the action of viral proteins of the second group.

  9. Use of Drosophila to study DNA repair

    International Nuclear Information System (INIS)

    Boyd, J.B.; Harris, P.V.; Sakaguchi, K.

    1988-01-01

    This paper discusses Drosophila, the premier metazoan organism for analyzing many fundamental features of eukaryotic gene regulation. The authors present adaptations of several approaches for studying DNA repair to an analysis of repair-defective mutants in Drosophila. A current understanding of Drosophila DNA repair is described

  10. Regulation of the Drosophila Enhancer of split and invected-engrailed gene complexes by sister chromatid cohesion proteins.

    Directory of Open Access Journals (Sweden)

    Cheri A Schaaf

    2009-07-01

    Full Text Available The cohesin protein complex was first recognized for holding sister chromatids together and ensuring proper chromosome segregation. Cohesin also regulates gene expression, but the mechanisms are unknown. Cohesin associates preferentially with active genes, and is generally absent from regions in which histone H3 is methylated by the Enhancer of zeste [E(z] Polycomb group silencing protein. Here we show that transcription is hypersensitive to cohesin levels in two exceptional cases where cohesin and the E(z-mediated histone methylation simultaneously coat the entire Enhancer of split and invected-engrailed gene complexes in cells derived from Drosophila central nervous system. These gene complexes are modestly transcribed, and produce seven of the twelve transcripts that increase the most with cohesin knockdown genome-wide. Cohesin mutations alter eye development in the same manner as increased Enhancer of split activity, suggesting that similar regulation occurs in vivo. We propose that cohesin helps restrain transcription of these gene complexes, and that deregulation of similarly cohesin-hypersensitive genes may underlie developmental deficits in Cornelia de Lange syndrome.

  11. A Sox Transcription Factor Is a Critical Regulator of Adult Stem Cell Proliferation in the Drosophila Intestine

    Directory of Open Access Journals (Sweden)

    Fanju W. Meng

    2015-11-01

    Full Text Available Adult organs and their resident stem cells are constantly facing the challenge of adapting cell proliferation to tissue demand, particularly in response to environmental stresses. Whereas most stress-signaling pathways are conserved between progenitors and differentiated cells, stem cells have the specific ability to respond by increasing their proliferative rate, using largely unknown mechanisms. Here, we show that a member of the Sox family of transcription factors in Drosophila, Sox21a, is expressed in intestinal stem cells (ISCs in the adult gut. Sox21a is essential for the proliferation of these cells during both normal epithelium turnover and repair. Its expression is induced in response to tissue damage, downstream of the Jun N-terminal kinase (JNK and extracellular signal-regulated kinase (ERK pathways, to promote ISC proliferation. Although short-lived, Sox21a mutant flies show no developmental defects, supporting the notion that this factor is a specific regulator of adult stem cell proliferation.

  12. DRhoGEF2 regulates cellular tension and cell pulsations in the Amnioserosa during Drosophila dorsal closure.

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    Dulce Azevedo

    Full Text Available Coordination of apical constriction in epithelial sheets is a fundamental process during embryogenesis. Here, we show that DRhoGEF2 is a key regulator of apical pulsation and constriction of amnioserosal cells during Drosophila dorsal closure. Amnioserosal cells mutant for DRhoGEF2 exhibit a consistent decrease in amnioserosa pulsations whereas overexpression of DRhoGEF2 in this tissue leads to an increase in the contraction time of pulsations. We probed the physical properties of the amnioserosa to show that the average tension in DRhoGEF2 mutant cells is lower than wild-type and that overexpression of DRhoGEF2 results in a tissue that is more solid-like than wild-type. We also observe that in the DRhoGEF2 overexpressing cells there is a dramatic increase of apical actomyosin coalescence that can contribute to the generation of more contractile forces, leading to amnioserosal cells with smaller apical surface than wild-type. Conversely, in DRhoGEF2 mutants, the apical actomyosin coalescence is impaired. These results identify DRhoGEF2 as an upstream regulator of the actomyosin contractile machinery that drives amnioserosa cells pulsations and apical constriction.

  13. Rapid DNA Synthesis During Early Drosophila Embryogenesis Is Sensitive to Maternal Humpty Dumpty Protein Function.

    Science.gov (United States)

    Lesly, Shera; Bandura, Jennifer L; Calvi, Brian R

    2017-11-01

    Problems with DNA replication cause cancer and developmental malformations. It is not fully understood how DNA replication is coordinated with development and perturbed in disease. We had previously identified the Drosophila gene humpty dumpty ( hd ), and showed that null alleles cause incomplete DNA replication, tissue undergrowth, and lethality. Animals homozygous for the missense allele, hd 272-9 , were viable, but adult females had impaired amplification of eggshell protein genes in the ovary, resulting in the maternal effects of thin eggshells and embryonic lethality. Here, we show that expression of an hd transgene in somatic cells of the ovary rescues amplification and eggshell synthesis but not embryo viability. The germline of these mothers remain mutant for the hd 272-9 allele, resulting in reduced maternal Hd protein and embryonic arrest during mitosis of the first few S/M nuclear cleavage cycles with chromosome instability and chromosome bridges. Epistasis analysis of hd with the rereplication mutation plutonium indicates that the chromosome bridges of hd embryos are the result of a failed attempt to segregate incompletely replicated sister chromatids. This study reveals that maternally encoded Humpty dumpty protein is essential for DNA replication and genome integrity during the little-understood embryonic S/M cycles. Moreover, the two hd 272-9 maternal-effect phenotypes suggest that ovarian gene amplification and embryonic cleavage are two time periods in development that are particularly sensitive to mild deficits in DNA replication function. This last observation has broader relevance for interpreting why mild mutations in the human ortholog of humpty dumpty and other DNA replication genes cause tissue-specific malformations of microcephalic dwarfisms. Copyright © 2017 by the Genetics Society of America.

  14. Live imaging of muscles in Drosophila metamorphosis: Towards high-throughput gene identification and function analysis.

    Science.gov (United States)

    Puah, Wee Choo; Wasser, Martin

    2016-03-01

    Time-lapse microscopy in developmental biology is an emerging tool for functional genomics. Phenotypic effects of gene perturbations can be studied non-invasively at multiple time points in chronological order. During metamorphosis of Drosophila melanogaster, time-lapse microscopy using fluorescent reporters allows visualization of alternative fates of larval muscles, which are a model for the study of genes related to muscle wasting. While doomed muscles enter hormone-induced programmed cell death, a smaller population of persistent muscles survives to adulthood and undergoes morphological remodeling that involves atrophy in early, and hypertrophy in late pupation. We developed a method that combines in vivo imaging, targeted gene perturbation and image analysis to identify and characterize genes involved in muscle development. Macrozoom microscopy helps to screen for interesting muscle phenotypes, while confocal microscopy in multiple locations over 4-5 days produces time-lapse images that are used to quantify changes in cell morphology. Performing a similar investigation using fixed pupal tissues would be too time-consuming and therefore impractical. We describe three applications of our pipeline. First, we show how quantitative microscopy can track and measure morphological changes of muscle throughout metamorphosis and analyze genes involved in atrophy. Second, our assay can help to identify genes that either promote or prevent histolysis of abdominal muscles. Third, we apply our approach to test new fluorescent proteins as live markers for muscle development. We describe mKO2 tagged Cysteine proteinase 1 (Cp1) and Troponin-I (TnI) as examples of proteins showing developmental changes in subcellular localization. Finally, we discuss strategies to improve throughput of our pipeline to permit genome-wide screens in the future. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

  15. [Functional analysis of Grp and Iris, the gag and env domesticated errantivirus genes, in the Drosophila melanogaster genome].

    Science.gov (United States)

    Makhnovskii, P A; Kuzmin, I V; Nefedova, L N; Kima, A I

    2016-01-01

    Drosophila melanogaster is the only invertebrate that contains endogenous retroviruses, which are called errantiviruses. Two domesticated genes, Grp and Iris, which originate from errantivirus gag and env, respectively, have been found in the D. melanogaster genome. The functions performed by the genes in Drosophila are still unclear. To identify the functions of domesticated gag and env in the D. melanogaster genome, expression of Iris and Grp was studied in strains differing by the presence or absence of the functional gypsy errantivirus. In addition, the expression levels were measured after injection of gram-positive and gram-negative bacteria, which activate different immune response pathways, and exposure to various abiotic stress factors. The presence of functional D. melanogaster retrovirus gypsy was found to increase the Grp expression level in somatic tissues of the carcass, while exerting no effect on the Iris expression level. Activation of the immune response in D. melanogaster by bacteria Bacillus cereus increased the Grp expression level and did not affect Iris expression. As for the effects of abiotic stress factors (oxidative stress, starvation, and heat and cold stress), the Grp expression level increased in response to starvation in D. melanogaster females, and the Iris expression level was downregulated in heat shock and oxidative stress. Based on the findings, Grp was assumed to play a direct role in the immune response in D. melanogaster; Iris is not involved in immune responses, but and apparently performs a cell function that is inhibited in stress.

  16. Metabolomic Studies in Drosophila.

    Science.gov (United States)

    Cox, James E; Thummel, Carl S; Tennessen, Jason M

    2017-07-01

    Metabolomic analysis provides a powerful new tool for studies of Drosophila physiology. This approach allows investigators to detect thousands of chemical compounds in a single sample, representing the combined contributions of gene expression, enzyme activity, and environmental context. Metabolomics has been used for a wide range of studies in Drosophila , often providing new insights into gene function and metabolic state that could not be obtained using any other approach. In this review, we survey the uses of metabolomic analysis since its entry into the field. We also cover the major methods used for metabolomic studies in Drosophila and highlight new directions for future research. Copyright © 2017 by the Genetics Society of America.

  17. Executive functions and self-regulation.

    Science.gov (United States)

    Hofmann, Wilhelm; Schmeichel, Brandon J; Baddeley, Alan D

    2012-03-01

    Self-regulation is a core aspect of adaptive human behavior that has been studied, largely in parallel, through the lenses of social and personality psychology as well as cognitive psychology. Here, we argue for more communication between these disciplines and highlight recent research that speaks to their connection. We outline how basic facets of executive functioning (working memory operations, behavioral inhibition, and task-switching) may subserve successful self-regulation. We also argue that temporary reductions in executive functions underlie many of the situational risk factors identified in the social psychological research on self-regulation and review recent evidence that the training of executive functions holds significant potential for improving poor self-regulation in problem populations. Copyright © 2012 Elsevier Ltd. All rights reserved.

  18. The Drosophila nicotinic acetylcholine receptor subunits Dα5 and Dα7 form functional homomeric and heteromeric ion channels

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    Lansdell Stuart J

    2012-06-01

    Full Text Available Abstract Background Nicotinic acetylcholine receptors (nAChRs play an important role as excitatory neurotransmitters in vertebrate and invertebrate species. In insects, nAChRs are the site of action of commercially important insecticides and, as a consequence, there is considerable interest in examining their functional properties. However, problems have been encountered in the successful functional expression of insect nAChRs, although a number of strategies have been developed in an attempt to overcome such difficulties. Ten nAChR subunits have been identified in the model insect Drosophila melanogaster (Dα1-Dα7 and Dβ1-Dβ3 and a similar number have been identified in other insect species. The focus of the present study is the Dα5, Dα6 and Dα7 subunits, which are distinguished by their sequence similarity to one another and also by their close similarity to the vertebrate α7 nAChR subunit. Results A full-length cDNA clone encoding the Drosophila nAChR Dα5 subunit has been isolated and the properties of Dα5-, Dα6- and Dα7-containing nAChRs examined in a variety of cell expression systems. We have demonstrated the functional expression, as homomeric nAChRs, of the Dα5 and Dα7 subunits in Xenopus oocytes by their co-expression with the molecular chaperone RIC-3. Also, using a similar approach, we have demonstrated the functional expression of a heteromeric ‘triplet’ nAChR (Dα5 + Dα6 + Dα7 with substantially higher apparent affinity for acetylcholine than is seen with other subunit combinations. In addition, specific cell-surface binding of [125I]-α-bungarotoxin was detected in both Drosophila and mammalian cell lines when Dα5 was co-expressed with Dα6 and RIC-3. In contrast, co-expression of additional subunits (including Dα7 with Dα5 and Dα6 prevented specific binding of [125I]-α-bungarotoxin in cell lines, suggesting that co-assembly with other nAChR subunits can block maturation of correctly folded nAChRs in

  19. Metabolic and functional phenotypic profiling of Drosophila melanogaster reveals reduced sex differentiation under stressful environmental conditions

    DEFF Research Database (Denmark)

    Orsted, Michael; Malmendal, Anders; Munoz, Joaquin

    2018-01-01

    Drosophila melanogaster (Diptera: Drosophilidae), and how this impacts the magnitude of sexual dimorphism. Experimental stressors that we exposed flies to during development were heat stress, poor nutrition, high acidity, high levels of ammonia and ethanol. Emerged male and female flies from the different...

  20. Functional characterization of ecto-5'-nucleotidases and apyrases in Drosophila melanogaster

    Czech Academy of Sciences Publication Activity Database

    Fencková, M.; Hobizalová, R.; Faltýnek Fric, Zdeněk; Doležal, T.

    2011-01-01

    Roč. 41, č. 12 (2011), s. 956-967 ISSN 0965-1748 Grant - others:GA ČR(CZ) GA204/09/1463 Institutional research plan: CEZ:AV0Z50070508 Keywords : Drosophila * ecto-5'-nucleotidase * apyrase Subject RIV: CE - Biochemistry Impact factor: 3.246, year: 2011

  1. Dual role of BMP signaling in the regulation of Drosophila intestinal stem cell self-renewal.

    Science.gov (United States)

    Tian, Aiguo; Jiang, Jin

    2017-10-02

    Many adult organs including Drosophila adult midguts rely on resident stem cells to replenish damaged cells during tissue homeostasis and regeneration. Previous studies have shown that, upon injury, intestinal stem cells (ISCs) in the midguts can increase proliferation and lineage differentiation to meet the demand for tissue repair. Our recent study has demonstrated that, in response to certain injury, midguts can expand ISC population size as an additional regenerative mechanism. We found that injury elicited by bleomycin feeding or bacterial infection increased the production of two BMP ligands (Dpp and Gbb) in enterocytes (ECs), leading to elevated BMP signaling in progenitor cells that drove an expansion of ISCs by promoting their symmetric self-renewing division. Interestingly, we also found that BMP signaling in ECs inhibits the production of Dpp and Gbb, and that this negative feedback mechanism is required to reset ISC pool size to the homeostatic state. Our findings suggest that BMP signaling exerts two opposing influences on stem cell activity depending on where it acts: BMP signaling in progenitor cells promotes ISC self-renewal while BMP signaling in ECs restricts ISC self-renewal by preventing excessive production of BMP ligands. Our results further suggest that transient expansion of ISC population in conjunction with increasing ISC proliferation provides a more effective strategy for tissue regeneration.

  2. C. elegans bicd-1, homolog of the Drosophila dynein accessory factor Bicaudal D, regulates the branching of PVD sensory neuron dendrites.

    Science.gov (United States)

    Aguirre-Chen, Cristina; Bülow, Hannes E; Kaprielian, Zaven

    2011-02-01

    The establishment of cell type-specific dendritic arborization patterns is a key phase in the assembly of neuronal circuitry that facilitates the integration and processing of synaptic and sensory input. Although studies in Drosophila and vertebrate systems have identified a variety of factors that regulate dendrite branch formation, the molecular mechanisms that control this process remain poorly defined. Here, we introduce the use of the Caenorhabditis elegans PVD neurons, a pair of putative nociceptors that elaborate complex dendritic arbors, as a tractable model for conducting high-throughput RNAi screens aimed at identifying key regulators of dendritic branch formation. By carrying out two separate RNAi screens, a small-scale candidate-based screen and a large-scale screen of the ~3000 genes on chromosome IV, we retrieved 11 genes that either promote or suppress the formation of PVD-associated dendrites. We present a detailed functional characterization of one of the genes, bicd-1, which encodes a microtubule-associated protein previously shown to modulate the transport of mRNAs and organelles in a variety of organisms. Specifically, we describe a novel role for bicd-1 in regulating dendrite branch formation and show that bicd-1 is likely to be expressed, and primarily required, in PVD neurons to control dendritic branching. We also present evidence that bicd-1 operates in a conserved pathway with dhc-1 and unc-116, components of the dynein minus-end-directed and kinesin-1 plus-end-directed microtubule-based motor complexes, respectively, and interacts genetically with the repulsive guidance receptor unc-5.

  3. Proteasome, but not autophagy, disruption results in severe eye and wing dysmorphia: a subunit- and regulator-dependent process in Drosophila.

    Science.gov (United States)

    Velentzas, Panagiotis D; Velentzas, Athanassios D; Pantazi, Asimina D; Mpakou, Vassiliki E; Zervas, Christos G; Papassideri, Issidora S; Stravopodis, Dimitrios J

    2013-01-01

    Proteasome-dependent and autophagy-mediated degradation of eukaryotic cellular proteins represent the two major proteostatic mechanisms that are critically implicated in a number of signaling pathways and cellular processes. Deregulation of functions engaged in protein elimination frequently leads to development of morbid states and diseases. In this context, and through the utilization of GAL4/UAS genetic tool, we herein examined the in vivo contribution of proteasome and autophagy systems in Drosophila eye and wing morphogenesis. By exploiting the ability of GAL4-ninaE. GMR and P{GawB}Bx(MS1096) genetic drivers to be strongly and preferentially expressed in the eye and wing discs, respectively, we proved that proteasomal integrity and ubiquitination proficiency essentially control fly's eye and wing development. Indeed, subunit- and regulator-specific patterns of severe organ dysmorphia were obtained after the RNAi-induced downregulation of critical proteasome components (Rpn1, Rpn2, α5, β5 and β6) or distinct protein-ubiquitin conjugators (UbcD6, but not UbcD1 and UbcD4). Proteasome deficient eyes presented with either rough phenotypes or strongly dysmorphic shapes, while transgenic mutant wings were severely folded and carried blistered structures together with loss of vein differentiation. Moreover, transgenic fly eyes overexpressing the UBP2-yeast deubiquitinase enzyme were characterized by an eyeless-like phenotype. Therefore, the proteasome/ubiquitin proteolytic activities are undoubtedly required for the normal course of eye and wing development. In contrast, the RNAi-mediated downregulation of critical Atg (1, 4, 7, 9 and 18) autophagic proteins revealed their non-essential, or redundant, functional roles in Drosophila eye and wing formation under physiological growth conditions, since their reduced expression levels could only marginally disturb wing's, but not eye's, morphogenetic organization and architecture. However, Atg9 proved indispensable for

  4. Proteasome, but not autophagy, disruption results in severe eye and wing dysmorphia: a subunit- and regulator-dependent process in Drosophila.

    Directory of Open Access Journals (Sweden)

    Panagiotis D Velentzas

    Full Text Available Proteasome-dependent and autophagy-mediated degradation of eukaryotic cellular proteins represent the two major proteostatic mechanisms that are critically implicated in a number of signaling pathways and cellular processes. Deregulation of functions engaged in protein elimination frequently leads to development of morbid states and diseases. In this context, and through the utilization of GAL4/UAS genetic tool, we herein examined the in vivo contribution of proteasome and autophagy systems in Drosophila eye and wing morphogenesis. By exploiting the ability of GAL4-ninaE. GMR and P{GawB}Bx(MS1096 genetic drivers to be strongly and preferentially expressed in the eye and wing discs, respectively, we proved that proteasomal integrity and ubiquitination proficiency essentially control fly's eye and wing development. Indeed, subunit- and regulator-specific patterns of severe organ dysmorphia were obtained after the RNAi-induced downregulation of critical proteasome components (Rpn1, Rpn2, α5, β5 and β6 or distinct protein-ubiquitin conjugators (UbcD6, but not UbcD1 and UbcD4. Proteasome deficient eyes presented with either rough phenotypes or strongly dysmorphic shapes, while transgenic mutant wings were severely folded and carried blistered structures together with loss of vein differentiation. Moreover, transgenic fly eyes overexpressing the UBP2-yeast deubiquitinase enzyme were characterized by an eyeless-like phenotype. Therefore, the proteasome/ubiquitin proteolytic activities are undoubtedly required for the normal course of eye and wing development. In contrast, the RNAi-mediated downregulation of critical Atg (1, 4, 7, 9 and 18 autophagic proteins revealed their non-essential, or redundant, functional roles in Drosophila eye and wing formation under physiological growth conditions, since their reduced expression levels could only marginally disturb wing's, but not eye's, morphogenetic organization and architecture. However, Atg9 proved

  5. Comparative analysis of behavioral and transcriptional variation underlying CO2 sensory neuron function and development in Drosophila.

    Science.gov (United States)

    Pan, Jia Wern; McLaughlin, Joi; Yang, Haining; Leo, Charles; Rambarat, Paula; Okuwa, Sumie; Monroy-Eklund, Anaïs; Clark, Sabrina; Jones, Corbin D; Volkan, Pelin Cayirlioglu

    2017-10-02

    Carbon dioxide is an important environmental cue for many insects, regulating many behaviors including some that have direct human impacts. To further improve our understanding of how this system varies among closely related insect species, we examined both the behavioral response to CO 2 as well as the transcriptional profile of key developmental regulators of CO 2 sensory neurons in the olfactory system across the Drosophila genus. We found that CO 2 generally evokes repulsive behavior across most of the Drosophilids we examined, but this behavior has been lost or reduced in several lineages. Comparisons of transcriptional profiles from the developing and adult antennae for subset these species suggest that behavioral differences in some species may be due to differences in the expression of the CO 2 co-receptor Gr63a. Furthermore, these differences in Gr63a expression are correlated with changes in the expression of a few genes known to be involved in the development of the CO 2 circuit, namely dac, an important regulator of sensilla fate for sensilla that house CO 2 ORNs, and mip120, a member of the MMB/dREAM epigenetic regulatory complex that regulates CO 2 receptor expression. In contrast, most of the other known structural, molecular, and developmental components of the peripheral Drosophila CO 2 olfactory system seem to be well-conserved across all examined lineages. These findings suggest that certain components of CO 2 sensory ORN development may be more evolutionarily labile, and may contribute to differences in CO 2 -evoked behavioral responses across species.

