Inhalation toxicity of indoor air pollutants in Drosophila melanogaster using integrated transcriptomics and computational behavior analyses

NASA Astrophysics Data System (ADS)

Eom, Hyun-Jeong; Liu, Yuedan; Kwak, Gyu-Suk; Heo, Muyoung; Song, Kyung Seuk; Chung, Yun Doo; Chon, Tae-Soo; Choi, Jinhee

2017-06-01

We conducted an inhalation toxicity test on the alternative animal model, Drosophila melanogaster, to investigate potential hazards of indoor air pollution. The inhalation toxicity of toluene and formaldehyde was investigated using comprehensive transcriptomics and computational behavior analyses. The ingenuity pathway analysis (IPA) based on microarray data suggests the involvement of pathways related to immune response, stress response, and metabolism in formaldehyde and toluene exposure based on hub molecules. We conducted a toxicity test using mutants of the representative genes in these pathways to explore the toxicological consequences of alterations of these pathways. Furthermore, extensive computational behavior analysis showed that exposure to either toluene or formaldehyde reduced most of the behavioral parameters of both wild-type and mutants. Interestingly, behavioral alteration caused by toluene or formaldehyde exposure was most severe in the p38b mutant, suggesting that the defects in the p38 pathway underlie behavioral alteration. Overall, the results indicate that exposure to toluene and formaldehyde via inhalation causes severe toxicity in Drosophila, by inducing significant alterations in gene expression and behavior, suggesting that Drosophila can be used as a potential alternative model in inhalation toxicity screening.

Inhalation toxicity of indoor air pollutants in Drosophila melanogaster using integrated transcriptomics and computational behavior analyses

PubMed Central

Eom, Hyun-Jeong; Liu, Yuedan; Kwak, Gyu-Suk; Heo, Muyoung; Song, Kyung Seuk; Chung, Yun Doo; Chon, Tae-Soo; Choi, Jinhee

2017-01-01

We conducted an inhalation toxicity test on the alternative animal model, Drosophila melanogaster, to investigate potential hazards of indoor air pollution. The inhalation toxicity of toluene and formaldehyde was investigated using comprehensive transcriptomics and computational behavior analyses. The ingenuity pathway analysis (IPA) based on microarray data suggests the involvement of pathways related to immune response, stress response, and metabolism in formaldehyde and toluene exposure based on hub molecules. We conducted a toxicity test using mutants of the representative genes in these pathways to explore the toxicological consequences of alterations of these pathways. Furthermore, extensive computational behavior analysis showed that exposure to either toluene or formaldehyde reduced most of the behavioral parameters of both wild-type and mutants. Interestingly, behavioral alteration caused by toluene or formaldehyde exposure was most severe in the p38b mutant, suggesting that the defects in the p38 pathway underlie behavioral alteration. Overall, the results indicate that exposure to toluene and formaldehyde via inhalation causes severe toxicity in Drosophila, by inducing significant alterations in gene expression and behavior, suggesting that Drosophila can be used as a potential alternative model in inhalation toxicity screening. PMID:28621308

A novel, tissue-specific, Drosophila homeobox gene.

PubMed

Barad, M; Jack, T; Chadwick, R; McGinnis, W

1988-07-01

The homeobox gene family of Drosophila appears to control a variety of position-specific patterning decisions during embryonic and imaginal development. Most of these patterning decisions determine groups of cells on the anterior-posterior axis of the Drosophila germ band. We have isolated a novel homeobox gene from Drosophila, designated H2.0. H2.0 has the most diverged homeobox so far characterized in metazoa, and, in contrast to all previously isolated homeobox genes, H2.0 exhibits a tissue-specific pattern of expression. The cells that accumulate transcripts for this novel gene correspond to the visceral musculature and its anlagen.

The P38alpha and P38delta MAP kinases may be gene therapy targets in the future treatment of severe burns.

PubMed

Wang, Shuyun; Huang, Qiaobing; Guo, Xiaohua; Brunk, Ulf T; Han, Jiahuai; Zhao, Keseng; Zhao, Ming

2010-08-01

Microvascular barrier damage, induced by thermal injury, imposes life-threatening problems owing to the pathophysiological consequences of plasma loss and impaired perfusion that finally may lead to multiple organ failure. The aim of the present study was to define the signaling role of selected mitogen-activated protein kinases (MAPKs) in general vessel hyperpermeability caused by burns and to look for a potential gene therapy. Rearrangement of cytoskeletons and cell tight junctions were evaluated by phalloidin labeling of actin and immunocytochemical demonstration of the ZO-1 protein, whereas blood vessel permeability was evaluated by a fluorescence ratio technique. The p38 MAPK inhibitor SB203580 largely blocked burn serum-induced stress-fiber formation and tight-junction damage. Using the adenoviral approach to transfect dominant negative forms of p38 MAPKs, we found that p38alpha and p38delta had similar effects. The in vivo part of the study showed that transfection of these two constructs significantly lowered general venular hyperpermeability and enhanced the survival of burned animals. Because the p38 MAPK pathway seems to play a crucial role in burn-induced vascular hyperpermeability, general transfection with p38 MAP dominant negative constructs might become a new therapeutic method to block burn-induced plasma leakage.

Dynamics of Wolbachia pipientis Gene Expression Across the Drosophila melanogaster Life Cycle

PubMed Central

Gutzwiller, Florence; Carmo, Catarina R.; Miller, Danny E.; Rice, Danny W.; Newton, Irene L. G.; Hawley, R. Scott; Teixeira, Luis; Bergman, Casey M.

2015-01-01

Symbiotic interactions between microbes and their multicellular hosts have manifold biological consequences. To better understand how bacteria maintain symbiotic associations with animal hosts, we analyzed genome-wide gene expression for the endosymbiotic α-proteobacteria Wolbachia pipientis across the entire life cycle of Drosophila melanogaster. We found that the majority of Wolbachia genes are expressed stably across the D. melanogaster life cycle, but that 7.8% of Wolbachia genes exhibit robust stage- or sex-specific expression differences when studied in the whole-organism context. Differentially-expressed Wolbachia genes are typically up-regulated after Drosophila embryogenesis and include many bacterial membrane, secretion system, and ankyrin repeat-containing proteins. Sex-biased genes are often organized as small operons of uncharacterized genes and are mainly up-regulated in adult Drosophila males in an age-dependent manner. We also systematically investigated expression levels of previously-reported candidate genes thought to be involved in host-microbe interaction, including those in the WO-A and WO-B prophages and in the Octomom region, which has been implicated in regulating bacterial titer and pathogenicity. Our work provides comprehensive insight into the developmental dynamics of gene expression for a widespread endosymbiont in its natural host context, and shows that public gene expression data harbor rich resources to probe the functional basis of the Wolbachia-Drosophila symbiosis and annotate the transcriptional outputs of the Wolbachia genome. PMID:26497146

Copper homeostasis gene discovery in Drosophila melanogaster.

PubMed

Norgate, Melanie; Southon, Adam; Zou, Sige; Zhan, Ming; Sun, Yu; Batterham, Phil; Camakaris, James

2007-06-01

Recent studies have shown a high level of conservation between Drosophila melanogaster and mammalian copper homeostasis mechanisms. These studies have also demonstrated the efficiency with which this species can be used to characterize novel genes, at both the cellular and whole organism level. As a versatile and inexpensive model organism, Drosophila is also particularly useful for gene discovery applications and thus has the potential to be extremely useful in identifying novel copper homeostasis genes and putative disease genes. In order to assess the suitability of Drosophila for this purpose, three screening approaches have been investigated. These include an analysis of the global transcriptional response to copper in both adult flies and an embryonic cell line using DNA microarray analysis. Two mutagenesis-based screens were also utilized. Several candidate copper homeostasis genes have been identified through this work. In addition, the results of each screen were carefully analyzed to identify any factors influencing efficiency and sensitivity. These are discussed here with the aim of maximizing the efficiency of future screens and the most suitable approaches are outlined. Building on this information, there is great potential for the further use of Drosophila for copper homeostasis gene discovery.

Human intellectual disability genes form conserved functional modules in Drosophila.

PubMed

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

2013-10-01

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.

Akirins, highly conserved nuclear proteins, required for NF-κB dependent gene expression in Drosophila and mice

PubMed Central

Goto, Akira; Matsushita, Kazufumi; Gesellchen, Viola; Chamy, Laure El; Kuttenkeuler, David; Takeuchi, Osamu; Hoffmann, Jules A.; Akira, Shizuo; Boutros, Michael; Reichhart, Jean-Marc

2009-01-01

During a genome-wide RNAi screen, we isolated CG8580 as a gene involved in the innate immune response of Drosophila. CG8580, which we named Akirin, acts in parallel with the NF-κB transcription factor downstream of the Imd pathway and was required for defense against Gram-negative bacteria. Akirin is highly conserved and the human genome contains two homologues, one of which was able to rescue the loss of function phenotype in Drosophila cells. Akirins had a strict nuclear localization. Knockout of both Akirin homologues in mice revealed that one had an essential function downstream of Toll-like receptor, tumor necrosis factor and interleukin 1-β (IL-1β) signaling pathways leading to the production of IL-6. Thus, Akirin is a conserved nuclear factor required for innate immune responses. PMID:18066067

Gene expression profile change and growth inhibition in Drosophila larvae treated with azadirachtin.

PubMed

Lai, Duo; Jin, Xiaoyong; Wang, Hao; Yuan, Mei; Xu, Hanhong

2014-09-20

Azadirachtin is a botanical insecticide that affects various biological processes. The effects of azadirachtin on the digital gene expression profile and growth inhibition in Drosophila larvae have not been investigated. In this study, we applied high-throughput sequencing technology to detect the differentially expressed genes of Drosophila larvae regulated by azadirachtin. A total of 15,322 genes were detected, and 28 genes were found to be significantly regulated by azadirachtin. Biological process and pathway analysis showed that azadirachtin affected starch and sucrose metabolism, defense response, signal transduction, instar larval or pupal development, and chemosensory behavior processes. The genes regulated by azadirachtin were mainly enriched in starch and sucrose metabolism. This study provided a general digital gene expression profile of dysregulated genes in response to azadirachtin and showed that azadirachtin provoked potent growth inhibitory effects in Drosophila larvae by regulating the genes of cuticular protein, amylase, and odorant-binding protein. Finally, we propose a potential mechanism underlying the dysregulation of the insulin/insulin-like growth factor signaling pathway by azadirachtin. Copyright © 2014 Elsevier B.V. All rights reserved.

Functional requirements driving the gene duplication in 12 Drosophila species.

PubMed

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.

A Comparative Study of WASP-67 b and HAT-P-38 b from WFC3 Data

NASA Astrophysics Data System (ADS)

Bruno, Giovanni; Lewis, Nikole K.; Stevenson, Kevin B.; Filippazzo, Joseph; Hill, Matthew; Fraine, Jonathan D.; Wakeford, Hannah R.; Deming, Drake; Kilpatrick, Brian; Line, Michael R.; Morley, Caroline V.; Collins, Karen A.; Conti, Dennis M.; Garlitz, Joseph; Rodriguez, Joseph E.

2018-02-01

Atmospheric temperature and planetary gravity are thought to be the main parameters affecting cloud formation in giant exoplanet atmospheres. Recent attempts to understand cloud formation have explored wide regions of the equilibrium temperature-gravity parameter space. In this study, we instead compare the case of two giant planets with nearly identical equilibrium temperature (T eq ∼ 1050 K) and gravity (g ∼ 10 m s‑1). During HST Cycle 23, we collected WFC3/G141 observations of the two planets, WASP-67 b and HAT-P-38 b. HAT-P-38 b, with mass 0.42 M J and radius 1.4 R J, exhibits a relatively clear atmosphere with a clear detection of water. We refine the orbital period of this planet with new observations, obtaining P = 4.6403294 ± 0.0000055 days. WASP-67 b, with mass 0.27 M J and radius 0.83 R J, shows a more muted water absorption feature than that of HAT-P-38 b, indicating either a higher cloud deck in the atmosphere or a more metal-rich composition. The difference in the spectra supports the hypothesis that giant exoplanet atmospheres carry traces of their formation history. Future observations in the visible and mid-infrared are needed to probe the aerosol properties and constrain the evolutionary scenario of these planets.

Gene expression profiles of Drosophila melanogaster exposed to an insecticidal extract of Piper nigrum.

PubMed

Jensen, Helen R; Scott, Ian M; Sims, Steve; Trudeau, Vance L; Arnason, John Thor

2006-02-22

Black pepper, Piper nigrum L. (Piperaceae), has insecticidal properties and could potentially be utilized as an alternative to synthetic insecticides. Piperine extracted from P. nigrum has a biphasic effect upon cytochrome P450 monooxygenase activity with an initial suppression followed by induction. In this study, an ethyl acetate extract of P. nigrum seeds was tested for insecticidal activity toward adult Musca domestica and Drosophila melanogaster. The effect of this same P. nigrum extract upon differential gene expression in D. melanogaster was investigated using cDNA microarray analysis of 7380 genes. Treatment of D. melanogaster with P. nigrum extract led to a greater than 2-fold upregulation of transcription of the cytochrome P450 phase I metabolism genes Cyp 6a8, Cyp 9b2, and Cyp 12d1 as well as the glutathione-S-transferase phase II metabolism gene Gst-S1. These data suggests a complex effect of P. nigrum upon toxin metabolism.

Human Intellectual Disability Genes Form Conserved Functional Modules in Drosophila

PubMed Central

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

Oestrogen exerts anti-inflammation via p38 MAPK/NF-κB cascade in adipocytes.

PubMed

Mu, Pan-Wei; Jiang, Ping; Wang, Man-Man; Chen, Yan-Ming; Zheng, Shu-Hui; Tan, Zhi; Jiang, Wei; Zeng, Long-Yi; Wang, Ting-Huai

Oestrogen has anti-inflammatory property in obesity. However, the mechanism is still not defined. To investigate the effect of oestrogen on LPS-induced monocyte chemoattractant protein-1 (MCP-1) production in adipocytes. Lipopolysaccharides (LPS) was used to imitate inflammatory responses and monocyte chemotactic protein-1 (MCP-1) was selected as an inflammatory marker to observe. 17β-Estradiol (E 2 ), SB203580 (SB), pyrrolidine dithiocarbamate (PDTC), pertussis toxin (PTX), wortmannin (WM), p65 siRNA and p38 MAPK siRNA were pre-treated respectively or together in LPS-induced MCP-1. Then p38 MAPK and NF-κB cascade were silenced successively to observe the change of each other. Lastly, oestrogen receptor (ER) α agonist, ERβ agonist and ER antagonist were utilised. LPS-induced MCP-1 largely impaired by pre-treatment with E 2 , SB, PDTC or silencing NF-κB subunit. E 2 inhibited LPS-induced MCP-1 in a time- and dose-dependent manner, which was related to the suppression of p65 translocation to nucleus. Furthermore, LPS rapidly activated p38 MAPK, while E 2 markedly inhibited this activation. It markedly attenuated LPS-stimulated p65 translocation to nucleus and MCP-1 production by transfecting with p38 MAPK siRNA or using p38 MAPK inhibitor. The oestrogen's inhibitory effect was mimicked by the ERα agonist, but not by the ERβ agonist. The inhibition of E 2 on p38 MAPK phosphorylation was prevented by ER antagonist. E 2 inhibits LPS-stimulated MCP-1 in adipocytes. This effect is related to the inhibition of p38 MAPK/NF-κB cascade, and ERα appears to be the dominant ER subtype in these events. Copyright © 2016 Asia Oceania Association for the Study of Obesity. Published by Elsevier Ltd. All rights reserved.

T-Box Genes in Drosophila Mesoderm Development.

PubMed

Reim, I; Frasch, M; Schaub, C

2017-01-01

In Drosophila there are eight genes encoding transcription factors of the T-box family, which are known to exert a variety of crucial developmental functions during ectodermal patterning processes, neuronal cell specification, mesodermal tissue development, and the development of extraembryonic tissues. In this review, we focus on the prominent roles of Drosophila T-box genes in mesodermal tissues. First, we describe the contributions of brachyenteron (byn) and optomotor-blind-related-gene-1 (org-1) to the development of the visceral mesoderm. Second, we provide an overview on the functions of the three Dorsocross paralogs (Doc1-3) and the two Tbx20-related paralogs (midline and H15) during Drosophila heart development. Third, we portray the roles of org-1 and midline/H15 in the specification of individual body wall and organ-attached muscles, including the function of org-1 in the transdifferentiation of certain heart-attached muscles during metamorphosis. The functional analysis of these evolutionarily conserved T-box genes, along with their interactions with other types of transcription factors and various signaling pathways, has provided key insights into the regulation of Drosophila visceral mesoderm, muscle, and heart development. © 2017 Elsevier Inc. All rights reserved.

Homophila: human disease gene cognates in Drosophila

PubMed Central

Chien, Samson; Reiter, Lawrence T.; Bier, Ethan; Gribskov, Michael

2002-01-01

Although many human genes have been associated with genetic diseases, knowing which mutations result in disease phenotypes often does not explain the etiology of a specific disease. Drosophila melanogaster provides a powerful system in which to use genetic and molecular approaches to investigate human genetic diseases. Homophila is an intergenomic resource linking the human and fly genomes in order to stimulate functional genomic investigations in Drosophila that address questions about genetic disease in humans. Homophila provides a comprehensive linkage between the disease genes compiled in Online Mendelian Inheritance in Man (OMIM) and the complete Drosophila genomic sequence. Homophila is a relational database that allows searching based on human disease descriptions, OMIM number, human or fly gene names, and sequence similarity, and can be accessed at http://homophila.sdsc.edu. PMID:11752278

The Drosophila genes CG14593 and CG30106 code for G-protein-coupled receptors specifically activated by the neuropeptides CCHamide-1 and CCHamide-2.

PubMed

Hansen, Karina K; Hauser, Frank; Williamson, Michael; Weber, Stine B; Grimmelikhuijzen, Cornelis J P

2011-01-07

Recently, a novel neuropeptide, CCHamide, was discovered in the silkworm Bombyx mori (L. Roller et al., Insect Biochem. Mol. Biol. 38 (2008) 1147-1157). We have now found that all insects with a sequenced genome have two genes, each coding for a different CCHamide, CCHamide-1 and -2. We have also cloned and deorphanized two Drosophila G-protein-coupled receptors (GPCRs) coded for by genes CG14593 and CG30106 that are selectively activated by Drosophila CCH-amide-1 (EC(50), 2×10(-9) M) and CCH-amide-2 (EC(50), 5×10(-9) M), respectively. Gene CG30106 (symbol synonym CG14484) has in a previous publication (E.C. Johnson et al., J. Biol. Chem. 278 (2003) 52172-52178) been wrongly assigned to code for an allatostatin-B receptor. This conclusion is based on our findings that the allatostatins-B do not activate the CG30106 receptor and on the recent findings from other research groups that the allatostatins-B activate an unrelated GPCR coded for by gene CG16752. Comparative genomics suggests that a duplication of the CCHamide neuropeptide signalling system occurred after the split of crustaceans and insects, about 410 million years ago, because only one CCHamide neuropeptide gene is found in the water flea Daphnia pulex (Crustacea) and the tick Ixodes scapularis (Chelicerata). Copyright © 2010 Elsevier Inc. All rights reserved.

Bee venom suppresses PMA-mediated MMP-9 gene activation via JNK/p38 and NF-kappaB-dependent mechanisms.

PubMed

Cho, Hyun-Ji; Jeong, Yun-Jeong; Park, Kwan-Kyu; Park, Yoon-Yub; Chung, Il-Kyung; Lee, Kwang-Gill; Yeo, Joo-Hong; Han, Sang-Mi; Bae, Young-Seuk; Chang, Young-Chae

2010-02-17

Bee venom has been used for the treatment of inflammatory diseases such as rheumatoid arthritis and for the relief of pain in traditional oriental medicine. The purpose of this study is to elucidate the effects of bee venom on MMP-9 expression and determine possible mechanisms by which bee venom relieves or prevents the expression of MMP-9 during invasion and metastasis of breast cancer cells. We examined the expression and activity of MMP-9 and possible signaling pathway affected in PMA-induced MCF-7 cells. Bee venom was obtained from the National Institute of Agricultural Science and Technology of Korea. Matrigel invasion assay, wound-healing assay, zymography assay, western blot assay, electrophoretic mobility shift assay and luciferase gene assay were used for assessment. Bee venom inhibited cell invasion and migration, and also suppressed MMP-9 activity and expression, processes related to tumor invasion and metastasis, in PMA-induced MCF-7 cells. Bee venom specifically suppressed the phosphorylation of p38/JNK and at the same time, suppressed the protein expression, DNA binding and promoter activity of NF-kappaB. The levels of phosphorylated ERK1/2 and c-Jun did not change. We also investigated MMP-9 inhibition by melittin, apamin and PLA(2), representative single component of bee venom. We confirmed that PMA-induced MMP-9 activity was significantly decreased by melittin, but not by apamin and phospholipase A(2). These data demonstrated that the expression of MMP-9 was abolished by melittin, the main component of bee venom. Bee venom inhibits PMA-induced MMP-9 expression and activity by inhibition of NF-kappaB via p38 MAPK and JNK signaling pathways in MCF-7 cells. These results indicate that bee venom can be a potential anti-metastatic and anti-invasive agent. This useful effect may lead to future clinical research on the anti-cancer properties of bee venom. Copyright 2009 Elsevier Ireland Ltd. All rights reserved.

Effect of the gene transformer of Anastrepha on the somatic sexual development of Drosophila.

PubMed

Ruiz, María-Fernanda; Sánchez, Lucas

2010-01-01

The gene transformer (tra) is the key regulatory memory device for sex determination in tephritid insects. The present manuscript addressed the question about the functional conservation of the tephritid Anastrepha Transformer protein to direct somatic sexual development in Drosophila (Drosophilidae). The transformer cDNA of Anastrepha encoding the putative full-length Tra protein was cloned in pUAST and introduced into Drosophila melanogaster. To express this protein, the GAL4-UAS system was used. The Anastrepha Tra protein induced the female-specific splicing of both dsx and fru pre-mRNAs in Drosophila XY male flies, so that these became transformed into females, though this transformation was incomplete (the sexually dimorphic foreleg basitarsus and the external terminalia were monitored). It was found that the degree of female transformation directly depended on the dose of Anastrepha tra and Drosophila transformer-2 (tra-2) genes, and that the Anastrepha Tra-Drosophila Tra2 complex is not as efficient as the Drosophila Tra-Tra2 complex at inducing the female-specific splicing of Drosophila dsx pre-mRNA. This can explain why the Anastrepha Tra protein cannot fully substitute for the endogenous Drosophila Tra protein.

Disruption of the Abdominal-B Promoter Tethering Element Results in a Loss of Long-Range Enhancer-Directed Hox Gene Expression in Drosophila

PubMed Central

Ho, Margaret C. W.; Schiller, Benjamin J.; Akbari, Omar S.; Bae, Esther; Drewell, Robert A.

2011-01-01

There are many examples within gene complexes of transcriptional enhancers interacting with only a subset of target promoters. A number of molecular mechanisms including promoter competition, insulators and chromatin looping are thought to play a role in regulating these interactions. At the Drosophila bithorax complex (BX-C), the IAB5 enhancer specifically drives gene expression only from the Abdominal-B (Abd-B) promoter, even though the enhancer and promoter are 55 kb apart and are separated by at least three insulators. In previous studies, we discovered that a 255 bp cis-regulatory module, the promoter tethering element (PTE), located 5′ of the Abd-B transcriptional start site is able to tether IAB5 to the Abd-B promoter in transgenic embryo assays. In this study we examine the functional role of the PTE at the endogenous BX-C using transposon-mediated mutagenesis. Disruption of the PTE by P element insertion results in a loss of enhancer-directed Abd-B expression during embryonic development and a homeotic transformation of abdominal segments. A partial deletion of the PTE and neighboring upstream genomic sequences by imprecise excision of the P element also results in a similar loss of Abd-B expression in embryos. These results demonstrate that the PTE is an essential component of the regulatory network at the BX-C and is required in vivo to mediate specific long-range enhancer-promoter interactions. PMID:21283702

Oxymatrine Inhibits Influenza A Virus Replication and Inflammation via TLR4, p38 MAPK and NF-κB Pathways.

PubMed

Dai, Jian-Ping; Wang, Qian-Wen; Su, Yun; Gu, Li-Ming; Deng, Hui-Xiong; Chen, Xiao-Xuan; Li, Wei-Zhong; Li, Kang-Sheng

2018-03-23

Oxymatrine (OMT) is a strong immunosuppressive agent that has been used in the clinic for many years. In the present study, by using plaque inhibition, luciferase reporter plasmids, qRT-PCR, western blotting, and ELISA assays, we have investigated the effect and mechanism of OMT on influenza A virus (IAV) replication and IAV-induced inflammation in vitro and in vivo. The results showed that OMT had excellent anti-IAV activity on eight IAV strains in vitro. OMT could significantly decrease the promoter activity of TLR3, TLR4, TLR7, MyD88, and TRAF6 genes, inhibit IAV-induced activations of Akt, ERK1/2, p38 MAPK, and NF-κB pathways, and suppress the expressions of inflammatory cytokines and MMP-2/-9. Activators of TLR4, p38 MAPK and NF-κB pathways could significantly antagonize the anti-IAV activity of OMT in vitro, including IAV replication and IAV-induced cytopathogenic effect (CPE). Furthermore, OMT could reduce the loss of body weight, significantly increase the survival rate of IAV-infected mice, decrease the lung index, pulmonary inflammation and lung viral titter, and improve pulmonary histopathological changes. In conclusion, OMT possesses anti-IAV and anti-inflammatory activities, the mechanism of action may be linked to its ability to inhibit IAV-induced activations of TLR4, p38 MAPK, and NF-κB pathways.

