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Mutant Flies (mutant + fly)
Selected AbstractsThe Drosophila nucleoporin gene nup154 is required for correct microfilament dynamics and cell death during oogenesisCYTOSKELETON, Issue 8 2007Maria Giovanna Riparbelli Abstract The Drosophila nucleoporin gene nup154 is required in both male and female germline for successful gametogenesis. Mutant flies lack differentiated sperm and lay abnormal eggs. We demonstrated that the egg phenotype was associated with specific alterations of the actin cytoskeleton at different stages of oogenesis. Actually, mutant egg chambers displayed an abnormal organization of both subcortical microfilaments and cytoplasmic actin bundles, that led to defective nurse cell dumping. TUNEL analysis also showed that the dumpless phenotype was associated with delayed apoptosis. The nup154 gene product was localized by conventional immunofluorescence microscopy to the nuclear envelope in a distinct punctuate pattern, characteristic of nuclear pore complex components. TEM analysis revealed that the protein was mainly distributed along filamentous structures that extended radially on the nuclear side of the pore, suggesting that Nup154 could be an integral component of the basket filaments associated with the nuclear pore complexes. We propose that Nup154 is necessary for correct nuclear pore complex functions and that the proper regulation of the actin cytoskeleton dynamics strongly relies upon nuclear pore integrity. Cell Motil. Cytoskeleton 2007. © 2007 Wiley-Liss, Inc. [source] Nucleolar colocalization of TAF1 and testis-specific TAFs during Drosophila spermatogenesisDEVELOPMENTAL DYNAMICS, Issue 10 2007Chad E. Metcalf Abstract In Drosophila, testis-specific TBP-associated factors (tTAFs) predominantly localize to spermatocyte nucleoli and regulate the transcription of genes necessary for spermatocyte entry into meiosis. tTAFs are paralogs of generally expressed TAF subunits of transcription factor IID (TFIID). Our recent observation that the generally expressed TAF1 isoform TAF1-2 is greatly enriched in testes prompted us to explore the functional relationship between general TAFs and tTAFs during spermatogenesis. Analysis by immunofluorescence microscopy revealed that among the general TFIID subunits examined (TATA-box binding protein [TBP], TAF1, TAF4, TAF5, and TAF9), only TAF1 colocalized with the tTAF Mia in spermatocyte nucleoli. Nucleolar localization of TAF1, but not Mia, was disrupted in tTAF mutant flies, and TAF1 dissociated from DNA prior to Mia as spermatocytes entered meiosis. Taken together, our results suggest stepwise assembly of a testis-specific TFIID complex (tTFIID) whereby a TAF1 isoform, presumably TAF1-2, is recruited to a core subassembly of tTAFs in spermatocyte nucleoli. Developmental Dynamics 236:2836,2843, 2007. © 2007 Wiley-Liss, Inc. [source] A succession of anesthetic endpoints in the Drosophila brainDEVELOPMENTAL NEUROBIOLOGY, Issue 11 2006Bruno van Swinderen Abstract General anesthetics abolish behavioral responsiveness in all animals, and in humans this is accompanied by loss of consciousness. Whether similar target mechanisms and behavioral endpoints exist across species remains controversial, although model organisms have been successfully used to study mechanisms of anesthesia. In Drosophila, a number of key mutants have been characterized as hypersensitive or resistant to general anesthetics by behavioral assays. In order to investigate general anesthesia in the Drosophila brain, local field potential (LFP) recordings were made during incremental exposures to isoflurane in wild-type and mutant flies. As in higher animals, general anesthesia in flies was found to involve a succession of distinct endpoints. At low doses, isoflurane uncoupled brain activity from ongoing movement, followed by a sudden attenuation in neural correlates of perception. Average LFP activity in the brain was more gradually attenuated with higher doses, followed by loss of movement behavior. Among mutants, a strong correspondence was found between behavioral and LFP sensitivities, thereby suggesting that LFP phenotypes are proximal to the anesthetic's mechanism of action. Finally, genetic and pharmacological analysis revealed that anesthetic sensitivities in the fly brain are, like other arousal states, influenced by dopaminergic activity. These results suggest that volatile anesthetics such as isoflurane may target the same processes that