  6. The lysosomal enzyme receptor protein (LERP is not essential, but is implicated in lysosomal function in Drosophila melanogaster

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    Medina Hasanagic

    2015-10-01

    Full Text Available The lysosomal enzyme receptor protein (LERP of Drosophila melanogaster is the ortholog of the mammalian cation-independent mannose 6-phosphate (Man 6-P receptor, which mediates trafficking of newly synthesized lysosomal acid hydrolases to lysosomes. However, flies lack the enzymes necessary to make the Man 6-P mark, and the amino acids implicated in Man 6-P binding by the mammalian receptor are not conserved in LERP. Thus, the function of LERP in sorting of lysosomal enzymes to lysosomes in Drosophila is unclear. Here, we analyze the consequence of LERP depletion in S2 cells and intact flies. RNAi-mediated knockdown of LERP in S2 cells had little or no effect on the cellular content or secretion of several lysosomal hydrolases. We generated a novel Lerp null mutation, LerpF6, which abolishes LERP protein expression. Lerp mutants have normal viability and fertility and display no overt phenotypes other than reduced body weight. Lerp mutant flies exhibit a 30–40% decrease in the level of several lysosomal hydrolases, and are hypersensitive to dietary chloroquine and starvation, consistent with impaired lysosome function. Loss of LERP also enhances an eye phenotype associated with defective autophagy. Our findings implicate Lerp in lysosome function and autophagy.

  7. Drosophila atonal fully rescues the phenotype of Math1 null mice: new functions evolve in new cellular contexts

    Science.gov (United States)

    Wang, Vincent Y.; Hassan, Bassem A.; Bellen, Hugo J.; Zoghbi, Huda Y.

    2002-01-01

    Many genes share sequence similarity between species, but their properties often change significantly during evolution. For example, the Drosophila genes engrailed and orthodenticle and the onychophoran gene Ultrabithorax only partially substitute for their mouse or Drosophila homologs. We have been analyzing the relationship between atonal (ato) in the fruit fly and its mouse homolog, Math1. In flies, ato acts as a proneural gene that governs the development of chordotonal organs (CHOs), which serve as stretch receptors in the body wall and joints and as auditory organs in the antennae. In the fly CNS, ato is important not for specification but for axonal arborization. Math1, in contrast, is required for the specification of cells in both the CNS and the PNS. Furthermore, Math1 serves a role in the development of secretory lineage cells in the gut, a function that does not parallel any known to be served by ato. We wondered whether ato and Math1 might be more functionally homologous than they appear, so we expressed Math1 in ato mutant flies and ato in Math1 null mice. To our surprise, the two proteins are functionally interchangeable.

  8. Reversible perturbations of gene regulation after genome editing in Drosophila cells.

    Directory of Open Access Journals (Sweden)

    Stefan Kunzelmann

    Full Text Available The prokaryotic phage defense CRISPR/cas-system has developed into a versatile toolbox for genome engineering and genetic studies in many organisms. While many efforts were spent on analyzing the consequences of off-target effects, only few studies addressed side-effects that occur due to the targeted manipulation of the genome. Here, we show that the CRISPR/cas9-mediated integration of an epitope tag in combination with a selection cassette can trigger an siRNA-mediated, epigenetic genome surveillance pathway in Drosophila melanogaster cells. After homology-directed insertion of the sequence coding for the epitope tag and the selection marker, a moderate level of siRNAs covering the inserted sequence and extending into the targeted locus was detected. This response affected protein levels less than two-fold and it persisted even after single cell cloning. However, removal of the selection cassette abolished the siRNA generation, demonstrating that this response is reversible. Consistently, marker-free genome engineering did not trigger the same surveillance mechanism. These two observations indicate that the selection cassette we employed induces an aberrant transcriptional arrangement and ultimately sets off the siRNA production. There have been prior concerns about undesirable effects induced by selection markers, but fortunately we were able to show that at least one of the epigenetic changes reverts as the marker gene is excised. Although the effects observed were rather weak (less than twofold de-repression upon ago2 or dcr-2 knock-down, we recommend that when selection markers are used during genome editing, a strategy for their subsequent removal should always be included.

  9. HIF- and Non-HIF-Regulated Hypoxic Responses Require the Estrogen-Related Receptor in Drosophila melanogaster

    Science.gov (United States)

    Li, Yan; Padmanabha, Divya; Gentile, Luciana B.; Dumur, Catherine I.; Beckstead, Robert B.; Baker, Keith D.

    2013-01-01

    Low-oxygen tolerance is supported by an adaptive response that includes a coordinate shift in metabolism and the activation of a transcriptional program that is driven by the hypoxia-inducible factor (HIF) pathway. The precise contribution of HIF-1a in the adaptive response, however, has not been determined. Here, we investigate how HIF influences hypoxic adaptation throughout Drosophila melanogaster development. We find that hypoxic-induced transcriptional changes are comprised of HIF-dependent and HIF-independent pathways that are distinct and separable. We show that normoxic set-points of carbohydrate metabolites are significantly altered in sima mutants and that these animals are unable to mobilize glycogen in hypoxia. Furthermore, we find that the estrogen-related receptor (dERR), which is a global regulator of aerobic glycolysis in larvae, is required for a competent hypoxic response. dERR binds to dHIFa and participates in the HIF-dependent transcriptional program in hypoxia. In addition, dERR acts in the absence of dHIFa in hypoxia and a significant portion of HIF-independent transcriptional responses can be attributed to dERR actions, including upregulation of glycolytic transcripts. These results indicate that competent hypoxic responses arise from complex interactions between HIF-dependent and -independent mechanisms, and that dERR plays a central role in both of these programs. PMID:23382692

  10. Integration of light and temperature in the regulation of circadian gene expression in Drosophila.

    Directory of Open Access Journals (Sweden)

    Catharine E Boothroyd

    2007-04-01

    Full Text Available Circadian clocks are aligned to the environment via synchronizing signals, or Zeitgebers, such as daily light and temperature cycles, food availability, and social behavior. In this study, we found that genome-wide expression profiles from temperature-entrained flies show a dramatic difference in the presence or absence of a thermocycle. Whereas transcript levels appear to be modified broadly by changes in temperature, there is a specific set of temperature-entrained circadian mRNA profiles that continue to oscillate in constant conditions. There are marked differences in the biological functions represented by temperature-driven or circadian regulation. The set of temperature-entrained circadian transcripts overlaps significantly with a previously defined set of transcripts oscillating in response to a photocycle. In follow-up studies, all thermocycle-entrained circadian transcript rhythms also responded to light/dark entrainment, whereas some photocycle-entrained rhythms did not respond to temperature entrainment. Transcripts encoding the clock components Period, Timeless, Clock, Vrille, PAR-domain protein 1, and Cryptochrome were all confirmed to be rhythmic after entrainment to a daily thermocycle, although the presence of a thermocycle resulted in an unexpected phase difference between period and timeless expression rhythms at the transcript but not the protein level. Generally, transcripts that exhibit circadian rhythms both in response to thermocycles and photocycles maintained the same mutual phase relationships after entrainment by temperature or light. Comparison of the collective temperature- and light-entrained circadian phases of these transcripts indicates that natural environmental light and temperature cycles cooperatively entrain the circadian clock. This interpretation is further supported by comparative analysis of the circadian phases observed for temperature-entrained and light-entrained circadian locomotor behavior. Taken

  11. Rasputin, the Drosophila homologue of the RasGAP SH3 binding protein, functions in ras- and Rho-mediated signaling.

    Science.gov (United States)

    Pazman, C; Mayes, C A; Fanto, M; Haynes, S R; Mlodzik, M

    2000-04-01

    The small GTPase Ras plays an important role in many cellular signaling processes. Ras activity is negatively regulated by GTPase activating proteins (GAPs). It has been proposed that RasGAP may also function as an effector of Ras activity. We have identified and characterized the Drosophila homologue of the RasGAP-binding protein G3BP encoded by rasputin (rin). rin mutants are viable and display defects in photoreceptor recruitment and ommatidial polarity in the eye. Mutations in rin/G3BP genetically interact with components of the Ras signaling pathway that function at the level of Ras and above, but not with Raf/MAPK pathway components. These interactions suggest that Rin is required as an effector in Ras signaling during eye development, supporting an effector role for RasGAP. The ommatidial polarity phenotypes of rin are similar to those of RhoA and the polarity genes, e.g. fz and dsh. Although rin/G3BP interacts genetically with RhoA, affecting both photoreceptor differentiation and polarity, it does not interact with the gain-of-function genotypes of fz and dsh. These data suggest that Rin is not a general component of polarity generation, but serves a function specific to Ras and RhoA signaling pathways.

  12. Anatomical Reconstruction and Functional Imaging Reveal an Ordered Array of Skylight Polarization Detectors in Drosophila.

    Science.gov (United States)

    Weir, Peter T; Henze, Miriam J; Bleul, Christiane; Baumann-Klausener, Franziska; Labhart, Thomas; Dickinson, Michael H

    2016-05-11

    Many insects exploit skylight polarization as a compass cue for orientation and navigation. In the fruit fly, Drosophila melanogaster, photoreceptors R7 and R8 in the dorsal rim area (DRA) of the compound eye are specialized to detect the electric vector (e-vector) of linearly polarized light. These photoreceptors are arranged in stacked pairs with identical fields of view and spectral sensitivities, but mutually orthogonal microvillar orientations. As in larger flies, we found that the microvillar orientation of the distal photoreceptor R7 changes in a fan-like fashion along the DRA. This anatomical arrangement suggests that the DRA constitutes a detector for skylight polarization, in which different e-vectors maximally excite different positions in the array. To test our hypothesis, we measured responses to polarized light of varying e-vector angles in the terminals of R7/8 cells using genetically encoded calcium indicators. Our data confirm a progression of preferred e-vector angles from anterior to posterior in the DRA, and a strict orthogonality between the e-vector preferences of paired R7/8 cells. We observed decreased activity in photoreceptors in response to flashes of light polarized orthogonally to their preferred e-vector angle, suggesting reciprocal inhibition between photoreceptors in the same medullar column, which may serve to increase polarization contrast. Together, our results indicate that the polarization-vision system relies on a spatial map of preferred e-vector angles at the earliest stage of sensory processing. The fly's visual system is an influential model system for studying neural computation, and much is known about its anatomy, physiology, and development. The circuits underlying motion processing have received the most attention, but researchers are increasingly investigating other functions, such as color perception and object recognition. In this work, we investigate the early neural processing of a somewhat exotic sense, called

  13. Ulysses transposable element of Drosophila shows high structural similarities to functional domains of retroviruses.

    Science.gov (United States)

    Evgen'ev, M B; Corces, V G; Lankenau, D H

    1992-06-05

    We have determined the DNA structure of the Ulysses transposable element of Drosophila virilis and found that this transposon is 10,653 bp and is flanked by two unusually large direct repeats 2136 bp long. Ulysses shows the characteristic organization of LTR-containing retrotransposons, with matrix and capsid protein domains encoded in the first open reading frame. In addition, Ulysses contains protease, reverse transcriptase, RNase H and integrase domains encoded in the second open reading frame. Ulysses lacks a third open reading frame present in some retrotransposons that could encode an env-like protein. A dendrogram analysis based on multiple alignments of the protease, reverse transcriptase, RNase H, integrase and tRNA primer binding site of all known Drosophila LTR-containing retrotransposon sequences establishes a phylogenetic relationship of Ulysses to other retrotransposons and suggests that Ulysses belongs to a new family of this type of elements.

  14. CREB Binding Protein Functions During Successive Stages of Eye Development in Drosophila

    OpenAIRE

    Kumar, Justin P.; Jamal, Tazeen; Doetsch, Alex; Turner, F. Rudolf; Duffy, Joseph B.

    2004-01-01

    During the development of the compound eye of Drosophila several signaling pathways exert both positive and inhibitory influences upon an array of nuclear transcription factors to produce a near-perfect lattice of unit eyes or ommatidia. Individual cells within the eye are exposed to many extracellular signals, express multiple surface receptors, and make use of a large complement of cell-subtype-specific DNA-binding transcription factors. Despite this enormous complexity, each cell will make...

  15. Hyperactive locomotion in a Drosophila model is a functional readout for the synaptic abnormalities underlying fragile X syndrome.

    Science.gov (United States)

    Kashima, Risa; Redmond, Patrick L; Ghatpande, Prajakta; Roy, Sougata; Kornberg, Thomas B; Hanke, Thomas; Knapp, Stefan; Lagna, Giorgio; Hata, Akiko

    2017-05-02

    Fragile X syndrome (FXS) is the most common cause of heritable intellectual disability and autism and affects ~1 in 4000 males and 1 in 8000 females. The discovery of effective treatments for FXS has been hampered by the lack of effective animal models and phenotypic readouts for drug screening. FXS ensues from the epigenetic silencing or loss-of-function mutation of the fragile X mental retardation 1 ( FMR1 ) gene, which encodes an RNA binding protein that associates with and represses the translation of target mRNAs. We previously found that the activation of LIM kinase 1 (LIMK1) downstream of augmented synthesis of bone morphogenetic protein (BMP) type 2 receptor (BMPR2) promotes aberrant synaptic development in mouse and Drosophila models of FXS and that these molecular and cellular markers were correlated in patients with FXS. We report that larval locomotion is augmented in a Drosophila FXS model. Genetic or pharmacological intervention on the BMPR2-LIMK pathway ameliorated the synaptic abnormality and locomotion phenotypes of FXS larvae, as well as hyperactivity in an FXS mouse model. Our study demonstrates that (i) the BMPR2-LIMK pathway is a promising therapeutic target for FXS and (ii) the locomotion phenotype of FXS larvae is a quantitative functional readout for the neuromorphological phenotype associated with FXS and is amenable to the screening novel FXS therapeutics. Copyright © 2017, American Association for the Advancement of Science.

  16. How gut transcriptional function of Drosophila melanogaster varies with the presence and composition of the gut microbiota.

    Science.gov (United States)

    Bost, Alyssa; Franzenburg, Soeren; Adair, Karen L; Martinson, Vincent G; Loeb, Greg; Douglas, Angela E

    2018-04-01

    Despite evidence from laboratory experiments that perturbation of the gut microbiota affects many traits of the animal host, our understanding of the effect of variation in microbiota composition on animals in natural populations is very limited. The core purpose of this study on the fruit fly Drosophila melanogaster was to identify the impact of natural variation in the taxonomic composition of gut bacterial communities on host traits, with the gut transcriptome as a molecular index of microbiota-responsive host traits. Use of the gut transcriptome was validated by demonstrating significant transcriptional differences between the guts of laboratory flies colonized with bacteria and maintained under axenic conditions. Wild Drosophila from six field collections made over two years had gut bacterial communities of diverse composition, dominated to varying extents by Acetobacteraceae and Enterobacteriaceae. The gut transcriptomes also varied among collections and differed markedly from those of laboratory flies. However, no overall relationship between variation in the wild fly transcriptome and taxonomic composition of the gut microbiota was evident at all taxonomic scales of bacteria tested for both individual fly genes and functional categories in Gene Ontology. We conclude that the interaction between microbiota composition and host functional traits may be confounded by uncontrolled variation in both ecological circumstance and host traits (e.g., genotype, age physiological condition) under natural conditions, and that microbiota effects on host traits identified in the laboratory should, therefore, be extrapolated to field population with great caution. © 2017 John Wiley & Sons Ltd.

  17. The ALS-associated proteins FUS and TDP-43 function together to affect Drosophila locomotion and life span

    Science.gov (United States)

    Wang, Ji-Wu; Brent, Jonathan R.; Tomlinson, Andrew; Shneider, Neil A.; McCabe, Brian D.

    2011-01-01

    The fatal adult motor neuron disease amyotrophic lateral sclerosis (ALS) shares some clinical and pathological overlap with frontotemporal dementia (FTD), an early-onset neurodegenerative disorder. The RNA/DNA-binding proteins fused in sarcoma (FUS; also known as TLS) and TAR DNA binding protein-43 (TDP-43) have recently been shown to be genetically and pathologically associated with familial forms of ALS and FTD. It is currently unknown whether perturbation of these proteins results in disease through mechanisms that are independent of normal protein function or via the pathophysiological disruption of molecular processes in which they are both critical. Here, we report that Drosophila mutants in which the homolog of FUS is disrupted exhibit decreased adult viability, diminished locomotor speed, and reduced life span compared with controls. These phenotypes were fully rescued by wild-type human FUS, but not ALS-associated mutant FUS proteins. A mutant of the Drosophila homolog of TDP-43 had similar, but more severe, deficits. Through cross-rescue analysis, we demonstrated that FUS acted together with and downstream of TDP-43 in a common genetic pathway in neurons. Furthermore, we found that these proteins associated with each other in an RNA-dependent complex. Our results establish that FUS and TDP-43 function together in vivo and suggest that molecular pathways requiring the combined activities of both of these proteins may be disrupted in ALS and FTD. PMID:21881207

  18. Insulin/IGF-regulated size scaling of neuroendocrine cells expressing the bHLH transcription factor Dimmed in Drosophila.

    Directory of Open Access Journals (Sweden)

    Jiangnan Luo

    Full Text Available Neurons and other cells display a large variation in size in an organism. Thus, a fundamental question is how growth of individual cells and their organelles is regulated. Is size scaling of individual neurons regulated post-mitotically, independent of growth of the entire CNS? Although the role of insulin/IGF-signaling (IIS in growth of tissues and whole organisms is well established, it is not known whether it regulates the size of individual neurons. We therefore studied the role of IIS in the size scaling of neurons in the Drosophila CNS. By targeted genetic manipulations of insulin receptor (dInR expression in a variety of neuron types we demonstrate that the cell size is affected only in neuroendocrine cells specified by the bHLH transcription factor DIMMED (DIMM. Several populations of DIMM-positive neurons tested displayed enlarged cell bodies after overexpression of the dInR, as well as PI3 kinase and Akt1 (protein kinase B, whereas DIMM-negative neurons did not respond to dInR manipulations. Knockdown of these components produce the opposite phenotype. Increased growth can also be induced by targeted overexpression of nutrient-dependent TOR (target of rapamycin signaling components, such as Rheb (small GTPase, TOR and S6K (S6 kinase. After Dimm-knockdown in neuroendocrine cells manipulations of dInR expression have significantly less effects on cell size. We also show that dInR expression in neuroendocrine cells can be altered by up or down-regulation of Dimm. This novel dInR-regulated size scaling is seen during postembryonic development, continues in the aging adult and is diet dependent. The increase in cell size includes cell body, axon terminations, nucleus and Golgi apparatus. We suggest that the dInR-mediated scaling of neuroendocrine cells is part of a plasticity that adapts the secretory capacity to changing physiological conditions and nutrient-dependent organismal growth.

  19. Ingestion of gallium phosphide nanowires has no adverse effect on Drosophila tissue function

    International Nuclear Information System (INIS)

    Adolfsson, Karl; Hammarin, Greger; Prinz, Christelle N; Schneider, Martina; Häcker, Udo

    2013-01-01

    Engineered nanoparticles have been under increasing scrutiny in recent years. High aspect ratio nanoparticles such as carbon nanotubes and nanowires have raised safety concerns due to their geometrical similarity to asbestos fibers. III–V epitaxial semiconductor nanowires are expected to be utilized in devices such as LEDs and solar cells and will thus be available to the public. In addition, clean-room staff fabricating and characterizing the nanowires are at risk of exposure, emphasizing the importance of investigating their possible toxicity. Here we investigated the effects of gallium phosphide nanowires on the fruit fly Drosophila melanogaster. Drosophila larvae and/or adults were exposed to gallium phosphide nanowires by ingestion with food. The toxicity and tissue interaction of the nanowires was evaluated by investigating tissue distribution, activation of immune response, genome-wide gene expression, life span, fecundity and somatic mutation rates. Our results show that gallium phosphide nanowires applied through the diet are not taken up into Drosophila tissues, do not elicit a measurable immune response or changes in genome-wide gene expression and do not significantly affect life span or somatic mutation rate. (paper)

  20. Intercellular calcium signaling is regulated by morphogens during Drosophila wing development

    OpenAIRE

    Chen, Danny; Levis, Megan; Arredondo-Walsh, Ninfamaria; Zartman, Jeremiah; Brodskiy, Pavel; Wu, Qinfeng; Huizar, Francisco; Soundarrajan, Dharsan; Narciso, Cody; Chen, Jianxu; Liang, Peixian

    2017-01-01

    Organ development is driven by a set of patterned inductive signals. However, how these signals are integrated to coordinate tissue patterning is still poorly understood. Calcium ions (Ca2+) are critical signaling components involved in signal integration and are regulated by a core Ca2+ signaling toolkit. Ca2+ signaling encodes a significant fraction of information in cells through both amplitude and frequency-dependent regulation of transcription factors and key regulatory enzymes. A range ...