Core Promoter Functions in the Regulation of Gene Expression of Drosophila Dorsal Target Genes*

PubMed Central

Zehavi, Yonathan; Kuznetsov, Olga; Ovadia-Shochat, Avital; Juven-Gershon, Tamar

2014-01-01

Developmental processes are highly dependent on transcriptional regulation by RNA polymerase II. The RNA polymerase II core promoter is the ultimate target of a multitude of transcription factors that control transcription initiation. Core promoters consist of core promoter motifs, e.g. the initiator, TATA box, and the downstream core promoter element (DPE), which confer specific properties to the core promoter. Here, we explored the importance of core promoter functions in the dorsal-ventral developmental gene regulatory network. This network includes multiple genes that are activated by different nuclear concentrations of Dorsal, an NFκB homolog transcription factor, along the dorsal-ventral axis. We show that over two-thirds of Dorsal target genes contain DPE sequence motifs, which is significantly higher than the proportion of DPE-containing promoters in Drosophila genes. We demonstrate that multiple Dorsal target genes are evolutionarily conserved and functionally dependent on the DPE. Furthermore, we have analyzed the activation of key Dorsal target genes by Dorsal, as well as by another Rel family transcription factor, Relish, and the dependence of their activation on the DPE motif. Using hybrid enhancer-promoter constructs in Drosophila cells and embryo extracts, we have demonstrated that the core promoter composition is an important determinant of transcriptional activity of Dorsal target genes. Taken together, our results provide evidence for the importance of core promoter composition in the regulation of Dorsal target genes. PMID:24634215

Excess amino acid polymorphism in mitochondrial DNA: contrasts among genes from Drosophila, mice, and humans.

PubMed

Rand, D M; Kann, L M

1996-07-01

Recent studies of mitochondrial DNA (mtDNA) variation in mammals and Drosophila have shown an excess of amino acid variation within species (replacement polymorphism) relative to the number of silent and replacement differences fixed between species. To examine further this pattern of nonneutral mtDNA evolution, we present sequence data for the ND3 and ND5 genes from 59 lines of Drosophila melanogaster and 29 lines of D. simulans. Of interest are the frequency spectra of silent and replacement polymorphisms, and potential variation among genes and taxa in the departures from neutral expectations. The Drosophila ND3 and ND5 data show no significant excess of replacement polymorphism using the McDonald-Kreitman test. These data are in contrast to significant departures from neutrality for the ND3 gene in mammals and other genes in Drosophila mtDNA (cytochrome b and ATPase 6). Pooled across genes, however, both Drosophila and human mtDNA show very significant excesses of amino acid polymorphism. Silent polymorphisms at ND5 show a significantly higher variance in frequency than replacement polymorphisms, and the latter show a significant skew toward low frequencies (Tajima's D = -1.954). These patterns are interpreted in light of the nearly neutral theory where mildly deleterious amino acid haplotypes are observed as ephemeral variants within species but do not contribute to divergence. The patterns of polymorphism and divergence at charge-altering amino acid sites are presented for the Drosophila ND5 gene to examine the evolution of functionally distinct mutations. Excess charge-altering polymorphism is observed at the carboxyl terminal and excess charge-altering divergence is detected at the amino terminal. While the mildly deleterious model fits as a net effect in the evolution of nonrecombining mitochondrial genomes, these data suggest that opposing evolutionary pressures may act on different regions of mitochondrial genes and genomes.

Transposable elements are enriched within or in close proximity to xenobiotic-metabolizing cytochrome P450 genes

PubMed Central

Chen, Song; Li, Xianchun

2007-01-01

Background Transposons, i.e. transposable elements (TEs), are the major internal spontaneous mutation agents for the variability of eukaryotic genomes. To address the general issue of whether transposons mediate genomic changes in environment-adaptation genes, we scanned two alleles per each of the six xenobiotic-metabolizing Helicoverpa zea cytochrome P450 loci, including CYP6B8, CYP6B27, CYP321A1, CYP321A2, CYP9A12v3 and CYP9A14, for the presence of transposon insertions by genome walking and sequence analysis. We also scanned thirteen Drosophila melanogaster P450s genes for TE insertions by in silico mapping and literature search. Results Twelve novel transposons, including LINEs (long interspersed nuclear elements), SINEs (short interspersed nuclear elements), MITEs (miniature inverted-repeat transposable elements), one full-length transib-like transposon, and one full-length Tcl-like DNA transpson, are identified from the alleles of the six H. zea P450 genes. The twelve transposons are inserted into the 5'flanking region, 3'flanking region, exon, or intron of the six environment-adaptation P450 genes. In D. melanogaster, seven out of the eight Drosophila P450s (CYP4E2, CYP6A2, CYP6A8, CYP6A9, CYP6G1, CYP6W1, CYP12A4, CYP12D1) implicated in insecticide resistance are associated with a variety of transposons. By contrast, all the five Drosophila P450s (CYP302A1, CYP306A1, CYP307A1, CYP314A1 and CYP315A1) involved in ecdysone biosynthesis and developmental regulation are free of TE insertions. Conclusion These results indicate that TEs are selectively retained within or in close proximity to xenobiotic-metabolizing P450 genes. PMID:17381843

Identification of unannotated exons of low abundance transcripts in Drosophila melanogaster and cloning of a new serine protease gene upregulated upon injury.

PubMed

Maia, Rafaela M; Valente, Valeria; Cunha, Marco A V; Sousa, Josane F; Araujo, Daniela D; Silva, Wilson A; Zago, Marco A; Dias-Neto, Emmanuel; Souza, Sandro J; Simpson, Andrew J G; Monesi, Nadia; Ramos, Ricardo G P; Espreafico, Enilza M; Paçó-Larson, Maria L

2007-07-24

The sequencing of the D.melanogaster genome revealed an unexpected small number of genes (~ 14,000) indicating that mechanisms acting on generation of transcript diversity must have played a major role in the evolution of complex metazoans. Among the most extensively used mechanisms that accounts for this diversity is alternative splicing. It is estimated that over 40% of Drosophila protein-coding genes contain one or more alternative exons. A recent transcription map of the Drosophila embryogenesis indicates that 30% of the transcribed regions are unannotated, and that 1/3 of this is estimated as missed or alternative exons of previously characterized protein-coding genes. Therefore, the identification of the variety of expressed transcripts depends on experimental data for its final validation and is continuously being performed using different approaches. We applied the Open Reading Frame Expressed Sequence Tags (ORESTES) methodology, which is capable of generating cDNA data from the central portion of rare transcripts, in order to investigate the presence of hitherto unnanotated regions of Drosophila transcriptome. Bioinformatic analysis of 1,303 Drosophila ORESTES clusters identified 68 sequences derived from unannotated regions in the current Drosophila genome version (4.3). Of these, a set of 38 was analysed by polyA+ northern blot hybridization, validating 17 (50%) new exons of low abundance transcripts. For one of these ESTs, we obtained the cDNA encompassing the complete coding sequence of a new serine protease, named SP212. The SP212 gene is part of a serine protease gene cluster located in the chromosome region 88A12-B1. This cluster includes the predicted genes CG9631, CG9649 and CG31326, which were previously identified as up-regulated after immune challenges in genomic-scale microarray analysis. In agreement with the proposal that this locus is co-regulated in response to microorganisms infection, we show here that SP212 is also up-regulated upon

Identification of unannotated exons of low abundance transcripts in Drosophila melanogaster and cloning of a new serine protease gene upregulated upon injury

PubMed Central

Maia, Rafaela M; Valente, Valeria; Cunha, Marco AV; Sousa, Josane F; Araujo, Daniela D; Silva, Wilson A; Zago, Marco A; Dias-Neto, Emmanuel; Souza, Sandro J; Simpson, Andrew JG; Monesi, Nadia; Ramos, Ricardo GP; Espreafico, Enilza M; Paçó-Larson, Maria L

2007-01-01

Background The sequencing of the D.melanogaster genome revealed an unexpected small number of genes (~ 14,000) indicating that mechanisms acting on generation of transcript diversity must have played a major role in the evolution of complex metazoans. Among the most extensively used mechanisms that accounts for this diversity is alternative splicing. It is estimated that over 40% of Drosophila protein-coding genes contain one or more alternative exons. A recent transcription map of the Drosophila embryogenesis indicates that 30% of the transcribed regions are unannotated, and that 1/3 of this is estimated as missed or alternative exons of previously characterized protein-coding genes. Therefore, the identification of the variety of expressed transcripts depends on experimental data for its final validation and is continuously being performed using different approaches. We applied the Open Reading Frame Expressed Sequence Tags (ORESTES) methodology, which is capable of generating cDNA data from the central portion of rare transcripts, in order to investigate the presence of hitherto unnanotated regions of Drosophila transcriptome. Results Bioinformatic analysis of 1,303 Drosophila ORESTES clusters identified 68 sequences derived from unannotated regions in the current Drosophila genome version (4.3). Of these, a set of 38 was analysed by polyA+ northern blot hybridization, validating 17 (50%) new exons of low abundance transcripts. For one of these ESTs, we obtained the cDNA encompassing the complete coding sequence of a new serine protease, named SP212. The SP212 gene is part of a serine protease gene cluster located in the chromosome region 88A12-B1. This cluster includes the predicted genes CG9631, CG9649 and CG31326, which were previously identified as up-regulated after immune challenges in genomic-scale microarray analysis. In agreement with the proposal that this locus is co-regulated in response to microorganisms infection, we show here that SP212 is also up

Evolution of Drosophila ribosomal protein gene core promoters.

PubMed

Ma, Xiaotu; Zhang, Kangyu; Li, Xiaoman

2009-03-01

The coordinated expression of ribosomal protein genes (RPGs) has been well documented in many species. Previous analyses of RPG promoters focus only on Fungi and mammals. Recognizing this gap and using a comparative genomics approach, we utilize a motif-finding algorithm that incorporates cross-species conservation to identify several significant motifs in Drosophila RPG promoters. As a result, significant differences of the enriched motifs in RPG promoter are found among Drosophila, Fungi, and mammals, demonstrating the evolutionary dynamics of the ribosomal gene regulatory network. We also report a motif present in similar numbers of RPGs among Drosophila species which does not appear to be conserved at the individual RPG gene level. A module-wise stabilizing selection theory is proposed to explain this observation. Overall, our results provide significant insight into the fast-evolving nature of transcriptional regulation in the RPG module.

Evolution of Drosophila ribosomal protein gene core promoters

PubMed Central

Ma, Xiaotu; Zhang, Kangyu; Li, Xiaoman

2011-01-01

The coordinated expression of ribosomal protein genes (RPGs) has been well documented in many species. Previous analyses of RPG promoters focus only on Fungi and mammals. Recognizing this gap and using a comparative genomics approach, we utilize a motif-finding algorithm that incorporates cross-species conservation to identify several significant motifs in Drosophila RPG promoters. As a result, significant differences of the enriched motifs in RPG promoter are found among Drosophila, Fungi, and mammals, demonstrating the evolutionary dynamics of the ribosomal gene regulatory network. We also report a motif present in similar numbers of RPGs among Drosophila species which does not appear to be conserved at the individual RPG gene level. A module-wise stabilizing selection theory is proposed to explain this observation. Overall, our results provide significant insight into the fast-evolving nature of transcriptional regulation in the RPG module. PMID:19059316

Conservation of regulatory sequences and gene expression patterns in the disintegrating Drosophila Hox gene complex

PubMed Central

Negre, Bárbara; Casillas, Sònia; Suzanne, Magali; Sánchez-Herrero, Ernesto; Akam, Michael; Nefedov, Michael; Barbadilla, Antonio; de Jong, Pieter; Ruiz, Alfredo

2005-01-01

Homeotic (Hox) genes are usually clustered and arranged in the same order as they are expressed along the anteroposterior body axis of metazoans. The mechanistic explanation for this colinearity has been elusive, and it may well be that a single and universal cause does not exist. The Hox-gene complex (HOM-C) has been rearranged differently in several Drosophila species, producing a striking diversity of Hox gene organizations. We investigated the genomic and functional consequences of the two HOM-C splits present in Drosophila buzzatii. Firstly, we sequenced two regions of the D. buzzatii genome, one containing the genes labial and abdominal A, and another one including proboscipedia, and compared their organization with that of D. melanogaster and D. pseudoobscura in order to map precisely the two splits. Then, a plethora of conserved noncoding sequences, which are putative enhancers, were identified around the three Hox genes closer to the splits. The position and order of these enhancers are conserved, with minor exceptions, between the three Drosophila species. Finally, we analyzed the expression patterns of the same three genes in embryos and imaginal discs of four Drosophila species with different Hox-gene organizations. The results show that their expression patterns are conserved despite the HOM-C splits. We conclude that, in Drosophila, Hox-gene clustering is not an absolute requirement for proper function. Rather, the organization of Hox genes is modular, and their clustering seems the result of phylogenetic inertia more than functional necessity. PMID:15867430

Hepatic p38α regulates gluconeogenesis by suppressing AMPK.

PubMed

Jing, Yanyan; Liu, Wei; Cao, Hongchao; Zhang, Duo; Yao, Xuan; Zhang, Shengjie; Xia, Hongfeng; Li, Dan; Wang, Yu-cheng; Yan, Jun; Hui, Lijian; Ying, Hao

2015-06-01

It is proposed that p38 is involved in gluconeogenesis, however, the genetic evidence is lacking and precise mechanisms remain poorly understood. We sought to delineate the role of hepatic p38α in gluconeogenesis during fasting by applying a loss-of-function genetic approach. We examined fasting glucose levels, performed pyruvate tolerance test, imaged G6Pase promoter activity, as well as determined the expression of gluconeogenic genes in mice with a targeted deletion of p38α in liver. Results were confirmed both in vivo and in vitro by using an adenoviral dominant-negative form of p38α (p38α-AF) and the constitutively active mitogen-activated protein kinase 6, respectively. Adenoviral dominant-negative form of AMP-activated protein kinase α (DN-AMPKα) was employed to test our proposed model. Mice lacking hepatic p38α exhibited reduced fasting glucose level and impaired gluconeogenesis. Interestingly, hepatic deficiency of p38α did not result in an alteration in CREB phosphorylation, but led to an increase in AMPKα phosphorylation. Adenoviral DN-AMPKα could abolish the effect of p38α-AF on gluconeogenesis. Knockdown of up-steam transforming growth factor β-activated kinase 1 decreased the AMPKα phosphorylation induced by p38α-AF, suggesting a negative feedback loop. Consistently, inverse correlations between p38 and AMPKα phosphorylation were observed during fasting and in diabetic mouse models. Importantly, adenoviral p38α-AF treatment ameliorated hyperglycemia in diabetic mice. Our study provides evidence that hepatic p38α functions as a negative regulator of AMPK signaling in maintaining gluconeogenesis, dysregulation of this regulatory network contributes to unrestrained gluconeogenesis in diabetes, and hepatic p38α could be a drug target for hyperglycemia. Copyright © 2015 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

Faster-X Evolution of Gene Expression in Drosophila

PubMed Central

Meisel, Richard P.; Malone, John H.; Clark, Andrew G.

2012-01-01

DNA sequences on X chromosomes often have a faster rate of evolution when compared to similar loci on the autosomes, and well articulated models provide reasons why the X-linked mode of inheritance may be responsible for the faster evolution of X-linked genes. We analyzed microarray and RNA–seq data collected from females and males of six Drosophila species and found that the expression levels of X-linked genes also diverge faster than autosomal gene expression, similar to the “faster-X” effect often observed in DNA sequence evolution. Faster-X evolution of gene expression was recently described in mammals, but it was limited to the evolutionary lineages shortly following the creation of the therian X chromosome. In contrast, we detect a faster-X effect along both deep lineages and those on the tips of the Drosophila phylogeny. In Drosophila males, the dosage compensation complex (DCC) binds the X chromosome, creating a unique chromatin environment that promotes the hyper-expression of X-linked genes. We find that DCC binding, chromatin environment, and breadth of expression are all predictive of the rate of gene expression evolution. In addition, estimates of the intraspecific genetic polymorphism underlying gene expression variation suggest that X-linked expression levels are not under relaxed selective constraints. We therefore hypothesize that the faster-X evolution of gene expression is the result of the adaptive fixation of beneficial mutations at X-linked loci that change expression level in cis. This adaptive faster-X evolution of gene expression is limited to genes that are narrowly expressed in a single tissue, suggesting that relaxed pleiotropic constraints permit a faster response to selection. Finally, we present a conceptional framework to explain faster-X expression evolution, and we use this framework to examine differences in the faster-X effect between Drosophila and mammals. PMID:23071459

Drosophila selenophosphate synthetase 1 regulates vitamin B6 metabolism: prediction and confirmation

PubMed Central

2011-01-01

Background There are two selenophosphate synthetases (SPSs) in higher eukaryotes, SPS1 and SPS2. Of these two isotypes, only SPS2 catalyzes selenophosphate synthesis. Although SPS1 does not contain selenophosphate synthesis activity, it was found to be essential for cell growth and embryogenesis in Drosophila. The function of SPS1, however, has not been elucidated. Results Differentially expressed genes in Drosophila SL2 cells were identified using two-way analysis of variance methods and clustered according to their temporal expression pattern. Gene ontology analysis was performed against differentially expressed genes and gene ontology terms related to vitamin B6 biosynthesis were found to be significantly affected at the early stage at which megamitochondria were not formed (day 3) after SPS1 knockdown. Interestingly, genes related to defense and amino acid metabolism were affected at a later stage (day 5) following knockdown. Levels of pyridoxal phosphate, an active form of vitamin B6, were decreased by SPS1 knockdown. Treatment of SL2 cells with an inhibitor of pyridoxal phosphate synthesis resulted in both a similar pattern of expression as that found by SPS1 knockdown and the formation of megamitochondria, the major phenotypic change observed by SPS1 knockdown. Conclusions These results indicate that SPS1 regulates vitamin B6 synthesis, which in turn impacts various cellular systems such as amino acid metabolism, defense and other important metabolic activities. PMID:21864351

p38 mitogen-activated protein kinase-induced nuclear factor kappa-light-chain-enhancer of activated B cell activity is required for neuroprotection in retinal ischemia/reperfusion injury.

PubMed

Jiang, Shao-Yun; Zou, Yuan-Yuan; Wang, Jian-Tao

2012-01-01

In our previous study, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) played a neuroprotective role in retinal ischemia/reperfusion (I/R) injury in rats. However, the mechanism of NF-κB neuroprotection is still unclear. We hypothesize that p38 mitogen-activated protein kinase (MAPK) is expressed and NF-κB activity induced by p38 MAPK plays a neuroprotective role through antiapoptotic genes (B-cell lymphoma [Bcl]-2 and Bcl-XL) in retinal cells in retinal I/R injury. Retinal ischemia was induced by elevating intraocular pressure in rats. After retinal I/R, the p38 MAPK, NF-κB p65, Bcl-2, and Bcl-XL mRNA levels were measured with real-time polymerase chain reaction. NF-κB p65 activity was assessed with NF-κB enzyme-linked immunosorbent assay in retinal I/R injury and after application of the p38 MAPK inhibitor (SB203580). Furthermore, SB203580 and NF-κB p65 short interfering RNA (siRNA) were used in retinal I/R injury to examine the effects on Bcl-2 and Bcl-XL levels and nucleosome release in the retina and cell survival in the ganglion cell layer. The mRNA levels of NF-κB p65 and p38 MAPK reached a peak at 6 h after retinal I/R and then decreased gradually. The mRNA levels of Bcl-2 and Bcl-XL significantly increased at 2, 4, and 6 h, peaked at 8 h, and decreased gradually, but remained at a higher level compared with the normal control, which was accompanied by an increase in NF-κB p65 in nuclear extracts. After application of SB203580, the increase in the NF-κB p65 levels in the nucleus induced with I/R was completely abolished, and the mRNA expression of Bcl-2 and Bcl-XL decreased significantly compared with the I/R controls. In addition, NF-κB p65 siRNA inhibited Bcl-2 and Bcl-XL expression. Inhibition of the p38 MAPK-NF-κB pathway (using SB203580 or NF-κB p65 siRNA) increased retinal nucleosome release and decreased the number of ganglion cells. These findings provide evidence of crosstalk between p38 MAPK and NF-κB p65 and

p38 mitogen-activated protein kinase–induced nuclear factor kappa-light-chain-enhancer of activated B cell activity is required for neuroprotection in retinal ischemia/reperfusion injury

PubMed Central

Jiang, Shao-Yun; Zou, Yuan-Yuan

2012-01-01

Purpose In our previous study, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) played a neuroprotective role in retinal ischemia/reperfusion (I/R) injury in rats. However, the mechanism of NF-κB neuroprotection is still unclear. We hypothesize that p38 mitogen-activated protein kinase (MAPK) is expressed and NF-κB activity induced by p38 MAPK plays a neuroprotective role through antiapoptotic genes (B-cell lymphoma [Bcl]-2 and Bcl-XL) in retinal cells in retinal I/R injury. Methods Retinal ischemia was induced by elevating intraocular pressure in rats. After retinal I/R, the p38 MAPK, NF-κB p65, Bcl-2, and Bcl-XL mRNA levels were measured with real-time polymerase chain reaction. NF-κB p65 activity was assessed with NF-κB enzyme-linked immunosorbent assay in retinal I/R injury and after application of the p38 MAPK inhibitor (SB203580). Furthermore, SB203580 and NF-κB p65 short interfering RNA (siRNA) were used in retinal I/R injury to examine the effects on Bcl-2 and Bcl-XL levels and nucleosome release in the retina and cell survival in the ganglion cell layer. Results The mRNA levels of NF-κB p65 and p38 MAPK reached a peak at 6 h after retinal I/R and then decreased gradually. The mRNA levels of Bcl-2 and Bcl-XL significantly increased at 2, 4, and 6 h, peaked at 8 h, and decreased gradually, but remained at a higher level compared with the normal control, which was accompanied by an increase in NF-κB p65 in nuclear extracts. After application of SB203580, the increase in the NF-κB p65 levels in the nucleus induced with I/R was completely abolished, and the mRNA expression of Bcl-2 and Bcl-XL decreased significantly compared with the I/R controls. In addition, NF-κB p65 siRNA inhibited Bcl-2 and Bcl-XL expression. Inhibition of the p38 MAPK-NF-κB pathway (using SB203580 or NF-κB p65 siRNA) increased retinal nucleosome release and decreased the number of ganglion cells. Conclusions These findings provide evidence of crosstalk

Low-dose radiation induces Drosophila innate immunity through Toll pathway activation.

PubMed

Seong, Ki Moon; Kim, Cha Soon; Lee, Byung-Sub; Nam, Seon Young; Yang, Kwang Hee; Kim, Ji-Young; Park, Joong-Jean; Min, Kyung-Jin; Jin, Young-Woo

2012-01-01

Numerous studies report that exposing certain organisms to low-dose radiation induces beneficial effects on lifespan, tumorigenesis, and immunity. By analyzing survival after bacterial infection and antimicrobial peptide gene expression in irradiated flies, we demonstrate that low-dose irradiation of Drosophila enhances innate immunity. Low-dose irradiation of flies significantly increased resistance against gram-positive and gram-negative bacterial infections, as well as expression of several antimicrobial peptide genes. Additionally, low-dose irradiation also resulted in a specific increase in expression of key proteins of the Toll signaling pathway and phosphorylated forms of p38 and JNK. These results indicate that innate immunity is activated after low-dose irradiation through Toll signaling pathway in Drosophila.

B7-H3 Augments Inflammatory Responses and Exacerbates Brain Damage via Amplifying NF-κB p65 and MAPK p38 Activation during Experimental Pneumococcal Meningitis.

PubMed

Chen, Xuqin; Li, Yan; Blankson, Siobhan; Liu, Min; Huang, Danping; Redmond, H Paul; Huang, Jing; Wang, Jiang Huai; Wang, Jian

2017-01-01

The costimulatory protein B7-H3 has been shown to play a contributory role in the development and progression of experimental pneumococcal meningitis by augmentation of the innate immunity-associated inflammatory response via a TLR2-dependent manner. This study aimed to clarify the component(s) of TLR2-mediated signal transduction pathways responsible for B7-H3-augmented inflammatory response and subsequent brain damage during experimental pneumococcal meningitis. Administration of B7-H3 did not augment expression of TLR2 and other TLR2 upstream components, but led to an enhanced formation of MyD88-IRAK immunocomplex in the brain of S. pneumoniae-infected mice. Furthermore, B7-H3 substantially augmented S. pneumoniae-induced activation of TLR2 downstream NF-κB p65 and MAPK p38 pathways in the brain of S. pneumoniae-infected mice. Notably, blockage of NF-κB p65 and/or MAPK p38 with their specific inhibitors strongly attenuated B7-H3-amplified inflammatory response with significantly reduced proinflammatory cytokine and chemokine production, and markedly ameliorated B7-H3-exacerbated disruption of blood-brain barrier and severity of disease status in S. pneumoniae-infected mice. These results indicate that targeting NF-κB p65 and/or MAPK p38 may represent a promising therapeutic option for amelioration of overwhelming inflammatory response-associated brain injury frequently observed during pneumococcal meningitis.

B7-H3 Augments Inflammatory Responses and Exacerbates Brain Damage via Amplifying NF-κB p65 and MAPK p38 Activation during Experimental Pneumococcal Meningitis

PubMed Central

Chen, Xuqin; Li, Yan; Blankson, Siobhan; Liu, Min; Huang, Danping; Redmond, H. Paul; Huang, Jing; Wang, Jiang Huai; Wang, Jian

2017-01-01

The costimulatory protein B7-H3 has been shown to play a contributory role in the development and progression of experimental pneumococcal meningitis by augmentation of the innate immunity-associated inflammatory response via a TLR2-dependent manner. This study aimed to clarify the component(s) of TLR2-mediated signal transduction pathways responsible for B7-H3-augmented inflammatory response and subsequent brain damage during experimental pneumococcal meningitis. Administration of B7-H3 did not augment expression of TLR2 and other TLR2 upstream components, but led to an enhanced formation of MyD88-IRAK immunocomplex in the brain of S. pneumoniae-infected mice. Furthermore, B7-H3 substantially augmented S. pneumoniae-induced activation of TLR2 downstream NF-κB p65 and MAPK p38 pathways in the brain of S. pneumoniae-infected mice. Notably, blockage of NF-κB p65 and/or MAPK p38 with their specific inhibitors strongly attenuated B7-H3-amplified inflammatory response with significantly reduced proinflammatory cytokine and chemokine production, and markedly ameliorated B7-H3-exacerbated disruption of blood-brain barrier and severity of disease status in S. pneumoniae-infected mice. These results indicate that targeting NF-κB p65 and/or MAPK p38 may represent a promising therapeutic option for amelioration of overwhelming inflammatory response-associated brain injury frequently observed during pneumococcal meningitis. PMID:28141831

The activation of p38 MAPK primarily contributes to UV-induced RhoB expression by recruiting the c-Jun and p300 to the distal CCAAT box of the RhoB promoter

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ahn, Jiwon; Department of Microbiology, Chungnam National University, Daejeon 305-764; Choi, Jeong-Hae

2011-06-03

Highlights: {yields} Regulation of transcriptional activation of RhoB is still unclear. {yields} We examine the effect of p38 MAPK inhibition, and c-Jun and RhoB depletion on UV-induced RhoB expression and apoptosis. {yields} We identify the regions of RhoB promoter necessary to confer UV responsiveness using pRhoB-luciferase reporter assays. {yields} c-Jun, ATF2 and p300 are dominantly associated with NF-Y on the distal CCAAT box. {yields} The activation of p38 MAPK primarily contribute to UV-induced RhoB expression by recruiting the c-Jun and p300 proteins on distal CCAAT box of RhoB promoter. -- Abstract: The Ras-related small GTP-binding protein RhoB is rapidly inducedmore » in response to genotoxic stresses caused by ionizing radiation. It is known that UV-induced RhoB expression results from the binding of activating transcription factor 2 (ATF2) via NF-Y to the inverted CCAAT box (-23) of the RhoB promoter. Here, we show that the association of c-Jun with the distal CCAAT box (-72) is primarily involved in UV-induced RhoB expression and p38 MAPK regulated RhoB induction through the distal CCAAT box. UV-induced RhoB expression and apoptosis were markedly attenuated by pretreatment with the p38 MAPK inhibitor. siRNA knockdown of RhoB, ATF2 and c-Jun resulted in decreased RhoB expression and eventually restored the growth of UV-irradiated Jurkat cells. In the reporter assay using luciferase under the RhoB promoter, inhibition of RhoB promoter activity by the p38 inhibitor and knockdown of c-Jun using siRNA occurred through the distal CCAAT box. Immunoprecipitation and DNA affinity protein binding assays revealed the association of c-Jun and p300 via NF-YA and the dissociation of histone deacetylase 1 (HDAC1) via c-Jun recruitment to the CCAAT boxes of the RhoB promoter. These results suggest that the activation of p38 MAPK primarily contributes to UV-induced RhoB expression by recruiting the c-Jun and p300 proteins to the distal CCAAT box of the RhoB promoter

Resources for Biological Annotation of the Drosophila Genome

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gerald M. Rubin

2005-08-08

This project supported seed money for the development of cDNA and genetic resources to support studies of the Drosophila melanogaster genome. Key publications supported by this work that provide additional detail: (1) ''The Drosophila gene collection: identification of putative full-length cDNAs for 70% of D. melanogaster genes''; and (2) ''The Berkeley Drosophila Genome Project gene disruption project: Single P-element insertions mutating 25% of vital Drosophila genes''.