sustain wakefulness and attention in the brain. LFP correlates of general anesthesia in Drosophila provide a powerful new approach to uncovering the nature of these processes. © 2006 Wiley Periodicals, Inc. J Neurobiol 66: 1195,1211, 2006 [source] The Drosophila PRR GNBP3 assembles effector complexes involved in antifungal defenses independently of its Toll -pathway activation functionEUROPEAN JOURNAL OF IMMUNOLOGY, Issue 5 2010Alexey A. Matskevich Abstract The Drosophila Toll -signaling pathway controls the systemic antifungal host response. Gram-negative binding protein 3 (GNBP3), a member of the ,-glucan recognition protein family senses fungal infections and activates this pathway. A second detection system perceives the activity of proteolytic fungal virulence factors and redundantly activates Toll. GNBP3hades mutant flies succumb more rapidly to Candida albicans and to entomopathogenic fungal infections than WT flies, despite normal triggering of the Toll pathway via the virulence detection system. These observations suggest that GNBP3 triggers antifungal defenses that are not dependent on activation of the Toll pathway. Here, we show that GNBP3 agglutinates fungal cells. Furthermore, it can activate melanization in a Toll -independent manner. Melanization is likely to be an essential defense against some fungal infections given that the entomopathogenic fungus Beauveria bassiana inhibits the activity of the main melanization enzymes, the phenol oxidases. Finally, we show that GNBP3 assembles "attack complexes", which comprise phenoloxidase and the necrotic serpin. We propose that Drosophila GNBP3 targets fungi immediately at the inception of the infection by bringing effector molecules in direct contact with the invading microorganisms. [source] Adenylyl cyclase encoded by AC78C participates in sugar perception in Drosophila melanogasterEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 10 2008Kohei Ueno Abstract In gustatory receptor neurons (GRNs) in Drosophila melanogaster, Gr5a and one of the Gr64s encode sugar receptors with seven transmembrane domains. Previously, we have shown that the responses to various sugars are depressed in DGs, mutant flies (Ueno et al., 2006). Because DGs, is a homolog of Gs, we hypothesized that the sugar receptors are coupled to adenylyl cyclase (AC) in Drosophila. The aim of this study was to identify the AC that participates in sugar perception. Here, we found that an AC inhibitor, MDL-12330A, depressed the response in GRNs to trehalose as well as sucrose; that an AC gene, AC78C, was expressed in the sugar-sensitive GRNs; that RNAi against AC78C depressed the electrical response in GRNs to sucrose; and that the sugar response in GRNs, as well as sugar intake in a behavioral assay in an AC78C mutant, was depressed at low sugar concentrations. We conclude that AC78C, via cAMP, participates in the sugar-taste signaling pathway at the low concentration range. [source] In vivo potentiation of human oestrogen receptor , by Cdk7-mediated phosphorylationGENES TO CELLS, Issue 10 2004Saya Ito Phosphorylation of the Ser118 residue in the N-terminal A/B domain of the human oestrogen receptor , (hER,) by mitogen-activated protein kinase (MAPK), stimulated via growth factor signalling pathways, is known to potentiate ER, ligand-induced transactivation function. Besides MAPK, cyclin dependent kinase 7 (Cdk7) in the TFIIH complex has also been found to potentiate hER, transactivation in vitro through Ser118 phosphorylation. To investigate an impact of Cdk7 on hER, transactivation in vivo, we assessed activity of hER, in a wild-type and cdk7 inactive mutant Drosophila that ectopically expressed hER, in the eye disc. Ectopic expression of the wild-type or mutant receptors, together with a green fluorescent protein (GFP) reporter gene, allowed us to demonstrate that hER, expressed in the fly tissues was transcriptionally functional and adequately responded to hER, ligands in the patterns similar to those observed in mammalian cells. Replacement of Ser118 with alanine in hER, (S118A mutant) significantly reduced the ligand-induced hER, transactivation function. Importantly, while in cdk7 inactive mutant Drosophila the wild-type hER, exhibited reduced response to the ligand; levels of transactivation by the hER, S118A mutant were not affected in these inactive cdk7 mutant flies. Furthermore, phosphorylation of hER, at Ser118 has been observed in vitro by both human and Drosophila Cdk7. Our findings