  1. Presynaptic DLG regulates synaptic function through the localization of voltage-activated Ca2+ Channels

    Science.gov (United States)

    Astorga, César; Jorquera, Ramón A.; Ramírez, Mauricio; Kohler, Andrés; López, Estefanía; Delgado, Ricardo; Córdova, Alex; Olguín, Patricio; Sierralta, Jimena

    2016-01-01

    The DLG-MAGUK subfamily of proteins plays a role on the recycling and clustering of glutamate receptors (GLUR) at the postsynaptic density. discs-large1 (dlg) is the only DLG-MAGUK gene in Drosophila and originates two main products, DLGA and DLGS97 which differ by the presence of an L27 domain. Combining electrophysiology, immunostaining and genetic manipulation at the pre and postsynaptic compartments we study the DLG contribution to the basal synaptic-function at the Drosophila larval neuromuscular junction. Our results reveal a specific function of DLGS97 in the regulation of the size of GLUR fields and their subunit composition. Strikingly the absence of any of DLG proteins at the presynaptic terminal disrupts the clustering and localization of the calcium channel DmCa1A subunit (Cacophony), decreases the action potential-evoked release probability and alters short-term plasticity. Our results show for the first time a crucial role of DLG proteins in the presynaptic function in vivo. PMID:27573697

  2. Presynaptic DLG regulates synaptic function through the localization of voltage-activated Ca(2+) Channels.

    Science.gov (United States)

    Astorga, César; Jorquera, Ramón A; Ramírez, Mauricio; Kohler, Andrés; López, Estefanía; Delgado, Ricardo; Córdova, Alex; Olguín, Patricio; Sierralta, Jimena

    2016-08-30

    The DLG-MAGUK subfamily of proteins plays a role on the recycling and clustering of glutamate receptors (GLUR) at the postsynaptic density. discs-large1 (dlg) is the only DLG-MAGUK gene in Drosophila and originates two main products, DLGA and DLGS97 which differ by the presence of an L27 domain. Combining electrophysiology, immunostaining and genetic manipulation at the pre and postsynaptic compartments we study the DLG contribution to the basal synaptic-function at the Drosophila larval neuromuscular junction. Our results reveal a specific function of DLGS97 in the regulation of the size of GLUR fields and their subunit composition. Strikingly the absence of any of DLG proteins at the presynaptic terminal disrupts the clustering and localization of the calcium channel DmCa1A subunit (Cacophony), decreases the action potential-evoked release probability and alters short-term plasticity. Our results show for the first time a crucial role of DLG proteins in the presynaptic function in vivo.

  3. A novel ecdysone receptor mediates steroid-regulated developmental events during the mid-third instar of Drosophila.

    Directory of Open Access Journals (Sweden)

    Benjamin F B Costantino

    2008-06-01

    Full Text Available The larval salivary gland of Drosophila melanogaster synthesizes and secretes glue glycoproteins that cement developing animals to a solid surface during metamorphosis. The steroid hormone 20-hydroxyecdysone (20E is an essential signaling molecule that modulates most of the physiological functions of the larval gland. At the end of larval development, it is known that 20E--signaling through a nuclear receptor heterodimer consisting of EcR and USP--induces the early and late puffing cascade of the polytene chromosomes and causes the exocytosis of stored glue granules into the lumen of the gland. It has also been reported that an earlier pulse of hormone induces the temporally and spatially specific transcriptional activation of the glue genes; however, the receptor responsible for triggering this response has not been characterized. Here we show that the coordinated expression of the glue genes midway through the third instar is mediated by 20E acting to induce genes of the Broad Complex (BRC through a receptor that is not an EcR/USP heterodimer. This result is novel because it demonstrates for the first time that at least some 20E-mediated, mid-larval, developmental responses are controlled by an uncharacterized receptor that does not contain an RXR-like component.

  4. Atg9 antagonizes TOR signaling to regulate intestinal cell growth and epithelial homeostasis in Drosophila.

    Science.gov (United States)

    Wen, Jung-Kun; Wang, Yi-Ting; Chan, Chih-Chiang; Hsieh, Cheng-Wen; Liao, Hsiao-Man; Hung, Chin-Chun; Chen, Guang-Chao

    2017-11-16

    Autophagy is essential for maintaining cellular homeostasis and survival under various stress conditions. Autophagy-related gene 9 (Atg9) encodes a multipass transmembrane protein thought to act as a membrane carrier for forming autophagosomes. However, the molecular regulation and physiological importance of Atg9 in animal development remain largely unclear. Here, we generated Atg9 null mutant flies and found that loss of Atg9 led to shortened lifespan, locomotor defects, and increased susceptibility to stress. Atg9 loss also resulted in aberrant adult midgut morphology with dramatically enlarged enterocytes. Interestingly, inhibiting the TOR signaling pathway rescued the midgut defects of the Atg9 mutants. In addition, Atg9 interacted with PALS1-associated tight junction protein (Patj), which associates with TSC2 to regulate TOR activity. Depletion of Atg9 caused a marked decrease in TSC2 levels. Our findings revealed an antagonistic relationship between Atg9 and TOR signaling in the regulation of cell growth and tissue homeostasis.

  5. Novel Triazole linked 2-phenyl benzoxazole derivatives induce apoptosis by inhibiting miR-2, miR-13 and miR-14 function in Drosophila melanogaster.

    Science.gov (United States)

    Mondal, Tanmoy; Lavanya, A V S; Mallick, Akash; Dadmala, Tulshiram L; Kumbhare, Ravindra M; Bhadra, Utpal; Bhadra, Manika Pal

    2017-06-01

    Apoptosis is an important phenomenon in multi cellular organisms for maintaining tissue homeostasis and embryonic development. Defect in apoptosis leads to a number of disorders like- autoimmune disorder, immunodeficiency and cancer. 21-22 nucleotides containing micro RNAs (miRNAs/miRs) function as a crucial regulator of apoptosis alike other cellular pathways. Recently, small molecules have been identified as a potent inducer of apoptosis. In this study, we have identified novel Triazole linked 2-phenyl benzoxazole derivatives (13j and 13h) as a negative regulator of apoptosis inhibiting micro RNAs (miR-2, miR-13 and miR-14) in a well established in vivo model Drosophila melanogaster where the process of apoptosis is very similar to human apoptosis. These compounds inhibit miR-2, miR-13 and miR-14 activity at their target sites, which induce an increased caspase activity, and in turn influence the caspase dependent apoptotic pathway. These two compounds also increase the mitochondrial reactive oxygen species (ROS) level to trigger apoptotic cell death.

  6. A splice isoform of DNedd4, DNedd4-long, negatively regulates neuromuscular synaptogenesis and viability in Drosophila.

    Directory of Open Access Journals (Sweden)

    Yunan Zhong

    Full Text Available Neuromuscular (NM synaptogenesis is a tightly regulated process. We previously showed that in flies, Drosophila Nedd4 (dNedd4/dNedd4S is required for proper NM synaptogenesis by promoting endocytosis of commissureless from the muscle surface, a pre-requisite step for muscle innervation. DNedd4 is an E3 ubiquitin ligase comprised of a C2-WW(x3-Hect domain architecture, which includes several splice isoforms, the most prominent ones are dNedd4-short (dNedd4S and dNedd4-long (dNedd4Lo.We show here that while dNedd4S is essential for NM synaptogenesis, the dNedd4Lo isoform inhibits this process and causes lethality. Our results reveal that unlike dNedd4S, dNedd4Lo cannot rescue the lethality of dNedd4 null (DNedd4(T121FS flies. Moreover, overexpression of UAS-dNedd4Lo specifically in wildtype muscles leads to NM synaptogenesis defects, impaired locomotion and larval lethality. These negative effects of dNedd4Lo are ameliorated by deletion of two regions (N-terminus and Middle region unique to this isoform, and by inactivating the catalytic activity of dNedd4Lo, suggesting that these unique regions, as well as catalytic activity, are responsible for the inhibitory effects of dNedd4Lo on synaptogenesis. In accord with these findings, we demonstrate by sqRT-PCR an increase in dNedd4S expression relative to the expression of dNedd4Lo during embryonic stages when synaptogenesis takes place.Our studies demonstrate that splice isoforms of the same dNedd4 gene can lead to opposite effects on NM synaptogenesis.

  7. Interactions between Type III receptor tyrosine phosphatases and growth factor receptor tyrosine kinases regulate tracheal tube formation in Drosophila

    Directory of Open Access Journals (Sweden)

    Mili Jeon

    2012-04-01

    The respiratory (tracheal system of the Drosophila melanogaster larva is an intricate branched network of air-filled tubes. Its developmental logic is similar in some ways to that of the vertebrate vascular system. We previously described a unique embryonic tracheal tubulogenesis phenotype caused by loss of both of the Type III receptor tyrosine phosphatases (RPTPs, Ptp4E and Ptp10D. In Ptp4E Ptp10D double mutants, the linear tubes in unicellular and terminal tracheal branches are converted into bubble-like cysts that incorporate apical cell surface markers. This tube geometry phenotype is modulated by changes in the activity or expression of the epidermal growth factor receptor (Egfr tyrosine kinase (TK. Ptp10D physically interacts with Egfr. Here we demonstrate that the Ptp4E Ptp10D phenotype is the consequence of the loss of negative regulation by the RPTPs of three growth factor receptor TKs: Egfr, Breathless and Pvr. Reducing the activity of any of the three kinases by tracheal expression of dominant-negative mutants suppresses cyst formation. By competing dominant-negative and constitutively active kinase mutants against each other, we show that the three RTKs have partially interchangeable activities, so that increasing the activity of one kinase can compensate for the effects of reducing the activity of another. This implies that SH2-domain downstream effectors that are required for the phenotype are likely to be able to interact with phosphotyrosine sites on all three receptor TKs. We also show that the phenotype involves increases in signaling through the MAP kinase and Rho GTPase pathways.

  8. Drosophila's contribution to stem cell research [v1; ref status: indexed, http://f1000r.es/5h7

    Directory of Open Access Journals (Sweden)

    Gyanesh Singh

    2015-06-01

    Full Text Available The discovery of Drosophila stem cells with striking similarities to mammalian stem cells has brought new hope for stem cell research. A recent development in Drosophila stem cell research is bringing wider opportunities for contemporary stem cell biologists. In this regard, Drosophila germ cells are becoming a popular model of stem cell research. In several cases, genes that controlled Drosophila stem cells were later discovered to have functional homologs in mammalian stem cells. Like mammals, Drosophila germline stem cells (GSCs are controlled by both intrinsic as well as external signals. Inside the Drosophila testes, germline and somatic stem cells form a cluster of cells (the hub. Hub cells depend on JAK-STAT signaling, and, in absence of this signal, they do not self-renew. In Drosophila, significant changes occur within the stem cell niche that contributes to a decline in stem cell number over time. In case of aging Drosophila, somatic niche cells show reduced DE-cadherin and unpaired (Upd proteins. Unpaired proteins are known to directly decrease stem cell number within the niches, and, overexpression of upd within niche cells restored GSCs in older males also . Stem cells in the midgut of Drosophila are also very promising. Reduced Notch signaling was found to increase the number of midgut progenitor cells. On the other hand, activation of the Notch pathway decreased proliferation of these cells. Further research in this area should lead to the discovery of additional factors that regulate stem and progenitor cells in Drosophila.

  9. The Nutrient-Responsive Hormone CCHamide-2 Controls Growth by Regulating Insulin-like Peptides in the Brain of Drosophila melanogaster.

    Science.gov (United States)

    Sano, Hiroko; Nakamura, Akira; Texada, Michael J; Truman, James W; Ishimoto, Hiroshi; Kamikouchi, Azusa; Nibu, Yutaka; Kume, Kazuhiko; Ida, Takanori; Kojima, Masayasu

    2015-05-01

    The coordination of growth with nutritional status is essential for proper development and physiology. Nutritional information is mostly perceived by peripheral organs before being relayed to the brain, which modulates physiological responses. Hormonal signaling ensures this organ-to-organ communication, and the failure of endocrine regulation in humans can cause diseases including obesity and diabetes. In Drosophila melanogaster, the fat body (adipose tissue) has been suggested to play an important role in coupling growth with nutritional status. Here, we show that the peripheral tissue-derived peptide hormone CCHamide-2 (CCHa2) acts as a nutrient-dependent regulator of Drosophila insulin-like peptides (Dilps). A BAC-based transgenic reporter revealed strong expression of CCHa2 receptor (CCHa2-R) in insulin-producing cells (IPCs) in the brain. Calcium imaging of brain explants and IPC-specific CCHa2-R knockdown demonstrated that peripheral-tissue derived CCHa2 directly activates IPCs. Interestingly, genetic disruption of either CCHa2 or CCHa2-R caused almost identical defects in larval growth and developmental timing. Consistent with these phenotypes, the expression of dilp5, and the release of both Dilp2 and Dilp5, were severely reduced. Furthermore, transcription of CCHa2 is altered in response to nutritional levels, particularly of glucose. These findings demonstrate that CCHa2 and CCHa2-R form a direct link between peripheral tissues and the brain, and that this pathway is essential for the coordination of systemic growth with nutritional availability. A mammalian homologue of CCHa2-R, Bombesin receptor subtype-3 (Brs3), is an orphan receptor that is expressed in the islet β-cells; however, the role of Brs3 in insulin regulation remains elusive. Our genetic approach in Drosophila melanogaster provides the first evidence, to our knowledge, that bombesin receptor signaling with its endogenous ligand promotes insulin production.

  10. Grasshopper Lazarillo, a GPI-anchored Lipocalin, increases Drosophila longevity and stress resistance, and functionally replaces its secreted homolog NLaz.

    Science.gov (United States)

    Ruiz, Mario; Wicker-Thomas, Claude; Sanchez, Diego; Ganfornina, Maria D

    2012-10-01

    Lazarillo (Laz) is a glycosyl-phosphatidylinositol (GPI)-linked glycoprotein first characterized in the developing nervous system of the grasshopper Schistocerca americana. It belongs to the Lipocalins, a functionally diverse family of mostly secreted proteins. In this work we test whether the protective capacity known for Laz homologs in flies and vertebrates (NLaz, GLaz and ApoD) is evolutionarily conserved in grasshopper Laz, and can be exerted from the plasma membrane in a cell-autonomous manner. First we demonstrate that extracellular forms of Laz have autocrine and paracrine protecting effects for oxidative stress-challenged Drosophila S2 cells. Then we assay the effects of overexpressing GPI-linked Laz in adult Drosophila and whether it rescues both known and novel phenotypes of NLaz null mutants. Local effects of GPI-linked Laz inside and outside the nervous system promote survival upon different stress forms, and extend lifespan and healthspan of the flies in a cell-type dependent manner. Outside the nervous system, expression in fat body cells but not in hemocytes results in protection. Within the nervous system, glial cell expression is more effective than neuronal expression. Laz actions are sexually dimorphic in some expression domains. Fat storage promotion and not modifications in hydrocarbon profiles or quantities explain the starvation-desiccation resistance caused by Laz overexpression. This effect is exerted when Laz is expressed ubiquitously or in dopaminergic cells, but not in hemocytes. Grasshopper Laz functionally restores the loss of NLaz, rescuing stress-sensitivity as well as premature accumulation of aging-related damage, monitored by advanced glycation end products (AGEs). However Laz does not rescue NLaz courtship behavioral defects. Finally, the presence of two new Lipocalins with predicted GPI-anchors in mosquitoes shows that the functional advantages of GPI-linkage have been commonly exploited by Lipocalins in the arthropodan lineage

  11. Drosophila Atlastin in motor neurons is required for locomotion and presynaptic function.

    Science.gov (United States)

    De Gregorio, Cristian; Delgado, Ricardo; Ibacache, Andrés; Sierralta, Jimena; Couve, Andrés

    2017-10-15

    Hereditary spastic paraplegias (HSPs) are characterized by spasticity and weakness of the lower limbs, resulting from length-dependent axonopathy of the corticospinal tracts. In humans, the HSP-related atlastin genes ATL1 - ATL3 catalyze homotypic membrane fusion of endoplasmic reticulum (ER) tubules. How defects in neuronal Atlastin contribute to axonal degeneration has not been explained satisfactorily. Using Drosophila , we demonstrate that downregulation or overexpression of Atlastin in motor neurons results in decreased crawling speed and contraction frequency in larvae, while adult flies show progressive decline in climbing ability. Broad expression in the nervous system is required to rescue the atlastin -null Drosophila mutant ( atl 2 ) phenotype. Importantly, both spontaneous release and the reserve pool of synaptic vesicles are affected. Additionally, axonal secretory organelles are abnormally distributed, whereas presynaptic proteins diminish at terminals and accumulate in distal axons, possibly in lysosomes. Our findings suggest that trafficking defects produced by Atlastin dysfunction in motor neurons result in redistribution of presynaptic components and aberrant mobilization of synaptic vesicles, stressing the importance of ER-shaping proteins and the susceptibility of motor neurons to their mutations or depletion. © 2017. Published by The Company of Biologists Ltd.

  12. Gain-of-function screen for genes that affect Drosophila muscle pattern formation.

    Directory of Open Access Journals (Sweden)

    Nicole Staudt

    2005-10-01

    Full Text Available This article reports the production of an EP-element insertion library with more than 3,700 unique target sites within the Drosophila melanogaster genome and its use to systematically identify genes that affect embryonic muscle pattern formation. We designed a UAS/GAL4 system to drive GAL4-responsive expression of the EP-targeted genes in developing apodeme cells to which migrating myotubes finally attach and in an intrasegmental pattern of cells that serve myotubes as a migration substrate on their way towards the apodemes. The results suggest that misexpression of more than 1.5% of the Drosophila genes can interfere with proper myotube guidance and/or muscle attachment. In addition to factors already known to participate in these processes, we identified a number of enzymes that participate in the synthesis or modification of protein carbohydrate side chains and in Ubiquitin modifications and/or the Ubiquitin-dependent degradation of proteins, suggesting that these processes are relevant for muscle pattern formation.

  13. Lipoproteins in Drosophila melanogaster—Assembly, Function, and Influence on Tissue Lipid Composition

    Science.gov (United States)

    Palm, Wilhelm; Sampaio, Julio L.; Brankatschk, Marko; Carvalho, Maria; Mahmoud, Ali; Shevchenko, Andrej; Eaton, Suzanne

    2012-01-01

    Interorgan lipid transport occurs via lipoproteins, and altered lipoprotein levels correlate with metabolic disease. However, precisely how lipoproteins affect tissue lipid composition has not been comprehensively analyzed. Here, we identify the major lipoproteins of Drosophila melanogaster and use genetics and mass spectrometry to study their assembly, interorgan trafficking, and influence on tissue lipids. The apoB-family lipoprotein Lipophorin (Lpp) is the major hemolymph lipid carrier. It is produced as a phospholipid-rich particle by the fat body, and its secretion requires Microsomal Triglyceride Transfer Protein (MTP). Lpp acquires sterols and most diacylglycerol (DAG) at the gut via Lipid Transfer Particle (LTP), another fat body-derived apoB-family lipoprotein. The gut, like the fat body, is a lipogenic organ, incorporating both de novo–synthesized and dietary fatty acids into DAG for export. We identify distinct requirements for LTP and Lpp-dependent lipid mobilization in contributing to the neutral and polar lipid composition of the brain and wing imaginal disc. These studies define major routes of interorgan lipid transport in Drosophila and uncover surprising tissue-specific differences in lipoprotein lipid utilization. PMID:22844248

  14. Adaptive Evolution of Gene Expression in Drosophila

    Directory of Open Access Journals (Sweden)

    Armita Nourmohammad

    2017-08-01

    Full Text Available Gene expression levels are important quantitative traits that link genotypes to molecular functions and fitness. In Drosophila, population-genetic studies have revealed substantial adaptive evolution at the genomic level, but the evolutionary modes of gene expression remain controversial. Here, we present evidence that adaptation dominates the evolution of gene expression levels in flies. We show that 64% of the observed expression divergence across seven Drosophila species are adaptive changes driven by directional selection. Our results are derived from time-resolved data of gene expression divergence across a family of related species, using a probabilistic inference method for gene-specific selection. Adaptive gene expression is stronger in specific functional classes, including regulation, sensory perception, sexual behavior, and morphology. Moreover, we identify a large group of genes with sex-specific adaptation of expression, which predominantly occurs in males. Our analysis opens an avenue to map system-wide selection on molecular quantitative traits independently of their genetic basis.

  15. Behavioral Teratogenesis in Drosophila melanogaster.

    Science.gov (United States)

    Mishra, Monalisa; Barik, Bedanta Kumar

    2018-01-01

    Developmental biology is a fascinating branch of science which helps us to understand the mechanism of development, thus the findings are used in various therapeutic approach. Drosophila melanogaster served as a model to find the key molecules that initiate and regulate the mechanism of development. Various genes, transcription factors, and signaling pathways helping in development are identified in Drosophila. Many toxic compounds, which can affect the development, are also recognized using Drosophila model. These compounds, which can affect the development, are named as a teratogen. Many teratogens identified using Drosophila may also act as a teratogen for a human being since 75% of conservation exist between the disease genes present in Drosophila and human. There are certain teratogens, which do not cause developmental defect if exposed during pregnancy, however; behavioral defect appears in later part of development. Such compounds are named as a behavioral teratogen. Thus, it is worthy to identify the potential behavioral teratogen using Drosophila model. Drosophila behavior is well studied in various developmental stages. This chapter describes various methods which can be employed to test behavioral teratogenesis in Drosophila.