Differential gene expression related to Nora virus infection of Drosophila melanogaster.

PubMed

Cordes, Ethan J; Licking-Murray, Kellie D; Carlson, Kimberly A

2013-08-01

Nora virus is a recently discovered RNA picorna-like virus that produces a persistent infection in Drosophila melanogaster, but the antiviral pathway or change in gene expression is unknown. We performed cDNA microarray analysis comparing the gene expression profiles of Nora virus infected and uninfected wild-type D. melanogaster. This analysis yielded 58 genes exhibiting a 1.5-fold change or greater and p-value less than 0.01. Of these genes, 46 were up-regulated and 12 down-regulated in response to infection. To validate the microarray results, qRT-PCR was performed with probes for Chorion protein 16 and Troponin C isoform 4, which show good correspondence with cDNA microarray results. Differential regulation of genes associated with Toll and immune-deficient pathways, cytoskeletal development, Janus Kinase-Signal Transducer and Activator of Transcription interactions, and a potential gut-specific innate immune response were found. This genome-wide expression profile of Nora virus infection of D. melanogaster can pinpoint genes of interest for further investigation of antiviral pathways employed, genetic mechanisms, sites of replication, viral persistence, and developmental effects. Copyright © 2013. Published by Elsevier B.V.

A Role for the p38 Mitogen-activated Protein Kinase Pathway in Myocardial Cell Growth, Sarcomeric Organization, and Cardiac-specific Gene Expression

PubMed Central

Zechner, Dietmar; Thuerauf, Donna J.; Hanford, Deanna S.; McDonough, Patrick M.; Glembotski, Christopher C.

1997-01-01

Three hallmark features of the cardiac hypertrophic growth program are increases in cell size, sarcomeric organization, and the induction of certain cardiac-specific genes. All three features of hypertrophy are induced in cultured myocardial cells by α1- adrenergic receptor agonists, such as phenylephrine (PE) and other growth factors that activate mitogen- activated protein kinases (MAPKs). In this study the MAPK family members extracellular signal–regulated kinase (ERK), c-jun NH2-terminal kinase (JNK), and p38 were activated by transfecting cultured cardiac myocytes with constructs encoding the appropriate kinases possessing gain-of-function mutations. Transfected cells were then analyzed for changes in cell size, sarcomeric organization, and induction of the genes for the A- and B-type natriuretic peptides (NPs), as well as the α-skeletal actin (α-SkA) gene. While activation of JNK and/or ERK with MEKK1COOH or Raf-1 BXB, respectively, augmented cell size and effected relatively modest increases in NP and α-SkA promoter activities, neither upstream kinase conferred sarcomeric organization. However, transfection with MKK6 (Glu), which specifically activated p38, augmented cell size, induced NP and α-Ska promoter activities by up to 130-fold, and elicited sarcomeric organization in a manner similar to PE. Moreover, all three growth features induced by MKK6 (Glu) or PE were blocked with the p38-specific inhibitor, SB 203580. These results demonstrate novel and potentially central roles for MKK6 and p38 in the regulation of myocardial cell hypertrophy. PMID:9314533

High rate of translocation-based gene birth on the Drosophila Y chromosome.

PubMed

Tobler, Ray; Nolte, Viola; Schlötterer, Christian

2017-10-31

The Y chromosome is a unique genetic environment defined by a lack of recombination and male-limited inheritance. The Drosophila Y chromosome has been gradually acquiring genes from the rest of the genome, with only seven Y-linked genes being gained over the past 63 million years (0.12 gene gains per million years). Using a next-generation sequencing (NGS)-powered genomic scan, we show that gene transfers to the Y chromosome are much more common than previously suspected: at least 25 have arisen across three Drosophila species over the past 5.4 million years (1.67 per million years for each lineage). The gene transfer rate is significantly lower in Drosophila melanogaster than in the Drosophila simulans clade, primarily due to Y-linked retrotranspositions being significantly more common in the latter. Despite all Y-linked gene transfers being evolutionarily recent (<1 million years old), only three showed evidence for purifying selection ( ω ≤ 0.14). Thus, although the resulting Y-linked functional gene acquisition rate (0.25 new genes per million years) is double the longer-term estimate, the fate of most new Y-linked genes is defined by rapid degeneration and pseudogenization. Our results show that Y-linked gene traffic, and the molecular mechanisms governing these transfers, can diverge rapidly between species, revealing the Drosophila Y chromosome to be more dynamic than previously appreciated. Our analytical method provides a powerful means to identify Y-linked gene transfers and will help illuminate the evolutionary dynamics of the Y chromosome in Drosophila and other species. Copyright © 2017 the Author(s). Published by PNAS.

DNA Repair in Drosophila: Mutagens, Models, and Missing Genes

PubMed Central

Sekelsky, Jeff

2017-01-01

The numerous processes that damage DNA are counterbalanced by a complex network of repair pathways that, collectively, can mend diverse types of damage. Insights into these pathways have come from studies in many different organisms, including Drosophila melanogaster. Indeed, the first ideas about chromosome and gene repair grew out of Drosophila research on the properties of mutations produced by ionizing radiation and mustard gas. Numerous methods have been developed to take advantage of Drosophila genetic tools to elucidate repair processes in whole animals, organs, tissues, and cells. These studies have led to the discovery of key DNA repair pathways, including synthesis-dependent strand annealing, and DNA polymerase theta-mediated end joining. Drosophila appear to utilize other major repair pathways as well, such as base excision repair, nucleotide excision repair, mismatch repair, and interstrand crosslink repair. In a surprising number of cases, however, DNA repair genes whose products play important roles in these pathways in other organisms are missing from the Drosophila genome, raising interesting questions for continued investigations. PMID:28154196

Quantitative analysis of bristle number in Drosophila mutants identifies genes involved in neural development

NASA Technical Reports Server (NTRS)

Norga, Koenraad K.; Gurganus, Marjorie C.; Dilda, Christy L.; Yamamoto, Akihiko; Lyman, Richard F.; Patel, Prajal H.; Rubin, Gerald M.; Hoskins, Roger A.; Mackay, Trudy F.; Bellen, Hugo J.

2003-01-01

BACKGROUND: The identification of the function of all genes that contribute to specific biological processes and complex traits is one of the major challenges in the postgenomic era. One approach is to employ forward genetic screens in genetically tractable model organisms. In Drosophila melanogaster, P element-mediated insertional mutagenesis is a versatile tool for the dissection of molecular pathways, and there is an ongoing effort to tag every gene with a P element insertion. However, the vast majority of P element insertion lines are viable and fertile as homozygotes and do not exhibit obvious phenotypic defects, perhaps because of the tendency for P elements to insert 5' of transcription units. Quantitative genetic analysis of subtle effects of P element mutations that have been induced in an isogenic background may be a highly efficient method for functional genome annotation. RESULTS: Here, we have tested the efficacy of this strategy by assessing the extent to which screening for quantitative effects of P elements on sensory bristle number can identify genes affecting neural development. We find that such quantitative screens uncover an unusually large number of genes that are known to function in neural development, as well as genes with yet uncharacterized effects on neural development, and novel loci. CONCLUSIONS: Our findings establish the use of quantitative trait analysis for functional genome annotation through forward genetics. Similar analyses of quantitative effects of P element insertions will facilitate our understanding of the genes affecting many other complex traits in Drosophila.

Negative regulation of P element excision by the somatic product and terminal sequences of P in drosophila melanogaster

USDA-ARS?s Scientific Manuscript database

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...

Antisense protein tyrosine phosphatase 1B reverses activation of p38 mitogen-activated protein kinase in liver of ob/ob mice.

PubMed

Gum, Rebecca J; Gaede, Lori L; Heindel, Matthew A; Waring, Jeffrey F; Trevillyan, James M; Zinker, Bradley A; Stark, Margery E; Wilcox, Denise; Jirousek, Michael R; Rondinone, Cristina M; Ulrich, Roger G

2003-06-01

Phosphorylation of stress-activated kinase p38, a MAPK family member, was increased in liver of ob/ob diabetic mice relative to lean littermates. Treatment of ob/ob mice with protein tyrosine phosphatase 1B (PTP1B) antisense oligonucleotides (ASO) reduced phosphorylation of p38 in liver-to below lean littermate levels-and normalized plasma glucose while reducing plasma insulin. Phosphorylation of ERK, but not JNK, was also decreased in ASO-treated mice. PTP1B ASO decreased TNFalpha protein levels and phosphorylation of the transcription factor cAMP response element binding protein (CREB) in liver, both of which can occur through decreased phosphorylation of p38 and both of which have been implicated in insulin resistance or hyperglycemia. Decreased p38 phosphorylation was not directly due to decreased phosphorylation of the kinases that normally phosphorylate p38-MKK3 and MKK6. Additionally, p38 phosphorylation was not enhanced in liver upon insulin stimulation of ASO-treated ob/ob mice (despite increased activation of other signaling molecules) corroborating that p38 is not directly affected via the insulin receptor. Instead, decreased phosphorylation of p38 may be due to increased expression of MAPK phosphatases, particularly the p38/ERK phosphatase PAC1 (phosphatase of activated cells). This study demonstrates that reduction of PTP1B protein using ASO reduces activation of p38 and its substrates TNFalpha and CREB in liver of diabetic mice, which correlates with decreased hyperglycemia and hyperinsulinemia.

Genes encoding cuticular proteins are components of the Nimrod gene cluster in Drosophila.

PubMed

Cinege, Gyöngyi; Zsámboki, János; Vidal-Quadras, Maite; Uv, Anne; Csordás, Gábor; Honti, Viktor; Gábor, Erika; Hegedűs, Zoltán; Varga, Gergely I B; Kovács, Attila L; Juhász, Gábor; Williams, Michael J; Andó, István; Kurucz, Éva

2017-08-01

The Nimrod gene cluster, located on the second chromosome of Drosophila melanogaster, is the largest synthenic unit of the Drosophila genome. Nimrod genes show blood cell specific expression and code for phagocytosis receptors that play a major role in fruit fly innate immune functions. We previously identified three homologous genes (vajk-1, vajk-2 and vajk-3) located within the Nimrod cluster, which are unrelated to the Nimrod genes, but are homologous to a fourth gene (vajk-4) located outside the cluster. Here we show that, unlike the Nimrod candidates, the Vajk proteins are expressed in cuticular structures of the late embryo and the late pupa, indicating that they contribute to cuticular barrier functions. Copyright © 2017 Elsevier Ltd. All rights reserved.

Birth of a new gene on the Y chromosome of Drosophila melanogaster

PubMed Central

Carvalho, Antonio Bernardo; Vicoso, Beatriz; Russo, Claudia A. M.; Swenor, Bonnielin; Clark, Andrew G.

2015-01-01

Contrary to the pattern seen in mammalian sex chromosomes, where most Y-linked genes have X-linked homologs, the Drosophila X and Y chromosomes appear to be unrelated. Most of the Y-linked genes have autosomal paralogs, so autosome-to-Y transposition must be the main source of Drosophila Y-linked genes. Here we show how these genes were acquired. We found a previously unidentified gene (flagrante delicto Y, FDY) that originated from a recent duplication of the autosomal gene vig2 to the Y chromosome of Drosophila melanogaster. Four contiguous genes were duplicated along with vig2, but they became pseudogenes through the accumulation of deletions and transposable element insertions, whereas FDY remained functional, acquired testis-specific expression, and now accounts for ∼20% of the vig2-like mRNA in testis. FDY is absent in the closest relatives of D. melanogaster, and DNA sequence divergence indicates that the duplication to the Y chromosome occurred ∼2 million years ago. Thus, FDY provides a snapshot of the early stages of the establishment of a Y-linked gene and demonstrates how the Drosophila Y has been accumulating autosomal genes. PMID:26385968

p38 mitogen-activated protein kinase up-regulates NF-{kappa}B transcriptional activation through RelA phosphorylation during stretch-induced myogenesis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ji, Guoping; Liu, Dongxu; Liu, Jing

2010-01-01

p38 MAPK and nuclear factor-B (NF-B) signaling pathways play an indispensable role in the control of skeletal myogenesis. The specific contribution of these signaling pathways to the response of myoblast to the mechanical stimulation and the molecular mechanisms underlying this response remain unresolved. Using an established in vitro model, we now show that p38 MAP kinase activity regulates the transcriptional activation of NF-{kappa}B in response to mechanical stimulation of myoblasts. Furthermore, SB203580 blocked stretch-induced NF-{kappa}B activation during myogenesis, not through down-regulation of degradation of I{kappa}B-{alpha}, and consequent translocation of the p65 subunit of NF-{kappa}B to the nucleus. It is likelymore » that stretch-induced NF-{kappa}B activation by phosphorylation of p65 NF-{kappa}B. Moreover, depletion of p38{alpha} using siRNA significantly reduces stretch-induced phosphorylation of RelA and NF-{kappa}B activity. These results provides the first evidence of a cross-talk between p38 MAPK and NF-{kappa}B signaling pathways during stretch-induced myogenesis, with phosphorylation of RelA being one of the effectors of this promyogenic mechanism. The {alpha} isoform of p38MAP kinase regulates the transcriptional activation of NF-{kappa}B following stimulation with cyclic stretch.« less

The Evolution of Olfactory Gene Families in Drosophila and the Genomic Basis of chemical-Ecological Adaptation in Drosophila suzukii

PubMed Central

Ramasamy, Sukanya; Ometto, Lino; Crava, Cristina M.; Revadi, Santosh; Kaur, Rupinder; Horner, David S.; Pisani, Davide; Dekker, Teun; Anfora, Gianfranco; Rota-Stabelli, Omar

2016-01-01

How the evolution of olfactory genes correlates with adaption to new ecological niches is still a debated topic. We explored this issue in Drosophila suzukii, an emerging model that reproduces on fresh fruit rather than in fermenting substrates like most other Drosophila. We first annotated the repertoire of odorant receptors (ORs), odorant binding proteins (OBPs), and antennal ionotropic receptors (aIRs) in the genomes of two strains of D. suzukii and of its close relative Drosophila biarmipes. We then analyzed these genes on the phylogeny of 14 Drosophila species: whereas ORs and OBPs are characterized by higher turnover rates in some lineages including D. suzukii, aIRs are conserved throughout the genus. Drosophila suzukii is further characterized by a non-random distribution of OR turnover on the gene phylogeny, consistent with a change in selective pressures. In D. suzukii, we found duplications and signs of positive selection in ORs with affinity for short-chain esters, and loss of function of ORs with affinity for volatiles produced during fermentation. These receptors—Or85a and Or22a—are characterized by divergent alleles in the European and American genomes, and we hypothesize that they may have been replaced by some of the duplicated ORs in corresponding neurons, a hypothesis reciprocally confirmed by electrophysiological recordings. Our study quantifies the evolution of olfactory genes in Drosophila and reveals an array of genomic events that can be associated with the ecological adaptations of D. suzukii. PMID:27435796

Versatile P(acman) BAC Libraries for Transgenesis Studies in Drosophila melanogaster

DOE Office of Scientific and Technical Information (OSTI.GOV)

Venken, Koen J.T.; Carlson, Joseph W.; Schulze, Karen L.

2009-04-21

We constructed Drosophila melanogaster BAC libraries with 21-kb and 83-kb inserts in the P(acman) system. Clones representing 12-fold coverage and encompassing more than 95percent of annotated genes were mapped onto the reference genome. These clones can be integrated into predetermined attP sites in the genome using Phi C31 integrase to rescue mutations. They can be modified through recombineering, for example to incorporate protein tags and assess expression patterns.

Prostate Cancer Diagnostics and Prognostics Based on Interphase Spatial Genome Positioning

DTIC Science & Technology

2016-03-01

the Drosophila melanogaster genome at the ...and van Steensel, B. (2006). 1176 Characterization of the Drosophila melanogaster genome at the nuclear lamina. Nat Genet 38, 1177 1005-1014. doi...region according to the gene distribution pattern in primary genomic sequence . J Cell Biol 174:27–38 Therizols P, Illingworth RS, Courilleau C,

Delimiting regulatory sequences of the Drosophila melanogaster Ddc gene.

PubMed Central

Hirsh, J; Morgan, B A; Scholnick, S B

1986-01-01

We delimited sequences necessary for in vivo expression of the Drosophila melanogaster dopa decarboxylase gene Ddc. The expression of in vitro-altered genes was assayed following germ line integration via P-element vectors. Sequences between -209 and -24 were necessary for normally regulated expression, although genes lacking these sequences could be expressed at 10 to 50% of wild-type levels at specific developmental times. These genes showed components of normal developmental expression, which suggests that they retain some regulatory elements. All Ddc genes lacking the normal immediate 5'-flanking sequences were grossly deficient in larval central nervous system expression. Thus, this upstream region must contain at least one element necessary for this expression. A mutated Ddc gene without a normal TATA boxlike sequence used the normal RNA start points, indicating that this sequences is not required for start point specificity. Images PMID:3099170

KDM4B/JMJD2B is a p53 target gene that modulates the amplitude of p53 response after DNA damage

PubMed Central

Moon, Eui Jung; Razorenova, Olga V.; Krieg, Adam J.; von Eyben, Rie

2017-01-01

Abstract The p53 tumor suppressor protein plays a critical role in orchestrating the genomic response to various stress signals by acting as a master transcriptional regulator. Differential gene activity is controlled by transcription factors but also dependent on the underlying chromatin structure, especially on covalent histone modifications. After screening different histone lysine methyltransferases and demethylases, we identified JMJD2B/KDM4B as a p53-inducible gene in response to DNA damage. p53 directly regulates JMJD2B gene expression by binding to a canonical p53-consensus motif in the JMJD2B promoter. JMJD2B induction attenuates the transcription of key p53 transcriptional targets including p21, PIG3 and PUMA, and this modulation is dependent on the catalytic capacity of JMJD2B. Conversely, JMJD2B silencing led to an enhancement of the DNA-damage driven induction of p21 and PIG3. These findings indicate that JMJD2B acts in an auto-regulatory loop by which p53, through JMJD2B activation, is able to influence its own transcriptional program. Functionally, exogenous expression of JMJD2B enhanced subcutaneous tumor growth of colon cancer cells in a p53-dependent manner, and genetic inhibition of JMJD2B impaired tumor growth in vivo. These studies provide new insights into the regulatory effect exerted by JMJD2B on tumor growth through the modulation of p53 target genes. PMID:28073943

The Evolution of Olfactory Gene Families in Drosophila and the Genomic Basis of chemical-Ecological Adaptation in Drosophila suzukii.

PubMed

Ramasamy, Sukanya; Ometto, Lino; Crava, Cristina M; Revadi, Santosh; Kaur, Rupinder; Horner, David S; Pisani, Davide; Dekker, Teun; Anfora, Gianfranco; Rota-Stabelli, Omar

2016-08-16

How the evolution of olfactory genes correlates with adaption to new ecological niches is still a debated topic. We explored this issue in Drosophila suzukii, an emerging model that reproduces on fresh fruit rather than in fermenting substrates like most other Drosophila We first annotated the repertoire of odorant receptors (ORs), odorant binding proteins (OBPs), and antennal ionotropic receptors (aIRs) in the genomes of two strains of D. suzukii and of its close relative Drosophila biarmipes We then analyzed these genes on the phylogeny of 14 Drosophila species: whereas ORs and OBPs are characterized by higher turnover rates in some lineages including D. suzukii, aIRs are conserved throughout the genus. Drosophila suzukii is further characterized by a non-random distribution of OR turnover on the gene phylogeny, consistent with a change in selective pressures. In D. suzukii, we found duplications and signs of positive selection in ORs with affinity for short-chain esters, and loss of function of ORs with affinity for volatiles produced during fermentation. These receptors-Or85a and Or22a-are characterized by divergent alleles in the European and American genomes, and we hypothesize that they may have been replaced by some of the duplicated ORs in corresponding neurons, a hypothesis reciprocally confirmed by electrophysiological recordings. Our study quantifies the evolution of olfactory genes in Drosophila and reveals an array of genomic events that can be associated with the ecological adaptations of D. suzukii. © The Author(s) 2016. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

P38 mitogen-activated protein kinase (p38 MAPK) overexpression in clinical staging of nasopharyngeal carcinoma

NASA Astrophysics Data System (ADS)

Farhat; Asnir, R. A.; Yudhistira, A.; Daulay, E. R.; Muzakkir, M. M.; Yulius, S.

2018-03-01

Molecular biological research on nasopharyngeal carcinoma has been widely practiced, such as VEGF, EGFR, COX-2 expression and so on. MAPK plays a role in cell growth such as proliferation, differentiation, and apoptosis, primarily contributing to gene expression, where p38 MAPK pathway mostly associate with anti-apoptosis and cause cell transformation. The aim of this study is to determine the expression of p38 MAPK in clinical stage of nasopharyngeal carcinoma so that the result can be helpful in prognosis and adjunctive therapy in nasopharyngeal carcinoma. The research design is descriptive. It was done in THT- KL Department of FK USU/RSUP Haji Adam Malik, Medan and Pathology Anatomical Department of FK USU. The study was conducted from December 2011 to May 2012. The Samples are all patients who diagnosed with nasopharyngeal carcinoma in oncology division of Otorhinolaryngology Department. p38 MAPK overexpression was found in 21 samples (70%) from 30 nasopharyngeal carcinoma samples. The elevated of p38 MAPK expression most found on T4 by eight samples (38.1%), N3 lymph node group by nine samples (42.9%), stage IV of clinical staging is as many as 15 samples (71.4%). p38 MAPK most expressed in stage IV clinical staging of patients with nasopharyngeal carcinoma.

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

PubMed

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.

BMPs Regulate msx Gene Expression in the Dorsal Neuroectoderm of Drosophila and Vertebrates by Distinct Mechanisms

PubMed Central

Esteves, Francisco F.; Taylor, Erika; Pyrowolakis, George; Fisher, Shannon; Bier, Ethan

2014-01-01

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. PMID:25210771

Gene Regulation Networks for Modeling Drosophila Development

NASA Technical Reports Server (NTRS)

Mjolsness, E.

1999-01-01

This chapter will very briefly introduce and review some computational experiments in using trainable gene regulation network models to simulate and understand selected episodes in the development of the fruit fly, Drosophila Melanogaster.

Mycobacterium tuberculosis PE13 (Rv1195) manipulates the host cell fate via p38-ERK-NF-κB axis and apoptosis.

PubMed

Li, Hui; Li, Qiming; Yu, Zhaoxiao; Zhou, Mingliang; Xie, Jianping

2016-07-01

PE/PPE family proteins are mycobacteria unique molecules, named after their N-terminal conserved PE (Pro-Glu) and PPE (Pro-Pro-Glu) domains. Mycobacterium tuberculosis (Mtb) PE family gene encoded cell surface proteins are previously reported to be involved in virulence and interaction with host. To explore the role of a novel PE member (PE13, Rv1195), M. smegmatis was used as surrogate host. The study showed that Rv1195 was a cell wall associated protein. Rv1195 can enhance the survival of recombinants under stress conditions such as H2O2, SDS, low pH. This is largely due to the upregulated transcription of Rv1195, since diverse stresses can increase the promoter activity of Rv1195 gene, consistent with enhanced survival within macrophages. Ms_Rv1195 infection also increased the production of interlukin-6 (IL-6) and IL-1β from macrophages, while decreased the secretion of suppressor of cytokine signaling 3 (SOCS3) in comparison with the vector-only control. The cell death was also precipitated by the Ms_Rv1195 infection. Inhibitors treatment showed that the p38-ERK-NF-κB axis was involved in the Rv1195 triggered change of IL-6 and IL-1β expression. In summary, we showed that PE13 (Rv1195) is a new PE family member actively engaged in the interaction between Mycobacterium and host, signaling through p38-ERK-NF-κB axis and apoptosis.

Drosophila Shaking-B protein forms gap junctions in paired Xenopus oocytes.

PubMed

Phelan, P; Stebbings, L A; Baines, R A; Bacon, J P; Davies, J A; Ford, C

1998-01-08

In most multicellular organisms direct cell-cell communication is mediated by the intercellular channels of gap junctions. These channels allow the exchange of ions and molecules that are believed to be essential for cell signalling during development and in some differentiated tissues. Proteins called connexins, which are products of a multigene family, are the structural components of vertebrate gap junctions. Surprisingly, molecular homologues of the connexins have not been described in any invertebrate. A separate gene family, which includes the Drosophila genes shaking-B and l(1)ogre, and the Caenorhabditis elegans genes unc-7 and eat-5, encodes transmembrane proteins with a predicted structure similar to that of the connexins. shaking-B and eat-5 are required for the formation of functional gap junctions. To test directly whether Shaking-B is a channel protein, we expressed it in paired Xenopus oocytes. Here we show that Shaking-B localizes to the membrane, and that its presence induces the formation of functional intercellular channels. To our knowledge, this is the first structural component of an invertebrate gap junction to be characterized.