demonstrate that Cdk7 is involved in regulation of the ligand-induced transactivation function of hER,in vivo via Ser118 phosphorylation. [source] Genetic Control of Acute Ethanol-Induced Behaviors in DrosophilaALCOHOLISM, Issue 8 2000Carol M. Singh Background: In most organisms in which acute ethanol exposure has been studied, it leads to similar changes in behavior. Generally, low ethanol doses activate the central nervous system, whereas high doses are sedative. Sensitivity to the acute intoxicating effects of ethanol is in part under genetic control in rodents and humans, and reduced sensitivity in humans predicts the development of alcoholism (Crabbe et al., 1994; Schuckit, 1994). We have established Drosophila melanogaster as a model organism to study the mechanisms that regulate acute sensitivity to ethanol. Methods: We measured the effects of ethanol vapor on Drosophila locomotor behaviors by using three different assays. Horizontal locomotion was quantified in a locomotor chamber, turning behavior was assayed in narrow tubes, and ethanol-induced loss of postural control was measured in an inebriometer. Mutants with altered sensitivity to the acute effects of ethanol were generated by treatment with ethyl methane sulfonate and isolated by selection in the inebriometer. We ascertained the effects of these mutations on ethanol pharmacokinetics by measuring ethanol levels in extracts of flies at various times during and after ethanol exposure. Results: Among nearly 30,000 potentially mutant flies tested, we isolated 19 mutant strains with reduced and 4 strains with increased sensitivity to the acute effects of ethanol as measured in the inebriometer. Of these mutants, four showed changes in ethanol absorption. Two mutants, named barfly and tipsy to reflect their reduced and increased ethanol sensitivity in the inebriometer, respectively, were analyzed for locomotor behaviors. Both mutants exhibited ethanol-induced hyperactivity that was indistinguishable from wild type. However, barfly and tipsy displayed reduced and increased sensitivity to the sedative effects of ethanol, respectively. Finally, both mutants showed an increased rate of ethanol-induced turning behavior. Conclusions: The effects of acute ethanol exposure on Drosophila locomotor behaviors are remarkably similar to those described for mammals. The analysis of mutants with altered sensitivity to ethanol revealed that the genetic pathways which regulate these responses are complex and that single genes can affect hyperactivity, turning, and sedation independently. [source] Interaction of proteins involved in ecdysone and juvenile hormone signal transduction,ARCHIVES OF INSECT BIOCHEMISTRY AND PHYSIOLOGY (ELECTRONIC), Issue 2 2009Kavita Bitra Abstract Ecdysteroids and juvenile hormones (JH) regulate a variety of developmental, physiological, behavioral, and metabolic processes. Ecdysteroids function through a heterodimeric complex of two nuclear receptors, ecdysone receptor (EcR) and ultraspiracle (USP). An 85 kDa protein identified in Drosophila melanogaster methoprene-tolerant (Met) mutant binds to JH III with high affinity, and the mutant flies are resistant to juvenile hormone analog (JHA), methoprene. Reporter assays using the yeast two-hybrid system were performed in order to study the molecular interactions between EcR, USP and Met. As expected, EcR fused to the B42 activation domain and USP fused to the LexA DNA binding domain interacted with each other and supported induction of the reporter gene in the presence of stable ecdysteroid analog, RG-102240 or steroids, muristerone A and ponasterone A. The USP:USP homodimers supported expression of the reporter gene in the absence of ligand, and there was no significant increase in the reporter activity after addition of a JHA, methoprene. Similarly, Met:Met homodimers as well as Met:EcR and Met:USP heterodimers induced reporter activity in the absence of ligand and addition of ecdysteroid or JH analogs did not increase the reporter activity regulated by either homodimers or heterodimers of Met protein. Two-hybrid assays in insect cells and in vitro pull-down assays confirmed the interaction of Met with EcR and USP. These data suggest that the proteins that are involved in signal transduction of ecdysteroids (EcR and USP) and juvenile hormones (Met) interact to mediate cross-talk between these two important hormones. Arch. Insect Biochem. Physiol. 2008. © 2008 Wiley-Liss, Inc. [source] | ||||||||||||||||||||||