  16. Phospho-regulated Drosophila adducin is a determinant of synaptic plasticity in a complex with Dlg and PIP2 at the larval neuromuscular junction

    Directory of Open Access Journals (Sweden)

    Simon Ji Hau Wang

    2014-11-01

    Full Text Available Adducin is a ubiquitously expressed actin- and spectrin-binding protein involved in cytoskeleton organization, and is regulated through phosphorylation of the myristoylated alanine-rich C-terminal kinase (MARCKS-homology domain by protein kinase C (PKC. We have previously shown that the Drosophila adducin, Hu-li tai shao (Hts, plays a role in larval neuromuscular junction (NMJ growth. Here, we find that the predominant isoforms of Hts at the NMJ contain the MARCKS-homology domain, which is important for interactions with Discs large (Dlg and phosphatidylinositol 4,5-bisphosphate (PIP2. Through the use of Proximity Ligation Assay (PLA, we show that the adducin-like Hts isoforms are in complexes with Dlg and PIP2 at the NMJ. We provide evidence that Hts promotes the phosphorylation and delocalization of Dlg at the NMJ through regulation of the transcript distribution of the PAR-1 and CaMKII kinases in the muscle. We also show that Hts interactions with Dlg and PIP2 are impeded through phosphorylation of the MARCKS-homology domain. These results are further evidence that Hts is a signaling-responsive regulator of synaptic plasticity in Drosophila.

  17. Regulation of Pattern Formation and Gene Amplification During Drosophila Oogenesis by the miR-318 microRNA

    DEFF Research Database (Denmark)

    Ge, Wanzhong; Deng, Qiannan; Guo, Ting

    2015-01-01

    Pattern formation during epithelial development requires the coordination of multiple signaling pathways. Here, we investigate the functions of an ovary-enriched miRNA, miR-318, in epithelial development during Drosophila oogenesis. miR-318 maternal loss-of-function mutants were female sterile...... and laid eggs with abnormal morphology. Removal of miR-318 disrupted the dorsal-anterior follicle cell patterning, resulting in abnormal dorsal appendages. miR-318 mutant females also produced thin and fragile eggshells, due to impaired chorion gene amplification. We provide evidence that the ecdysone......RNAs in maintaining cell fate and promoting the developmental transition in the female follicular epithelium....

  18. Drosophila Polo regulates the spindle assembly checkpoint through Mps1-dependent BubR1 phosphorylation.

    Science.gov (United States)

    Conde, Carlos; Osswald, Mariana; Barbosa, João; Moutinho-Santos, Tatiana; Pinheiro, Diana; Guimarães, Sofia; Matos, Irina; Maiato, Helder; Sunkel, Claudio E

    2013-06-12

    Maintenance of genomic stability during eukaryotic cell division relies on the spindle assembly checkpoint (SAC) that prevents mitotic exit until all chromosomes are properly attached to the spindle. Polo is a mitotic kinase proposed to be involved in SAC function, but its role has remained elusive. We demonstrate that Polo and Aurora B functional interdependency comprises a positive feedback loop that promotes Mps1 kinetochore localization and activity. Expression of constitutively active Polo restores normal Mps1 kinetochore levels even after Aurora B inhibition, highlighting a role for Polo in Mps1 recruitment to unattached kinetochores downstream of Aurora B. We also show that Mps1 kinetochore localization is required for BubR1 hyperphosphorylation and formation of the 3F3/2 phosphoepitope. This is essential to allow recruitment of Cdc20 to unattached kinetochores and the assembly of anaphase-promoting complex/cyclosome-inhibitory complexes to levels that ensure long-term SAC activity. We propose a model in which Polo controls Mps1-dependent BubR1 phosphorylation to promote Cdc20 kinetochore recruitment and sustained SAC function.

  19. bullwinkle and shark regulate dorsal-appendage morphogenesis in Drosophila oogenesis.

    Science.gov (United States)

    Tran, David H; Berg, Celeste A

    2003-12-01

    bullwinkle (bwk) regulates embryonic anteroposterior patterning and, through a novel germline-to-soma signal, morphogenesis of the eggshell dorsal appendages. We screened for dominant modifiers of the bullwinkle mooseantler eggshell phenotype and identified shark, which encodes an SH2-domain, ankyrin-repeat tyrosine kinase. At the onset of dorsal-appendage formation, shark is expressed in a punctate pattern in the squamous stretch cells overlying the nurse cells. Confocal microscopy with cell-type-specific markers demonstrates that the stretch cells act as a substrate for the migrating dorsal-appendage-forming cells and extend cellular projections towards them. Mosaic analyses reveal that shark is required in follicle cells for cell migration and chorion deposition. Proper shark RNA expression in the stretch cells requires bwk activity, while restoration of shark expression in the stretch cells suppresses the bwk dorsal-appendage phenotype. These results suggest that shark plays an important downstream role in the bwk-signaling pathway. Candidate testing implicates Src42A in a similar role, suggesting conservation with a vertebrate signaling pathway involving non-receptor tyrosine kinases.

  20. Negative regulation of P element excision by the somatic product and terminal sequences of P in drosophila melanogaster

    Science.gov (United States)

    A transient in vivo P element excision assay was used to test the regulatory properties of putative repressor-encoding plasmids in Drosophila melanogaster embryos. The somatic expression of an unmodified transposase transcription unit under the control of a heat shock gene promoter (phsn) effectivel...

  1. Viability, longevity, and egg production of Drosophila melanogaster are regulated by the miR-282 microDNA

    Czech Academy of Sciences Publication Activity Database

    Vilmos, P.; Bujna, Á.; Szuperák, M.; Havelda, Z.; Várallyay, É.; Szabad, J.; Kučerová, Lucie; Somogyi, K.; Kristó, I.; Lukácsovich, T.; Jankovics, F.; Henn, L.; Erdélyi, M.

    2013-01-01

    Roč. 195, č. 2 (2013), s. 469-480 ISSN 0016-6731 Grant - others:Hungarian National Science Foundation(HU) NK84121; Hungarian National Science Foundation(HU) K108538 Institutional support: RVO:60077344 Keywords : Drosophila melanogaster Subject RIV: ED - Physiology Impact factor: 4.389, year: 2012

  2. Combinatorial action of Grainyhead, Extradenticle and Notch in regulating Hox mediated apoptosis in Drosophila larval CNS.

    Science.gov (United States)

    Khandelwal, Risha; Sipani, Rashmi; Govinda Rajan, Sriivatsan; Kumar, Raviranjan; Joshi, Rohit

    2017-10-01

    Hox mediated neuroblast apoptosis is a prevalent way to pattern larval central nervous system (CNS) by different Hox genes, but the mechanism of this apoptosis is not understood. Our studies with Abdominal-A (Abd-A) mediated larval neuroblast (pNB) apoptosis suggests that AbdA, its cofactor Extradenticle (Exd), a helix-loop-helix transcription factor Grainyhead (Grh), and Notch signaling transcriptionally contribute to expression of RHG family of apoptotic genes. We find that Grh, AbdA, and Exd function together at multiple motifs on the apoptotic enhancer. In vivo mutagenesis of these motifs suggest that they are important for the maintenance of the activity of the enhancer rather than its initiation. We also find that Exd function is independent of its known partner homothorax in this apoptosis. We extend some of our findings to Deformed expressing region of sub-esophageal ganglia where pNBs undergo a similar Hox dependent apoptosis. We propose a mechanism where common players like Exd-Grh-Notch work with different Hox genes through region specific enhancers to pattern respective segments of larval central nervous system.

  3. Aging and Autophagic Function Influences the Progressive Decline of Adult Drosophila Behaviors.

    Directory of Open Access Journals (Sweden)

    Eric P Ratliff

    Full Text Available Multiple neurological disorders are characterized by the abnormal accumulation of protein aggregates and the progressive impairment of complex behaviors. Our Drosophila studies demonstrate that middle-aged wild-type flies (WT, ~4-weeks exhibit a marked accumulation of neural aggregates that is commensurate with the decline of the autophagy pathway. However, enhancing autophagy via neuronal over-expression of Atg8a (Atg8a-OE reduces the age-dependent accumulation of aggregates. Here we assess basal locomotor activity profiles for single- and group-housed male and female WT flies and observed that only modest behavioral changes occurred by 4-weeks of age, with the noted exception of group-housed male flies. Male flies in same-sex social groups exhibit a progressive increase in nighttime activity. Infrared videos show aged group-housed males (4-weeks are engaged in extensive bouts of courtship during periods of darkness, which is partly repressed during lighted conditions. Together, these nighttime courtship behaviors were nearly absent in young WT flies and aged Atg8a-OE flies. Previous studies have indicated a regulatory role for olfaction in male courtship partner choice. Coincidently, the mRNA expression profiles of several olfactory genes decline with age in WT flies; however, they are maintained in age-matched Atg8a-OE flies. Together, these results suggest that middle-aged male flies develop impairments in olfaction, which could contribute to the dysregulation of courtship behaviors during dark time periods. Combined, our results demonstrate that as Drosophila age, they develop early behavior defects that are coordinate with protein aggregate accumulation in the nervous system. In addition, the nighttime activity behavior is preserved when neuronal autophagy is maintained (Atg8a-OE flies. Thus, environmental or genetic factors that modify autophagic capacity could have a positive impact on neuronal aging and complex behaviors.

  4. Drosophila Clock Is Required in Brain Pacemaker Neurons to Prevent Premature Locomotor Aging Independently of Its Circadian Function.

    Directory of Open Access Journals (Sweden)

    Alexandra Vaccaro

    2017-01-01

    Full Text Available Circadian clocks control many self-sustained rhythms in physiology and behavior with approximately 24-hour periodicity. In many organisms, oxidative stress and aging negatively impact the circadian system and sleep. Conversely, loss of the clock decreases resistance to oxidative stress, and may reduce lifespan and speed up brain aging and neurodegeneration. Here we examined the effects of clock disruptions on locomotor aging and longevity in Drosophila. We found that lifespan was similarly reduced in three arrhythmic mutants (ClkAR, cyc0 and tim0 and in wild-type flies under constant light, which stops the clock. In contrast, ClkAR mutants showed significantly faster age-related locomotor deficits (as monitored by startle-induced climbing than cyc0 and tim0, or than control flies under constant light. Reactive oxygen species accumulated more with age in ClkAR mutant brains, but this did not appear to contribute to the accelerated locomotor decline of the mutant. Clk, but not Cyc, inactivation by RNA interference in the pigment-dispersing factor (PDF-expressing central pacemaker neurons led to similar loss of climbing performance as ClkAR. Conversely, restoring Clk function in these cells was sufficient to rescue the ClkAR locomotor phenotype, independently of behavioral rhythmicity. Accelerated locomotor decline of the ClkAR mutant required expression of the PDF receptor and correlated to an apparent loss of dopaminergic neurons in the posterior protocerebral lateral 1 (PPL1 clusters. This neuronal loss was rescued when the ClkAR mutation was placed in an apoptosis-deficient background. Impairing dopamine synthesis in a single pair of PPL1 neurons that innervate the mushroom bodies accelerated locomotor decline in otherwise wild-type flies. Our results therefore reveal a novel circadian-independent requirement for Clk in brain circadian neurons to maintain a subset of dopaminergic cells and avoid premature locomotor aging in Drosophila.

  5. Regulation of satellite cell function in sarcopenia

    Directory of Open Access Journals (Sweden)

    Stephen E Alway

    2014-09-01

    Full Text Available The mechanisms contributing to sarcopenia include reduced satellite cell (myogenic stem cell function that is impacted by the environment (niche of these cells. Satellite cell function is affected by oxidative stress, which is elevated in aged muscles, and this along with changes in largely unknown systemic factors, likely contribute to the manner in which satellite cells respond to stressors such as exercise, disuse or rehabilitation in sarcopenic muscles. Nutritional intervention provides one therapeutic strategy to improve the satellite cell niche and systemic factors, with the goal of improving satellite cell function in aging muscles. Although many elderly persons consume various nutraceuticals with the hope of improving health, most of these compounds have not been thoroughly tested, and the impacts that they might have on sarcopenia, and satellite cell function are not clear. This review discusses data pertaining to the satellite cell responses and function in aging skeletal muscle, and the impact that three compounds: resveratrol, green tea catechins and β-Hydroxy-β-methylbutyrate have on regulating satellite cell function and therefore contributing to reducing sarcopenia or improving muscle mass after disuse in aging. The data suggest that these nutraceutical compounds improve satellite cell function during rehabilitative loading in animal models of aging after disuse (i.e., muscle regeneration. While these compounds have not been rigorously tested in humans, the data from animal models of aging provide a strong basis for conducting additional focused work to determine if these or other nutraceuticals can offset the muscle losses, or improve regeneration in sarcopenic muscles of older humans via improving satellite cell function.

  6. Regulation of Satellite Cell Function in Sarcopenia

    Science.gov (United States)

    Alway, Stephen E.; Myers, Matthew J.; Mohamed, Junaith S.

    2014-01-01

    The mechanisms contributing to sarcopenia include reduced satellite cell (myogenic stem cell) function that is impacted by the environment (niche) of these cells. Satellite cell function is affected by oxidative stress, which is elevated in aged muscles, and this along with changes in largely unknown systemic factors, likely contribute to the manner in which satellite cells respond to stressors such as exercise, disuse, or rehabilitation in sarcopenic muscles. Nutritional intervention provides one therapeutic strategy to improve the satellite cell niche and systemic factors, with the goal of improving satellite cell function in aging muscles. Although many elderly persons consume various nutraceuticals with the hope of improving health, most of these compounds have not been thoroughly tested, and the impacts that they might have on sarcopenia and satellite cell function are not clear. This review discusses data pertaining to the satellite cell responses and function in aging skeletal muscle, and the impact that three compounds: resveratrol, green tea catechins, and β-Hydroxy-β-methylbutyrate have on regulating satellite cell function and therefore contributing to reducing sarcopenia or improving muscle mass after disuse in aging. The data suggest that these nutraceutical compounds improve satellite cell function during rehabilitative loading in animal models of aging after disuse (i.e., muscle regeneration). While these compounds have not been rigorously tested in humans, the data from animal models of aging provide a strong basis for conducting additional focused work to determine if these or other nutraceuticals can offset the muscle losses, or improve regeneration in sarcopenic muscles of older humans via improving satellite cell function. PMID:25295003

  7. Genome-wide miRNA screening reveals miR-310 family members negatively regulate the immune response in Drosophila melanogaster via co-targeting Drosomycin.

    Science.gov (United States)

    Li, Yao; Li, Shengjie; Li, Ruimin; Xu, Jiao; Jin, Ping; Chen, Liming; Ma, Fei

    2017-03-01

    Although innate immunity mediated by Toll signaling has been extensively studied in Drosophila melanogaster, the role of miRNAs in regulating the Toll-mediated immune response remains largely unknown. In this study, following Gram-positive bacterial challenge, we identified 93 differentially expressed miRNAs via genome-wide miRNA screening. These miRNAs were regarded as immune response related (IRR). Eight miRNAs were confirmed to be involved in the Toll-mediated immune response upon Gram-positive bacterial infection through genetic screening of 41 UAS-miRNA lines covering 60 miRNAs of the 93 IRR miRNAs. Interestingly, four out of these eight miRNAs, miR-310, miR-311, miR-312 and miR-313, are clustered miRNAs and belong to the miR-310 family. These miR-310 family members were shown to target and regulate the expression of Drosomycin, an antimicrobial peptide produced by Toll signaling. Taken together, our study implies important regulatory roles of miRNAs in the Toll-mediated innate immune response of Drosophila upon Gram-positive bacterial infection. Copyright © 2016 Elsevier Ltd. All rights reserved.

  8. Investigating the Regulation of Estrogen Receptor-Mediated Transcription

    National Research Council Canada - National Science Library

    Thackray, Varykina

    2002-01-01

    ...-mediated regulation of specific target genes are still lacking. We have developed an estrogen responsive system in the fruit fly, Drosophila melanogaster in order to explore the functional interactions between ER and other cellular proteins...

  9. Investigating the Regulation of Estrogen Receptor-Mediated Transcription

    National Research Council Canada - National Science Library

    Thackray, Varykina

    2001-01-01

    ...-mediated regulation of specific target genes are still lacking. We have developed an estrogen responsive system in the fruit fly, Drosophila melanogaster in order to explore the functional interactions between ER and other cellular proteins...

  10. Autophagy in Drosophila: From Historical Studies to Current Knowledge

    Science.gov (United States)

    Mulakkal, Nitha C.; Nagy, Peter; Takats, Szabolcs; Tusco, Radu; Juhász, Gábor; Nezis, Ioannis P.

    2014-01-01

    The discovery of evolutionarily conserved Atg genes required for autophagy in yeast truly revolutionized this research field and made it possible to carry out functional studies on model organisms. Insects including Drosophila are classical and still popular models to study autophagy, starting from the 1960s. This review aims to summarize past achievements and our current knowledge about the role and regulation of autophagy in Drosophila, with an outlook to yeast and mammals. The basic mechanisms of autophagy in fruit fly cells appear to be quite similar to other eukaryotes, and the role that this lysosomal self-degradation process plays in Drosophila models of various diseases already made it possible to recognize certain aspects of human pathologies. Future studies in this complete animal hold great promise for the better understanding of such processes and may also help finding new research avenues for the treatment of disorders with misregulated autophagy. PMID:24949430

  11. Autophagy in Drosophila: From Historical Studies to Current Knowledge

    Directory of Open Access Journals (Sweden)

    Nitha C. Mulakkal

    2014-01-01

    Full Text Available The discovery of evolutionarily conserved Atg genes required for autophagy in yeast truly revolutionized this research field and made it possible to carry out functional studies on model organisms. Insects including Drosophila are classical and still popular models to study autophagy, starting from the 1960s. This review aims to summarize past achievements and our current knowledge about the role and regulation of autophagy in Drosophila, with an outlook to yeast and mammals. The basic mechanisms of autophagy in fruit fly cells appear to be quite similar to other eukaryotes, and the role that this lysosomal self-degradation process plays in Drosophila models of various diseases already made it possible to recognize certain aspects of human pathologies. Future studies in this complete animal hold great promise for the better understanding of such processes and may also help finding new research avenues for the treatment of disorders with misregulated autophagy.

  12. The Drosophila gene CheB42a is a novel modifier of Deg/ENaC channel function.

    Directory of Open Access Journals (Sweden)

    Yehuda Ben-Shahar

    2010-02-01

    Full Text Available Degenerin/epithelial Na(+ channels (DEG/ENaC represent a diverse family of voltage-insensitive cation channels whose functions include Na(+ transport across epithelia, mechanosensation, nociception, salt sensing, modification of neurotransmission, and detecting the neurotransmitter FMRFamide. We previously showed that the Drosophila melanogaster Deg/ENaC gene lounge lizard (llz is co-transcribed in an operon-like locus with another gene of unknown function, CheB42a. Because operons often encode proteins in the same biochemical or physiological pathway, we hypothesized that CHEB42A and LLZ might function together. Consistent with this hypothesis, we found both genes expressed in cells previously implicated in sensory functions during male courtship. Furthermore, when coexpressed, LLZ coprecipitated with CHEB42A, suggesting that the two proteins form a complex. Although LLZ expressed either alone or with CHEB42A did not generate ion channel currents, CHEB42A increased current amplitude of another DEG/ENaC protein whose ligand (protons is known, acid-sensing ion channel 1a (ASIC1a. We also found that CHEB42A was cleaved to generate a secreted protein, suggesting that CHEB42A may play an important role in the extracellular space. These data suggest that CHEB42A is a modulatory subunit for sensory-related Deg/ENaC signaling. These results are consistent with operon-like transcription of CheB42a and llz and explain the similar contributions of these genes to courtship behavior.

  13. The octopamine receptor Octβ2R regulates ovulation in Drosophila melanogaster.

    Directory of Open Access Journals (Sweden)

    Junghwa Lim

    Full Text Available Oviposition is induced upon mating in most insects. Ovulation is a primary step in oviposition, representing an important target to control insect pests and vectors, but limited information is available on the underlying mechanism. Here we report that the beta adrenergic-like octopamine receptor Octβ2R serves as a key signaling molecule for ovulation and recruits protein kinase A and Ca(2+/calmodulin-sensitive kinase II as downstream effectors for this activity. We found that the octβ2r homozygous mutant females are sterile. They displayed normal courtship, copulation, sperm storage and post-mating rejection behavior but were unable to lay eggs. We have previously shown that octopamine neurons in the abdominal ganglion innervate the oviduct epithelium. Consistently, restored expression of Octβ2R in oviduct epithelial cells was sufficient to reinstate ovulation and full fecundity in the octβ2r mutant females, demonstrating that the oviduct epithelium is a major site of Octβ2R's function in oviposition. We also found that overexpression of the protein kinase A catalytic subunit or Ca(2+/calmodulin-sensitive protein kinase II led to partial rescue of octβ2r's sterility. This suggests that Octβ2R activates cAMP as well as additional effectors including Ca(2+/calmodulin-sensitive protein kinase II for oviposition. All three known beta adrenergic-like octopamine receptors stimulate cAMP production in vitro. Octβ1R, when ectopically expressed in the octβ2r's oviduct epithelium, fully reinstated ovulation and fecundity. Ectopically expressed Octβ3R, on the other hand, partly restored ovulation and fecundity while OAMB-K3 and OAMB-AS that increase Ca(2+ levels yielded partial rescue of ovulation but not fecundity deficit. These observations suggest that Octβ2R have distinct signaling capacities in vivo and activate multiple signaling pathways to induce egg laying. The findings reported here narrow the knowledge gap and offer insight into novel

  14. The dopaminergic system in the aging brain of Drosophila

    Directory of Open Access Journals (Sweden)

    Katherine E White

    2010-12-01

    Full Text Available Drosophila models of Parkinson’s disease are characterised by two principal phenotypes: the specific loss of dopaminergic neurons in the aging brain and defects in motor behavior. However, an age-related analysis of these baseline parameters in wildtype Drosophila is lacking. Here we analysed the dopaminergic system and motor behavior in aging Drosophila. Dopaminergic neurons in the adult brain can be grouped into bilateral symmetric clusters, each comprising a stereotypical number of cells. Analysis of TH>mCD8::GFP and cell type-specific MARCM clones revealed that dopaminergic neurons show cluster-specific, stereotypical projection patterns with terminal arborization in target regions that represent distinct functional areas of the adult brain. Target areas include the mushroom bodies, involved in memory formation and motivation, and the central complex, involved in the control of motor behavior, indicating that similar to the mammalian brain, dopaminergic neurons in the fly brain are involved in the regulation of specific behaviors. Behavioral analysis revealed that Drosophila show an age-related decline in startle-induced locomotion and negative geotaxis. Motion tracking however, revealed that walking activity and exploration behavior, but not centrophobism increase at late stages of life. Analysis of TH>Dcr2, mCD8::GFP revealed a specific effect of Dcr2 expression on walking activity but not on exploratory or centrophobic behavior, indicating that the siRNA pathway may modulate distinct dopaminergic behaviors in Drosophila. Moreover, dopaminergic neurons were maintained between early- and late life, as quantified by TH>mCD8::GFP and anti-TH labelling, indicating that adult onset, age-related degeneration of dopaminergic neurons does not occur in the aging brain of Drosophila. Taken together, our data establish baseline parameters in Drosophila for the study of Parkinson’s disease as well as other disorders affecting dopaminergic neurons

  15. Gene expression profiling of brakeless mutant Drosophila embryos.

    Science.gov (United States)

    Crona, Filip; Singla, Bhumica; Mannervik, Mattias

    2015-12-01

    The transcriptional co-regulator Brakeless performs many important functions during Drosophila development, but few target genes have been identified. Here we use Affymetrix microarrays to identify Brakeless-regulated genes in 2-4 h old Drosophila embryos. Robust multi-array analysis (RMA) and statistical tests revealed 240 genes that changed their expression more than 1.5 fold. We find that up- and down-regulated genes fall into distinct gene ontology categories. In our associated study [2] we demonstrate that both up- and down-regulated genes can be direct Brakeless targets. Our results indicate that the co-repressor and co-activator activities of Brakeless may result in distinct biological responses. The microarray data complies with MIAME guidelines and is deposited in GEO under accession number GSE60048.