MiMIC: a highly versatile transposon insertion resource for engineering Drosophila melanogaster genes

PubMed Central

Venken, Koen J. T.; Schulze, Karen L.; Haelterman, Nele A.; Pan, Hongling; He, Yuchun; Evans-Holm, Martha; Carlson, Joseph W.; Levis, Robert W.; Spradling, Allan C.; Hoskins, Roger A.; Bellen, Hugo J.

2011-01-01

We demonstrate the versatility of a collection of insertions of the transposon Minos mediated integration cassette (MiMIC), in Drosophila melanogaster. MiMIC contains a gene-trap cassette and the yellow+ marker flanked by two inverted bacteriophage ΦC31 attP sites. MiMIC integrates almost at random in the genome to create sites for DNA manipulation. The attP sites allow the replacement of the intervening sequence of the transposon with any other sequence through recombinase mediated cassette exchange (RMCE). We can revert insertions that function as gene traps and cause mutant phenotypes to wild type by RMCE and modify insertions to control GAL4 or QF overexpression systems or perform lineage analysis using the Flp system. Insertions within coding introns can be exchanged with protein-tag cassettes to create fusion proteins to follow protein expression and perform biochemical experiments. The applications of MiMIC vastly extend the Drosophila melanogaster toolkit. PMID:21985007

Comparative genome sequencing of Drosophila pseudoobscura: Chromosomal, gene, and cis-element evolution

PubMed Central

Richards, Stephen; Liu, Yue; Bettencourt, Brian R.; Hradecky, Pavel; Letovsky, Stan; Nielsen, Rasmus; Thornton, Kevin; Hubisz, Melissa J.; Chen, Rui; Meisel, Richard P.; Couronne, Olivier; Hua, Sujun; Smith, Mark A.; Zhang, Peili; Liu, Jing; Bussemaker, Harmen J.; van Batenburg, Marinus F.; Howells, Sally L.; Scherer, Steven E.; Sodergren, Erica; Matthews, Beverly B.; Crosby, Madeline A.; Schroeder, Andrew J.; Ortiz-Barrientos, Daniel; Rives, Catharine M.; Metzker, Michael L.; Muzny, Donna M.; Scott, Graham; Steffen, David; Wheeler, David A.; Worley, Kim C.; Havlak, Paul; Durbin, K. James; Egan, Amy; Gill, Rachel; Hume, Jennifer; Morgan, Margaret B.; Miner, George; Hamilton, Cerissa; Huang, Yanmei; Waldron, Lenée; Verduzco, Daniel; Clerc-Blankenburg, Kerstin P.; Dubchak, Inna; Noor, Mohamed A.F.; Anderson, Wyatt; White, Kevin P.; Clark, Andrew G.; Schaeffer, Stephen W.; Gelbart, William; Weinstock, George M.; Gibbs, Richard A.

2005-01-01

We have sequenced the genome of a second Drosophila species, Drosophila pseudoobscura, and compared this to the genome sequence of Drosophila melanogaster, a primary model organism. Throughout evolution the vast majority of Drosophila genes have remained on the same chromosome arm, but within each arm gene order has been extensively reshuffled, leading to a minimum of 921 syntenic blocks shared between the species. A repetitive sequence is found in the D. pseudoobscura genome at many junctions between adjacent syntenic blocks. Analysis of this novel repetitive element family suggests that recombination between offset elements may have given rise to many paracentric inversions, thereby contributing to the shuffling of gene order in the D. pseudoobscura lineage. Based on sequence similarity and synteny, 10,516 putative orthologs have been identified as a core gene set conserved over 25–55 million years (Myr) since the pseudoobscura/melanogaster divergence. Genes expressed in the testes had higher amino acid sequence divergence than the genome-wide average, consistent with the rapid evolution of sex-specific proteins. Cis-regulatory sequences are more conserved than random and nearby sequences between the species—but the difference is slight, suggesting that the evolution of cis-regulatory elements is flexible. Overall, a pattern of repeat-mediated chromosomal rearrangement, and high coadaptation of both male genes and cis-regulatory sequences emerges as important themes of genome divergence between these species of Drosophila. PMID:15632085

The Drosophila ETV5 Homologue Ets96B: Molecular Link between Obesity and Bipolar Disorder.

PubMed

Williams, Michael J; Klockars, Anica; Eriksson, Anders; Voisin, Sarah; Dnyansagar, Rohit; Wiemerslage, Lyle; Kasagiannis, Anna; Akram, Mehwish; Kheder, Sania; Ambrosi, Valerie; Hallqvist, Emilie; Fredriksson, Robert; Schiöth, Helgi B

2016-06-01

Several reports suggest obesity and bipolar disorder (BD) share some physiological and behavioural similarities. For instance, obese individuals are more impulsive and have heightened reward responsiveness, phenotypes associated with BD, while bipolar patients become obese at a higher rate and earlier age than people without BD; however, the molecular mechanisms of such an association remain obscure. Here we demonstrate, using whole transcriptome analysis, that Drosophila Ets96B, homologue of obesity-linked gene ETV5, regulates cellular systems associated with obesity and BD. Consistent with a role in obesity and BD, loss of nervous system Ets96B during development increases triacylglyceride concentration, while inducing a heightened startle-response, as well as increasing hyperactivity and reducing sleep. Of notable interest, mouse Etv5 and Drosophila Ets96B are expressed in dopaminergic-rich regions, and loss of Ets96B specifically in dopaminergic neurons recapitulates the metabolic and behavioural phenotypes. Moreover, our data indicate Ets96B inhibits dopaminergic-specific neuroprotective systems. Additionally, we reveal that multiple SNPs in human ETV5 link to body mass index (BMI) and BD, providing further evidence for ETV5 as an important and novel molecular intermediate between obesity and BD. We identify a novel molecular link between obesity and bipolar disorder. The Drosophila ETV5 homologue Ets96B regulates the expression of cellular systems with links to obesity and behaviour, including the expression of a conserved endoplasmic reticulum molecular chaperone complex known to be neuroprotective. Finally, a connection between the obesity-linked gene ETV5 and bipolar disorder emphasizes a functional relationship between obesity and BD at the molecular level.

The pink gene encodes the Drosophila orthologue of the human Hermansky-Pudlak syndrome 5 (HPS5) gene.

PubMed

Syrzycka, Monika; McEachern, Lori A; Kinneard, Jennifer; Prabhu, Kristel; Fitzpatrick, Kathleen; Schulze, Sandra; Rawls, John M; Lloyd, Vett K; Sinclair, Donald A R; Honda, Barry M

2007-06-01

Hermansky-Pudlak syndrome (HPS) consists of a set of human autosomal recessive disorders, with symptoms resulting from defects in genes required for protein trafficking in lysosome-related organelles such as melanosomes and platelet dense granules. A number of human HPS genes and rodent orthologues have been identified whose protein products are key components of 1 of 4 different protein complexes (AP-3 or BLOC-1, -2, and -3) that are key participants in the process. Drosophila melanogaster has been a key model organism in demonstrating the in vivo significance of many genes involved in protein trafficking pathways; for example, mutations in the "granule group" genes lead to changes in eye colour arising from improper protein trafficking to pigment granules in the developing eye. An examination of the chromosomal positioning of Drosophila HPS gene orthologues suggested that CG9770, the Drosophila HPS5 orthologue, might correspond to the pink locus. Here we confirm this gene assignment, making pink the first eye colour gene in flies to be identified as a BLOC complex gene.

Colonization of heterochromatic genes by transposable elements in Drosophila.

PubMed

Dimitri, Patrizio; Junakovic, Nikolaj; Arcà, Bruno

2003-04-01

As a further step toward understanding transposable element-host genome interactions, we investigated the molecular anatomy of introns from five heterochromatic and 22 euchromatic protein-coding genes of Drosophila melanogaster. A total of 79 kb of intronic sequences from heterochromatic genes and 355 kb of intronic sequences from euchromatic genes have been used in Blast searches against Drosophila transposable elements (TEs). The results show that TE-homologous sequences belonging to 19 different families represent about 50% of intronic DNA from heterochromatic genes. In contrast, only 0.1% of the euchromatic intron DNA exhibits homology to known TEs. Intraspecific and interspecific size polymorphisms of introns were found, which are likely to be associated with changes in TE-related sequences. Together, the enrichment in TEs and the apparent dynamic state of heterochromatic introns suggest that TEs contribute significantly to the evolution of genes located in heterochromatin.

Evidence for the Involvement of p38 MAPK Activation in Barnacle Larval Settlement

PubMed Central

He, Li-Sheng; Xu, Ying; Matsumura, Kiyotaka; Zhang, Yu; Zhang, Gen; Qi, Shu-Hua; Qian, Pei-Yuan

2012-01-01

The barnacle Balanus ( = Amphibalanus) amphitrite is a major marine fouling animal. Understanding the molecular mechanism of larval settlement in this species is critical for anti-fouling research. In this study, we cloned one isoform of p38 MAPK (Bar-p38 MAPK) from this species, which shares the significant characteristic of containing a TGY motif with other species such as yeast, Drosophila and humans. The activation of p38 MAPK was detected by an antibody that recognizes the conserved dual phosphorylation sites of TGY. The results showed that phospho-p38 MAPK (pp38 MAPK) was more highly expressed at the cyprid stage, particularly in aged cyprids, in comparison to other stages, including the nauplius and juvenile stages. Immunostaining showed that Bar-p38 MAPK and pp38 MAPK were mainly located at the cyprid antennules, and especially the third and fourth segments, which are responsible for substratum exploration during settlement. The expression and localization patterns of Bar-p38 MAPK suggest its involvement in larval settlement. This postulation was also supported by the larval settlement bioassay with the p38 MAPK inhibitor SB203580. Behavioral analysis by live imaging revealed that the larvae were still capable of exploring the surface of the substratum after SB203580 treatment. This shows that the effect of p38 MAPK on larval settlement might be by regulating the secretion of permanent proteinaceous substances. Furthermore, the level of pp38 MAPK dramatically decreased after full settlement, suggesting that Bar-p38 MAPK maybe plays a role in larval settlement rather than metamorphosis. Finally, we found that Bar-p38 MAPK was highly activated when larvae confronted extracts of adult barnacle containing settlement cues, whereas larvae pre-treated with SB203580 failed to respond to the crude adult extracts. PMID:23115639

Lack of robustness of life extension associated with several single-gene P element mutations in Drosophila melanogaster.

PubMed

Mockett, Robin J; Nobles, Amber C

2013-10-01

The hypothesis tested in this study was that single-gene mutations found previously to extend the life span of Drosophila melanogaster could do so consistently in both long-lived y w and standard w (1118) genetic backgrounds. GAL4 drivers were used to express upstream activation sequence (UAS)-responder transgenes globally or in the nervous system. Transgenes associated with oxidative damage prevention (UAS-hSOD1 and UAS-GCLc) or removal (EP-UAS-Atg8a and UAS-dTOR (FRB) ) failed to increase mean life spans in any expression pattern in either genetic background. Flies containing a UAS-EGFP-bMSRA (C) transgene associated with protein repair were found not to exhibit life extension or detectable enhanced green fluorescent protein (EGFP) activity. The presence of UAS-responder transgenes was confirmed by PCR amplification and sequencing at the 5' and 3' end of each insertion. These results cast doubt on the robustness of life extension in flies carrying single-gene mutations and suggest that the effects of all such mutations should be tested independently in multiple genetic backgrounds and laboratory environments.

Structure-based design, synthesis, and biological evaluation of imidazo[1,2-b]pyridazine-based p38 MAP kinase inhibitors.

PubMed

Kaieda, Akira; Takahashi, Masashi; Takai, Takafumi; Goto, Masayuki; Miyazaki, Takahiro; Hori, Yuri; Unno, Satoko; Kawamoto, Tomohiro; Tanaka, Toshimasa; Itono, Sachiko; Takagi, Terufumi; Hamada, Teruki; Shirasaki, Mikio; Okada, Kengo; Snell, Gyorgy; Bragstad, Ken; Sang, Bi-Ching; Uchikawa, Osamu; Miwatashi, Seiji

2018-02-01

We identified novel potent inhibitors of p38 MAP kinase using structure-based design strategy. X-ray crystallography showed that when p38 MAP kinase is complexed with TAK-715 (1) in a co-crystal structure, Phe169 adopts two conformations, where one interacts with 1 and the other shows no interaction with 1. Our structure-based design strategy shows that these two conformations converge into one via enhanced protein-ligand hydrophobic interactions. According to the strategy, we focused on scaffold transformation to identify imidazo[1,2-b]pyridazine derivatives as potent inhibitors of p38 MAP kinase. Among the herein described and evaluated compounds, N-oxide 16 exhibited potent inhibition of p38 MAP kinase and LPS-induced TNF-α production in human monocytic THP-1 cells, and significant in vivo efficacy in rat collagen-induced arthritis models. In this article, we report the discovery of potent, selective and orally bioavailable imidazo[1,2-b]pyridazine-based p38 MAP kinase inhibitors with pyridine N-oxide group. Copyright © 2018 Elsevier Ltd. All rights reserved.

The gene transformer-2 of Sciara (Diptera, Nematocera) and its effect on Drosophila sexual development.

PubMed

Martín, Iker; Ruiz, María F; Sánchez, Lucas

2011-03-15

The gene transformer-2, which is involved in sex determination, has been studied in Drosophila, Musca, Ceratitis, Anastrepha and Lucilia. All these members of Diptera belong to the suborder Brachycera. In this work, it is reported the isolation and characterisation of genes transformer-2 of the dipterans Sciara ocellaris and Bradysia coprophila (formerly Sciara coprophila), which belong to the much less extensively analysed Sciaridae Family of the Suborder Nematocera, which is paraphyletic with respect to Suborder Brachycera. The transformer-2 genes of the studied Sciara species were found to be transcribed in both sexes during development and adult life, in both the soma and germ lines. They produced a single primary transcript, which follows the same alternative splicing in both sexes, giving rise to different mRNAs isoforms. In S. ocellaris the most abundant mRNA isoform encoded a full-length protein of 251 amino acids, while that of B. coprophila encoded a protein of 246 amino acids. Both showed the features of the SR protein family. The less significant mRNA isoforms of both species encoded truncated, presumably non-functional Transformer-2 proteins. The comparison of the functional Sciara Transformer-2 proteins among themselves and those of other insects revealed the greatest degree of conservation in the RRM domain and linker region. In contrast, the RS1 and RS2 domains showed extensive variation with respect to their number of amino acids and their arginine-serine (RS) dipeptide content. The expression of S. ocellaris Transformer-2 protein in Drosophila XX pseudomales lacking the endogenous transformer-2 function caused their partial feminisation. The transformer-2 genes of both Sciaridae species encode a single protein in both sexes that shares the characteristics of the Transformer-2 proteins of other insects. These proteins showed conserved sex-determination function in Drosophila; i.e., they were able to form a complex with the endogenous Drosophila

The gene transformer-2 of Sciara (Diptera, Nematocera) and its effect on Drosophila sexual development

PubMed Central

2011-01-01

Background The gene transformer-2, which is involved in sex determination, has been studied in Drosophila, Musca, Ceratitis, Anastrepha and Lucilia. All these members of Diptera belong to the suborder Brachycera. In this work, it is reported the isolation and characterisation of genes transformer-2 of the dipterans Sciara ocellaris and Bradysia coprophila (formerly Sciara coprophila), which belong to the much less extensively analysed Sciaridae Family of the Suborder Nematocera, which is paraphyletic with respect to Suborder Brachycera. Results The transformer-2 genes of the studied Sciara species were found to be transcribed in both sexes during development and adult life, in both the soma and germ lines. They produced a single primary transcript, which follows the same alternative splicing in both sexes, giving rise to different mRNAs isoforms. In S. ocellaris the most abundant mRNA isoform encoded a full-length protein of 251 amino acids, while that of B. coprophila encoded a protein of 246 amino acids. Both showed the features of the SR protein family. The less significant mRNA isoforms of both species encoded truncated, presumably non-functional Transformer-2 proteins. The comparison of the functional Sciara Transformer-2 proteins among themselves and those of other insects revealed the greatest degree of conservation in the RRM domain and linker region. In contrast, the RS1 and RS2 domains showed extensive variation with respect to their number of amino acids and their arginine-serine (RS) dipeptide content. The expression of S. ocellaris Transformer-2 protein in Drosophila XX pseudomales lacking the endogenous transformer-2 function caused their partial feminisation. Conclusions The transformer-2 genes of both Sciaridae species encode a single protein in both sexes that shares the characteristics of the Transformer-2 proteins of other insects. These proteins showed conserved sex-determination function in Drosophila; i.e., they were able to form a complex

A novel role of Drosophila cytochrome P450-4e3 in permethrin insecticide tolerance.

PubMed

Terhzaz, Selim; Cabrero, Pablo; Brinzer, Robert A; Halberg, Kenneth A; Dow, Julian A T; Davies, Shireen-A

2015-12-01

The exposure of insects to xenobiotics, such as insecticides, triggers a complex defence response necessary for survival. This response includes the induction of genes that encode key Cytochrome P450 monooxygenase detoxification enzymes. Drosophila melanogaster Malpighian (renal) tubules are critical organs in the detoxification and elimination of these foreign compounds, so the tubule response induced by dietary exposure to the insecticide permethrin was examined. We found that expression of the gene encoding Cytochrome P450-4e3 (Cyp4e3) is significantly up-regulated by Drosophila fed on permethrin and that manipulation of Cyp4e3 levels, specifically in the principal cells of the Malpighian tubules, impacts significantly on the survival of permethrin-fed flies. Both dietary exposure to permethrin and Cyp4e3 knockdown cause a significant elevation of oxidative stress-associated markers in the tubules, including H2O2 and lipid peroxidation byproduct, HNE (4-hydroxynonenal). Thus, Cyp4e3 may play an important role in regulating H2O2 levels in the endoplasmic reticulum (ER) where it resides, and its absence triggers a JAK/STAT and NF-κB-mediated stress response, similar to that observed in cells under ER stress. This work increases our understanding of the molecular mechanisms of insecticide detoxification and provides further evidence of the oxidative stress responses induced by permethrin metabolism. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

A Test for Gene Flow among Sympatric and Allopatric Hawaiian Picture-Winged Drosophila.

PubMed

Kang, Lin; Garner, Harold R; Price, Donald K; Michalak, Pawel

2017-06-01

The Hawaiian Drosophila are one of the most species-rich endemic groups in Hawaii and a spectacular example of adaptive radiation. Drosophila silvestris and D. heteroneura are two closely related picture-winged Drosophila species that occur sympatrically on Hawaii Island and are known to hybridize in nature, yet exhibit highly divergent behavioral and morphological traits driven largely through sexual selection. Their closest-related allopatric species, D. planitibia from Maui, exhibits hybrid male sterility and reduced behavioral reproductive isolation when crossed experimentally with D. silvestris or D. heteroneura. A modified four-taxon test for gene flow was applied to recently obtained genomes of the three Hawaiian Drosophila species. The analysis indicates recent gene flow in sympatry, but also, although less extensive, between allopatric species. This study underscores the prevalence of gene flow, even in taxonomic groups considered classic examples of allopatric speciation on islands. The potential confounding effects of gene flow in phylogenetic and population genetics inference are discussed, as well as the implications for conservation.

Cytogenetic mapping of the Muller F element genes in Drosophila willistoni group.

PubMed

Pita, Sebastián; Panzera, Yanina; Lúcia da Silva Valente, Vera; de Melo, Zilpa das Graças Silva; Garcia, Carolina; Garcia, Ana Cristina Lauer; Montes, Martín Alejandro; Rohde, Claudia

2014-10-01

Comparative genomics in Drosophila began in 1940, when Muller stated that the ancestral haploid karyotype of this genus is constituted by five acrocentric chromosomes and one dot chromosome, named A to F elements. In some species of the willistoni group such as Drosophila willistoni and D. insularis, the F element, instead of a dot chromosome, has been incorporated into the E element, forming chromosome III (E + F fusion). The aim of this study was to investigate the scope of the E + F fusion in the willistoni group, evaluating six other species. Fluorescent in situ hybridization was used to locate two genes of the F element previously studied-cubitus interruptus (ci) and eyeless (ey)-in species of the willistoni and bocainensis subgroups. Moreover, polytene chromosome photomaps corresponding to the F element (basal portion of chromosome III) were constructed for each species studied. In D. willistoni, D. paulistorum and D. equinoxialis, the ci gene was located in subSectction 78B and the ey gene in 78C. In D. tropicalis, ci was located in subSection 76B and ey in 76C. In species of the bocainensis subgroup, ci and ey were localized, respectively, at subsections 76B and 76C in D. nebulosa and D. capricorni, and 76A and 76C in D. fumipennis. Despite the differences in the subsection numbers, all species showed the same position for ci and ey. The results confirm the synteny of E + F fusion in willistoni and bocainensis subgroups, and allow estimating the occurrence of this event at 15 Mya, at least.

Phylogeny, integration and expression of sigma virus-like genes in Drosophila.

PubMed

Ballinger, Matthew J; Bruenn, Jeremy A; Taylor, Derek J

2012-10-01

The recent and surprising discovery of widespread NIRVs (non-retroviral integrated RNA viruses) has highlighted the importance of genomic interactions between non-retroviral RNA viruses and their eukaryotic hosts. Among the viruses with integrated representatives are the rhabdoviruses, a family of negative sense single-stranded RNA viruses. We identify sigma virus-like NIRVs of Drosophila spp. that represent unique cases where NIRVs are closely related to exogenous RNA viruses in a model host organism. We have used a combination of bioinformatics and laboratory methods to explore the evolution and expression of sigma virus-like NIRVs in Drosophila. Recent integrations in Drosophila provide a promising experimental system to study functionality of NIRVs. Moreover, the genomic architecture of recent NIRVs provides an unusual evolutionary window on the integration mechanism. For example, we found that a sigma virus-like polymerase associated protein (P) gene appears to have been integrated by template switching of the blastopia-like LTR retrotransposon. The sigma virus P-like NIRV is present in multiple retroelement fused open reading frames on the X and 3R chromosomes of Drosophila yakuba - the X-linked copy is transcribed to produce an RNA product in adult flies. We present the first account of sigma virus-like NIRVs and the first example of NIRV expression in a model animal system, and therefore provide a platform for further study of the possible functions of NIRVs in animal hosts. Copyright © 2012 Elsevier Inc. All rights reserved.

Occurrence of the structural enterocin A, P, B, L50B genes in enterococci of different origin.

PubMed

Strompfová, Viola; Lauková, Andrea; Simonová, Monika; Marcináková, Miroslava

2008-12-10

Enterococci are well-known producers of antimicrobial peptides--bacteriocins (enterocins) and the number of characterized enterocins has been significantly increased. Recently, enterocins are of great interest for their potential as biopreservatives in food or feed while research on enterocins as alternative antimicrobials in humans and animals is only at the beginning. The present study provides a survey about the occurrence of enterocin structural genes A, P, B, L50B in a target of 427 strains of Enterococcus faecium (368) and Enterococcus faecalis (59) species from different sources (animal isolates, food and feed) performed by PCR method. Based on our results, 234 strains possessed one or more enterocin structural gene(s). The genes of enterocin P and enterocin A were the most frequently detected structural genes among the PCR positive strains (170 and 155 strains, respectively). Different frequency of the enterocin genes occurrence was detected in strains according to their origin; the strains from horses and silage showed the highest frequency of enterocin genes presence. All possible combinations of the tested genes occurred at least twice except the combination of the gene of enterocin B and L50B which possessed neither strain. The gene of enterocin A was exclusively detected among E. faecium strains, while the gene of enterocin P, B, L50B were detected in strains of both species E. faecium and E. faecalis. In conclusion, a high-frequency and variability of enterocin structural genes exists among enterococci of different origin what offers a big possibility to find effective bacteriocin-producing strains for their application in veterinary medicine.

Monitoring autophagic flux using Ref(2)P, the Drosophila p62 ortholog.

PubMed

DeVorkin, Lindsay; Gorski, Sharon M

2014-09-02

Human p62, also known as Sequestome-1 (SQSTM1), is a multifunctional scaffold protein that contains many domains, including a Phox/Bem1P (PB1) multimerization domain, an ubiquitin-associated (UBA) domain, and a light chain 3 (LC3) recognition sequence. p62 binds ubiquitinated proteins and targets them for degradation by the proteasome. In addition, p62 directly binds LC3; this may serve as a mechanism to deliver ubiquitinated proteins for degradation by autophagy. During this process, p62 itself is degraded. The inhibition of autophagy leads to the accumulation of p62, indicating that it can be used as a marker of autophagic flux. Ref(2)P (refractory to sigma P), the Drosophila ortholog of p62, is also required for the formation of ubiquitinated protein aggregates. Ref(2)P contains a putative LC3-interacting region, and genetic inhibition of autophagy in Drosophila leads to the accumulation of Ref(2)P protein levels. Thus, like p62, Ref(2)P may serve as a marker of autophagic flux. Here we provide two procedures to examine Ref(2)P protein levels in Drosophila ovaries. © 2014 Cold Spring Harbor Laboratory Press.

Acidic Food pH Increases Palatability and Consumption and Extends Drosophila Lifespan.

PubMed

Deshpande, Sonali A; Yamada, Ryuichi; Mak, Christine M; Hunter, Brooke; Soto Obando, Alina; Hoxha, Sany; Ja, William W

2015-12-01

Despite the prevalent use of Drosophila as a model in studies of nutrition, the effects of fundamental food properties, such as pH, on animal health and behavior are not well known. We examined the effect of food pH on adult Drosophila lifespan, feeding behavior, and microbiota composition and tested the hypothesis that pH-mediated changes in palatability and total consumption are required for modulating longevity. We measured the effect of buffered food (pH 5, 7, or 9) on male gustatory responses (proboscis extension), total food intake, and male and female lifespan. The effect of food pH on germfree male lifespan was also assessed. Changes in fly-associated microbial composition as a result of food pH were determined by 16S ribosomal RNA gene sequencing. Male gustatory responses, total consumption, and male and female longevity were additionally measured in the taste-defective Pox neuro (Poxn) mutant and its transgenic rescue control. An acidic diet increased Drosophila gustatory responses (40-230%) and food intake (5-50%) and extended survival (10-160% longer median lifespan) compared with flies on either neutral or alkaline pH food. Alkaline food pH shifted the composition of fly-associated bacteria and resulted in greater lifespan extension (260% longer median survival) after microbes were eliminated compared with flies on an acidic (50%) or neutral (130%) diet. However, germfree flies lived longer on an acidic diet (5-20% longer median lifespan) compared with those on either neutral or alkaline pH food. Gustatory responses, total consumption, and longevity were unaffected by food pH in Poxn mutant flies. Food pH can directly influence palatability and feeding behavior and affect parameters such as microbial growth to ultimately affect Drosophila lifespan. Fundamental food properties altered by dietary or drug interventions may therefore contribute to changes in animal physiology, metabolism, and survival. © 2015 American Society for Nutrition.