  16. Short neuropeptide F acts as a functional neuromodulator for olfactory memory in Kenyon cells of Drosophila mushroom bodies.

    Science.gov (United States)

    Knapek, Stephan; Kahsai, Lily; Winther, Asa M E; Tanimoto, Hiromu; Nässel, Dick R

    2013-03-20

    In insects, many complex behaviors, including olfactory memory, are controlled by a paired brain structure, the so-called mushroom bodies (MB). In Drosophila, the development, neuroanatomy, and function of intrinsic neurons of the MB, the Kenyon cells, have been well characterized. Until now, several potential neurotransmitters or neuromodulators of Kenyon cells have been anatomically identified. However, whether these neuroactive substances of the Kenyon cells are functional has not been clarified yet. Here we show that a neuropeptide precursor gene encoding four types of short neuropeptide F (sNPF) is required in the Kenyon cells for appetitive olfactory memory. We found that activation of Kenyon cells by expressing a thermosensitive cation channel (dTrpA1) leads to a decrease in sNPF immunoreactivity in the MB lobes. Targeted expression of RNA interference against the sNPF precursor in Kenyon cells results in a highly significant knockdown of sNPF levels. This knockdown of sNPF in the Kenyon cells impairs sugar-rewarded olfactory memory. This impairment is not due to a defect in the reflexive sugar preference or odor response. Consistently, knockdown of sNPF receptors outside the MB causes deficits in appetitive memory. Altogether, these results suggest that sNPF is a functional neuromodulator released by Kenyon cells.

  17. Sex-specific influences of mtDNA mitotype and diet on mitochondrial functions and physiological traits in Drosophila melanogaster.

    Directory of Open Access Journals (Sweden)

    Wen C Aw

    Full Text Available Here we determine the sex-specific influence of mtDNA type (mitotype and diet on mitochondrial functions and physiology in two Drosophila melanogaster lines. In many species, males and females differ in aspects of their energy production. These sex-specific influences may be caused by differences in evolutionary history and physiological functions. We predicted the influence of mtDNA mutations should be stronger in males than females as a result of the organelle's maternal mode of inheritance in the majority of metazoans. In contrast, we predicted the influence of diet would be greater in females due to higher metabolic flexibility. We included four diets that differed in their protein: carbohydrate (P:C ratios as they are the two-major energy-yielding macronutrients in the fly diet. We assayed four mitochondrial function traits (Complex I oxidative phosphorylation, reactive oxygen species production, superoxide dismutase activity, and mtDNA copy number and four physiological traits (fecundity, longevity, lipid content, and starvation resistance. Traits were assayed at 11 d and 25 d of age. Consistent with predictions we observe that the mitotype influenced males more than females supporting the hypothesis of a sex-specific selective sieve in the mitochondrial genome caused by the maternal inheritance of mitochondria. Also, consistent with predictions, we found that the diet influenced females more than males.

  18. BMAA neurotoxicity in Drosophila.

    Science.gov (United States)

    Zhou, Xianchong; Escala, Wilfredo; Papapetropoulos, Spyridon; Bradley, Walter G; Zhai, R Grace

    2009-01-01

    We report the establishment of an in vivo model using the fruit fly Drosophila melanogaster to investigate the toxic effects of L-BMAA. We found that dietary intake of BMAA reduced the lifespan as well as the neurological functions of flies. Furthermore, we have developed an HPLC method to reliably detect both free and protein-bound BMAA in fly tissue extracts.

  19. Cancer in Drosophila

    DEFF Research Database (Denmark)

    Herranz, Héctor; Eichenlaub, Teresa; Cohen, Stephen M

    2016-01-01

    Cancer genomics has greatly increased our understanding of the complexity of the genetic and epigenetic changes found in human tumors. Understanding the functional relationships among these elements calls for the use of flexible genetic models. We discuss the use of Drosophila models to study...

  20. Identified peptidergic neurons in the Drosophila brain regulate insulin-producing cells, stress responses and metabolism by coexpressed short neuropeptide F and corazonin.

    Science.gov (United States)

    Kapan, Neval; Lushchak, Oleh V; Luo, Jiangnan; Nässel, Dick R

    2012-12-01

    Insulin/IGF-like signaling regulates the development, growth, fecundity, metabolic homeostasis, stress resistance and lifespan in worms, flies and mammals. Eight insulin-like peptides (DILP1-8) are found in Drosophila. Three of these (DILP2, 3 and 5) are produced by a set of median neurosecretory cells (insulin-producing cells, IPCs) in the brain. Activity in the IPCs of adult flies is regulated by glucose and several neurotransmitters and neuropeptides. One of these, short neuropeptide F (sNPF), regulates food intake, growth and Dilp transcript levels in IPCs via the sNPF receptor (sNPFR1) expressed on IPCs. Here we identify a set of brain neurons that utilizes sNPF to activate the IPCs. These sNPF-expressing neurons (dorsal lateral peptidergic neurons, DLPs) also produce the neuropeptide corazonin (CRZ) and have axon terminations impinging on IPCs. Knockdown of either sNPF or CRZ in DLPs extends survival in flies exposed to starvation and alters carbohydrate and lipid metabolism. Expression of sNPF in DLPs in the sNPF mutant background is sufficient to rescue wild-type metabolism and response to starvation. Since CRZ receptor RNAi in IPCs affects starvation resistance and metabolism, similar to peptide knockdown in DLPs, it is likely that also CRZ targets the IPCs. Knockdown of sNPF, but not CRZ in DLPs decreases transcription of Dilp2 and 5 in the brain, suggesting different mechanisms of action on IPCs of the two co-released peptides. Our findings indicate that sNPF and CRZ co-released from a small set of neurons regulate IPCs, stress resistance and metabolism in adult Drosophila.

  1. Identification of Novel Ovarian Cancer Oncogenes that Function by Regulating Exosome Function

    Science.gov (United States)

    2017-09-01

    Novel Ovarian Cancer Oncogenes that Function by Regulating Exosome Function September 2017 x 1Sep2016...31Aug2017 Email: mbirrer@partners.org 6 Identification of Novel Ovarian Cancer Oncogenes that Function by Regulating Exosome Function xx

  2. Chemical mutagens, transposons, and transgenes to interrogate gene function in Drosophila melanogaster.

    Science.gov (United States)

    Venken, Koen J T; Bellen, Hugo J

    2014-06-15

    The study of genetics, genes, and chromosomal inheritance was initiated by Thomas Morgan in 1910, when the first visible mutations were identified in fruit flies. The field expanded upon the work initiated by Herman Muller in 1926 when he used X-rays to develop the first balancer chromosomes. Today, balancers are still invaluable to maintain mutations and transgenes but the arsenal of tools has expanded vastly and numerous new methods have been developed, many relying on the availability of the genome sequence and transposable elements. Forward genetic screens based on chemical mutagenesis or transposable elements have resulted in the unbiased identification of many novel players involved in processes probed by specific phenotypic assays. Reverse genetic approaches have relied on the availability of a carefully selected set of transposon insertions spread throughout the genome to allow the manipulation of the region in the vicinity of each insertion. Lastly, the ability to transform Drosophila with single copy transgenes using transposons or site-specific integration using the ΦC31 integrase has allowed numerous manipulations, including the ability to create and integrate genomic rescue constructs, generate duplications, RNAi knock-out technology, binary expression systems like the GAL4/UAS system as well as other methods. Here, we will discuss the most useful methodologies to interrogate the fruit fly genome in vivo focusing on chemical mutagenesis, transposons and transgenes. Genome engineering approaches based on nucleases and RNAi technology are discussed in following chapters. Copyright © 2014 Elsevier Inc. All rights reserved.

  3. Peptide regulators of peripheral taste function.

    Science.gov (United States)

    Dotson, Cedrick D; Geraedts, Maartje C P; Munger, Steven D

    2013-03-01

    The peripheral sensory organ of the gustatory system, the taste bud, contains a heterogeneous collection of sensory cells. These taste cells can differ in the stimuli to which they respond and the receptors and other signaling molecules they employ to transduce and encode those stimuli. This molecular diversity extends to the expression of a varied repertoire of bioactive peptides that appear to play important functional roles in signaling taste information between the taste cells and afferent sensory nerves and/or in processing sensory signals within the taste bud itself. Here, we review studies that examine the expression of bioactive peptides in the taste bud and the impact of those peptides on taste functions. Many of these peptides produced in taste buds are known to affect appetite, satiety or metabolism through their actions in the brain, pancreas and other organs, suggesting a functional link between the gustatory system and the neural and endocrine systems that regulate feeding and nutrient utilization. Copyright © 2013 Elsevier Ltd. All rights reserved.

  4. Copper Sensing Function of Drosophila Metal-Responsive Transcription Factor-1 Is Mediated By a Tetranuclear Cu(I) Cluster

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    Chen, X.; Hua, H.; Balamurugan, K.; Kong, X.; Zhang, L.; George, G.N.; Georgiev, O.; Schaffner, W.; Giedroc, D.P.

    2009-05-12

    Drosophila melanogaster MTF-1 (dMTF-1) is a copper-responsive transcriptional activator that mediates resistance to Cu, as well as Zn and Cd. Here, we characterize a novel cysteine-rich domain which is crucial for sensing excess intracellular copper by dMTF-1. Transgenic flies expressing mutant dMTF-1 containing alanine substitutions of two, four or six cysteine residues within the sequence {sup 547}CNCTNCKCDQTKSCHGGDC{sup 565} are significantly or completely impaired in their ability to protect flies from copper toxicity and fail to up-regulate MtnA (metallothionein) expression in response to excess Cu. In contrast, these flies exhibit wild-type survival in response to copper deprivation thus revealing that the cysteine cluster domain is required only for sensing Cu load by dMTF-1. Parallel studies show that the isolated cysteine cluster domain is required to protect a copper-sensitive S. cerevisiae ace1 strain from copper toxicity. Cu(I) ligation by a Cys-rich domain peptide fragment drives the cooperative assembly of a polydentate [Cu{sub 4}-S{sub 6}] cage structure, characterized by a core of trigonally S{sub 3} coordinated Cu(I) ions bound by bridging thiolate ligands. While reminiscent of Cu{sub 4}-L{sub 6} (L = ligand) tetranuclear clusters in copper regulatory transcription factors of yeast, the absence of significant sequence homology is consistent with convergent evolution of a sensing strategy particularly well suited for Cu(I).

  5. The Drosophila melanogaster DmCK2beta transcription unit encodes for functionally non-redundant protein isoforms.

    Science.gov (United States)

    Jauch, Eike; Wecklein, Heike; Stark, Felix; Jauch, Mandy; Raabe, Thomas

    2006-06-07

    Genes encoding for the two evolutionary highly conserved subunits of a heterotetrameric protein kinase CK2 holoenzyme are present in all examined eukaryotic genomes. Depending on the organism, multiple transcription units encoding for a catalytically active CK2alpha subunit and/or a regulatory CK2beta subunit may exist. The phosphotransferase activity of members of the protein kinase CK2alpha family is thought to be independent of second messengers but is modulated by interaction with CK2beta-like proteins. In the genome of Drosophila melanogaster, one gene encoding for a CK2alpha subunit and three genes encoding for CK2beta-like proteins are present. The X-linked DmCK2beta transcription unit encodes for several CK2beta protein isoforms due to alternative splicing of its primary transcript. We addressed the question whether CK2beta-like proteins are redundant in function. Our in vivo experiments show that variations of the very C-terminal tail of CK2beta isoforms encoded by the X-linked DmCK2beta transcription unit influence their functional properties. In addition, we find that CK2beta-like proteins encoded by the autosomal D. melanogaster genes CK2betates and CK2beta' cannot fully substitute for a loss of CK2beta isoforms encoded by DmCK2beta.

  6. TRPA1 channels in Drosophila and honey bee ectoparasitic mites share heat sensitivity and temperature-related physiological functions

    Directory of Open Access Journals (Sweden)

    Guangda Peng

    2016-10-01

    Full Text Available The transient receptor potential cation channel, subfamily A, member 1 (TRPA1 is conserved between many arthropods, and in some has been shown to function as a chemosensor for noxious compounds. Activation of arthropod TRPA1 channels by temperature fluctuations has been tested in only a few insect species, and all of them were shown to be activated by heat. The recent identification of chemosensitive TRPA1 channels from two honey bee ectoparasitic mite species (VdTRPA1 and TmTRPA1 have provided an opportunity to study the temperature-dependent activation and the temperature-associated physiological functions of TRPA1 channels in non-insect arthropods. We found that both mite TRPA1 channels are heat sensitive and capable of rescuing the temperature-related behavioral defects of a Drosophila melanogaster trpA1 mutant. These results suggest that heat-sensitivity of TRPA1 could be conserved between many arthropods despite its amino acid sequence diversity. Nevertheless, the ankyrin repeats (ARs 6 and 7 are well-conserved between six heat-sensitive arthropod TRPA1 channels and have critical roles for the heat activation of VdTRPA1.

  7. Drosophila Protein Kinase CK2: Genetics, Regulatory Complexity and Emerging Roles during Development

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    Mohna Bandyopadhyay

    2016-12-01

    Full Text Available CK2 is a Ser/Thr protein kinase that is highly conserved amongst all eukaryotes. It is a well-known oncogenic kinase that regulates vital cell autonomous functions and animal development. Genetic studies in the fruit fly Drosophila are providing unique insights into the roles of CK2 in cell signaling, embryogenesis, organogenesis, neurogenesis, and the circadian clock, and are revealing hitherto unknown complexities in CK2 functions and regulation. Here, we review Drosophila CK2 with respect to its structure, subunit diversity, potential mechanisms of regulation, developmental abnormalities linked to mutations in the gene encoding CK2 subunits, and emerging roles in multiple aspects of eye development. We examine the Drosophila CK2 “interaction map” and the eye-specific “transcriptome” databases, which raise the prospect that this protein kinase has many additional targets in the developing eye. We discuss the possibility that CK2 functions during early retinal neurogenesis in Drosophila and mammals bear greater similarity than has been recognized, and that this conservation may extend to other developmental programs. Together, these studies underscore the immense power of the Drosophila model organism to provide new insights and avenues to further investigate developmentally relevant targets of this protein kinase.

  8. Conservation of cardiac L-type Ca2+ channels and their regulation in Drosophila: A novel genetically-pliable channelopathic model.

    Science.gov (United States)

    Limpitikul, Worawan B; Viswanathan, Meera C; O'Rourke, Brian; Yue, David T; Cammarato, Anthony

    2018-04-21

    Dysregulation of L-type Ca 2+ channels (LTCCs) underlies numerous cardiac pathologies. Understanding their modulation with high fidelity relies on investigating LTCCs in their native environment with intact interacting proteins. Such studies benefit from genetic manipulation of endogenous channels in cardiomyocytes, which often proves cumbersome in mammalian models. Drosophila melanogaster, however, offers a potentially efficient alternative as it possesses a relatively simple heart, is genetically pliable, and expresses well-conserved genes. Fluorescence in situ hybridization confirmed an abundance of Ca-α1D and Ca-α1T mRNA in fly myocardium, which encode subunits that specify hetero-oligomeric channels homologous to mammalian LTCCs and T-type Ca 2+ channels, respectively. Cardiac-specific knockdown of Ca-α1D via interfering RNA abolished cardiac contraction, suggesting Ca-α1D (i.e. A1D) represents the primary functioning Ca 2+ channel in Drosophila hearts. Moreover, we successfully isolated viable single cardiomyocytes and recorded Ca 2+ currents via patch clamping, a feat never before accomplished with the fly model. The profile of Ca 2+ currents recorded in individual cells when Ca 2+ channels were hypomorphic, absent, or under selective LTCC blockage by nifedipine, additionally confirmed the predominance of A1D current across all activation voltages. T-type current, activated at more negative voltages, was also detected. Lastly, A1D channels displayed Ca 2+ -dependent inactivation, a critical negative feedback mechanism of LTCCs, and the current through them was augmented by forskolin, an activator of the protein kinase A pathway. In sum, the Drosophila heart possesses a conserved compendium of Ca 2+ channels, suggesting that the fly may serve as a robust and effective platform for studying cardiac channelopathies. Copyright © 2018 Elsevier Ltd. All rights reserved.

  9. FIG4 regulates lysosome membrane homeostasis independent of phosphatase function.

    Science.gov (United States)

    Bharadwaj, Rajnish; Cunningham, Kathleen M; Zhang, Ke; Lloyd, Thomas E

    2016-02-15

    FIG4 is a phosphoinositide phosphatase that is mutated in several diseases including Charcot-Marie-Tooth Disease 4J (CMT4J) and Yunis-Varon syndrome (YVS). To investigate the mechanism of disease pathogenesis, we generated Drosophila models of FIG4-related diseases. Fig4 null mutant animals are viable but exhibit marked enlargement of the lysosomal compartment in muscle cells and neurons, accompanied by an age-related decline in flight ability. Transgenic animals expressing Drosophila Fig4 missense mutations corresponding to human pathogenic mutations can partially rescue lysosomal expansion phenotypes, consistent with these mutations causing decreased FIG4 function. Interestingly, Fig4 mutations predicted to inactivate FIG4 phosphatase activity rescue lysosome expansion phenotypes, and mutations in the phosphoinositide (3) phosphate kinase Fab1 that performs the reverse enzymatic reaction also causes a lysosome expansion phenotype. Since FIG4 and FAB1 are present together in the same biochemical complex, these data are consistent with a model in which FIG4 serves a phosphatase-independent biosynthetic function that is essential for lysosomal membrane homeostasis. Lysosomal phenotypes are suppressed by genetic inhibition of Rab7 or the HOPS complex, demonstrating that FIG4 functions after endosome-to-lysosome fusion. Furthermore, disruption of the retromer complex, implicated in recycling from the lysosome to Golgi, does not lead to similar phenotypes as Fig4, suggesting that the lysosomal defects are not due to compromised retromer-mediated recycling of endolysosomal membranes. These data show that FIG4 plays a critical noncatalytic function in maintaining lysosomal membrane homeostasis, and that this function is disrupted by mutations that cause CMT4J and YVS. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  10. Heterochromatin protein 1 (HP1a positively regulates euchromatic gene expression through RNA transcript association and interaction with hnRNPs in Drosophila.

    Directory of Open Access Journals (Sweden)

    Lucia Piacentini

    2009-10-01

    Full Text Available Heterochromatin Protein 1 (HP1a is a well-known conserved protein involved in heterochromatin formation and gene silencing in different species including humans. A general model has been proposed for heterochromatin formation and epigenetic gene silencing in different species that implies an essential role for HP1a. According to the model, histone methyltransferase enzymes (HMTases methylate the histone H3 at lysine 9 (H3K9me, creating selective binding sites for itself and the chromodomain of HP1a. This complex is thought to form a higher order chromatin state that represses gene activity. It has also been found that HP1a plays a role in telomere capping. Surprisingly, recent studies have shown that HP1a is present at many euchromatic sites along polytene chromosomes of Drosophila melanogaster, including the developmental and heat-shock-induced puffs, and that this protein can be removed from these sites by in vivo RNase treatment, thus suggesting an association of HP1a with the transcripts of many active genes. To test this suggestion, we performed an extensive screening by RIP-chip assay (RNA-immunoprecipitation on microarrays, and we found that HP1a is associated with transcripts of more than one hundred euchromatic genes. An expression analysis in HP1a mutants shows that HP1a is required for positive regulation of these genes. Cytogenetic and molecular assays show that HP1a also interacts with the well known proteins DDP1, HRB87F, and PEP, which belong to different classes of heterogeneous nuclear ribonucleoproteins (hnRNPs involved in RNA processing. Surprisingly, we found that all these hnRNP proteins also bind heterochromatin and are dominant suppressors of position effect variegation. Together, our data show novel and unexpected functions for HP1a and hnRNPs proteins. All these proteins are in fact involved both in RNA transcript processing and in heterochromatin formation. This suggests that, in general, similar epigenetic mechanisms

  11. Dopamine modulates metabolic rate and temperature sensitivity in Drosophila melanogaster.

    Directory of Open Access Journals (Sweden)

    Taro Ueno

    Full Text Available Homeothermal animals, such as mammals, maintain their body temperature by heat generation and heat dissipation, while poikilothermal animals, such as insects, accomplish it by relocating to an environment of their favored temperature. Catecholamines are known to regulate thermogenesis and metabolic rate in mammals, but their roles in other animals are poorly understood. The fruit fly, Drosophila melanogaster, has been used as a model system for the genetic studies of temperature preference behavior. Here, we demonstrate that metabolic rate and temperature sensitivity of some temperature sensitive behaviors are regulated by dopamine in Drosophila. Temperature-sensitive molecules like dTrpA1 and shi(ts induce temperature-dependent behavioral changes, and the temperature at which the changes are induced were lowered in the dopamine transporter-defective mutant, fumin. The mutant also displays a preference for lower temperatures. This thermophobic phenotype was rescued by the genetic recovery of the dopamine transporter in dopamine neurons. Flies fed with a dopamine biosynthesis inhibitor (3-iodo-L-tyrosine, which diminishes dopamine signaling, exhibited preference for a higher temperature. Furthermore, we found that the metabolic rate is up-regulated in the fumin mutant. Taken together, dopamine has functions in the temperature sensitivity of behavioral changes and metabolic rate regulation in Drosophila, as well as its previously reported functions in arousal/sleep regulation.