SUMO-Enriched Proteome for Drosophila Innate Immune Response

PubMed Central

Handu, Mithila; Kaduskar, Bhagyashree; Ravindranathan, Ramya; Soory, Amarendranath; Giri, Ritika; Elango, Vijay Barathi; Gowda, Harsha; Ratnaparkhi, Girish S.

2015-01-01

Small ubiquitin-like modifier (SUMO) modification modulates the expression of defense genes in Drosophila, activated by the Toll/nuclear factor-κB and immune-deficient/nuclear factor-κB signaling networks. We have, however, limited understanding of the SUMO-modulated regulation of the immune response and lack information on SUMO targets in the immune system. In this study, we measured the changes to the SUMO proteome in S2 cells in response to a lipopolysaccharide challenge and identified 1619 unique proteins in SUMO-enriched lysates. A confident set of 710 proteins represents the immune-induced SUMO proteome and analysis suggests that specific protein domains, cellular pathways, and protein complexes respond to immune stress. A small subset of the confident set was validated by in-bacto SUMOylation and shown to be bona-fide SUMO targets. These include components of immune signaling pathways such as Caspar, Jra, Kay, cdc42, p38b, 14-3-3ε, as well as cellular proteins with diverse functions, many being components of protein complexes, such as prosß4, Rps10b, SmD3, Tango7, and Aats-arg. Caspar, a human FAF1 ortholog that negatively regulates immune-deficient signaling, is SUMOylated at K551 and responds to treatment with lipopolysaccharide in cultured cells. Our study is one of the first to describe SUMO proteome for the Drosophila immune response. Our data and analysis provide a global framework for the understanding of SUMO modification in the host response to pathogens. PMID:26290570

SUMO-Enriched Proteome for Drosophila Innate Immune Response.

PubMed

Handu, Mithila; Kaduskar, Bhagyashree; Ravindranathan, Ramya; Soory, Amarendranath; Giri, Ritika; Elango, Vijay Barathi; Gowda, Harsha; Ratnaparkhi, Girish S

2015-08-18

Small ubiquitin-like modifier (SUMO) modification modulates the expression of defense genes in Drosophila, activated by the Toll/nuclear factor-κB and immune-deficient/nuclear factor-κB signaling networks. We have, however, limited understanding of the SUMO-modulated regulation of the immune response and lack information on SUMO targets in the immune system. In this study, we measured the changes to the SUMO proteome in S2 cells in response to a lipopolysaccharide challenge and identified 1619 unique proteins in SUMO-enriched lysates. A confident set of 710 proteins represents the immune-induced SUMO proteome and analysis suggests that specific protein domains, cellular pathways, and protein complexes respond to immune stress. A small subset of the confident set was validated by in-bacto SUMOylation and shown to be bona-fide SUMO targets. These include components of immune signaling pathways such as Caspar, Jra, Kay, cdc42, p38b, 14-3-3ε, as well as cellular proteins with diverse functions, many being components of protein complexes, such as prosß4, Rps10b, SmD3, Tango7, and Aats-arg. Caspar, a human FAF1 ortholog that negatively regulates immune-deficient signaling, is SUMOylated at K551 and responds to treatment with lipopolysaccharide in cultured cells. Our study is one of the first to describe SUMO proteome for the Drosophila immune response. Our data and analysis provide a global framework for the understanding of SUMO modification in the host response to pathogens. Copyright © 2015 Handu et al.

Third Chromosome Balancer Inversions Disrupt Protein-Coding Genes and Influence Distal Recombination Events in Drosophila melanogaster

PubMed Central

Miller, Danny E.; Cook, Kevin R.; Arvanitakis, Alexandra V.; Hawley, R. Scott

2016-01-01

Balancer chromosomes are multiply inverted chromosomes that suppress meiotic crossing over and prevent the recovery of crossover products. Balancers are commonly used in Drosophila melanogaster to maintain deleterious alleles and in stock construction. They exist for all three major chromosomes, yet the molecular location of the breakpoints and the exact nature of many of the mutations carried by the second and third chromosome balancers has not been available. Here, we precisely locate eight of 10 of the breakpoints on the third chromosome balancer TM3, six of eight on TM6, and nine of 11 breakpoints on TM6B. We find that one of the inversion breakpoints on TM3 bisects the highly conserved tumor suppressor gene p53—a finding that may have important consequences for a wide range of studies in Drosophila. We also identify evidence of single and double crossovers between several TM3 and TM6B balancers and their normal-sequence homologs that have created genetic diversity among these chromosomes. Overall, this work demonstrates the practical importance of precisely identifying the position of inversion breakpoints of balancer chromosomes and characterizing the mutant alleles carried by them. PMID:27172211

Lactobacillus acidophilus Induces Cytokine and Chemokine Production via NF-κB and p38 Mitogen-Activated Protein Kinase Signaling Pathways in Intestinal Epithelial Cells

PubMed Central

Lü, Xuena; Man, Chaoxin; Han, Linlin; Shan, Yi; Qu, Xingguang; Liu, Ying; Yang, Shiqin; Xue, Yuqing; Zhang, Yinghua

2012-01-01

Intestinal epithelial cells can respond to certain bacteria by producing an array of cytokines and chemokines which are associated with host immune responses. Lactobacillus acidophilus NCFM is a characterized probiotic, originally isolated from human feces. This study aimed to test the ability of L. acidophilus NCFM to stimulate cytokine and chemokine production in intestinal epithelial cells and to elucidate the mechanisms involved in their upregulation. In experiments using intestinal epithelial cell lines and mouse models, we observed that L. acidophilus NCFM could rapidly but transiently upregulate a number of effector genes encoding cytokines and chemokines such as interleukin 1α (IL-1α), IL-1β, CCL2, and CCL20 and that cytokines showed lower expression levels with L. acidophilus NCFM treatment than chemokines. Moreover, L. acidophilus NCFM could activate a pathogen-associated molecular pattern receptor, Toll-like receptor 2 (TLR2), in intestinal epithelial cell lines. The phosphorylation of NF-κB p65 and p38 mitogen-activated protein kinase (MAPK) in intestinal epithelial cell lines was also enhanced by L. acidophilus NCFM. Furthermore, inhibitors of NF-κB (pyrrolidine dithiocarbamate [PDTC]) and p38 MAPK (SB203580) significantly reduced cytokine and chemokine production in the intestinal epithelial cell lines stimulated by L. acidophilus NCFM, suggesting that both NF-κB and p38 MAPK signaling pathways were important for the production of cytokines and chemokines induced by L. acidophilus NCFM. PMID:22357649

Diagnostic Significance of p38 Isoforms (p38α, p38β, p38γ, p38δ) in Head and Neck Squamous Cell Carcinoma: Comparative Serum Level Evaluation and Design of Novel Peptide Inhibitor Targeting the Same.

PubMed

Sahu, Vishal; Nigam, Lokesh; Agnihotri, Vertica; Gupta, Abhishek; Shekhar, Shashank; Subbarao, Naidu; Bhaskar, Suman; Dey, Sharmistha

2018-05-09

The p38 mitogen-activated protein kinase (MAPKs) play a crucial role in the production of pro-inflammatory cytokines and over-expression of it increase cytokines which promote cancer. Among four isoforms, p38α has been well studied in head and neck squamous cell carcinoma (HNSCC) and other cancers as a therapeutic target.p38δ has recently emerged as a potential disease-specific drug target. Elevated serum p38α level in HNSCC was reported earlier from our lab. This study aims to estimate the levels of p38 MAPK-isoforms in the serum of HNSCC and design peptide inhibitor targeting the same. Levels of p38 MAPK isoforms in the serum of HNSCC and healthy controls were quantified by surface plasmon resonance technology. The peptide inhibitor for p38 MAPK was designed by molecular modeling using Grid-based Ligand Docking with Energetics tools and compared with known specific inhibitors. We have observed highly elevated levels of all four isoforms of p38 MAPK in serum of HNSCC patients compared to the control group. Further, serum p38α, p38β, and p38δ levels were down regulated after therapy in follow up patients, while p38γ showed no response to the therapy. Present study screened designed peptide WFYH as a specific inhibitor against p38δ. The specific inhibitor of p38δ was found to have no effect on p38α due to great structural difference at ATP binding pocket. In this study, first time estimated the levels of p38 MAPK isoforms in the serum of HNSCC. It can be concluded that p38 MAPK isoforms can be a diagnostic and prognostic marker for HNSCC and p38δ as a therapeutic target.

Drosophila CLOCK target gene characterization: implications for circadian tissue-specific gene expression

PubMed Central

Abruzzi, Katharine Compton; Rodriguez, Joseph; Menet, Jerome S.; Desrochers, Jennifer; Zadina, Abigail; Luo, Weifei; Tkachev, Sasha; Rosbash, Michael

2011-01-01

CLOCK (CLK) is a master transcriptional regulator of the circadian clock in Drosophila. To identify CLK direct target genes and address circadian transcriptional regulation in Drosophila, we performed chromatin immunoprecipitation (ChIP) tiling array assays (ChIP–chip) with a number of circadian proteins. CLK binding cycles on at least 800 sites with maximal binding in the early night. The CLK partner protein CYCLE (CYC) is on most of these sites. The CLK/CYC heterodimer is joined 4–6 h later by the transcriptional repressor PERIOD (PER), indicating that the majority of CLK targets are regulated similarly to core circadian genes. About 30% of target genes also show cycling RNA polymerase II (Pol II) binding. Many of these generate cycling RNAs despite not being documented in prior RNA cycling studies. This is due in part to different RNA isoforms and to fly head tissue heterogeneity. CLK has specific targets in different tissues, implying that important CLK partner proteins and/or mechanisms contribute to gene-specific and tissue-specific regulation. PMID:22085964

A Rehabilitation of the Genetic Map of the 84b-D Region in DROSOPHILA MELANOGASTER

PubMed Central

Cavener, Douglas R.; Otteson, Deborah C.; Kaufman, Thomas C.

1986-01-01

A reanalysis of the 84B3 to 84B3,5 region of the polytene chromosomes of Drosophila melanogaster has led to the identification and localization of 16 genes. These genes include 11 vital loci, four genes exhibiting nonlethal visible mutant phenotypes and one gene encoding a nonessential enzyme. The identity of the gene products of two of the vital genes has been determined to be α-tubulin and glucose dehydrogenase (Gld). Three newly identified genes, sticking (stk), half out ( hat) and trapped (ted), as well as Gld are required for eclosion. Among the nonessential genes are roughened eye (roe) and ruffed eye (rue), which affect eye texture. The roe phenotype is greatly enhanced by deletions that simultaneously remove roe and an unidentified locus in 84E. Mutations in another nonessential gene, rotund (rn), are characterized by pattern deletions of most adult appendages. PMID:3095179

A hobo transgene that encodes the P-element transposase in Drosophila melanogaster: autoregulation and cytotype control of transposase activity.

PubMed Central

Simmons, Michael J; Haley, Kevin J; Grimes, Craig D; Raymond, John D; Niemi, Jarad B

2002-01-01

Drosophila were genetically transformed with a hobo transgene that contains a terminally truncated but otherwise complete P element fused to the promoter from the Drosophila hsp70 gene. Insertions of this H(hsp/CP) transgene on either of the major autosomes produced the P transposase in both the male and female germlines, but not in the soma. Heat-shock treatments significantly increased transposase activity in the female germline; in the male germline, these treatments had little effect. The transposase activity of two insertions of the H(hsp/CP) transgene was not significantly greater than their separate activities, and one insertion of this transgene reduced the transposase activity of P(ry(+), Delta2-3)99B, a stable P transgene, in the germline as well as in the soma. These observations suggest that, through alternate splicing, the H(hsp/CP) transgene produces a repressor that feeds back negatively to regulate transposase expression or function in both the somatic and germline tissues. The H(hsp/CP) transgenes are able to induce gonadal dysgenesis when the transposase they encode has P-element targets to attack. However, this ability and the ability to induce P-element excisions are repressed by the P cytotype, a chromosomal/cytoplasmic state that regulates P elements in the germline. PMID:12019234

Immune genes and divergent antimicrobial peptides in flies of the subgenus Drosophila.

PubMed

Hanson, Mark A; Hamilton, Phineas T; Perlman, Steve J

2016-10-24

Drosophila is an important model for studying the evolution of animal immunity, due to the powerful genetic tools developed for D. melanogaster. However, Drosophila is an incredibly speciose lineage with a wide range of ecologies, natural histories, and diverse natural enemies. Surprisingly little functional work has been done on immune systems of species other than D. melanogaster. In this study, we examine the evolution of immune genes in the speciose subgenus Drosophila, which diverged from the subgenus Sophophora (that includes D. melanogaster) approximately 25-40 Mya. We focus on D. neotestacea, a woodland species used to study interactions between insects and parasitic nematodes, and combine recent transcriptomic data with infection experiments to elucidate aspects of host immunity. We found that the vast majority of genes involved in the D. melanogaster immune response are conserved in D. neotestacea, with a few interesting exceptions, particularly in antimicrobial peptides (AMPs); until recently, AMPs were not thought to evolve rapidly in Drosophila. Unexpectedly, we found a distinct diptericin in subgenus Drosophila flies that appears to have evolved under diversifying (positive) selection. We also describe the presence of the AMP drosocin, which was previously thought to be restricted to the subgenus Sophophora, in the subgenus Drosophila. We challenged two subgenus Drosophila species, D. neotestacea and D. virilis with bacterial and fungal pathogens and quantified AMP expression. While diptericin in D. virilis was induced by exposure to gram-negative bacteria, it was not induced in D. neotestacea, showing that conservation of immune genes does not necessarily imply conservation of the realized immune response. Our study lends support to the idea that invertebrate AMPs evolve rapidly, and that Drosophila harbor a diverse repertoire of AMPs with potentially important functional consequences.

Cytochrome P450-Dependent Metabolism of Caffeine in Drosophila melanogaster

PubMed Central

Coelho, Alexandra; Fraichard, Stephane; Le Goff, Gaëlle; Faure, Philippe; Artur, Yves; Ferveur, Jean-François; Heydel, Jean-Marie

2015-01-01

Caffeine (1, 3, 7-trimethylxanthine), an alkaloid produced by plants, has antioxidant and insecticide properties that can affect metabolism and cognition. In vertebrates, the metabolites derived from caffeine have been identified, and their functions have been characterized. However, the metabolites of caffeine in insects remain unknown. Thus, using radiolabelled caffeine, we have identified some of the primary caffeine metabolites produced in the body of Drosophila melanogaster males, including theobromine, paraxanthine and theophylline. In contrast to mammals, theobromine was the predominant metabolite (paraxanthine in humans; theophylline in monkeys; 1, 3, 7-trimethyluric acid in rodents). A transcriptomic screen of Drosophila flies exposed to caffeine revealed the coordinated variation of a large set of genes that encode xenobiotic-metabolizing proteins, including several cytochromes P450s (CYPs) that were highly overexpressed. Flies treated with metyrapone—an inhibitor of CYP enzymes—showed dramatically decreased caffeine metabolism, indicating that CYPs are involved in this process. Using interference RNA genetic silencing, we measured the metabolic and transcriptomic effect of three candidate CYPs. Silencing of CYP6d5 completely abolished theobromine synthesis, whereas CYP6a8 and CYP12d1 silencing induced different consequences on metabolism and gene expression. Therefore, we characterized several metabolic products and some enzymes potentially involved in the degradation of caffeine. In conclusion, this pioneer approach to caffeine metabolism in insects opens novel perspectives for the investigation of the physiological effects of caffeine metabolites. It also indicates that caffeine could be used as a biomarker to evaluate CYP phenotypes in Drosophila and other insects. PMID:25671424

Interleukin-5 regulates genes involved in B-cell terminal maturation.

PubMed

Horikawa, Keisuke; Takatsu, Kiyoshi

2006-08-01

Interleukin (IL)-5 induces CD38-activated splenic B cells to differentiate into immunoglobulin M-secreting cells and undergo micro to gamma 1 class switch recombination (CSR) at the DNA level, resulting in immunoglobulin G1 (IgG1) production. Interestingly, IL-4, a well-known IgG1-inducing factor does not induce immunoglobulin production or micro to gamma 1 CSR in CD38-activated B cells. In the present study, we implemented complementary DNA microarrays to investigate the contribution of IL-5-induced gene expression in CD38-stimulated B cells to immunoglobulin-secreting cell differentiation and micro to gamma 1 CSR. IL-5 and IL-4 stimulation of CD38-activated B cells induced the expression of 418 and 289 genes, respectively, that consisted of several clusters. Surprisingly, IL-5-inducible 78 genes were redundantly regulated by IL-4. IL-5 and IL-4 also suppressed the gene expression of 319 and 325 genes, respectively, 97 of which were overlapped. Genes critically regulated by IL-5 include immunoglobulin-related genes such as J chain and immunoglobulinkappa, and genes involved in B-cell maturation such as BCL6, activation-induced cytidine deaminase (Aid) and B lymphocyte-induced maturation protein-1 (Blimp-1) and tend to be induced slowly after IL-5 stimulation. Intriguingly, among genes, the retroviral induction of Blimp-1 and Aid in CD38-activated B cells could induce IL-4-dependent maturation to Syndecan-1+ antibody-secreting cells and micro to gamma 1 CSR, respectively, in CD38-activated B cells. Taken together, preferential Aid and Blimp-1 expression plays a critical role in IL-5-induced immunoglobulin-secreting cell differentiation and micro to gamma 1 CSR in CD38-activated B cells.

FlyAtlas: database of gene expression in the tissues of Drosophila melanogaster

PubMed Central

Robinson, Scott W.; Herzyk, Pawel; Dow, Julian A. T.; Leader, David P.

2013-01-01

The FlyAtlas resource contains data on the expression of the genes of Drosophila melanogaster in different tissues (currently 25—17 adult and 8 larval) obtained by hybridization of messenger RNA to Affymetrix Drosophila Genome 2 microarrays. The microarray probe sets cover 13 250 Drosophila genes, detecting 12 533 in an unambiguous manner. The data underlying the original web application (http://flyatlas.org) have been restructured into a relational database and a Java servlet written to provide a new web interface, FlyAtlas 2 (http://flyatlas.gla.ac.uk/), which allows several additional queries. Users can retrieve data for individual genes or for groups of genes belonging to the same or related ontological categories. Assistance in selecting valid search terms is provided by an Ajax ‘autosuggest’ facility that polls the database as the user types. Searches can also focus on particular tissues, and data can be retrieved for the most highly expressed genes, for genes of a particular category with above-average expression or for genes with the greatest difference in expression between the larval and adult stages. A novel facility allows the database to be queried with a specific gene to find other genes with a similar pattern of expression across the different tissues. PMID:23203866

FlyAtlas: database of gene expression in the tissues of Drosophila melanogaster.

PubMed

Robinson, Scott W; Herzyk, Pawel; Dow, Julian A T; Leader, David P

2013-01-01

The FlyAtlas resource contains data on the expression of the genes of Drosophila melanogaster in different tissues (currently 25-17 adult and 8 larval) obtained by hybridization of messenger RNA to Affymetrix Drosophila Genome 2 microarrays. The microarray probe sets cover 13,250 Drosophila genes, detecting 12,533 in an unambiguous manner. The data underlying the original web application (http://flyatlas.org) have been restructured into a relational database and a Java servlet written to provide a new web interface, FlyAtlas 2 (http://flyatlas.gla.ac.uk/), which allows several additional queries. Users can retrieve data for individual genes or for groups of genes belonging to the same or related ontological categories. Assistance in selecting valid search terms is provided by an Ajax 'autosuggest' facility that polls the database as the user types. Searches can also focus on particular tissues, and data can be retrieved for the most highly expressed genes, for genes of a particular category with above-average expression or for genes with the greatest difference in expression between the larval and adult stages. A novel facility allows the database to be queried with a specific gene to find other genes with a similar pattern of expression across the different tissues.

Metal-metal interaction mediates the iron induction of Drosophila MtnB

DOE Office of Scientific and Technical Information (OSTI.GOV)

Qiang, Wenjia; Huang, Yunpeng; Wan, Zhihui

Metallothionein (MT) protein families are a class of small and universal proteins rich in cysteine residues. They are synthesized in response to heavy metal stresses to sequester the toxic ions by metal-thiolate bridges. Five MT family members, namely MtnA, MtnB, MtnC, MtnD and MtnE, have been discovered and identified in Drosophila. These five isoforms of MTs are regulated by metal responsive transcription factor dMTF-1 and play differentiated but overlapping roles in detoxification of metal ions. Previous researches have shown that Drosophila MtnB responds to copper (Cu), cadmium (Cd) and zinc (Zn). Interestingly in this study we found that Drosophila MtnBmore » expression also responds to elevated iron levels in the diet. Further investigations revealed that MtnB plays limited roles in iron detoxification, and the direct binding of MtnB to ferrous iron in vitro is also weak. The induction of MtnB by iron turns out to be mediated by iron interference of other metals, because EDTA at even a partial concentration of that of iron can suppress this induction. Indeed, in the presence of iron, zinc homeostasis is altered, as reflected by expression changes of zinc transporters dZIP1 and dZnT1. Thus, iron-mediated MtnB induction appears resulting from interrupted homeostasis of other metals such as zinc, which in turns induced MtnB expression. Metal-metal interaction may more widely exist than we expected. - Highlights: • Metallothionein B expression is regulated by iron in Drosophila melanogaster. • MtnB has limited physiological roles in iron detoxification. • Binding affinity of MtnB to iron is weak in vitro. • Induction of Drosophila MtnB by iron is mediated indirectly through metal-metal interaction.« less

Naringin protects ultraviolet B-induced skin damage by regulating p38 MAPK signal pathway.

PubMed

Ren, Xiaolin; Shi, Yuling; Zhao, Di; Xu, Mengyu; Li, Xiaolong; Dang, Yongyan; Ye, Xiyun

2016-05-01

Naringin is a bioflavonoid and has free radical scavenging and anti-inflammatory properties. We examined the effects of naringin on skin after ultraviolet radiation B (UVB) irradiation and the signal pathways by in vitro and in vivo assay. HaCaT cells pretreated with naringin significantly inhibited UVB induced-cell apoptosis and production of intracellular reactive oxygen species (ROS). The expressions of interleukin-1β (IL-1β), interleukin-6 (IL-6), interleukin-8 (IL-8) and cyclooxygenase-2 (COX-2) in HaCaT cells pretreated with naringin were decreased compared with the only UVB group. Also, the activation of p38 induced by UVB in HaCaT cells was reversed by naringin treatments. The inhibition function of naringin on p38 activity was more obvious than JNK. In vivo, topical treatments with naringin prevented the increase of epidermal thickness, IL-6 production, cell apoptosis and the overexpression of COX-2 in BALB/c mice skin irradiated with UVB. Naringin treatment also markedly blocked the activation of p38 in response to UVB stimulation in the mouse skin. Naringin can effectively protect against UVB-induced keratinocyte apoptosis and skin damage by inhibiting ROS production, COX-2 overexpression and strong inflammation reactions. It seemed that naringin played its role against UVB-induced skin damage through inhibition of mitogen-activated protein kinase (MAPK)/p38 activation. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

Acidic Food pH Increases Palatability and Consumption and Extends Drosophila Lifespan12

PubMed Central

Deshpande, Sonali A; Yamada, Ryuichi; Mak, Christine M; Hunter, Brooke; Obando, Alina Soto; Hoxha, Sany; Ja, William W

2015-01-01

Background: Despite the prevalent use of Drosophila as a model in studies of nutrition, the effects of fundamental food properties, such as pH, on animal health and behavior are not well known. Objectives: We examined the effect of food pH on adult Drosophila lifespan, feeding behavior, and microbiota composition and tested the hypothesis that pH-mediated changes in palatability and total consumption are required for modulating longevity. Methods: We measured the effect of buffered food (pH 5, 7, or 9) on male gustatory responses (proboscis extension), total food intake, and male and female lifespan. The effect of food pH on germfree male lifespan was also assessed. Changes in fly-associated microbial composition as a result of food pH were determined by 16S ribosomal RNA gene sequencing. Male gustatory responses, total consumption, and male and female longevity were additionally measured in the taste-defective Pox neuro (Poxn) mutant and its transgenic rescue control. Results: An acidic diet increased Drosophila gustatory responses (40–230%) and food intake (5–50%) and extended survival (10–160% longer median lifespan) compared with flies on either neutral or alkaline pH food. Alkaline food pH shifted the composition of fly-associated bacteria and resulted in greater lifespan extension (260% longer median survival) after microbes were eliminated compared with flies on an acidic (50%) or neutral (130%) diet. However, germfree flies lived longer on an acidic diet (5–20% longer median lifespan) compared with those on either neutral or alkaline pH food. Gustatory responses, total consumption, and longevity were unaffected by food pH in Poxn mutant flies. Conclusions: Food pH can directly influence palatability and feeding behavior and affect parameters such as microbial growth to ultimately affect Drosophila lifespan. Fundamental food properties altered by dietary or drug interventions may therefore contribute to changes in animal physiology, metabolism, and

An RNAi Screen for Genes Involved in Nanoscale Protrusion Formation on Corneal Lens in Drosophila melanogaster.

PubMed

Minami, Ryunosuke; Sato, Chiaki; Yamahama, Yumi; Kubo, Hideo; Hariyama, Takahiko; Kimura, Ken-Ichi

2016-12-01

The "moth-eye" structure, which is observed on the surface of corneal lens in several insects, supports anti-reflective and self-cleaning functions due to nanoscale protrusions known as corneal nipples. Although the morphology and function of the "moth-eye" structure, are relatively well studied, the mechanism of protrusion formation from cell-secreted substances is unknown. In Drosophila melanogaster, a compound eye consists of approximately 800 facets, the surface of which is formed by the corneal lens with nanoscale protrusions. In the present study, we sought to identify genes involved in "moth-eye" structure, formation in order to elucidate the developmental mechanism of the protrusions in Drosophila. We re-examined the aberrant patterns in classical glossy-eye mutants by scanning electron microscope and classified the aberrant patterns into groups. Next, we screened genes encoding putative structural cuticular proteins and genes involved in cuticular formation using eye specific RNAi silencing methods combined with the Gal4/UAS expression system. We identified 12 of 100 candidate genes, such as cuticular proteins family genes (Cuticular protein 23B and Cuticular protein 49Ah), cuticle secretion-related genes (Syntaxin 1A and Sec61 ββ subunit), ecdysone signaling and biosynthesis-related genes (Ecdysone receptor, Blimp-1, and shroud), and genes involved in cell polarity/cell architecture (Actin 5C, shotgun, armadillo, discs large1, and coracle). Although some of the genes we identified may affect corneal protrusion formation indirectly through general patterning defects in eye formation, these initial findings have encouraged us to more systematically explore the precise mechanisms underlying the formation of nanoscale protrusions in Drosophila.

Dopamine Dynamics and Signaling in Drosophila: An Overview of Genes, Drugs and Behavioral Paradigms

PubMed Central

Yamamoto, Shinya; Seto, Elaine S.

2014-01-01

Changes in dopamine (DA) signaling have been implicated in a number of human neurologic and psychiatric disorders. Similarly, defects in DA signaling in the fruit fly, Drosophila melanogaster, have also been associated with several behavioral defects. As most genes involved in DA synthesis, transport, secretion, and signaling are conserved between species, Drosophila is a powerful genetic model organism to study the regulation of DA signaling in vivo. In this review, we will provide an overview of the genes and drugs that regulate DA biology in Drosophila. Furthermore, we will discuss the behavioral paradigms that are regulated by DA signaling in flies. By analyzing the genes and neuronal circuits that govern such behaviors using sophisticated genetic, pharmacologic, electrophysiologic, and imaging approaches in Drosophila, we will likely gain a better understanding about how this neuromodulator regulates motor tasks and cognition in humans. PMID:24770636

Rapidly Evolving Toll-3/4 Genes Encode Male-Specific Toll-Like Receptors in Drosophila.

PubMed

Levin, Tera C; Malik, Harmit S

2017-09-01

Animal Toll-like receptors (TLRs) have evolved through a pattern of duplication and divergence. Whereas mammalian TLRs directly recognize microbial ligands, Drosophila Tolls bind endogenous ligands downstream of both developmental and immune signaling cascades. Here, we find that most Toll genes in Drosophila evolve slowly with little gene turnover (gains/losses), consistent with their important roles in development and indirect roles in microbial recognition. In contrast, we find that the Toll-3/4 genes have experienced an unusually rapid rate of gene gains and losses, resulting in lineage-specific Toll-3/4s and vastly different gene repertoires among Drosophila species, from zero copies (e.g., D. mojavensis) to nineteen copies (e.g., D. willistoni). In D. willistoni, we find strong evidence for positive selection in Toll-3/4 genes, localized specifically to an extracellular region predicted to overlap with the binding site of Spätzle, the only known ligand of insect Tolls. However, because Spätzle genes are not experiencing similar selective pressures, we hypothesize that Toll-3/4s may be rapidly evolving because they bind to a different ligand, akin to TLRs outside of insects. We further find that most Drosophila Toll-3/4 genes are either weakly expressed or expressed exclusively in males, specifically in the germline. Unlike other Toll genes in D. melanogaster, Toll-3, and Toll-4 have apparently escaped from essential developmental roles, as knockdowns have no substantial effects on viability or male fertility. Based on these findings, we propose that the Toll-3/4 genes represent an exceptionally rapidly evolving lineage of Drosophila Toll genes, which play an unusual, as-yet-undiscovered role in the male germline. © The Author 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

Rapidly Evolving Toll-3/4 Genes Encode Male-Specific Toll-Like Receptors in Drosophila

PubMed Central

Levin, Tera C.; Malik, Harmit S.

2017-01-01

Abstract Animal Toll-like receptors (TLRs) have evolved through a pattern of duplication and divergence. Whereas mammalian TLRs directly recognize microbial ligands, Drosophila Tolls bind endogenous ligands downstream of both developmental and immune signaling cascades. Here, we find that most Toll genes in Drosophila evolve slowly with little gene turnover (gains/losses), consistent with their important roles in development and indirect roles in microbial recognition. In contrast, we find that the Toll-3/4 genes have experienced an unusually rapid rate of gene gains and losses, resulting in lineage-specific Toll-3/4s and vastly different gene repertoires among Drosophila species, from zero copies (e.g., D. mojavensis) to nineteen copies (e.g., D. willistoni). In D. willistoni, we find strong evidence for positive selection in Toll-3/4 genes, localized specifically to an extracellular region predicted to overlap with the binding site of Spätzle, the only known ligand of insect Tolls. However, because Spätzle genes are not experiencing similar selective pressures, we hypothesize that Toll-3/4s may be rapidly evolving because they bind to a different ligand, akin to TLRs outside of insects. We further find that most Drosophila Toll-3/4 genes are either weakly expressed or expressed exclusively in males, specifically in the germline. Unlike other Toll genes in D. melanogaster, Toll-3, and Toll-4 have apparently escaped from essential developmental roles, as knockdowns have no substantial effects on viability or male fertility. Based on these findings, we propose that the Toll-3/4 genes represent an exceptionally rapidly evolving lineage of Drosophila Toll genes, which play an unusual, as-yet-undiscovered role in the male germline. PMID:28541576

Escherichia coli K-12 and B contain functional bacteriophage P2 ogr genes.

PubMed Central

Slettan, A; Gebhardt, K; Kristiansen, E; Birkeland, N K; Lindqvist, B H

1992-01-01

The bacteriophage P2 ogr gene encodes an essential 72-amino-acid protein which acts as a positive regulator of P2 late transcription. A P2 ogr deletion phage, which depends on the supply of Ogr protein in trans for lytic growth on Escherichia coli C, has previously been constructed. E. coli B and K-12 were found to support the growth of the ogr-defective P2 phage because of the presence of functional ogr genes located in cryptic P2-like prophages in these strains. The cryptic ogr genes were cloned and sequenced. Compared with the P2 wild-type ogr gene, the ogr genes in the B and K-12 strains are conserved, containing mostly silent base substitutions. One of the base substitutions in the K-12 ogr gene results in replacement of an alanine with valine at position 57 in the Ogr protein but does not seem to affect the function of Ogr as a transcriptional activator. The cryptic ogr genes are constitutively transcribed, apparently at a higher level than the wild-type ogr gene in a P2 lysogen. Images PMID:1597424

Inhibition of p38 mitogen-activated protein kinase impairs influenza virus-induced primary and secondary host gene responses and protects mice from lethal H5N1 infection.

PubMed

Börgeling, Yvonne; Schmolke, Mirco; Viemann, Dorothee; Nordhoff, Carolin; Roth, Johannes; Ludwig, Stephan

2014-01-03

Highly pathogenic avian influenza viruses (HPAIV) induce severe inflammation in poultry and men. One characteristic of HPAIV infections is the induction of a cytokine burst that strongly contributes to viral pathogenicity. This cell-intrinsic hypercytokinemia seems to involve hyperinduction of p38 mitogen-activated protein kinase. Here we investigate the role of p38 MAPK signaling in the antiviral response against HPAIV in mice as well as in human endothelial cells, the latter being a primary source of cytokines during systemic infections. Global gene expression profiling of HPAIV-infected endothelial cells in the presence of the p38-specific inhibitor SB 202190 revealed that inhibition of p38 MAPK leads to reduced expression of IFNβ and other cytokines after H5N1 and H7N7 infection. More than 90% of all virus-induced genes were either partially or fully dependent on p38 signaling. Moreover, promoter analysis confirmed a direct impact of p38 on the IFNβ promoter activity. Furthermore, upon treatment with IFN or conditioned media from HPAIV-infected cells, p38 controls interferon-stimulated gene expression by coregulating STAT1 by phosphorylation at serine 727. In vivo inhibition of p38 MAPK greatly diminishes virus-induced cytokine expression concomitant with reduced viral titers, thereby protecting mice from lethal infection. These observations show that p38 MAPK acts on two levels of the antiviral IFN response. Initially the kinase regulates IFN induction and, at a later stage, p38 controls IFN signaling and thereby expression of IFN-stimulated genes. Thus, inhibition of MAP kinase p38 may be an antiviral strategy that protects mice from lethal influenza by suppressing excessive cytokine expression.

Patterns of Nucleotide Diversity at the Regions Encompassing the Drosophila Insulin-Like Peptide (dilp) Genes: Demography vs. Positive Selection in Drosophila melanogaster

PubMed Central

Guirao-Rico, Sara; Aguadé, Montserrat

2013-01-01

In Drosophila, the insulin-signaling pathway controls some life history traits, such as fertility and lifespan, and it is considered to be the main metabolic pathway involved in establishing adult body size. Several observations concerning variation in body size in the Drosophila genus are suggestive of its adaptive character. Genes encoding proteins in this pathway are, therefore, good candidates to have experienced adaptive changes and to reveal the footprint of positive selection. The Drosophila insulin-like peptides (DILPs) are the ligands that trigger the insulin-signaling cascade. In Drosophila melanogaster, there are several peptides that are structurally similar to the single mammalian insulin peptide. The footprint of recent adaptive changes on nucleotide variation can be unveiled through the analysis of polymorphism and divergence. With this aim, we have surveyed nucleotide sequence variation at the dilp1-7 genes in a natural population of D. melanogaster. The comparison of polymorphism in D. melanogaster and divergence from D. simulans at different functional classes of the dilp genes provided no evidence of adaptive protein evolution after the split of the D. melanogaster and D. simulans lineages. However, our survey of polymorphism at the dilp gene regions of D. melanogaster has provided some evidence for the action of positive selection at or near these genes. The regions encompassing the dilp1-4 genes and the dilp6 gene stand out as likely affected by recent adaptive events. PMID:23308258

Unusual Variability of the Drosophila Melanogaster Ref(2)p Protein Which Controls the Multiplication of Sigma Rhabdovirus

PubMed Central

Dru, P.; Bras, F.; Dezelee, S.; Gay, P.; Petitjean, A. M.; Pierre-Deneubourg, A.; Teninges, D.; Contamine, D.

1993-01-01

The ref(2)P gene of Drosophila melanogaster was identified by the discovery of two alleles, P(o) and P(p), respectively, permissive and restrictive for sigma rhabdovirus multiplication. A surprising variability of this gene was first noticed by the observation of size differences between the transcripts of permissive and restrictive alleles. In this paper, another restrictive allele, P(n), clearly distinct from P(p), is described: it exhibits a weaker antiviral effect than P(p) and differs from P(p) by its molecular structure. Five types of alleles were distinguished on the basis of their molecular structure, as revealed by S1 nuclease analysis of 17 D. melanogaster strains; three alleles were permissive and two restrictive. Comparison of the sequences of four haplotypes revealed numerous point mutations, two deletions (21 and 24 bp) and a complex event involving a 3-bp deletion, all affected the coding region. The unusual variability of the ref(2)P locus was confirmed by the high ratio of amino acid replacements to synonymous mutations (7:1), as compared to that of other genes, such as the Adh (2:42). Nevertheless, nucleotide sequence comparison with the Drosophila erecta ref(2)P gene shows that selective pressures are exerted to maintain the existence of a functional protein. The effects of this high variability on the ref(2)P protein are discussed in relation to its specific antiviral properties and to its function in D. melanogaster, where it is required for male fertility. PMID:8462852

Nur77 attenuates endothelin-1 expression via downregulation of NF-κB and p38 MAPK in A549 cells and in an ARDS rat model.

PubMed

Jiang, Yujie; Zeng, Yi; Huang, Xia; Qin, Yueqiu; Luo, Weigui; Xiang, Shulin; Sooranna, Suren R; Pinhu, Liao

2016-12-01

Acute respiratory distress syndrome (ARDS) is characterized by inflammatory injury to the alveolar and capillary barriers that results in impaired gas exchange and severe acute respiratory failure. Nuclear orphan receptor Nur77 has emerged as a regulator of gene expression in inflammation, and its role in the pathogenesis of ARDS is not clear. The objective of this study is to investigate the potential role of Nur77 and its underlying mechanism in the regulation of endothelin-1 (ET-1) expression in lipopolysaccharide (LPS)-induced A549 cells and an ARDS rat model. We demonstrate that LPS induced Nur77 expression and nuclear export in A549 cells. Overexpression of Nur77 markedly decreased basal and LPS-induced ET-1 expression in A549 cells, whereas knockdown of Nur77 increased the ET-1 expression. LPS-induced phosphorylation and nuclear translocation of NF-κB and p38 MAPK were blocked by Nur77 overexpression and augmented by Nur77 knockdown in A549 cells. In vivo, LPS induced Nur77 expression in lung in ARDS rats. Pharmacological activation of Nur77 by cytosporone B (CsnB) inhibited ET-1 expression in ARDS rats, decreased LPS-induced phosphorylation of NF-κB and p38 MAPK, and relieved lung, liver, and kidney injury. Pharmacological deactivation of Nur77 by 1,1-bis-(3'-indolyl)-1-(p-hydroxyphenyl)methane (DIM-C-pPhOH, C-DIM8) had no effect on ET-1 expression and lung injury. These results indicated that Nur77 decreases ET-1 expression by suppressing NF-κB and p38 MAPK in LPS-stimulated A549 cells in vitro, and, in an LPS-induced ARDS rat model, CsnB reduced ET-1 expression and lung injury in ARDS rats. Copyright © 2016 the American Physiological Society.

Rosiglitazone attenuates NF-{kappa}B-dependent ICAM-1 and TNF-{alpha} production caused by homocysteine via inhibiting ERK{sub 1/2}/p38MAPK activation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bai, Yong-Ping; Liu, Yu-Hui; Chen, Jia

2007-08-17

Previous studies demonstrated an important interaction between nuclear factor-kappaB (NF-{kappa}B) activation and homocysteine (Hcy)-induced cytokines expression in endothelial cells and vascular smooth muscle cells. However, the underlying mechanism remains illusive. In this study, we investigated the effects of Hcy on NF-{kappa}B-mediated sICAM-1, TNF-{alpha} production and the possible involvement of ERK{sub 1/2}/p38MAPK pathway. The effects of rosiglitazone intervention were also examined. Our results show that Hcy increased the levels of sICAM-1 and TNF-{alpha} in cultured human umbilical vein endothelial cells (HUVECs) in a time- and concentration-dependent manner. This effect was significantly depressed by rosiglitazone and different inhibitors (PDTC, NF-{kappa}B inhibitor; PD98059,more » MEK inhibitor; SB203580, p38MAPK specific inhibitor; and staurosporine, PKC inhibitor). Next, we investigated the effect of Hcy on ERK{sub 1/2}/p38MAPK pathway and NF-{kappa}B activity in HUVECs. The results show that Hcy activated both ERK{sub 1/2}/p38MAPK pathway and NF-{kappa}B-DNA-binding activity. These effects were markedly inhibited by rosiglitazone as well as other inhibitors (SB203580, PD98059, and PDTC). Further, the pretreatment of staurosporine abrogated ERK{sub 1/2}/p38MAPK phosphorylation, suggesting that Hcy-induced ERK{sub 1/2}/p38MAPK activation is associated with PKC activity. Our results provide evidence that Hcy-induced NF-{kappa}B activation was mediated by activation of ERK{sub 1/2}/p38MAPK pathway involving PKC activity. Rosiglitazone reduces the NF-{kappa}B-mediated sICAM-1 and TNF-{alpha} production induced by Hcy via inhibition of ERK{sub 1/2}/p38MAPK pa0011thw.« less

Investigation of Seasonal and Latitudinal Effects on the Expression of Clock Genes in Drosophila

NASA Astrophysics Data System (ADS)

Hosseini, Seyede Sanaz; Nazarimehr, Fahimeh; Jafari, Sajad

The primary goal in this work is to develop a dynamical model capturing the influence of seasonal and latitudinal variations on the expression of Drosophila clock genes. To this end, we study a specific dynamical system with strange attractors that exhibit changes of Drosophila activity in a range of latitudes and across different seasons. Bifurcations of this system are analyzed to peruse the effect of season and latitude on the behavior of clock genes. Existing experimental data collected from the activity of Drosophila melanogaster corroborate the dynamical model.

Common genes regulate food and ethanol intake in Drosophila.

PubMed

Sekhon, Morgan L; Lamina, Omoteniola; Hogan, Kerry E; Kliethermes, Christopher L

2016-06-01

The abuse liability of alcohol (ethanol) is believed to result in part from its actions on neurobiological substrates that underlie the motivation toward food and other natural reinforcers, and a growing body of evidence indicates that these substrates are broadly conserved among animal phyla. Understanding the extent to which the substrates regulating ethanol and food intake overlap is an important step toward developing therapeutics that selectively reduce ethanol intake. In the current experiments, we measured food and ethanol intake in Recombinant Inbred (RI) lines of Drosophila melanogaster using several assays, and then calculated genetic correlations to estimate the degree to which common genes might underlie behavior in these assays. We found that food intake and ethanol intake as measured in the capillary assay are genetically correlated traits in D. melanogaster, as well as in a panel of 11 Drosophila species that we tested subsequently. RI line differences in food intake in a dyed food assay were genetically unrelated to ethanol intake in the capillary assay or to ethanol preference measured using an olfactory trap apparatus. Using publicly available gene expression data, we found that expression profiles across the RI lines of a number of genes (including the D2-like dopamine receptor, DOPA decarboxylase, and fruitless) correlated with the RI line differences in food and ethanol intake we measured, while the expression profiles of other genes, including NPF, and the NPF and 5-HT2 receptors, correlated only with ethanol intake or preference. Our results suggest that food and ethanol intake are regulated by some common genes in Drosophila, but that other genes regulate ethanol intake independently of food intake. These results have implications toward the development of therapeutics that preferentially reduce ethanol intake. Copyright © 2016 Elsevier Inc. All rights reserved.

Cardiomyocyte Regulation of Systemic Lipid Metabolism by the Apolipoprotein B-Containing Lipoproteins in Drosophila

PubMed Central

Ishikawa, Zachary

2017-01-01

The heart has emerged as an important organ in the regulation of systemic lipid homeostasis; however, the underlying mechanism remains poorly understood. Here, we show that Drosophila cardiomyocytes regulate systemic lipid metabolism by producing apolipoprotein B-containing lipoproteins (apoB-lipoproteins), essential lipid carriers that are so far known to be generated only in the fat body. In a Drosophila genetic screen, we discovered that when haplo-insufficient, microsomal triglyceride transfer protein (mtp), required for the biosynthesis of apoB-lipoproteins, suppressed the development of diet-induced obesity. Tissue-specific inhibition of Mtp revealed that whereas knockdown of mtp only in the fat body decreases systemic triglyceride (TG) content on normal food diet (NFD) as expected, knockdown of mtp only in the cardiomyocytes also equally decreases systemic TG content on NFD, suggesting that the cardiomyocyte- and fat body-derived apoB-lipoproteins serve similarly important roles in regulating whole-body lipid metabolism. Unexpectedly, on high fat diet (HFD), knockdown of mtp in the cardiomyocytes, but not in fat body, protects against the gain in systemic TG levels. We further showed that inhibition of the Drosophila apoB homologue, apolipophorin or apoLpp, another gene essential for apoB-lipoprotein biosynthesis, affects systemic TG levels similarly to that of Mtp inhibition in the cardiomyocytes on NFD or HFD. Finally, we determined that HFD differentially alters Mtp and apoLpp expression in the cardiomyocytes versus the fat body, culminating in higher Mtp and apoLpp levels in the cardiomyocytes than in fat body and possibly underlying the predominant role of cardiomyocyte-derived apoB-lipoproteins in lipid metabolic regulation. Our findings reveal a novel and significant function of heart-mediated apoB-lipoproteins in controlling lipid homeostasis. PMID:28095410

Identification of FGF-dependent genes in the Drosophila tracheal system.

PubMed

Stahl, Markus; Schuh, Reinhard; Adryan, Boris

2007-01-01

The embryonic development of the tracheal system of the fruit fly Drosophila provides a paradigm for genetic studies of branching morphogenesis. Efforts of many laboratories have identified Branchless (Bnl, a fibroblast growth factor homologue) and Breathless (Btl, the receptor homologue) as crucial factors at many stages of tracheal system development. The downstream targets of the Bnl/Btl signalling cascade, however, remain mostly unknown. Misexpression of the bnl gene results in specific tracheal phenotypes that lead to larval death. We characterised the transcriptional profiles of targeted over-expression of bnl in the embryonic trachea and of loss-of-function bnl(P1) mutant embryos. Gene expression data was mapped to high-throughput in situ hybridisation based ImaGO-annotation. Thus, we identified and confirmed by quantitative PCR 13 Bnl-dependent genes that are expressed in cells within and outside of the tracheal system.

Putative synaptic genes defined from a Drosophila whole body developmental transcriptome by a machine learning approach.

PubMed

Pazos Obregón, Flavio; Papalardo, Cecilia; Castro, Sebastián; Guerberoff, Gustavo; Cantera, Rafael

2015-09-15

Assembly and function of neuronal synapses require the coordinated expression of a yet undetermined set of genes. Although roughly a thousand genes are expected to be important for this function in Drosophila melanogaster, just a few hundreds of them are known so far. In this work we trained three learning algorithms to predict a "synaptic function" for genes of Drosophila using data from a whole-body developmental transcriptome published by others. Using statistical and biological criteria to analyze and combine the predictions, we obtained a gene catalogue that is highly enriched in genes of relevance for Drosophila synapse assembly and function but still not recognized as such. The utility of our approach is that it reduces the number of genes to be tested through hypothesis-driven experimentation.

A screen for immunity genes evolving under positive selection in Drosophila.

PubMed

Jiggins, F M; Kim, K W

2007-05-01

Genes involved in the immune system tend to have higher rates of adaptive evolution than other genes in the genome, probably because they are coevolving with pathogens. We have screened a sample of Drosophila genes to identify those evolving under positive selection. First, we identified rapidly evolving immunity genes by comparing 140 loci in Drosophila erecta and D. yakuba. Secondly, we resequenced 23 of the fastest evolving genes from the independent species pair D. melanogaster and D. simulans, and identified those under positive selection using a McDonald-Kreitman test. There was strong evidence of adaptive evolution in two serine proteases (persephone and spirit) and a homolog of the Anopheles serpin SRPN6, and weaker evidence in another serine protease and the death domain protein dFADD. These results add to mounting evidence that immune signalling pathway molecules often evolve rapidly, possibly because they are sites of host-parasite coevolution.

Fisetin Inhibits Migration and Invasion of Human Cervical Cancer Cells by Down-Regulating Urokinase Plasminogen Activator Expression through Suppressing the p38 MAPK-Dependent NF-κB Signaling Pathway

PubMed Central

Chou, Ruey-Hwang; Hsieh, Shu-Ching; Yu, Yung-Luen; Huang, Min-Hsien; Huang, Yi-Chang; Hsieh, Yi-Hsien

2013-01-01

Fisetin (3,3’,4’,7-tetrahydroxyflavone), a naturally occurring flavonoid, has been reported to inhibit proliferation and induce apoptosis in several cancer types. However, its effect on the anti-metastatic potential of cervical cancer cells remains unclear. In the present study, we found that fisetin inhibits the invasion and migration of cervical cancer cells. The expression and activity of urokinase plasminogen activator (uPA) was significantly suppressed by fisetin in a dose-dependent manner. We also demonstrated that fisetin reduces the phosphorylation of p38 MAPK, but not that of ERK1/2, JNK1/2, or AKT. Addition of a p38 MAPK inhibitor, SB203580, further enhanced the inhibitory effect of fisetin on the expression and activity of uPA and the invasion and motility in cervical cancer cells. Fisetin suppressed the TPA (tetradecanoylphorbol-13-acetate)-induced activation of p38 MAPK and uPA, and inhibited the TPA-enhanced migratory and invasive abilities. Furthermore, the promoter activity of the uPA gene was dramatically repressed by fisetin, which disrupted the nuclear translocation of NF-κB and its binding amount on the promoter of the uPA gene, and these suppressive effects could be further enhanced by SB203580. This study provides strong evidence for the molecular mechanism of fisetin in inhibiting the aggressive phenotypes by repression of uPA via interruption of p38 MAPK-dependent NF-κB signaling pathway in cervical cancer cells and thus contributes insight to the potential of using fisetin as a therapeutic strategy against cervical cancer by inhibiting migration and invasion. PMID:23940799

Evolutionarily conserved heterogeneous nuclear ribonucleoprotein (hnRNP) A/B proteins functionally interact with human and Drosophila TAR DNA-binding protein 43 (TDP-43).

PubMed

Romano, Maurizio; Buratti, Emanuele; Romano, Giulia; Klima, Raffaella; Del Bel Belluz, Lisa; Stuani, Cristiana; Baralle, Francisco; Feiguin, Fabian

2014-03-07

Human TDP-43 represents the main component of neuronal inclusions found in patients with neurodegenerative diseases, especially frontotemporal lobar degeneration and amyotrophic lateral sclerosis. In vitro and in vivo studies have shown that the TAR DNA-binding protein 43 (TDP-43) Drosophila ortholog (TBPH) can biochemically and functionally overlap the properties of the human factor. The recent direct implication of the human heterogeneous nuclear ribonucleoproteins (hnRNPs) A2B1 and A1, known TDP-43 partners, in the pathogenesis of multisystem proteinopathy and amyotrophic lateral sclerosis supports the hypothesis that the physical and functional interplay between TDP-43 and hnRNP A/B orthologs might play a crucial role in the pathogenesis of neurodegenerative diseases. To test this hypothesis and further validate the fly system as a useful model to study this type of diseases, we have now characterized human TDP-43 and Drosophila TBPH similarity in terms of protein-protein interaction pathways. In this work we show that TDP-43 and TBPH share the ability to associate in vitro with Hrp38/Hrb98DE/CG9983, the fruit fly ortholog of the human hnRNP A1/A2 factors. Interestingly, the protein regions of TDP-43 and Hrp38 responsible for reciprocal interactions are conserved through evolution. Functionally, experiments in HeLa cells demonstrate that TDP-43 is necessary for the inhibitory activity of Hrp38 on splicing. Finally, Drosophila in vivo studies show that Hrp38 deficiency produces locomotive defects and life span shortening in TDP-43 with and without animals. These results suggest that hnRNP protein levels can play a modulatory role on TDP-43 functions.

Systemic Activin signaling independently regulates sugar homeostasis, cellular metabolism, and pH balance in Drosophila melanogaster

PubMed Central

Ghosh, Arpan C.; O’Connor, Michael B.