  12. Drosophila PINK1 and parkin loss-of-function mutants display a range of non-motor Parkinson's disease phenotypes.

    Science.gov (United States)

    Julienne, Hannah; Buhl, Edgar; Leslie, David S; Hodge, James J L

    2017-08-01

    Parkinson's disease (PD) is more commonly associated with its motor symptoms and the related degeneration of dopamine (DA) neurons. However, it is becoming increasingly clear that PD patients also display a wide range of non-motor symptoms, including memory deficits and disruptions of their sleep-wake cycles. These have a large impact on their quality of life, and often precede the onset of motor symptoms, but their etiology is poorly understood. The fruit fly Drosophila has already been successfully used to model PD, and has been used extensively to study relevant non-motor behaviours in other contexts, but little attention has yet been paid to modelling non-motor symptoms of PD in this genetically tractable organism. We examined memory performance and circadian rhythms in flies with loss-of-function mutations in two PD genes: PINK1 and parkin. We found learning and memory abnormalities in both mutant genotypes, as well as a weakening of circadian rhythms that is underpinned by electrophysiological changes in clock neurons. Our study paves the way for further work that may help us understand the mechanisms underlying these neglected aspects of PD, thus identifying new targets for treatments to address these non-motor problems specifically and perhaps even to halt disease progression in its prodromal phase. Crown Copyright © 2017. Published by Elsevier Inc. All rights reserved.

  13. Data Integration and Applications of Functional Gene Networks in Drosophila Melanogaster

    Science.gov (United States)

    Costello, James Christopher

    2009-01-01

    Understanding the function of every gene in the genome is a central goal in the biological sciences. This includes full characterization of a genes phenotypic effects, molecular interactions, the evolutionary forces that shape its function(s), and how these functions interrelate. Despite a long history and considerable effort to understand all…

  14. Dendritic Actin Cytoskeleton: Structure, Functions, and Regulations

    Directory of Open Access Journals (Sweden)

    Anja Konietzny

    2017-05-01

    Full Text Available Actin is a versatile and ubiquitous cytoskeletal protein that plays a major role in both the establishment and the maintenance of neuronal polarity. For a long time, the most prominent roles that were attributed to actin in neurons were the movement of growth cones, polarized cargo sorting at the axon initial segment, and the dynamic plasticity of dendritic spines, since those compartments contain large accumulations of actin filaments (F-actin that can be readily visualized using electron- and fluorescence microscopy. With the development of super-resolution microscopy in the past few years, previously unknown structures of the actin cytoskeleton have been uncovered: a periodic lattice consisting of actin and spectrin seems to pervade not only the whole axon, but also dendrites and even the necks of dendritic spines. Apart from that striking feature, patches of F-actin and deep actin filament bundles have been described along the lengths of neurites. So far, research has been focused on the specific roles of actin in the axon, while it is becoming more and more apparent that in the dendrite, actin is not only confined to dendritic spines, but serves many additional and important functions. In this review, we focus on recent developments regarding the role of actin in dendrite morphology, the regulation of actin dynamics by internal and external factors, and the role of F-actin in dendritic protein trafficking.

  15. Gustatory Processing in Drosophila melanogaster.

    Science.gov (United States)

    Scott, Kristin

    2018-01-07

    The ability to identify nutrient-rich food and avoid toxic substances is essential for an animal's survival. Although olfaction and vision contribute to food detection, the gustatory system acts as a final checkpoint control for food acceptance or rejection. The vinegar fly Drosophila melanogaster tastes many of the same stimuli as mammals and provides an excellent model system for comparative studies of taste detection. The relative simplicity of the fly brain and behaviors, along with the molecular genetic and functional approaches available in this system, allow the examination of gustatory neural circuits from sensory input to motor output. This review discusses the molecules and cells that detect taste compounds in the periphery and the circuits that process taste information in the brain. These studies are providing insight into how the detection of taste compounds regulates feeding decisions.

  16. Drosophila Syd-1, liprin-α, and protein phosphatase 2A B' subunit Wrd function in a linear pathway to prevent ectopic accumulation of synaptic materials in distal axons.

    Science.gov (United States)

    Li, Long; Tian, Xiaolin; Zhu, Mingwei; Bulgari, Dinara; Böhme, Mathias A; Goettfert, Fabian; Wichmann, Carolin; Sigrist, Stephan J; Levitan, Edwin S; Wu, Chunlai

    2014-06-18

    During synaptic development, presynaptic differentiation occurs as an intrinsic property of axons to form specialized areas of plasma membrane [active zones (AZs)] that regulate exocytosis and endocytosis of synaptic vesicles. Genetic and biochemical studies in vertebrate and invertebrate model systems have identified a number of proteins involved in AZ assembly. However, elucidating the molecular events of AZ assembly in a spatiotemporal manner remains a challenge. Syd-1 (synapse defective-1) and Liprin-α have been identified as two master organizers of AZ assembly. Genetic and imaging analyses in invertebrates show that Syd-1 works upstream of Liprin-α in synaptic assembly through undefined mechanisms. To understand molecular pathways downstream of Liprin-α, we performed a proteomic screen of Liprin-α-interacting proteins in Drosophila brains. We identify Drosophila protein phosphatase 2A (PP2A) regulatory subunit B' [Wrd (Well Rounded)] as a Liprin-α-interacting protein, and we demonstrate that it mediates the interaction of Liprin-α with PP2A holoenzyme and the Liprin-α-dependent synaptic localization of PP2A. Interestingly, loss of function in syd-1, liprin-α, or wrd shares a common defect in which a portion of synaptic vesicles, dense-core vesicles, and presynaptic cytomatrix proteins ectopically accumulate at the distal, but not proximal, region of motoneuron axons. Strong genetic data show that a linear syd-1/liprin-α/wrd pathway in the motoneuron antagonizes glycogen synthase kinase-3β kinase activity to prevent the ectopic accumulation of synaptic materials. Furthermore, we provide data suggesting that the syd-1/liprin-α/wrd pathway stabilizes AZ specification at the nerve terminal and that such a novel function is independent of the roles of syd-1/liprin-α in regulating the morphology of the T-bar structural protein BRP (Bruchpilot). Copyright © 2014 the authors 0270-6474/14/348474-14$15.00/0.

  17. Drosophila Syd-1, Liprin-α, and Protein Phosphatase 2A B′ Subunit Wrd Function in a Linear Pathway to Prevent Ectopic Accumulation of Synaptic Materials in Distal Axons

    Science.gov (United States)

    Li, Long; Tian, Xiaolin; Zhu, Mingwei; Bulgari, Dinara; Böhme, Mathias A.; Goettfert, Fabian; Wichmann, Carolin; Sigrist, Stephan J.; Levitan, Edwin S.

    2014-01-01

    During synaptic development, presynaptic differentiation occurs as an intrinsic property of axons to form specialized areas of plasma membrane [active zones (AZs)] that regulate exocytosis and endocytosis of synaptic vesicles. Genetic and biochemical studies in vertebrate and invertebrate model systems have identified a number of proteins involved in AZ assembly. However, elucidating the molecular events of AZ assembly in a spatiotemporal manner remains a challenge. Syd-1 (synapse defective-1) and Liprin-α have been identified as two master organizers of AZ assembly. Genetic and imaging analyses in invertebrates show that Syd-1 works upstream of Liprin-α in synaptic assembly through undefined mechanisms. To understand molecular pathways downstream of Liprin-α, we performed a proteomic screen of Liprin-α-interacting proteins in Drosophila brains. We identify Drosophila protein phosphatase 2A (PP2A) regulatory subunit B′ [Wrd (Well Rounded)] as a Liprin-α-interacting protein, and we demonstrate that it mediates the interaction of Liprin-α with PP2A holoenzyme and the Liprin-α-dependent synaptic localization of PP2A. Interestingly, loss of function in syd-1, liprin-α, or wrd shares a common defect in which a portion of synaptic vesicles, dense-core vesicles, and presynaptic cytomatrix proteins ectopically accumulate at the distal, but not proximal, region of motoneuron axons. Strong genetic data show that a linear syd-1/liprin-α/wrd pathway in the motoneuron antagonizes glycogen synthase kinase-3β kinase activity to prevent the ectopic accumulation of synaptic materials. Furthermore, we provide data suggesting that the syd-1/liprin-α/wrd pathway stabilizes AZ specification at the nerve terminal and that such a novel function is independent of the roles of syd-1/liprin-α in regulating the morphology of the T-bar structural protein BRP (Bruchpilot). PMID:24948803

  18. Sex comb on midleg (Scm) is a functional link between PcG-repressive complexes in Drosophila.

    Science.gov (United States)

    Kang, Hyuckjoon; McElroy, Kyle A; Jung, Youngsook Lucy; Alekseyenko, Artyom A; Zee, Barry M; Park, Peter J; Kuroda, Mitzi I

    2015-06-01

    The Polycomb group (PcG) proteins are key regulators of development in Drosophila and are strongly implicated in human health and disease. How PcG complexes form repressive chromatin domains remains unclear. Using cross-linked affinity purifications of BioTAP-Polycomb (Pc) or BioTAP-Enhancer of zeste [E(z)], we captured all PcG-repressive complex 1 (PRC1) or PRC2 core components and Sex comb on midleg (Scm) as the only protein strongly enriched with both complexes. Although previously not linked to PRC2, we confirmed direct binding of Scm and PRC2 using recombinant protein expression and colocalization of Scm with PRC1, PRC2, and H3K27me3 in embryos and cultured cells using ChIP-seq (chromatin immunoprecipitation [ChIP] combined with deep sequencing). Furthermore, we found that RNAi knockdown of Scm and overexpression of the dominant-negative Scm-SAM (sterile α motif) domain both affected the binding pattern of E(z) on polytene chromosomes. Aberrant localization of the Scm-SAM domain in long contiguous regions on polytene chromosomes revealed its independent ability to spread on chromatin, consistent with its previously described ability to oligomerize in vitro. Pull-downs of BioTAP-Scm captured PRC1 and PRC2 and additional repressive complexes, including PhoRC, LINT, and CtBP. We propose that Scm is a key mediator connecting PRC1, PRC2, and transcriptional silencing. Combined with previous structural and genetic analyses, our results strongly suggest that Scm coordinates PcG complexes and polymerizes to produce broad domains of PcG silencing. © 2015 Kang et al.; Published by Cold Spring Harbor Laboratory Press.

  19. Functional Conservation of the Glide/Gcm Regulatory Network Controlling Glia, Hemocyte, and Tendon Cell Differentiation in Drosophila

    Science.gov (United States)

    Cattenoz, Pierre B.; Popkova, Anna; Southall, Tony D.; Aiello, Giuseppe; Brand, Andrea H.; Giangrande, Angela

    2016-01-01

    High-throughput screens allow us to understand how transcription factors trigger developmental processes, including cell specification. A major challenge is identification of their binding sites because feedback loops and homeostatic interactions may mask the direct impact of those factors in transcriptome analyses. Moreover, this approach dissects the downstream signaling cascades and facilitates identification of conserved transcriptional programs. Here we show the results and the validation of a DNA adenine methyltransferase identification (DamID) genome-wide screen that identifies the direct targets of Glide/Gcm, a potent transcription factor that controls glia, hemocyte, and tendon cell differentiation in Drosophila. The screen identifies many genes that had not been previously associated with Glide/Gcm and highlights three major signaling pathways interacting with Glide/Gcm: Notch, Hedgehog, and JAK/STAT, which all involve feedback loops. Furthermore, the screen identifies effector molecules that are necessary for cell-cell interactions during late developmental processes and/or in ontogeny. Typically, immunoglobulin (Ig) domain–containing proteins control cell adhesion and axonal navigation. This shows that early and transiently expressed fate determinants not only control other transcription factors that, in turn, implement a specific developmental program but also directly affect late developmental events and cell function. Finally, while the mammalian genome contains two orthologous Gcm genes, their function has been demonstrated in vertebrate-specific tissues, placenta, and parathyroid glands, begging questions on the evolutionary conservation of the Gcm cascade in higher organisms. Here we provide the first evidence for the conservation of Gcm direct targets in humans. In sum, this work uncovers novel aspects of cell specification and sets the basis for further understanding of the role of conserved Gcm gene regulatory cascades. PMID:26567182

  20. Rudimentary expression of RYamide in Drosophila melanogaster relative to other Drosophila species points to a functional decline of this neuropeptide gene.

    Science.gov (United States)

    Veenstra, Jan A; Khammassi, Hela

    2017-04-01

    RYamides are arthropod neuropeptides with unknown function. In 2011 two RYamides were isolated from D. melanogaster as the ligands for the G-protein coupled receptor CG5811. The D. melanogaster gene encoding these neuropeptides is highly unusual, as there are four RYamide encoding exons in the current genome assembly, but an exon encoding a signal peptide is absent. Comparing the D. melanogaster gene structure with those from other species, including D. virilis, suggests that the gene is degenerating. RNAseq data from 1634 short sequence read archives at NCBI containing more than 34 billion spots yielded numerous individual spots that correspond to the RYamide encoding exons, of which a large number include the intron-exon boundary at the start of this exon. Although 72 different sequences have been spliced onto this RYamide encoding exon, none codes for the signal peptide of this gene. Thus, the RNAseq data for this gene reveal only noise and no signal. The very small quantities of peptide recovered during isolation and the absence of credible RNAseq data, indicates that the gene is very little expressed, while the RYamide gene structure in D. melanogaster suggests that it might be evolving into a pseudogene. Yet, the identification of the peptides it encodes clearly shows it is still functional. Using region specific antisera, we could localize numerous neurons and enteroendocrine cells in D. willistoni, D. virilis and D. pseudoobscura, but only two adult abdominal neurons in D. melanogaster. Those two neurons project to and innervate the rectal papillae, suggesting that RYamides may be involved in the regulation of water homeostasis. Copyright © 2017 Elsevier Ltd. All rights reserved.

  1. Primary structure and functional characterization of a Drosophila dopamine receptor with high homology to human D1/5 receptors.

    Science.gov (United States)

    Gotzes, F; Balfanz, S; Baumann, A

    1994-01-01

    Members of the superfamily of G-protein coupled receptors share significant similarities in sequence and transmembrane architecture. We have isolated a Drosophila homologue of the mammalian dopamine receptor family using a low stringency hybridization approach. The deduced amino acid sequence is approximately 70% homologous to the human D1/D5 receptors. When expressed in HEK 293 cells, the Drosophila receptor stimulates cAMP production in response to dopamine application. This effect was mimicked by SKF 38393, a specific D1 receptor agonist, but inhibited by dopaminergic antagonists such as butaclamol and flupentixol. In situ hybridization revealed that the Drosophila dopamine receptor is highly expressed in the somata of the optic lobes. This suggests that the receptor might be involved in the processing of visual information and/or visual learning in invertebrates.

  2. Early gene Broad complex plays a key role in regulating the immune response triggered by ecdysone in the Malpighian tubules of Drosophila melanogaster.

    Science.gov (United States)

    Verma, Puja; Tapadia, Madhu G

    2015-08-01

    In insects, humoral response to injury is accomplished by the production of antimicrobial peptides (AMPs) which are secreted in the hemolymph to eliminate the pathogen. Drosophila Malpighian tubules (MTs), however, are unique immune organs that show constitutive expression of AMPs even in unchallenged conditions and the onset of immune response is developmental stage dependent. Earlier reports have shown ecdysone positively regulates immune response after pathogenic challenge however, a robust response requires prior potentiation by the hormone. Here we provide evidence to show that MTs do not require prior potentiation with ecdysone hormone for expression of AMPs and they respond to ecdysone very fast even without immune challenge, although the different AMPs Diptericin, Cecropin, Attacin, Drosocin show differential expression in response to ecdysone. We show that early gene Broad complex (BR-C) could be regulating the IMD pathway by activating Relish and physically interacting with it to activate AMPs expression. BR-C depletion from Malpighian tubules renders the flies susceptible to infection. We also show that in MTs ecdysone signaling is transduced by EcR-B1 and B2. In the absence of ecdysone signaling the IMD pathway associated genes are down regulated and activation and translocation of transcription factor Relish is also affected. Copyright © 2015 Elsevier Ltd. All rights reserved.

  3. Single Nucleotide Polymorphism Markers for Genetic Mapping in Drosophila melanogaster

    OpenAIRE

    Hoskins, Roger A.; Phan, Alexander C.; Naeemuddin, Mohammed; Mapa, Felipa A.; Ruddy, David A.; Ryan, Jessica J.; Young, Lynn M.; Wells, Trent; Kopczynski, Casey; Ellis, Michael C.

    2001-01-01

    For nearly a century, genetic analysis in Drosophila melanogaster has been a powerful tool for analyzing gene function, yet Drosophila lacks the molecular genetic mapping tools that recently have revolutionized human, mouse, and plant genetics. Here, we describe the systematic characterization of a dense set of molecular markers in Drosophila by using a sequence tagged site-based physical map of the genome. We identify 474 biallelic markers in standard laboratory strains of Drosophila that sp...

  4. The fruitless gene is required for the proper formation of axonal tracts in the embryonic central nervous system of Drosophila

    NARCIS (Netherlands)

    Song, Ho-Juhn; Billeter, Jean-Christophe; Reynaud, Enrique; Carlo, Troy; Spana, Eric P; Perrimon, Norbert; Goodwin, Stephen F; Baker, Bruce S; Taylor, Barbara J

    2002-01-01

    The fruitless (fru) gene in Drosophila melanogaster is a multifunctional gene that has sex-specific functions in the regulation of male sexual behavior and sex-nonspecific functions affecting adult viability and external morphology. While much attention has focused on fru's sex-specific roles, less

  5. Matrix regulators in neural stem cell functions.

    Science.gov (United States)

    Wade, Anna; McKinney, Andrew; Phillips, Joanna J

    2014-08-01

    Neural stem/progenitor cells (NSPCs) reside within a complex and dynamic extracellular microenvironment, or niche. This niche regulates fundamental aspects of their behavior during normal neural development and repair. Precise yet dynamic regulation of NSPC self-renewal, migration, and differentiation is critical and must persist over the life of an organism. In this review, we summarize some of the major components of the NSPC niche and provide examples of how cues from the extracellular matrix regulate NSPC behaviors. We use proteoglycans to illustrate the many diverse roles of the niche in providing temporal and spatial regulation of cellular behavior. The NSPC niche is comprised of multiple components that include; soluble ligands, such as growth factors, morphogens, chemokines, and neurotransmitters, the extracellular matrix, and cellular components. As illustrated by proteoglycans, a major component of the extracellular matrix, the NSPC, niche provides temporal and spatial regulation of NSPC behaviors. The factors that control NSPC behavior are vital to understand as we attempt to modulate normal neural development and repair. Furthermore, an improved understanding of how these factors regulate cell proliferation, migration, and differentiation, crucial for malignancy, may reveal novel anti-tumor strategies. This article is part of a Special Issue entitled Matrix-mediated cell behaviour and properties. Copyright © 2014 Elsevier B.V. All rights reserved.

  6. Conserved family of glycerol kinase loci in Drosophila melanogaster

    Science.gov (United States)

    Martinez Agosto, Julian A.; McCabe, Edward R.B.

    2009-01-01

    Glycerol kinase (GK) is an enzyme that catalyzes the formation of glycerol 3-phosphate from ATP and glycerol, the rate-limiting step in glycerol utilization. We analyzed the genome of the model organism Drosophila melanogaster and identified five GK orthologs, including two loci with sequence homology to the mammalian Xp21 GK protein. Using a combination of sequence analysis and evolutionary comparisons of orthologs between species, we characterized functional domains in the protein required for GK activity. Our findings include additional conserved domains that suggest novel nuclear and mitochondrial functions for glycerol kinase in apoptosis and transcriptional regulation. Investigation of GK function in Drosophila will inform us about the role of this enzyme in development and will provide us with a tool to examine genetic modifiers of human metabolic disorders. PMID:16545593

  7. TIF-IA-dependent regulation of ribosome synthesis in drosophila muscle is required to maintain systemic insulin signaling and larval growth.