2014-01-01

The ability to maintain cellular and physiological metabolic homeostasis is key for the survival of multicellular organisms in changing environmental conditions. However, our understanding of extracellular signaling pathways that modulate metabolic processes remains limited. In this study we show that the Activin-like ligand Dawdle (Daw) is a major regulator of systemic metabolic homeostasis and cellular metabolism in Drosophila. We find that loss of canonical Smad signaling downstream of Daw leads to defects in sugar and systemic pH homeostasis. Although Daw regulates sugar homeostasis by positively influencing insulin release, we find that the effect of Daw on pH balance is independent of its role in insulin signaling and is caused by accumulation of organic acids that are primarily tricarboxylic acid (TCA) cycle intermediates. RNA sequencing reveals that a number of TCA cycle enzymes and nuclear-encoded mitochondrial genes including genes involved in oxidative phosphorylation and β-oxidation are up-regulated in the daw mutants, indicating either a direct or indirect role of Daw in regulating these genes. These findings establish Activin signaling as a major metabolic regulator and uncover a functional link between TGF-β signaling, insulin signaling, and metabolism in Drosophila. PMID:24706779

Minos-insertion mutant of the Drosophila GBA gene homologue showed abnormal phenotypes of climbing ability, sleep and life span with accumulation of hydroxy-glucocerebroside.

PubMed

Kawasaki, Haruhisa; Suzuki, Takahiro; Ito, Kumpei; Takahara, Tsubasa; Goto-Inoue, Naoko; Setou, Mitsutoshi; Sakata, Kazuki; Ishida, Norio

2017-05-30

Gaucher's disease in humans is considered a deficiency of glucocerebrosidase (GlcCerase) that result in the accumulation of its substrate, glucocerebroside (GlcCer). Although mouse models of Gaucher's disease have been reported from several laboratories, these models are limited due to the perinatal lethality of GlcCerase gene. Here, we examined phenotypes of Drosophila melanogaster homologues genes of the human Gaucher's disease gene by using Minos insertion. One of two Minos insertion mutants to unknown function gene (CG31414) accumulates the hydroxy-GlcCer in whole body of Drosophila melanogaster. This mutant showed abnormal phenotypes of climbing ability and sleep, and short lifespan. These abnormal phenotypes are very similar to that of Gaucher's disease in human. In contrast, another Minos insertion mutant (CG31148) and its RNAi line did not show such severe phenotype as observed in CG31414 gene mutation. The data suggests that Drosophila CG31414 gene mutation might be useful for unraveling the molecular mechanism of Gaucher's disease. Copyright © 2017 Elsevier B.V. All rights reserved.

Drosophila FoxP Mutants Are Deficient in Operant Self-Learning

PubMed Central

Mendoza, Ezequiel; Colomb, Julien; Rybak, Jürgen; Pflüger, Hans-Joachim; Zars, Troy

2014-01-01

Intact function of the Forkhead Box P2 (FOXP2) gene is necessary for normal development of speech and language. This important role has recently been extended, first to other forms of vocal learning in animals and then also to other forms of motor learning. The homology in structure and in function among the FoxP gene members raises the possibility that the ancestral FoxP gene may have evolved as a crucial component of the neural circuitry mediating motor learning. Here we report that genetic manipulations of the single Drosophila orthologue, dFoxP, disrupt operant self-learning, a form of motor learning sharing several conceptually analogous features with language acquisition. Structural alterations of the dFoxP locus uncovered the role of dFoxP in operant self-learning and habit formation, as well as the dispensability of dFoxP for operant world-learning, in which no motor learning occurs. These manipulations also led to subtle alterations in the brain anatomy, including a reduced volume of the optic glomeruli. RNAi-mediated interference with dFoxP expression levels copied the behavioral phenotype of the mutant flies, even in the absence of mRNA degradation. Our results provide evidence that motor learning and language acquisition share a common ancestral trait still present in extant invertebrates, manifest in operant self-learning. This ‘deep’ homology probably traces back to before the split between vertebrate and invertebrate animals. PMID:24964149

A damped oscillator imposes temporal order on posterior gap gene expression in Drosophila.

PubMed

Verd, Berta; Clark, Erik; Wotton, Karl R; Janssens, Hilde; Jiménez-Guri, Eva; Crombach, Anton; Jaeger, Johannes

2018-02-01

Insects determine their body segments in two different ways. Short-germband insects, such as the flour beetle Tribolium castaneum, use a molecular clock to establish segments sequentially. In contrast, long-germband insects, such as the vinegar fly Drosophila melanogaster, determine all segments simultaneously through a hierarchical cascade of gene regulation. Gap genes constitute the first layer of the Drosophila segmentation gene hierarchy, downstream of maternal gradients such as that of Caudal (Cad). We use data-driven mathematical modelling and phase space analysis to show that shifting gap domains in the posterior half of the Drosophila embryo are an emergent property of a robust damped oscillator mechanism, suggesting that the regulatory dynamics underlying long- and short-germband segmentation are much more similar than previously thought. In Tribolium, Cad has been proposed to modulate the frequency of the segmentation oscillator. Surprisingly, our simulations and experiments show that the shift rate of posterior gap domains is independent of maternal Cad levels in Drosophila. Our results suggest a novel evolutionary scenario for the short- to long-germband transition and help explain why this transition occurred convergently multiple times during the radiation of the holometabolan insects.

A damped oscillator imposes temporal order on posterior gap gene expression in Drosophila

PubMed Central

Verd, Berta; Clark, Erik; Wotton, Karl R.; Janssens, Hilde; Jiménez-Guri, Eva; Crombach, Anton

2018-01-01

Insects determine their body segments in two different ways. Short-germband insects, such as the flour beetle Tribolium castaneum, use a molecular clock to establish segments sequentially. In contrast, long-germband insects, such as the vinegar fly Drosophila melanogaster, determine all segments simultaneously through a hierarchical cascade of gene regulation. Gap genes constitute the first layer of the Drosophila segmentation gene hierarchy, downstream of maternal gradients such as that of Caudal (Cad). We use data-driven mathematical modelling and phase space analysis to show that shifting gap domains in the posterior half of the Drosophila embryo are an emergent property of a robust damped oscillator mechanism, suggesting that the regulatory dynamics underlying long- and short-germband segmentation are much more similar than previously thought. In Tribolium, Cad has been proposed to modulate the frequency of the segmentation oscillator. Surprisingly, our simulations and experiments show that the shift rate of posterior gap domains is independent of maternal Cad levels in Drosophila. Our results suggest a novel evolutionary scenario for the short- to long-germband transition and help explain why this transition occurred convergently multiple times during the radiation of the holometabolan insects. PMID:29451884

Identifying candidate genes affecting developmental time in Drosophila melanogaster: pervasive pleiotropy and gene-by-environment interaction

PubMed Central

Mensch, Julián; Lavagnino, Nicolás; Carreira, Valeria Paula; Massaldi, Ana; Hasson, Esteban; Fanara, Juan José

2008-01-01

Background Understanding the genetic architecture of ecologically relevant adaptive traits requires the contribution of developmental and evolutionary biology. The time to reach the age of reproduction is a complex life history trait commonly known as developmental time. In particular, in holometabolous insects that occupy ephemeral habitats, like fruit flies, the impact of developmental time on fitness is further exaggerated. The present work is one of the first systematic studies of the genetic basis of developmental time, in which we also evaluate the impact of environmental variation on the expression of the trait. Results We analyzed 179 co-isogenic single P[GT1]-element insertion lines of Drosophila melanogaster to identify novel genes affecting developmental time in flies reared at 25°C. Sixty percent of the lines showed a heterochronic phenotype, suggesting that a large number of genes affect this trait. Mutant lines for the genes Merlin and Karl showed the most extreme phenotypes exhibiting a developmental time reduction and increase, respectively, of over 2 days and 4 days relative to the control (a co-isogenic P-element insertion free line). In addition, a subset of 42 lines selected at random from the initial set of 179 lines was screened at 17°C. Interestingly, the gene-by-environment interaction accounted for 52% of total phenotypic variance. Plastic reaction norms were found for a large number of developmental time candidate genes. Conclusion We identified components of several integrated time-dependent pathways affecting egg-to-adult developmental time in Drosophila. At the same time, we also show that many heterochronic phenotypes may arise from changes in genes involved in several developmental mechanisms that do not explicitly control the timing of specific events. We also demonstrate that many developmental time genes have pleiotropic effects on several adult traits and that the action of most of them is sensitive to temperature during

Uncoupling neurogenic gene networks in the Drosophila embryo.

PubMed

Rogers, William A; Goyal, Yogesh; Yamaya, Kei; Shvartsman, Stanislav Y; Levine, Michael S

2017-04-01

The EGF signaling pathway specifies neuronal identities in the Drosophila embryo by regulating developmental patterning genes such as intermediate neuroblasts defective ( ind ). EGFR is activated in the ventral midline and neurogenic ectoderm by the Spitz ligand, which is processed by the Rhomboid protease. CRISPR/Cas9 was used to delete defined rhomboid enhancers mediating expression at each site of Spitz processing. Surprisingly, the neurogenic ectoderm, not the ventral midline, was found to be the dominant source of EGF patterning activity. We suggest that Drosophila is undergoing an evolutionary transition in central nervous system (CNS)-organizing activity from the ventral midline to the neurogenic ectoderm. © 2017 Rogers et al.; Published by Cold Spring Harbor Laboratory Press.

Characterization of CG6178 gene product with high sequence similarity to firefly luciferase in Drosophila melanogaster.

PubMed

Oba, Yuichi; Ojika, Makoto; Inouye, Satoshi

2004-03-31

This is the first identification of a long-chain fatty acyl-CoA synthetase in Drosophila by enzymatic characterization. The gene product of CG6178 (CG6178) in Drosophila melanogaster genome, which has a high sequence similarity to firefly luciferase, has been expressed and characterized. CG6178 showed long-chain fatty acyl-CoA synthetic activity in the presence of ATP, CoA and Mg(2+), suggesting a fatty acyl adenylate is an intermediate. Recently, it was revealed that firefly luciferase has two catalytic functions, monooxygenase (luciferase) and AMP-mediated CoA ligase (fatty acyl-CoA synthetase). However, unlike firefly luciferase, CG6178 did not show luminescence activity in the presence of firefly luciferin, ATP, CoA and Mg(2+). The enzymatic properties of CG6178 including substrate specificity, pH dependency and optimal temperature were close to those of firefly luciferase and rat fatty acyl-CoA synthetase. Further, phylogenic analyses strongly suggest that the firefly luciferase gene may have evolved from a fatty acyl-CoA synthetase gene as a common ancestral gene.

The NSL Complex Regulates Housekeeping Genes in Drosophila

PubMed Central

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

Identification of Genes Associated with Resilience/Vulnerability to Sleep Deprivation and Starvation in Drosophila

PubMed Central

Thimgan, Matthew S.; Seugnet, Laurent; Turk, John; Shaw, Paul J.

2015-01-01

, Seugnet L, Turk J, Shaw PJ. Identification of genes associated with resilience/vulnerability to sleep deprivation and starvation in Drosophila. SLEEP 2015;38(5):801–814. PMID:25409104

Deciphering the combinatorial architecture of a Drosophila homeotic gene enhancer

PubMed Central

Drewell, Robert A.; Nevarez, Michael J.; Kurata, Jessica S.; Winkler, Lauren N.; Li, Lily; Dresch, Jacqueline M.

2013-01-01

Summary In Drosophila, the 330 kb bithorax complex regulates cellular differentiation along the anterio-posterior axis during development in the thorax and abdomen and is comprised of three homeotic genes: Ultrabithorax, abdominal-A, and Abdominal-B. The expression of each of these genes is in turn controlled through interactions between transcription factors and a number of cis-regulatory modules in the neighboring intergenic regions. In this study, we examine how the sequence architecture of transcription factor binding sites mediates the functional activity of one of these cis-regulatory modules. Using computational, mathematical modeling and experimental molecular genetic approaches we investigate the IAB7b enhancer, which regulates Abdominal-B expression specifically in the presumptive seventh and ninth abdominal segments of the early embryo. A cross-species comparison of the IAB7b enhancer reveals an evolutionarily conserved signature motif containing two FUSHI-TARAZU activator transcription factor binding sites. We find that the transcriptional repressors KNIRPS, KRUPPEL and GIANT are able to restrict reporter gene expression to the posterior abdominal segments, using different molecular mechanisms including short-range repression and competitive binding. Additionally, we show the functional importance of the spacing between the two FUSHI-TARAZU binding sites and discuss the potential importance of cooperativity for transcriptional activation. Our results demonstrate that the transcriptional output of the IAB7b cis-regulatory module relies on a complex set of combinatorial inputs mediated by specific transcription factor binding and that the sequence architecture at this enhancer is critical to maintain robust regulatory function. PMID:24514265

Fast and Efficient Drosophila melanogaster Gene Knock-Ins Using MiMIC Transposons

PubMed Central

Vilain, Sven; Vanhauwaert, Roeland; Maes, Ine; Schoovaerts, Nils; Zhou, Lujia; Soukup, Sandra; da Cunha, Raquel; Lauwers, Elsa; Fiers, Mark; Verstreken, Patrik

2014-01-01

Modern molecular genetics studies necessitate the manipulation of genes in their endogenous locus, but most of the current methodologies require an inefficient donor-dependent homologous recombination step to locally modify the genome. Here we describe a methodology to efficiently generate Drosophila knock-in alleles by capitalizing on the availability of numerous genomic MiMIC transposon insertions carrying recombinogenic attP sites. Our methodology entails the efficient PhiC31-mediated integration of a recombination cassette flanked by unique I-SceI and/or I-CreI restriction enzyme sites into an attP-site. These restriction enzyme sites allow for double-strand break−mediated removal of unwanted flanking transposon sequences, while leaving the desired genomic modifications or recombination cassettes. As a proof-of-principle, we mutated LRRK, tau, and sky by using different MiMIC elements. We replaced 6 kb of genomic DNA encompassing the tau locus and 35 kb encompassing the sky locus with a recombination cassette that permits easy integration of DNA at these loci and we also generated a functional LRRKHA knock in allele. Given that ~92% of the Drosophila genes are located within the vicinity (<35 kb) of a MiMIC element, our methodology enables the efficient manipulation of nearly every locus in the fruit fly genome without the need for inefficient donor-dependent homologous recombination events. PMID:25298537

Fast and efficient Drosophila melanogaster gene knock-ins using MiMIC transposons.

PubMed

Vilain, Sven; Vanhauwaert, Roeland; Maes, Ine; Schoovaerts, Nils; Zhou, Lujia; Soukup, Sandra; da Cunha, Raquel; Lauwers, Elsa; Fiers, Mark; Verstreken, Patrik

2014-10-08

Modern molecular genetics studies necessitate the manipulation of genes in their endogenous locus, but most of the current methodologies require an inefficient donor-dependent homologous recombination step to locally modify the genome. Here we describe a methodology to efficiently generate Drosophila knock-in alleles by capitalizing on the availability of numerous genomic MiMIC transposon insertions carrying recombinogenic attP sites. Our methodology entails the efficient PhiC31-mediated integration of a recombination cassette flanked by unique I-SceI and/or I-CreI restriction enzyme sites into an attP-site. These restriction enzyme sites allow for double-strand break-mediated removal of unwanted flanking transposon sequences, while leaving the desired genomic modifications or recombination cassettes. As a proof-of-principle, we mutated LRRK, tau, and sky by using different MiMIC elements. We replaced 6 kb of genomic DNA encompassing the tau locus and 35 kb encompassing the sky locus with a recombination cassette that permits easy integration of DNA at these loci and we also generated a functional LRRK(HA) knock in allele. Given that ~92% of the Drosophila genes are located within the vicinity (<35 kb) of a MiMIC element, our methodology enables the efficient manipulation of nearly every locus in the fruit fly genome without the need for inefficient donor-dependent homologous recombination events. Copyright © 2014 Vilain et al.

Functional conservation of the human EXT1 tumor suppressor gene and its Drosophila homolog tout velu.

PubMed

Dasgupta, Ujjaini; Dixit, Bharat L; Rusch, Melissa; Selleck, Scott; The, Inge

2007-08-01

Heparan sulfate proteoglycans play a vital role in signaling of various growth factors in both Drosophila and vertebrates. In Drosophila, mutations in the tout velu (ttv) gene, a homolog of the mammalian EXT1 tumor suppressor gene, leads to abrogation of glycosaminoglycan (GAG) biosynthesis. This impairs distribution and signaling activities of various morphogens such as Hedgehog (Hh), Wingless (Wg), and Decapentaplegic (Dpp). Mutations in members of the exostosin (EXT) gene family lead to hereditary multiple exostosis in humans leading to bone outgrowths and tumors. In this study, we provide genetic and biochemical evidence that the human EXT1 (hEXT1) gene is conserved through species and can functionally complement the ttv mutation in Drosophila. The hEXT1 gene was able to rescue a ttv null mutant to adulthood and restore GAG biosynthesis.

Drosophila Melanogaster Mitochondrial DNA: Gene Organization and Evolutionary Considerations

PubMed Central

Garesse, R.

1988-01-01

The sequence of a 8351-nucleotide mitochondrial DNA (mtDNA) fragment has been obtained extending the knowledge of the Drosophila melanogaster mitochondrial genome to 90% of its coding region. The sequence encodes seven polypeptides, 12 tRNAs and the 3' end of the 16S rRNA and CO III genes. The gene organization is strictly conserved with respect to the Drosophila yakuba mitochondrial genome, and different from that found in mammals and Xenopus. The high A + T content of D. melanogaster mitochondrial DNA is reflected in a reiterative codon usage, with more than 90% of the codons ending in T or A, G + C rich codons being practically absent. The average level of homology between the D. melanogaster and D. yakuba sequences is very high (roughly 94%), although insertion and deletions have been detected in protein, tRNA and large ribosomal genes. The analysis of nucleotide changes reveals a similar frequency for transitions and transversions, and reflects a strong bias against G+C on both strands. The predominant type of transition is strand specific. PMID:3130291

Keratinocyte p38δ loss inhibits Ras-induced tumor formation, while systemic p38δ loss enhances skin inflammation in the early phase of chemical carcinogenesis in mouse skin.

PubMed

Kiss, Alexi; Koppel, Aaron C; Anders, Joanna; Cataisson, Christophe; Yuspa, Stuart H; Blumenberg, Miroslav; Efimova, Tatiana

2016-05-01

p38δ expression and/or activity are increased in human cutaneous malignancies, including invasive squamous cell carcinoma (SCC) and head and neck SCC, but the role of p38δ in cutaneous carcinogenesis has not been well-defined. We have reported that mice with germline loss of p38δ exhibited a reduced susceptibility to skin tumor development compared with wild-type mice in the two-stage 7,12-dimethylbenz(a)anthracene (DMBA)/12-O-tetradecanoylphorbol-13-acetate (TPA) chemical skin carcinogenesis model. Here, we report that p38δ gene ablation inhibited the growth of tumors generated from v-ras(Ha) -transformed keratinocytes in skin orthografts to nude mice, indicating that keratinocyte-intrinsic p38δ is required for Ras-induced tumorigenesis. Gene expression profiling of v-ras(Ha) -transformed p38δ-null keratinocytes revealed transcriptional changes associated with cellular responses linked to tumor suppression, such as reduced proliferation and increased differentiation, cell adhesion, and cell communications. Notably, a short-term DMBA/TPA challenge, modeling the initial stages of chemical skin carcinogenesis treatment, elicited an enhanced inflammation in p38δ-null skin compared with skin of wild-type mice, as assessed by measuring the expression of pro-inflammatory cytokines, including IL-1β, IL-6, IL-17, and TNFα. Additionally, p38δ-null skin and p38δ-null keratinocytes exhibited increased p38α activation and signaling in response to acute inflammatory challenges, suggesting a role for p38α in stimulating the elevated inflammatory response in p38δ-null skin during the initial phases of the DMBA/TPA treatment compared with similarly treated p38δ(+/+) skin. Altogether, our results indicate that p38δ signaling regulates skin carcinogenesis not only by keratinocyte cell-autonomous mechanisms, but also by influencing the interaction between between the epithelial compartment of the developing skin tumor and its stromal microenvironment. © 2015 Wiley

Cloning of the cDNA for a human homologue of the Drosophila white gene and mapping to chromosome 21q22.3

DOE Office of Scientific and Technical Information (OSTI.GOV)

Haiming Chen; Lalioti, M.D.; Perrin, G.

1996-07-01

In an effort to contribute to the transcript map of human chromosome 21 and the understanding of the pathophysiology of trisomy 21, we have used exon trapping to identify fragments of chromosome 21 genes. Two trapped exons, from pools of chromosome 21-specific cosmids, showed homology to the Drosophila white (w) gene. We subsequently cloned the corresponding cDNA for a human homologue of the Drosophila w gene (hW) from human retina and fetal brain cDNA libraries. The gene belongs to the ATP-binding cassette transporter gene family and is homologous to Drosophila w (and to 2 genes from other species) and tomore » a lesser extent to Drosophila brown (bw) and scarlet (st) genes that are all involved in the transport of eye pigment precursor molecules. A DNA polymorphism with 62% heterozygosity due to variation of a poly (T) region in the 3{prime} UTR of the hW has been identified and used for the incorporation of this gene to the genetic map of chromosome 21. The hW is located at 21q22.3 between DNA markers D21S212 and D21S49 in a P1 clone that also contains marker BCEI. The gene is expressed at various levels in many human tissues. The contributions of this gene to the Down syndrome phenotypes, to human eye color, and to the resulting phenotypes of null or missense mutations are presently unknown. 56 refs., 8 figs., 1 tab.« less

Gene expression variations during Drosophila metamorphosis in space: The GENE experiment in the Spanish cervantes missions to the ISS

NASA Astrophysics Data System (ADS)

Herranz, Raul; Benguria, Alberto; Medina, Javier; Gasset, Gilbert; van Loon, Jack J.; Zaballos, Angel; Marco, Roberto

2005-08-01

The ISS expedition 8, a Soyuz Mission, flew to the International Space Station (ISS) to replace the two- member ISS crew during October 2003. During this crew exchanging flight, the Spanish Cervantes Scientific Mission took place. In it some biological experiments were performed among them three proposed by our Team. The third member of the expedition, the Spanish born ESA astronaut Pedro Duque, returned within the Soyuz 7 capsule carrying the experiment containing transport box after almost 11 days in microgravity. In one of the three experiments, the GENE experiment, we intended to determine how microgravity affects the gene expression pattern of Drosophila with one of the current more powerful technologies , a complete Drosophila melanogaster genome microarray (AffymetrixTM, version 1.0). Due to the constrains in the current ISS experiments, we decided to limit our experiment to the organism rebuilding processes that occurs during Drosophila metamorphosis. In addition to the ISS samples, several control experiments have been performed including a 1g Ground control parallel to the ISS flight samples, a Random Position Machine microgravity simulated control and a parallel Hypergravity (10g) experiment. Extracted RNA from the samples was used to test the differences in gene expression during Drosophila development. A preliminary analysis of the results indicates that around five hundred genes change their expression profiles, many of them belonging to particular ontology classification groups.

A novel acetylcholinesterase gene in mosquitoes codes for the insecticide target and is non-homologous to the ace gene in Drosophila.

PubMed Central

Weill, Mylène; Fort, Philippe; Berthomieu, Arnaud; Dubois, Marie Pierre; Pasteur, Nicole; Raymond, Michel

2002-01-01

Acetylcholinesterase (AChE) is the target of two major insecticide families, organophosphates (OPs) and carbamates. AChE insensitivity is a frequent resistance mechanism in insects and responsible mutations in the ace gene were identified in two Diptera, Drosophila melanogaster and Musca domestica. However, for other insects, the ace gene cloned by homology with Drosophila does not code for the insensitive AChE in resistant individuals, indicating the existence of a second ace locus. We identified two AChE loci in the genome of Anopheles gambiae, one (ace-1) being a new locus and the other (ace-2) being homologous to the gene previously described in Drosophila. The gene ace-1 has no obvious homologue in the Drosophila genome and was found in 15 mosquito species investigated. In An. gambiae, ace-1 and ace-2 display 53% similarity at the amino acid level and an overall phylogeny indicates that they probably diverged before the differentiation of insects. Thus, both genes are likely to be present in the majority of insects and the absence of ace-1 in Drosophila is probably due to a secondary loss. In one mosquito (Culex pipiens), ace-1 was found to be tightly linked with insecticide resistance and probably encodes the AChE OP target. These results have important implications for the design of new insecticides, as the target AChE is thus encoded by distinct genes in different insect groups, even within the Diptera: ace-2 in at least the Drosophilidae and Muscidae and ace-1 in at least the Culicidae. Evolutionary scenarios leading to such a peculiar situation are discussed. PMID:12396499

big bang gene modulates gut immune tolerance in Drosophila.

PubMed

Bonnay, François; Cohen-Berros, Eva; Hoffmann, Martine; Kim, Sabrina Y; Boulianne, Gabrielle L; Hoffmann, Jules A; Matt, Nicolas; Reichhart, Jean-Marc

2013-02-19

Chronic inflammation of the intestine is detrimental to mammals. Similarly, constant activation of the immune response in the gut by the endogenous flora is suspected to be harmful to Drosophila. Therefore, the innate immune response in the gut of Drosophila melanogaster is tightly balanced to simultaneously prevent infections by pathogenic microorganisms and tolerate the endogenous flora. Here we describe the role of the big bang (bbg) gene, encoding multiple membrane-associated PDZ (PSD-95, Discs-large, ZO-1) domain-containing protein isoforms, in the modulation of the gut immune response. We show that in the adult Drosophila midgut, BBG is present at the level of the septate junctions, on the apical side of the enterocytes. In the absence of BBG, these junctions become loose, enabling the intestinal flora to trigger a constitutive activation of the anterior midgut immune response. This chronic epithelial inflammation leads to a reduced lifespan of bbg mutant flies. Clearing the commensal flora by antibiotics prevents the abnormal activation of the gut immune response and restores a normal lifespan. We now provide genetic evidence that Drosophila septate junctions are part of the gut immune barrier, a function that is evolutionarily conserved in mammals. Collectively, our data suggest that septate junctions are required to maintain the subtle balance between immune tolerance and immune response in the Drosophila gut, which represents a powerful model to study inflammatory bowel diseases.

Identification of Genes That Interact With Drosophila liquid facets

PubMed Central

Eun, Suk Ho; Lea, Kristi; Overstreet, Erin; Stevens, Samuel; Lee, Ji-Hoon; Fischer, Janice A.