    Directory of Open Access Journals (Sweden)

    Abhishek Ghosh

    2014-10-01

    Full Text Available The conserved TOR kinase signaling network links nutrient availability to cell, tissue and body growth in animals. One important growth-regulatory target of TOR signaling is ribosome biogenesis. Studies in yeast and mammalian cell culture have described how TOR controls rRNA synthesis-a limiting step in ribosome biogenesis-via the RNA Polymerase I transcription factor TIF-IA. However, the contribution of TOR-dependent ribosome synthesis to tissue and body growth in animals is less clear. Here we show in Drosophila larvae that ribosome synthesis in muscle is required non-autonomously to maintain normal body growth and development. We find that amino acid starvation and TOR inhibition lead to reduced levels of TIF-IA, and decreased rRNA synthesis in larval muscle. When we mimic this decrease in muscle ribosome synthesis using RNAi-mediated knockdown of TIF-IA, we observe delayed larval development and reduced body growth. This reduction in growth is caused by lowered systemic insulin signaling via two endocrine responses: reduced expression of Drosophila insulin-like peptides (dILPs from the brain and increased expression of Imp-L2-a secreted factor that binds and inhibits dILP activity-from muscle. We also observed that maintaining TIF-IA levels in muscle could partially reverse the starvation-mediated suppression of systemic insulin signaling. Finally, we show that activation of TOR specifically in muscle can increase overall body size and this effect requires TIF-IA function. These data suggest that muscle ribosome synthesis functions as a nutrient-dependent checkpoint for overall body growth: in nutrient rich conditions, TOR is required to maintain levels of TIF-IA and ribosome synthesis to promote high levels of systemic insulin, but under conditions of starvation stress, reduced muscle ribosome synthesis triggers an endocrine response that limits systemic insulin signaling to restrict growth and maintain homeostasis.

  8. MicroRNA miR-308 regulates dMyc through a negative feedback loop in Drosophila

    Directory of Open Access Journals (Sweden)

    Kaveh Daneshvar

    2012-10-01

    The abundance of Myc protein must be exquisitely controlled to avoid growth abnormalities caused by too much or too little Myc. An intriguing mode of regulation exists in which Myc protein itself leads to reduction in its abundance. We show here that dMyc binds to the miR-308 locus and increases its expression. Using our gain-of-function approach, we show that an increase in miR-308 causes a destabilization of dMyc mRNA and reduced dMyc protein levels. In vivo knockdown of miR-308 confirmed the regulation of dMyc levels in embryos. This regulatory loop is crucial for maintaining appropriate dMyc levels and normal development. Perturbation of the loop, either by elevated miR-308 or elevated dMyc, caused lethality. Combining elevated levels of both, therefore restoring balance between miR-308 and dMyc levels, resulted in lower apoptotic activity and suppression of lethality. These results reveal a sensitive feedback mechanism that is crucial to prevent the pathologies caused by abnormal levels of dMyc.

  9. Family roles as family functioning regulators

    OpenAIRE

    STEPANYAN ARMINE

    2015-01-01

    The author examines the problems of formation and functioning of family roles. Having social roots, family roles appear on individual level by performing the social function of the formation of family as a social institute.

  10. Longitudinal Associations among Child Maltreatment, Social Functioning, and Cortisol Regulation

    Science.gov (United States)

    Alink, Lenneke R. A.; Cicchetti, Dante; Kim, Jungmeen; Rogosch, Fred A.

    2012-01-01

    Child maltreatment increases the risk for impaired social functioning and cortisol regulation. However, the longitudinal interplay among these factors is still unclear. This study aimed to shed light on the effect of maltreatment on social functioning and cortisol regulation over time. The sample consisted of 236 children (mean age 7.64 years, SD…

  11. Neuronal Cbl Controls Biosynthesis of Insulin-Like Peptides in Drosophila melanogaster

    Science.gov (United States)

    Yu, Yue; Sun, Ying; He, Shengqi; Yan, Cheng; Rui, Liangyou; Li, Wenjun

    2012-01-01

    The Cbl family proteins function as both E3 ubiquitin ligases and adaptor proteins to regulate various cellular signaling events, including the insulin/insulin-like growth factor 1 (IGF1) and epidermal growth factor (EGF) pathways. These pathways play essential roles in growth, development, metabolism, and survival. Here we show that in Drosophila melanogaster, Drosophila Cbl (dCbl) regulates longevity and carbohydrate metabolism through downregulating the production of Drosophila insulin-like peptides (dILPs) in the brain. We found that dCbl was highly expressed in the brain and knockdown of the expression of dCbl specifically in neurons by RNA interference increased sensitivity to oxidative stress or starvation, decreased carbohydrate levels, and shortened life span. Insulin-producing neuron-specific knockdown of dCbl resulted in similar phenotypes. dCbl deficiency in either the brain or insulin-producing cells upregulated the expression of dilp genes, resulting in elevated activation of the dILP pathway, including phosphorylation of Drosophila Akt and Drosophila extracellular signal-regulated kinase (dERK). Genetic interaction analyses revealed that blocking Drosophila epidermal growth factor receptor (dEGFR)-dERK signaling in pan-neurons or insulin-producing cells by overexpressing a dominant-negative form of dEGFR abolished the effect of dCbl deficiency on the upregulation of dilp genes. Furthermore, knockdown of c-Cbl in INS-1 cells, a rat β-cell line, also increased insulin biosynthesis and glucose-stimulated secretion in an ERK-dependent manner. Collectively, these results suggest that neuronal dCbl regulates life span, stress responses, and metabolism by suppressing dILP production and the EGFR-ERK pathway mediates the dCbl action. Cbl suppression of insulin biosynthesis is evolutionarily conserved, raising the possibility that Cbl may similarly exert its physiological actions through regulating insulin production in β cells. PMID:22778134

  12. Epigenetic telomere protection by Drosophila DNA damage response pathways.

    Science.gov (United States)

    Oikemus, Sarah R; Queiroz-Machado, Joana; Lai, KuanJu; McGinnis, Nadine; Sunkel, Claudio; Brodsky, Michael H

    2006-05-01

    Analysis of terminal deletion chromosomes indicates that a sequence-independent mechanism regulates protection of Drosophila telomeres. Mutations in Drosophila DNA damage response genes such as atm/tefu, mre11, or rad50 disrupt telomere protection and localization of the telomere-associated proteins HP1 and HOAP, suggesting that recognition of chromosome ends contributes to telomere protection. However, the partial telomere protection phenotype of these mutations limits the ability to test if they act in the epigenetic telomere protection mechanism. We examined the roles of the Drosophila atm and atr-atrip DNA damage response pathways and the nbs homolog in DNA damage responses and telomere protection. As in other organisms, the atm and atr-atrip pathways act in parallel to promote telomere protection. Cells lacking both pathways exhibit severe defects in telomere protection and fail to localize the protection protein HOAP to telomeres. Drosophila nbs is required for both atm- and atr-dependent DNA damage responses and acts in these pathways during DNA repair. The telomere fusion phenotype of nbs is consistent with defects in each of these activities. Cells defective in both the atm and atr pathways were used to examine if DNA damage response pathways regulate telomere protection without affecting telomere specific sequences. In these cells, chromosome fusion sites retain telomere-specific sequences, demonstrating that loss of these sequences is not responsible for loss of protection. Furthermore, terminally deleted chromosomes also fuse in these cells, directly implicating DNA damage response pathways in the epigenetic protection of telomeres. We propose that recognition of chromosome ends and recruitment of HP1 and HOAP by DNA damage response proteins is essential for the epigenetic protection of Drosophila telomeres. Given the conserved roles of DNA damage response proteins in telomere function, related mechanisms may act at the telomeres of other organisms.

  13. Conformational regulation of urokinase receptor function

    DEFF Research Database (Denmark)

    Gårdsvoll, Henrik; Jacobsen, Benedikte; Kriegbaum, Mette C

    2011-01-01

    PA per se into the hydrophobic ligand binding cavity of uPAR that modulates the function of this receptor. Based on these data, we now propose a model in which the inherent interdomain mobility in uPAR plays a major role in modulating its function. Particularly one uPAR conformation, which is stabilized...

  14. Analysis of Rca1 function at the G1-S transition in Drosophila melanogaster

    OpenAIRE

    Querings, Silvia

    2008-01-01

    Tight control of APC/C-Cdh1Fzr activity is essential for progression through mitosis and establishment of the G1 phase. Rca1 is a nuclear protein that inhibits the APC/C-Cdh1Fzr complex during G2 to allow cyclin accumulation and subsequent entry into mitosis. In this thesis, a localisation study of Rca1 was performed revealing that a nuclear localisation sequence (NLS) and other domains in the protein mediate efficient nuclear accumulation. Besides its function in G2, Rca1 expression can prom...

  15. Structure of PCNA from Drosophila melanogaster

    International Nuclear Information System (INIS)

    Wang, Ke; Shi, Zhubing; Zhang, Min; Cheng, Dianlin

    2013-01-01

    Proliferating cell nuclear antigen (PCNA) plays essential roles in DNA replication, DNA repair, cell-cycle regulation and chromatin metabolism. The PCNA from Drosophila melanogaster (DmPCNA) has been purified and crystallized. Proliferating cell nuclear antigen (PCNA) plays essential roles in DNA replication, DNA repair, cell-cycle regulation and chromatin metabolism. The PCNA from Drosophila melanogaster (DmPCNA) was purified and crystallized. The crystal of DmPCNA diffracted to 2.0 Å resolution and belonged to space group H3, with unit-cell parameters a = b = 151.16, c = 38.28 Å. The structure of DmPCNA was determined by molecular replacement. DmPCNA forms a symmetric homotrimer in a head-to-tail manner. An interdomain connector loop (IDCL) links the N- and C-terminal domains. Additionally, the N-terminal and C-terminal domains contact each other through hydrophobic associations. Compared with human PCNA, the IDCL of DmPCNA has conformational changes, which may explain their difference in function. This work provides a structural basis for further functional and evolutionary studies of PCNA

  16. Regulators of Long-Term Memory Revealed by Mushroom Body-Specific Gene Expression Profiling in Drosophila melanogaster.

    Science.gov (United States)

    Widmer, Yves F; Bilican, Adem; Bruggmann, Rémy; Sprecher, Simon G

    2018-06-20

    Memory formation is achieved by genetically tightly controlled molecular pathways that result in a change of synaptic strength and synapse organization. While for short-term memory traces rapidly acting biochemical pathways are in place, the formation of long-lasting memories requires changes in the transcriptional program of a cell. Although many genes involved in learning and memory formation have been identified, little is known about the genetic mechanisms required for changing the transcriptional program during different phases of long-term memory formation. With Drosophila melanogaster as a model system we profiled transcriptomic changes in the mushroom body, a memory center in the fly brain, at distinct time intervals during appetitive olfactory long-term memory formation using the targeted DamID technique. We describe the gene expression profiles during these phases and tested 33 selected candidate genes for deficits in long-term memory formation using RNAi knockdown. We identified 10 genes that enhance or decrease memory when knocked-down in the mushroom body. For vajk-1 and hacd1 , the two strongest hits, we gained further support for their crucial role in appetitive learning and forgetting. These findings show that profiling gene expression changes in specific cell-types harboring memory traces provides a powerful entry point to identify new genes involved in learning and memory. The presented transcriptomic data may further be used as resource to study genes acting at different memory phases. Copyright © 2018, Genetics.

  17. The developmental transcriptome of Drosophila melanogaster

    Energy Technology Data Exchange (ETDEWEB)

    University of Connecticut; Graveley, Brenton R.; Brooks, Angela N.; Carlson, Joseph W.; Duff, Michael O.; Landolin, Jane M.; Yang, Li; Artieri, Carlo G.; van Baren, Marijke J.; Boley, Nathan; Booth, Benjamin W.; Brown, James B.; Cherbas, Lucy; Davis, Carrie A.; Dobin, Alex; Li, Renhua; Lin, Wei; Malone, John H.; Mattiuzzo, Nicolas R.; Miller, David; Sturgill, David; Tuch, Brian B.; Zaleski, Chris; Zhang, Dayu; Blanchette, Marco; Dudoit, Sandrine; Eads, Brian; Green, Richard E.; Hammonds, Ann; Jiang, Lichun; Kapranov, Phil; Langton, Laura; Perrimon, Norbert; Sandler, Jeremy E.; Wan, Kenneth H.; Willingham, Aarron; Zhang, Yu; Zou, Yi; Andrews, Justen; Bicke, Peter J.; Brenner, Steven E.; Brent, Michael R.; Cherbas, Peter; Gingeras, Thomas R.; Hoskins, Roger A.; Kaufman, Thomas C.; Oliver, Brian; Celniker, Susan E.

    2010-12-02

    Drosophila melanogaster is one of the most well studied genetic model organisms; nonetheless, its genome still contains unannotated coding and non-coding genes, transcripts, exons and RNA editing sites. Full discovery and annotation are pre-requisites for understanding how the regulation of transcription, splicing and RNA editing directs the development of this complex organism. Here we used RNA-Seq, tiling microarrays and cDNA sequencing to explore the transcriptome in 30 distinct developmental stages. We identified 111,195 new elements, including thousands of genes, coding and non-coding transcripts, exons, splicing and editing events, and inferred protein isoforms that previously eluded discovery using established experimental, prediction and conservation-based approaches. These data substantially expand the number of known transcribed elements in the Drosophila genome and provide a high-resolution view of transcriptome dynamics throughout development. Drosophila melanogaster is an important non-mammalian model system that has had a critical role in basic biological discoveries, such as identifying chromosomes as the carriers of genetic information and uncovering the role of genes in development. Because it shares a substantial genic content with humans, Drosophila is increasingly used as a translational model for human development, homeostasis and disease. High-quality maps are needed for all functional genomic elements. Previous studies demonstrated that a rich collection of genes is deployed during the life cycle of the fly. Although expression profiling using microarrays has revealed the expression of, 13,000 annotated genes, it is difficult to map splice junctions and individual base modifications generated by RNA editing using such approaches. Single-base resolution is essential to define precisely the elements that comprise the Drosophila transcriptome. Estimates of the number of transcript isoforms are less accurate than estimates of the number of genes

  18. Long-term functional side-effects of stimulants and sedatives in Drosophila melanogaster.

    Directory of Open Access Journals (Sweden)

    Kennedy Matsagas

    2009-08-01

    Full Text Available Small invertebrate animals, such as nematodes and fruit flies, are increasingly being used to test candidate drugs both for specific therapeutic purposes and for long-term health effects. Some of the protocols used in these experiments feature such experimental design features as lifelong virginity and very low densities. By contrast, the ability of both fruit flies and nematodes to resist stress is frequently correlated with their longevity and other functional measures, suggesting that low-stress assays are not necessarily the only useful protocol for testing the long-term effects of drugs.Here we report an alternative protocol for fruit fly drug-testing that maximizes reproductive opportunities and other types of interaction, with moderately high population densities. We validate this protocol using two types of experimental tests: 1. We show that this protocol detects previously well-established genetic differences between outbred fruit fly populations. 2. We show that this protocol is able to distinguish among the long-term effects of similar types of drugs within two broad categories, stimulants and tranquilizers.Large-scale fly drug testing can be conducted using mixed-sex high-density cage assays. We find that the commonly-used stimulants caffeine and theobromine differ dramatically in their chronic functional effects, theobromine being more benign. Likewise, we find that two generic pharmaceutical tranquilizers, lithium carbonate and valproic acid, differ dramatically in their chronic effects, lithium being more benign. However, these findings do not necessarily apply to human subjects, and we thus do not recommend the use of any one substance over any other.

  19. Long-term functional side-effects of stimulants and sedatives in Drosophila melanogaster.

    Science.gov (United States)

    Matsagas, Kennedy; Lim, David B; Horwitz, Marc; Rizza, Cristina L; Mueller, Laurence D; Villeponteau, Bryant; Rose, Michael R

    2009-08-11

    Small invertebrate animals, such as nematodes and fruit flies, are increasingly being used to test candidate drugs both for specific therapeutic purposes and for long-term health effects. Some of the protocols used in these experiments feature such experimental design features as lifelong virginity and very low densities. By contrast, the ability of both fruit flies and nematodes to resist stress is frequently correlated with their longevity and other functional measures, suggesting that low-stress assays are not necessarily the only useful protocol for testing the long-term effects of drugs. Here we report an alternative protocol for fruit fly drug-testing that maximizes reproductive opportunities and other types of interaction, with moderately high population densities. We validate this protocol using two types of experimental tests: 1. We show that this protocol detects previously well-established genetic differences between outbred fruit fly populations. 2. We show that this protocol is able to distinguish among the long-term effects of similar types of drugs within two broad categories, stimulants and tranquilizers. Large-scale fly drug testing can be conducted using mixed-sex high-density cage assays. We find that the commonly-used stimulants caffeine and theobromine differ dramatically in their chronic functional effects, theobromine being more benign. Likewise, we find that two generic pharmaceutical tranquilizers, lithium carbonate and valproic acid, differ dramatically in their chronic effects, lithium being more benign. However, these findings do not necessarily apply to human subjects, and we thus do not recommend the use of any one substance over any other.

  20. Identification of ER Proteins Involved in the Functional Organisation of the Early Secretory Pathway in Drosophila Cells by a Targeted RNAi Screen

    Science.gov (United States)

    Kondylis, Vangelis; Tang, Yang; Fuchs, Florian; Boutros, Michael; Rabouille, Catherine

    2011-01-01

    Background In Drosophila, the early secretory apparatus comprises discrete paired Golgi stacks in close proximity to exit sites from the endoplasmic reticulum (tER sites), thus forming tER-Golgi units. Although many components involved in secretion have been identified, the structural components sustaining its organisation are less known. Here we set out to identify novel ER resident proteins involved in the of tER-Golgi unit organisation. Results To do so, we designed a novel screening strategy combining a bioinformatics pre-selection with an RNAi screen. We first selected 156 proteins exhibiting known or related ER retention/retrieval signals from a list of proteins predicted to have a signal sequence. We then performed a microscopy-based primary and confirmation RNAi screen in Drosophila S2 cells directly scoring the organisation of the tER-Golgi units. We identified 49 hits, most of which leading to an increased number of smaller tER-Golgi units (MG for “more and smaller Golgi”) upon depletion. 16 of them were validated and characterised, showing that this phenotype was not due to an inhibition in secretion, a block in G2, or ER stress. Interestingly, the MG phenotype was often accompanied by an increase in the cell volume. Out of 6 proteins, 4 were localised to the ER. Conclusions This work has identified novel proteins involved in the organisation of the Drosophila early secretory pathway. It contributes to the effort of assigning protein functions to gene annotation in the secretory pathway, and analysis of the MG hits revealed an enrichment of ER proteins. These results suggest a link between ER localisation, aspects of cell metabolism and tER-Golgi structural organisation. PMID:21383842

  1. Human SOD1 ALS Mutations in a Drosophila Knock-In Model Cause Severe Phenotypes and Reveal Dosage-Sensitive Gain- and Loss-of-Function Components.

    Science.gov (United States)

    Şahin, Aslı; Held, Aaron; Bredvik, Kirsten; Major, Paxton; Achilli, Toni-Marie; Kerson, Abigail G; Wharton, Kristi; Stilwell, Geoff; Reenan, Robert

    2017-02-01

    Amyotrophic Lateral Sclerosis (ALS) is the most common adult-onset motor neuron disease and familial forms can be caused by numerous dominant mutations of the copper-zinc superoxide dismutase 1 (SOD1) gene. Substantial efforts have been invested in studying SOD1-ALS transgenic animal models; yet, the molecular mechanisms by which ALS-mutant SOD1 protein acquires toxicity are not well understood. ALS-like phenotypes in animal models are highly dependent on transgene dosage. Thus, issues of whether the ALS-like phenotypes of these models stem from overexpression of mutant alleles or from aspects of the SOD1 mutation itself are not easily deconvolved. To address concerns about levels of mutant SOD1 in disease pathogenesis, we have genetically engineered four human ALS-causing SOD1 point mutations (G37R, H48R, H71Y, and G85R) into the endogenous locus of Drosophila SOD1 (dsod) via ends-out homologous recombination and analyzed the resulting molecular, biochemical, and behavioral phenotypes. Contrary to previous transgenic models, we have recapitulated ALS-like phenotypes without overexpression of the mutant protein. Drosophila carrying homozygous mutations rendering SOD1 protein enzymatically inactive (G85R, H48R, and H71Y) exhibited neurodegeneration, locomotor deficits, and shortened life span. The mutation retaining enzymatic activity (G37R) was phenotypically indistinguishable from controls. While the observed mutant dsod phenotypes were recessive, a gain-of-function component was uncovered through dosage studies and comparisons with age-matched dsod null animals, which failed to show severe locomotor defects or nerve degeneration. We conclude that the Drosophila knock-in model captures important aspects of human SOD1-based ALS and provides a powerful and useful tool for further genetic studies. Copyright © 2017 by the Genetics Society of America.

  2. FIG4 regulates lysosome membrane homeostasis independent of phosphatase function

    OpenAIRE

    Bharadwaj, Rajnish; Cunningham, Kathleen M.; Zhang, Ke; Lloyd, Thomas E.

    2015-01-01

    FIG4 is a phosphoinositide phosphatase that is mutated in several diseases including Charcot-Marie-Tooth Disease 4J (CMT4J) and Yunis-Varon syndrome (YVS). To investigate the mechanism of disease pathogenesis, we generated Drosophila models of FIG4-related diseases. Fig4 null mutant animals are viable but exhibit marked enlargement of the lysosomal compartment in muscle cells and neurons, accompanied by an age-related decline in flight ability. Transgenic animals expressing Drosophila Fig4 mi...

  3. Knockdown of SCF(Skp2 function causes double-parked accumulation in the nucleus and DNA re-replication in Drosophila plasmatocytes.

    Directory of Open Access Journals (Sweden)

    Paul T Kroeger

    Full Text Available In Drosophila, circulating hemocytes are derived from the cephalic mesoderm during the embryonic wave of hematopoiesis. These cells are contributed to the larva and persist through metamorphosis into the adult. To analyze this population of hemocytes, we considered data from a previously published RNAi screen in the hematopoietic niche, which suggested several members of the SCF complex play a role in lymph gland development. eater-Gal4;UAS-GFP flies were crossed to UAS-RNAi lines to knockdown the function of all known SCF complex members in a plasmatocyte-specific fashion, in order to identify which members are novel regulators of plasmatocytes. This specific SCF complex contains five core members: Lin-19-like, SkpA, Skp2, Roc1a and complex activator Nedd8. The complex was identified by its very distinctive large cell phenotype. Furthermore, these large cells stained for anti-P1, a plasmatocyte-specific antibody. It was also noted that the DNA in these cells appeared to be over-replicated. Gamma-tubulin and DAPI staining suggest the cells are undergoing re-replication as they had multiple centrioles and excessive DNA content. Further experimentation determined enlarged cells were BrdU-positive indicating they have progressed through S-phase. To determine how these cells become enlarged and undergo re-replication, cell cycle proteins were analyzed by immunofluorescence. This analysis identified three proteins that had altered subcellular localization in these enlarged cells: Cyclin E, Geminin and Double-parked. Previous research has shown that Double-parked must be degraded to exit S-phase, otherwise the DNA will undergo re-replication. When Double-parked was titrated from the nucleus by an excess of its inhibitor, geminin, the enlarged cells and aberrant protein localization phenotypes were partially rescued. The data in this report suggests that the SCF(Skp2 complex is necessary to ubiquitinate Double-parked during plasmatocyte cell division

  4. bHLH-O proteins balance the self-renewal and differentiation of Drosophila neural stem cells by regulating Earmuff expression.

    Science.gov (United States)

    Li, Xiaosu; Chen, Rui; Zhu, Sijun

    2017-11-15

    Balancing self-renewal and differentiation of stem cells requires differential expression of self-renewing factors in two daughter cells generated from the asymmetric division of the stem cells. In Drosophila type II neural stem cell (or neuroblast, NB) lineages, the expression of the basic helix-loop-helix-Orange (bHLH-O) family proteins, including Deadpan (Dpn) and E(spl) proteins, is required for maintaining the self-renewal and identity of type II NBs, whereas the absence of these self-renewing factors is essential for the differentiation of intermediate neural progenitors (INPs) generated from type II NBs. Here, we demonstrate that Dpn maintains type II NBs by suppressing the expression of Earmuff (Erm). We provide evidence that Dpn and E(spl) proteins suppress Erm by directly binding to C-sites and N-boxes in the cis-regulatory region of erm. Conversely, the absence of bHLH-O proteins in INPs allows activation of erm and Erm-mediated maturation of INPs. Our results further suggest that Pointed P1 (PntP1) mediates the dedifferentiation of INPs resulting from the loss of Erm or overexpression of Dpn or E(spl) proteins. Taken together, these findings reveal mechanisms underlying the regulation of the maintenance of type II NBs and differentiation of INPs through the differential expression of bHLH-O family proteins. Copyright © 2017 Elsevier Inc. All rights reserved.

  5. Drosophila homologue of Diaphanous 1 (DIAPH1) controls the metastatic potential of colon cancer cells by regulating microtubule-dependent adhesion.

    Science.gov (United States)

    Lin, Yuan-Na; Bhuwania, Ridhirama; Gromova, Kira; Failla, Antonio Virgilio; Lange, Tobias; Riecken, Kristoffer; Linder, Stefan; Kneussel, Matthias; Izbicki, Jakob R; Windhorst, Sabine

    2015-07-30

    Drosophila homologue of Diaphanous 1 (DIAPH1) regulates actin polymerization and microtubule (MT) stabilization upon stimulation with lysophosphatidic acid (LPA). Recently, we showed strongly reduced lung metastasis of DIAPH1-depleted colon cancer cells but we found accumulations of DIAPH1-depleted cells in bone marrow. Here, we analyzed possible organ- or tissue-specific metastasis of DIAPH1-depleted HCT-116 cells. Our data confirmed that depletion of DIAPH1 strongly inhibited lung metastasis and revealed that, in contrast to control cells, DIAPH1-depleted cells did not form metastases in further organs. Detailed mechanistic analysis on cells that were not stimulated with LPA to activate the cytoskeleton-modulating activity of DIAPH1, revealed that even under basal conditions DIAPH1 was essential for cellular adhesion to collagen. In non-stimulated cells DIAPH1 did not control actin dynamics but, interestingly, was essential for stabilization of microtubules (MTs). Additionally, DIAPH1 controlled directed vesicle trafficking and with this, local clustering of the adhesion protein integrin-β1 at the plasma membrane. Therefore, we conclude that under non-stimulating conditions DIAPH1 controls cellular adhesion by stabilizing MTs required for local clustering of integrin-β1 at the plasma membrane. Thus, blockade of DIAPH1-tubulin interaction may be a promising approach to inhibit one of the earliest steps in the metastatic cascade of colon cancer.

  6. Regeneration of Drosophila sensory neuron axons and dendrites is regulated by the Akt pathway involving Pten and microRNA bantam

    Science.gov (United States)

    Song, Yuanquan; Ori-McKenney, Kassandra M.; Zheng, Yi; Han, Chun; Jan, Lily Yeh; Jan, Yuh Nung

    2012-01-01

    Both cell-intrinsic and extrinsic pathways govern axon regeneration, but only a limited number of factors have been identified and it is not clear to what extent axon regeneration is evolutionarily conserved. Whether dendrites also regenerate is unknown. Here we report that, like the axons of mammalian sensory neurons, the axons of certain Drosophila dendritic arborization (da) neurons are capable of substantial regeneration in the periphery but not in the CNS, and activating the Akt pathway enhances axon regeneration in the CNS. Moreover, those da neurons capable of axon regeneration also display dendrite regeneration, which is cell type-specific, developmentally regulated, and associated with microtubule polarity reversal. Dendrite regeneration is restrained via inhibition of the Akt pathway in da neurons by the epithelial cell-derived microRNA bantam but is facilitated by cell-autonomous activation of the Akt pathway. Our study begins to reveal mechanisms for dendrite regeneration, which depends on both extrinsic and intrinsic factors, including the PTEN–Akt pathway that is also important for axon regeneration. We thus established an important new model system—the fly da neuron regeneration model that resembles the mammalian injury model—with which to study and gain novel insights into the regeneration machinery. PMID:22759636

  7. Identification of synaptic targets of Drosophila pumilio.

    Directory of Open Access Journals (Sweden)

    Gengxin Chen

    2008-02-01

    Full Text Available Drosophila Pumilio (Pum protein is a translational regulator involved in embryonic patterning and germline development. Recent findings demonstrate that Pum also plays an important role in the nervous system, both at the neuromuscular junction (NMJ and in long-term memory formation. In neurons, Pum appears to play a role in homeostatic control of excitability via down regulation of para, a voltage gated sodium channel, and may more generally modulate local protein synthesis in neurons via translational repression of eIF-4E. Aside from these, the biologically relevant targets of Pum in the nervous system remain largely unknown. We hypothesized that Pum might play a role in regulating the local translation underlying synapse-specific modifications during memory formation. To identify relevant translational targets, we used an informatics approach to predict Pum targets among mRNAs whose products have synaptic localization. We then used both in vitro binding and two in vivo assays to functionally confirm the fidelity of this informatics screening method. We find that Pum strongly and specifically binds to RNA sequences in the 3'UTR of four of the predicted target genes, demonstrating the validity of our method. We then demonstrate that one of these predicted target sequences, in the 3'UTR of discs large (dlg1, the Drosophila PSD95 ortholog, can functionally substitute for a canonical NRE (Nanos response element in vivo in a heterologous functional assay. Finally, we show that the endogenous dlg1 mRNA can be regulated by Pumilio in a neuronal context, the adult mushroom bodies (MB, which is an anatomical site of memory storage.

  8. Function and regulation of the Mediator complex.

    Science.gov (United States)

    Conaway, Ronald C; Conaway, Joan Weliky

    2011-04-01

    Over the past few years, advances in biochemical and genetic studies of the structure and function of the Mediator complex have shed new light on its subunit architecture and its mechanism of action in transcription by RNA polymerase II (pol II). The development of improved methods for reconstitution of recombinant Mediator subassemblies is enabling more in-depth analyses of basic features of the mechanisms by which Mediator interacts with and controls the activity of pol II and the general initiation factors. The discovery and characterization of multiple, functionally distinct forms of Mediator characterized by the presence or absence of the Cdk8 kinase module have led to new insights into how Mediator functions in both Pol II transcription activation and repression. Finally, progress in studies of the mechanisms by which the transcriptional activation domains (ADs) of DNA binding transcription factors target Mediator have brought to light unexpected complexities in the way Mediator participates in signal transduction. Copyright © 2011 Elsevier Ltd. All rights reserved.

  9. Glucocorticoid regulation of astrocytic fate and function.

    Directory of Open Access Journals (Sweden)

    Shuang Yu

    Full Text Available Glial loss in the hippocampus has been suggested as a factor in the pathogenesis of stress-related brain disorders that are characterized by dysregulated glucocorticoid (GC secretion. However, little is known about the regulation of astrocytic fate by GC. Here, we show that astrocytes derived from the rat hippocampus undergo growth inhibition and display moderate activation of caspase 3 after exposure to GC. Importantly, the latter event, observed both in situ and in primary astrocytic cultures is not followed by either early- or late-stage apoptosis, as monitored by stage I or stage II DNA fragmentation. Thus, unlike hippocampal granule neurons, astrocytes are resistant to GC-induced apoptosis; this resistance is due to lower production of reactive oxygen species (ROS and a greater buffering capacity against the cytotoxic actions of ROS. We also show that GC influence hippocampal cell fate by inducing the expression of astrocyte-derived growth factors implicated in the control of neural precursor cell proliferation. Together, our results suggest that GC instigate a hitherto unknown dialog between astrocytes and neural progenitors, adding a new facet to understanding how GC influence the cytoarchitecture of the hippocampus.

  10. TOR signaling pathway and autophagy are involved in the regulation of circadian rhythms in behavior and plasticity of L2 interneurons in the brain of Drosophila melanogaster.

    Science.gov (United States)

    Kijak, Ewelina; Pyza, Elżbieta

    2017-01-01

    Drosophila melanogaster is a common model used to study circadian rhythms in behavior and circadian clocks. However, numerous circadian rhythms have also been detected in non-clock neurons, especially in the first optic neuropil (lamina) of the fly's visual system. Such rhythms have been observed in the number of synapses and in the structure of interneurons, which exhibit changes in size and shape in a circadian manner. Although the patterns of these changes are known, the mechanism remains unclear. In the present study, we investigated the role of the TOR signaling pathway and autophagy in regulating circadian rhythms based on the behavior and structural plasticity of the lamina L2 monopolar cell dendritic trees. In addition, we examined the cyclic expression of the TOR signaling pathway (Tor, Pi3K class 1, Akt1) and autophagy (Atg5 and Atg7) genes in the fly's brain. We observed that Tor, Atg5 and Atg7 exhibit rhythmic expressions in the brain of wild-type flies in day/night conditions (LD 12:12) that are abolished in per01 clock mutants. The silencing of Tor in per expressing cells shortens a period of the locomotor activity rhythm of flies. In addition, silencing of the Tor and Atg5 genes in L2 cells disrupts the circadian plasticity of the L2 cell dendritic trees measured in the distal lamina. In turn, silencing of the Atg7 gene in L2 cells changes the pattern of this rhythm. Our results indicate that the TOR signaling pathway and autophagy are involved in the regulation of circadian rhythms in the behavior and plasticity of neurons in the brain of adult flies.

  11. Transforming Growth Factor β/Activin Signaling Functions as a Sugar-Sensing Feedback Loop to Regulate Digestive Enzyme Expression

    Directory of Open Access Journals (Sweden)

    Wen-bin Alfred Chng

    2014-10-01

    Full Text Available Summary: Organisms need to assess their nutritional state and adapt their digestive capacity to the demands for various nutrients. Modulation of digestive enzyme production represents a rational step to regulate nutriment uptake. However, the role of digestion in nutrient homeostasis has been largely neglected. In this study, we analyzed the mechanism underlying glucose repression of digestive enzymes in the adult Drosophila midgut. We demonstrate that glucose represses the expression of many carbohydrases and lipases. Our data reveal that the consumption of nutritious sugars stimulates the secretion of the transforming growth factor β (TGF-β ligand, Dawdle, from the fat body. Dawdle then acts via circulation to activate TGF-β/Activin signaling in the midgut, culminating in the repression of digestive enzymes that are highly expressed during starvation. Thus, our study not only identifies a mechanism that couples sugar sensing with digestive enzyme expression but points to an important role of TGF-β/Activin signaling in sugar metabolism. : Organisms modulate their digestive processes to reflect their nutritional state. In this study, Chng et al. demonstrate that the TGF-β/Activin pathway functions as a carbohydrate-sensing mechanism in the adult Drosophila midgut to regulate digestive enzyme expression. They show that the TGF-β ligand, Dawdle, and the canonical TGF-β/Activin signaling are essential to couple carbohydrate sensing with digestive enzyme expression. Thus, their study highlights an unexpected function of TGF-β/Activin signaling that is beyond their established roles in development and immunity.

  12. Emerging Regulation and Function of Betatrophin

    Directory of Open Access Journals (Sweden)

    Yi-Hsin Tseng

    2014-12-01

    Full Text Available Betatrophin, also known as TD26/RIFL/lipasin/ANGPTL8/C19orf80, is a novel protein predominantly expressed in human liver. To date, several betatrophin orthologs have been identified in mammals. Increasing evidence has revealed an association between betatrophin expression and serum lipid profiles, particularly in patients with obesity or diabetes. Stimulators of betatrophin, such as insulin, thyroid hormone, irisin and caloric intake, are usually relevant to energy expenditure or thermogenesis. In murine models, serum triglyceride levels as well as pancreatic cell proliferation are potently enhanced by betatrophin. Intriguingly, conflicting phenomena have also been reported that betatrophin suppresses hepatic triglyceride levels, suggesting that betatrophin function is mediated by complex regulatory processes. However, its precise physiological role remains unclear at present. In this review, we have summarized the current findings on betatrophin and their implications.

  13. Emerging regulation and function of betatrophin.

    Science.gov (United States)

    Tseng, Yi-Hsin; Yeh, Yung-Hsin; Chen, Wei-Jan; Lin, Kwang-Huei

    2014-12-18

    Betatrophin, also known as TD26/RIFL/lipasin/ANGPTL8/C19orf80, is a novel protein predominantly expressed in human liver. To date, several betatrophin orthologs have been identified in mammals. Increasing evidence has revealed an association between betatrophin expression and serum lipid profiles, particularly in patients with obesity or diabetes. Stimulators of betatrophin, such as insulin, thyroid hormone, irisin and caloric intake, are usually relevant to energy expenditure or thermogenesis. In murine models, serum triglyceride levels as well as pancreatic cell proliferation are potently enhanced by betatrophin. Intriguingly, conflicting phenomena have also been reported that betatrophin suppresses hepatic triglyceride levels, suggesting that betatrophin function is mediated by complex regulatory processes. However, its precise physiological role remains unclear at present. In this review, we have summarized the current findings on betatrophin and their implications.

  14. Neurotrophin signaling endosomes; biogenesis, regulation, and functions

    Science.gov (United States)

    Yamashita, Naoya; Kuruvilla, Rejji

    2016-01-01

    In the nervous system, communication between neurons and their post-synaptic target cells is critical for the formation, refinement and maintenance of functional neuronal connections. Diffusible signals secreted by target tissues, exemplified by the family of neurotrophins, impinge on nerve terminals to influence diverse developmental events including neuronal survival and axonal growth. Key mechanisms of action of target-derived neurotrophins include the cell biological processes of endocytosis and retrograde trafficking of their Trk receptors from growth cones to cell bodies. In this review, we summarize the molecular mechanisms underlying this endosome-mediated signaling, focusing on the instructive role of neurotrophin signaling itself in directing its own trafficking. Recent studies have linked impaired neurotrophin trafficking to neurodevelopmental disorders, highlighting the relevance of neurotrophin endosomes in human health. PMID:27327126

  15. Exercise and the Regulation of Immune Functions.

    Science.gov (United States)

    Simpson, Richard J; Kunz, Hawley; Agha, Nadia; Graff, Rachel

    2015-01-01

    Exercise has a profound effect on the normal functioning of the immune system. It is generally accepted that prolonged periods of intensive exercise training can depress immunity, while regular moderate intensity exercise is beneficial. Single bouts of exercise evoke a striking leukocytosis and a redistribution of effector cells between the blood compartment and the lymphoid and peripheral tissues, a response that is mediated by increased hemodynamics and the release of catecholamines and glucocorticoids following the activation of the sympathetic nervous system and the hypothalamic-pituitary-adrenal axis. Single bouts of prolonged exercise may impair T-cell, NK-cell, and neutrophil function, alter the Type I and Type II cytokine balance, and blunt immune responses to primary and recall antigens in vivo. Elite athletes frequently report symptoms associated with upper respiratory tract infections (URTI) during periods of heavy training and competition that may be due to alterations in mucosal immunity, particularly reductions in secretory immunoglobulin A. In contrast, single bouts of moderate intensity exercise are "immuno-enhancing" and have been used to effectively increase vaccine responses in "at-risk" patients. Improvements in immunity due to regular exercise of moderate intensity may be due to reductions in inflammation, maintenance of thymic mass, alterations in the composition of "older" and "younger" immune cells, enhanced immunosurveillance, and/or the amelioration of psychological stress. Indeed, exercise is a powerful behavioral intervention that has the potential to improve immune and health outcomes in the elderly, the obese, and patients living with cancer and chronic viral infections such as HIV. © 2015 Elsevier Inc. All rights reserved.

  16. Many ways to be small: different environmental regulators of size generate distinct scaling relationships in Drosophila melanogaster

    OpenAIRE

    Shingleton, Alexander W.; Estep, Chad M.; Driscoll, Michael V.; Dworkin, Ian

    2009-01-01

    Static allometries, the scaling relationship between body and trait size, describe the shape of animals in a population or species, and are generated in response to variation in genetic or environmental regulators of size. In principle, allometries may vary with the different size regulators that generate them, which can be problematic since allometric differences are also used to infer patterns of selection on morphology. We test this hypothesis by examining the patterns of scaling in Drosop...

  17. Unraveling the Neurobiology of Sleep and Sleep Disorders Using Drosophila.

    Science.gov (United States)

    Chakravarti, L; Moscato, E H; Kayser, M S

    2017-01-01

    Sleep disorders in humans are increasingly appreciated to be not only widespread but also detrimental to multiple facets of physical and mental health. Recent work has begun to shed light on the mechanistic basis of sleep disorders like insomnia, restless legs syndrome, narcolepsy, and a host of others, but a more detailed genetic and molecular understanding of how sleep goes awry is lacking. Over the past 15 years, studies in Drosophila have yielded new insights into basic questions regarding sleep function and regulation. More recently, powerful genetic approaches in the fly have been applied toward studying primary human sleep disorders and other disease states associated with dysregulated sleep. In this review, we discuss the contribution of Drosophila to the landscape of sleep biology, examining not only fundamental advances in sleep neurobiology but also how flies have begun to inform pathological sleep states in humans. © 2017 Elsevier Inc. All rights reserved.

  18. Olfactory memory traces in Drosophila.

    Science.gov (United States)

    Berry, Jacob; Krause, William C; Davis, Ronald L

    2008-01-01

    In Drosophila, the fruit fly, coincident exposure to an odor and an aversive electric shock can produce robust behavioral memory. This behavioral memory is thought to be regulated by cellular memory traces within the central nervous system of the fly. These molecular, physiological, or structural changes in neurons, induced by pairing odor and shock, regulate behavior by altering the neurons' response to the learned environment. Recently, novel in vivo functional imaging techniques have allowed researchers to observe cellular memory traces in intact animals. These investigations have revealed interesting temporal and spatial dynamics of cellular memory traces. First, a short-term cellular memory trace was discovered that exists in the antennal lobe, an early site of olfactory processing. This trace represents the recruitment of new synaptic activity into the odor representation and forms for only a short period of time just after training. Second, an intermediate-term cellular memory trace was found in the dorsal paired medial neuron, a neuron thought to play a role in stabilizing olfactory memories. Finally, a long-term protein synthesis-dependent cellular memory trace was discovered in the mushroom bodies, a structure long implicated in olfactory learning and memory. Therefore, it appears that aversive olfactory associations are encoded by multiple cellular memory traces that occur in different regions of the brain with different temporal domains.

  19. Adaptive Evolution of Gene Expression in Drosophila.

    Science.gov (United States)

    Nourmohammad, Armita; Rambeau, Joachim; Held, Torsten; Kovacova, Viera; Berg, Johannes; Lässig, Michael

    2017-08-08

    Gene expression levels are important quantitative traits that link genotypes to molecular functions and fitness. In Drosophila, population-genetic studies have revealed substantial adaptive evolution at the genomic level, but the evolutionary modes of gene expression remain controversial. Here, we present evidence that adaptation dominates the evolution of gene expression levels in flies. We show that 64% of the observed expression divergence across seven Drosophila species are adaptive changes driven by directional selection. Our results are derived from time-resolved data of gene expression divergence across a family of related species, using a probabilistic inference method for gene-specific selection. Adaptive gene expression is stronger in specific functional classes, including regulation, sensory perception, sexual behavior, and morphology. Moreover, we identify a large group of genes with sex-specific adaptation of expression, which predominantly occurs in males. Our analysis opens an avenue to map system-wide selection on molecular quantitative traits independently of their genetic basis. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.