2007-01-01

We have performed mutagenesis screens of the Drosophila X chromosome and the autosomes for dominant enhancers of the rough eye resulting from overexpression of liquid facets. The liquid facets gene encodes the homolog of vertebrate endocytic Epsin, an endocytic adapter protein. In Drosophila, Liquid facets is a core component of the Notch signaling pathway required in the signaling cells for ligand endocytosis and signaling. Why ligand internalization by the signaling cells is essential for signaling is a mystery. The requirement for Liquid facets is a hint at the answer, and the genes identified in this screen provide further clues. Mutant alleles of clathrin heavy chain, Rala, split ends, and auxilin were identified as enhancers. We describe the mutant alleles and mutant phenotypes of Rala and aux. We discuss the relevance of all of these genetic interactions to the function of Liquid facets in Notch signaling. PMID:17179082

Unusual Properties of Regulatory DNA from the Drosophila Engrailed Gene: Three ``pairing-Sensitive'' Sites within a 1.6-Kb Region

PubMed Central

Kassis, J. A.

1994-01-01

We have previously shown that a 2-kb fragment of engrailed DNA can suppress expression of a linked marker gene, white, in the P element vector CaSpeR. This suppression is dependent on the presence of two copies of engrailed DNA-containing P elements (P[en]) in proximity in the Drosophila genome (either in cis or in trans). In this study, the 2-kb fragment was dissected and found to contain three fragments of DNA which could mediate white suppression [called ``pairing-sensitive sites'' (PS)]. A PS site was also identified in regulatory DNA from the Drosophila escargot gene. The eye colors of six different P[en] insertions in the escargot gene suggest an interaction between P[en]-encoded and genome-encoded PS sites. I hypothesize that white gene expression from P[en] is repressed by the formation of a protein complex which is initiated at the engrailed PS sites and also requires interactions with flanking genomic DNA. Genes were sought which influence the function of PS sites. Mutations in some Polycomb and trithorax group genes were found to affect the eye color from some P[en] insertion sites. However, different mutations affected expression from different P[en] insertion sites and no one mutation was found to affect expression from all P[en] insertion sites examined. These results suggest that white expression from P[en] is not directly regulated by members of the Polycomb and trithorax group genes, but in some cases can be influenced by them. I propose that engrailed PS sites normally act to promote interactions between distantly located engrailed regulatory sites and the engrailed promoter. PMID:8005412

Mechanisms of gap gene expression canalization in the Drosophila blastoderm.

PubMed

Gursky, Vitaly V; Panok, Lena; Myasnikova, Ekaterina M; Manu; Samsonova, Maria G; Reinitz, John; Samsonov, Alexander M

2011-01-01

Extensive variation in early gap gene expression in the Drosophila blastoderm is reduced over time because of gap gene cross regulation. This phenomenon is a manifestation of canalization, the ability of an organism to produce a consistent phenotype despite variations in genotype or environment. The canalization of gap gene expression can be understood as arising from the actions of attractors in the gap gene dynamical system. In order to better understand the processes of developmental robustness and canalization in the early Drosophila embryo, we investigated the dynamical effects of varying spatial profiles of Bicoid protein concentration on the formation of the expression border of the gap gene hunchback. At several positions on the anterior-posterior axis of the embryo, we analyzed attractors and their basins of attraction in a dynamical model describing expression of four gap genes with the Bicoid concentration profile accounted as a given input in the model equations. This model was tested against a family of Bicoid gradients obtained from individual embryos. These gradients were normalized by two independent methods, which are based on distinct biological hypotheses and provide different magnitudes for Bicoid spatial variability. We showed how the border formation is dictated by the biological initial conditions (the concentration gradient of maternal Hunchback protein) being attracted to specific attracting sets in a local vicinity of the border. Different types of these attracting sets (point attractors or one dimensional attracting manifolds) define several possible mechanisms of border formation. The hunchback border formation is associated with intersection of the spatial gradient of the maternal Hunchback protein and a boundary between the attraction basins of two different point attractors. We demonstrated how the positional variability for hunchback is related to the corresponding variability of the basin boundaries. The observed reduction in

Drosophila melanogaster mounts a unique immune response to the Rhabdovirus sigma virus.

PubMed

Tsai, C W; McGraw, E A; Ammar, E-D; Dietzgen, R G; Hogenhout, S A

2008-05-01

Rhabdoviruses are important pathogens of humans, livestock, and plants that are often vectored by insects. Rhabdovirus particles have a characteristic bullet shape with a lipid envelope and surface-exposed transmembrane glycoproteins. Sigma virus (SIGMAV) is a member of the Rhabdoviridae and is a naturally occurring disease agent of Drosophila melanogaster. The infection is maintained in Drosophila populations through vertical transmission via germ cells. We report here the nature of the Drosophila innate immune response to SIGMAV infection as revealed by quantitative reverse transcription-PCR analysis of differentially expressed genes identified by microarray analysis. We have also compared and contrasted the immune response of the host with respect to two nonenveloped viruses, Drosophila C virus (DCV) and Drosophila X virus (DXV). We determined that SIGMAV infection upregulates expression of the peptidoglycan receptor protein genes PGRP-SB1 and PGRP-SD and the antimicrobial peptide (AMP) genes Diptericin-A, Attacin-A, Attacin-B, Cecropin-A1, and Drosocin. SIGMAV infection did not induce PGRP-SA and the AMP genes Drosomycin-B, Metchnikowin, and Defensin that are upregulated in DCV and/or DXV infections. Expression levels of the Toll and Imd signaling cascade genes are not significantly altered by SIGMAV infection. These results highlight shared and unique aspects of the Drosophila immune response to the three viruses and may shed light on the nature of the interaction with the host and the evolution of these associations.

Drosophila melanogaster Mounts a Unique Immune Response to the Rhabdovirus Sigma virus▿

PubMed Central

Tsai, C. W.; McGraw, E. A.; Ammar, E.-D.; Dietzgen, R. G.; Hogenhout, S. A.

2008-01-01

Rhabdoviruses are important pathogens of humans, livestock, and plants that are often vectored by insects. Rhabdovirus particles have a characteristic bullet shape with a lipid envelope and surface-exposed transmembrane glycoproteins. Sigma virus (SIGMAV) is a member of the Rhabdoviridae and is a naturally occurring disease agent of Drosophila melanogaster. The infection is maintained in Drosophila populations through vertical transmission via germ cells. We report here the nature of the Drosophila innate immune response to SIGMAV infection as revealed by quantitative reverse transcription-PCR analysis of differentially expressed genes identified by microarray analysis. We have also compared and contrasted the immune response of the host with respect to two nonenveloped viruses, Drosophila C virus (DCV) and Drosophila X virus (DXV). We determined that SIGMAV infection upregulates expression of the peptidoglycan receptor protein genes PGRP-SB1 and PGRP-SD and the antimicrobial peptide (AMP) genes Diptericin-A, Attacin-A, Attacin-B, Cecropin-A1, and Drosocin. SIGMAV infection did not induce PGRP-SA and the AMP genes Drosomycin-B, Metchnikowin, and Defensin that are upregulated in DCV and/or DXV infections. Expression levels of the Toll and Imd signaling cascade genes are not significantly altered by SIGMAV infection. These results highlight shared and unique aspects of the Drosophila immune response to the three viruses and may shed light on the nature of the interaction with the host and the evolution of these associations. PMID:18378641

DEAF-1 regulates immunity gene expression in Drosophila.

PubMed

Reed, Darien E; Huang, Xinhua M; Wohlschlegel, James A; Levine, Michael S; Senger, Kate

2008-06-17

Immunity genes are activated in the Drosophila fat body by Rel and GATA transcription factors. Here, we present evidence that an additional regulatory factor, deformed epidermal autoregulatory factor-1 (DEAF-1), also contributes to the immune response and is specifically important for the induction of two genes encoding antimicrobial peptides, Metchnikowin (Mtk) and Drosomycin (Drs). The systematic mutagenesis of a minimal Mtk 5' enhancer identified a sequence motif essential for both a response to LPS preparations in S2 cells and activation in the larval fat body in response to bacterial infection. Using affinity chromatography coupled to multidimensional protein identification technology (MudPIT), we identified DEAF-1 as a candidate regulator. DEAF-1 activates the expression of Mtk and Drs promoter-luciferase fusion genes in S2 cells. SELEX assays and footprinting data indicate that DEAF-1 binds to and activates Mtk and Drs regulatory DNAs via a TTCGGBT motif. The insertion of this motif into the Diptericin (Dpt) regulatory region confers DEAF-1 responsiveness to this normally DEAF-1-independent enhancer. The coexpression of DEAF-1 with Dorsal, Dif, and Relish results in the synergistic activation of transcription. We propose that DEAF-1 is a regulator of Drosophila immunity.

Regulation of nuclear factor κB (NF-κB) transcriptional activity via p65 acetylation by the chaperonin containing TCP1 (CCT).

PubMed

Pejanovic, Nadja; Hochrainer, Karin; Liu, Tao; Aerne, Birgit L; Soares, Miguel P; Anrather, Josef

2012-01-01

The NF-κB family member p65 is central to inflammation and immunity. The purpose of this study was to identify and characterize evolutionary conserved genes modulating p65 transcriptional activity. Using an RNAi screening approach, we identified chaperonin containing TCP1 subunit η (CCTη) as a regulator of Drosophila NF-κB proteins, Dorsal and Dorsal-related immunity factor (Dif). CCTη was also found to regulate NF-κB-driven transcription in mammalian cells, acting in a promoter-specific context, downstream of IκB kinase (IKK). CCTη knockdown repressed IκBα and CXCL2/MIP2 transcription during the early phase of NF-κB activation while impairing the termination of CCL5/RANTES and CXCL10/IP10 transcription. The latter effect was associated with increased DNA binding and reduced p65 acetylation, presumably by altering the activity of histone acetyltransferase CREB-binding protein (CBP). We identified p65 lysines (K) 122 and 123 as target residues mediating the CCTη-driven termination of NF-κB-dependent transcription. We propose that CCTη regulates NF-κB activity in a manner that resolves inflammation.

Prediction of gene expression in embryonic structures of Drosophila melanogaster.

PubMed

Samsonova, Anastasia A; Niranjan, Mahesan; Russell, Steven; Brazma, Alvis

2007-07-01

Understanding how sets of genes are coordinately regulated in space and time to generate the diversity of cell types that characterise complex metazoans is a major challenge in modern biology. The use of high-throughput approaches, such as large-scale in situ hybridisation and genome-wide expression profiling via DNA microarrays, is beginning to provide insights into the complexities of development. However, in many organisms the collection and annotation of comprehensive in situ localisation data is a difficult and time-consuming task. Here, we present a widely applicable computational approach, integrating developmental time-course microarray data with annotated in situ hybridisation studies, that facilitates the de novo prediction of tissue-specific expression for genes that have no in vivo gene expression localisation data available. Using a classification approach, trained with data from microarray and in situ hybridisation studies of gene expression during Drosophila embryonic development, we made a set of predictions on the tissue-specific expression of Drosophila genes that have not been systematically characterised by in situ hybridisation experiments. The reliability of our predictions is confirmed by literature-derived annotations in FlyBase, by overrepresentation of Gene Ontology biological process annotations, and, in a selected set, by detailed gene-specific studies from the literature. Our novel organism-independent method will be of considerable utility in enriching the annotation of gene function and expression in complex multicellular organisms.

Prediction of Gene Expression in Embryonic Structures of Drosophila melanogaster

PubMed Central

Samsonova, Anastasia A; Niranjan, Mahesan; Russell, Steven; Brazma, Alvis

2007-01-01

Understanding how sets of genes are coordinately regulated in space and time to generate the diversity of cell types that characterise complex metazoans is a major challenge in modern biology. The use of high-throughput approaches, such as large-scale in situ hybridisation and genome-wide expression profiling via DNA microarrays, is beginning to provide insights into the complexities of development. However, in many organisms the collection and annotation of comprehensive in situ localisation data is a difficult and time-consuming task. Here, we present a widely applicable computational approach, integrating developmental time-course microarray data with annotated in situ hybridisation studies, that facilitates the de novo prediction of tissue-specific expression for genes that have no in vivo gene expression localisation data available. Using a classification approach, trained with data from microarray and in situ hybridisation studies of gene expression during Drosophila embryonic development, we made a set of predictions on the tissue-specific expression of Drosophila genes that have not been systematically characterised by in situ hybridisation experiments. The reliability of our predictions is confirmed by literature-derived annotations in FlyBase, by overrepresentation of Gene Ontology biological process annotations, and, in a selected set, by detailed gene-specific studies from the literature. Our novel organism-independent method will be of considerable utility in enriching the annotation of gene function and expression in complex multicellular organisms. PMID:17658945

Differential gene expression related to Nora virus infection of Drosophila melanogaster

PubMed Central

Cordes, Ethan J.; Licking-Murray, Kellie D; Carlson, Kimberly A.

2013-01-01

Nora virus is a recently discovered RNA picorna-like virus that produces a persistent infection in Drosophila melanogaster, but the antiviral pathway or change in gene expression is unknown. We performed cDNA microarray analysis comparing the gene expression profiles of Nora virus infected and uninfected wild-type D. melanogaster. This analysis yielded 58 genes exhibiting a 1.5-fold change or greater and p-value less than 0.01. Of these genes, 46 were up-regulated and 12 down-regulated in response to infection. To validate the microarray results, qRT-PCR was performed with probes for Chorion protein 16 and Troponin C isoform 4, which show good correspondence with cDNA microarray results. Differential regulation of genes associated with Toll and immune-deficient pathways, cytoskeletal development, Janus Kinase-Signal Transducer and Activator of Transcription interactions, and a potential gut-specific innate immune response were found. This genome-wide expression profile of Nora virus infection of D. melanogaster can pinpoint genes of interest for further investigation of antiviral pathways employed, genetic mechanisms, sites of replication, viral persistence, and developmental effects. PMID:23603562

Methylation of CpG island of p14(ARK), p15(INK4b) and p16(INK4a) genes in coke oven workers.

PubMed

Zhang, H; Li, X; Ge, L; Yang, J; Sun, J; Niu, Q

2015-02-01

To detect the blood genomic DNA methylation in coke oven workers and find a possible early screening index for occupational lung cancer, 74 coke oven workers as the exposed group and 47 water pump workers as the controls were surveyed, and urine samples and peripheral blood mononuclear cells (PBMCs) were collected. Airborne benzo[a]pyrene (B[a]P) levels in workplace and urinary 1-hydroxypyrene (1-OH-Py) levels were determined by high-performance liquid chromatography. DNA damage of PBMCs and the p14(ARK), p15(INK4b) and p16(INK4a) gene CpG island methylation in the promoter region were detected by comet assay and methylation-specific polymerase chain reaction techniques, respectively. Results show that compared with the controls, concentration of airborne B[a]Ps was elevated in the coke plant, and urinary 1-OH-Py's level and DNA olive tail moment in comet assay were significantly increased in the coke oven workers, and p14(ARK), p15(INK4b) and p16(INK4a) gene methylation rates were also significantly increased. With the working years and urinary 1-OH-Py's level, the rates of p14(ARK) and p16(INK4a) gene methylation were significantly increased while that of p15(INK4b) gene methylation displayed no statistical change. We conclude that PBMCs' p14(ARK) and p16(INK4a) gene methylation may be used for screening and warning lung cancer in coke oven workers. © The Author(s) 2015.

Interaction between sleep and the immune response in Drosophila: a role for the NFkappaB relish.

PubMed

Williams, Julie A; Sathyanarayanan, Sriram; Hendricks, Joan C; Sehgal, Amita

2007-04-01

The regulation of sleep is poorly understood. While some molecules, including those involved in inflammatory/immune responses, have been implicated in the control of sleep, their role in this process remains unclear. The Drosophila model for sleep provides a powerful system to identify and test the role of sleep-relevant molecules. We conducted an unbiased screen for molecular candidates involved in sleep regulation by analyzing genome-wide changes in gene expression associated with sleep deprivation in Drosophila. To further examine a role of immune-related genes identified in the screen, we performed molecular assays, analysis of sleep behavior in relevant mutant and transgenic flies, and quantitative analysis of the immune response following sleep deprivation. A major class of genes that increased expression with sleep deprivation was that involved in the immune response. We found that immune genes were also upregulated during baseline conditions in the cyc01 sleep mutant. Since the expression of an NFkappaB, Relish, a central player in the inflammatory response, was increased with all manipulations that reduced sleep, we focused on this gene. Flies deficient in, but not lacking, Relish expression exhibited reduced levels of nighttime sleep, supporting a role for Relish in the control of sleep. This mutant phenotype was rescued by expression of a Relish transgene in fat bodies, which are the major site of inflammatory responses in Drosophila. Finally, sleep deprivation also affected the immune response, such that flies deprived of sleep for several hours were more resistant to bacterial infection than those flies not deprived of sleep. These results demonstrate a conserved interaction between sleep and the immune system. Genetic manipulation of an immune component alters sleep, and likewise, acute sleep deprivation alters the immune response.

Interaction Between Sleep and the Immune Response in Drosophila: A Role for the NFκB Relish

PubMed Central

Williams, Julie A.; Sathyanarayanan, Sriram; Hendricks, Joan C.; Sehgal, Amita

2010-01-01

Study Objectives The regulation of sleep is poorly understood. While some molecules, including those involved in inflammatory/immune responses, have been implicated in the control of sleep, their role in this process remains unclear. The Drosophila model for sleep provides a powerful system to identify and test the role of sleep-relevant molecules. Design We conducted an unbiased screen for molecular candidates involved in sleep regulation by analyzing genome-wide changes in gene expression associated with sleep deprivation in Drosophila. To further examine a role of immune-related genes identified in the screen, we performed molecular assays, analysis of sleep behavior in relevant mutant and transgenic flies, and quantitative analysis of the immune response following sleep deprivation. Results A major class of genes that increased expression with sleep deprivation was that involved in the immune response. We found that immune genes were also upregulated during baseline conditions in the cyc01 sleep mutant. Since the expression of an NFκB, Relish, a central player in the inflammatory response, was increased with all manipulations that reduced sleep, we focused on this gene. Flies deficient in, but not lacking, Relish expression exhibited reduced levels of nighttime sleep, supporting a role for Relish in the control of sleep. This mutant phenotype was rescued by expression of a Relish transgene in fat bodies, which are the major site of inflammatory responses in Drosophila. Finally, sleep deprivation also affected the immune response, such that flies deprived of sleep for several hours were more resistant to bacterial infection than those flies not deprived of sleep. Conclusion These results demonstrate a conserved interaction between sleep and the immune system. Genetic manipulation of an immune component alters sleep, and likewise, acute sleep deprivation alters the immune response. PMID:17520783

SlbZIP38, a Tomato bZIP Family Gene Downregulated by Abscisic Acid, Is a Negative Regulator of Drought and Salt Stress Tolerance

PubMed Central

Pan, Yanglu; Hu, Xin; Li, Chunyan; Xu, Xing; Su, Chenggang; Li, Jinhua; Song, Hongyuan; Zhang, Xingguo; Pan, Yu

2017-01-01

The basic leucine zipper (bZIP) transcription factors have crucial roles in plant stress responses. In this study, the bZIP family gene SlbZIP38 (GenBank accession No: XM004239373) was isolated from a tomato (Solanum lycopersicum cv. Ailsa Craig) mature leaf cDNA library. The DNA sequence of SlbZIP38 encodes a protein of 484 amino acids, including a highly conserved bZIP DNA-binding domain in the C-terminal region. We found that SlbZIP38 was differentially expressed in various organs of the tomato plant and was downregulated by drought, salt stress, and abscisic acid (ABA). However, overexpression of SlbZIP38 significantly decreased drought and salt stress tolerance in tomatoes (Ailsa Craig). The findings that SlbZIP38 overexpression reduced the chlorophyll and free proline content in leaves but increased the malondialdehyde content may explain the reduced drought and salt tolerance observed in these lines. These results suggest that SlbZIP38 is a negative regulator of drought and salt resistance that acts by modulating ABA signaling. PMID:29261143

Molecular Structure and Transformation of the Glucose Dehydrogenase Gene in Drosophila Melanogaster

PubMed Central

Whetten, R.; Organ, E.; Krasney, P.; Cox-Foster, D.; Cavener, D.

1988-01-01

We have precisely mapped and sequenced the three 5' exons of the Drosophila melanogaster Gld gene and have identified the start sites for transcription and translation. The first exon is composed of 335 nucleotides and does not contain any putative translation start codons. The second exon is separated from the first exon by 8 kb and contains the Gld translation start codon. The inferred amino acid sequence of the amino terminus contains two unusual features: three tandem repeats of serine-alanine, and a relatively high density of cysteine residues. P element-mediated transformation experiments demonstrated that a 17.5-kb genomic fragment contains the functional and regulatory components of the Gld gene. PMID:3143620

The Tribolium homeotic gene Abdominal is homologous to abdominal-A of the Drosophila bithorax complex

NASA Technical Reports Server (NTRS)

Stuart, J. J.; Brown, S. J.; Beeman, R. W.; Denell, R. E.; Spooner, B. S. (Principal Investigator)

1993-01-01

The Abdominal gene is a member of the single homeotic complex of the beetle, Tribolium castaneum. An integrated developmental genetic and molecular analysis shows that Abdominal is homologous to the abdominal-A gene of the bithorax complex of Drosophila. abdominal-A mutant embryos display strong homeotic transformations of the anterior abdomen (parasegments 7-9) to PS6, whereas developmental commitments in the posterior abdomen depend primarily on Abdominal-B. In beetle embryos lacking Abdominal function, parasegments throughout the abdomen are transformed to PS6. This observation demonstrates the general functional significance of parasegmental expression among insects and shows that the control of determinative decisions in the posterior abdomen by homeotic selector genes has undergone considerable evolutionary modification.

The Tribolium homeotic gene Abdominal is homologous to abdominal-A of the Drosophila bithorax complex.

PubMed

Stuart, J J; Brown, S J; Beeman, R W; Denell, R E

1993-01-01

The Abdominal gene is a member of the single homeotic complex of the beetle, Tribolium castaneum. An integrated developmental genetic and molecular analysis shows that Abdominal is homologous to the abdominal-A gene of the bithorax complex of Drosophila. abdominal-A mutant embryos display strong homeotic transformations of the anterior abdomen (parasegments 7-9) to PS6, whereas developmental commitments in the posterior abdomen depend primarily on Abdominal-B. In beetle embryos lacking Abdominal function, parasegments throughout the abdomen are transformed to PS6. This observation demonstrates the general functional significance of parasegmental expression among insects and shows that the control of determinative decisions in the posterior abdomen by homeotic selector genes has undergone considerable evolutionary modification.

Artemisia leaf extract induces apoptosis in human endometriotic cells through regulation of the p38 and NFκB pathways.

PubMed

Kim, Ji-Hyun; Jung, Seung-Hyun; Yang, Yeong-In; Ahn, Ji-Hye; Cho, Jin-Gyeong; Lee, Kyung-Tae; Baek, Nam-In; Choi, Jung-Hye

2013-02-13

Artemisia leaves have long been used for the treatment of gynecological disorders, including infertility and dysmenorrhea, which can be commonly caused by endometriosis. In the present study, we investigated the effect of Artemisia princeps extract (APE) on the cell growth and apoptosis of human endometriotic cells. MTT assays and FACS analysis using PI and Annexin staining were performed to study cell viability, cell cycle progression, and apoptosis. We also explored the mechanism of APE-induced effects by evaluating the activation of caspases, Akt, p38, and NFκB. The expressions of XIAP, Bcl-2, and Bcl-xL were measured by real-time RT-PCR and Western blot analyses. APE significantly inhibited the cell viability of 11Z and 12Z human endometriotic epithelial cells. Interestingly, endometriotic cells were more sensitive to APE treatment than immortalized endometrial cells (HES). Treatment with APE induced apoptosis of 11Z cells in a time-dependent manner, as shown by accumulation of sub G1 and apoptotic cell populations. In addition, treatment with APE stimulated the activation of caspase -3, -8, and -9 in a dose- and time-dependent manner. Furthermore, p38 was activated by APE treatment, and the p38 inhibitor SB203580 markedly inhibited APE-induced cell death in 11Z cells. Moreover, treatment with APE suppressed the activation of NFκB and the expressions of anti-apoptotic factors such as XIAP, Bcl-2, and Bcl-xL. These results indicate that APE is a potential anti-endometriotic agent, acting to induce apoptosis of endometrial cells through the modulation of the p38 and NFκB pathways. Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.

Evolutionary Genomics of Genes Involved in Olfactory Behavior in the Drosophila melanogaster Species Group

PubMed Central

Lavagnino, Nicolás; Serra, François; Arbiza, Leonardo; Dopazo, Hernán; Hasson, Esteban

2012-01-01

Previous comparative genomic studies of genes involved in olfactory behavior in Drosophila focused only on particular gene families such as odorant receptor and/or odorant binding proteins. However, olfactory behavior has a complex genetic architecture that is orchestrated by many interacting genes. In this paper, we present a comparative genomic study of olfactory behavior in Drosophila including an extended set of genes known to affect olfactory behavior. We took advantage of the recent burst of whole genome sequences and the development of powerful statistical tools to analyze genomic data and test evolutionary and functional hypotheses of olfactory genes in the six species of the Drosophila melanogaster species group for which whole genome sequences are available. Our study reveals widespread purifying selection and limited incidence of positive selection on olfactory genes. We show that the pace of evolution of olfactory genes is mostly independent of the life cycle stage, and of the number of life cycle stages, in which they participate in olfaction. However, we detected a relationship between evolutionary rates and the position that the gene products occupy in the olfactory system, genes occupying central positions tend to be more constrained than peripheral genes. Finally, we demonstrate that specialization to one host does not seem to be associated with bursts of adaptive evolution in olfactory genes in D. sechellia and D. erecta, the two specialists species analyzed, but rather different lineages have idiosyncratic evolutionary histories in which both historical and ecological factors have been involved. PMID:22346339

Definition of Cis-Acting Elements Regulating Expression of the Drosophila Melanogaster Ninae Opsin Gene by Oligonucleotide-Directed Mutagenesis

PubMed Central

Mismer, D.; Rubin, G. M.

1989-01-01

We have analyzed the cis-acting regulatory sequences of the Rh1 (ninaE) gene in Drosophila melanogaster by P-element-mediated germline transformation of indicator genes transcribed from mutant ninaE promoter sequences. We have previously shown that a 200-bp region extending from -120 to +67 relative to the transcription start site is sufficient to obtain eye-specific expression from the ninaE promoter. In the present study, 22 different 4-13-bp sequences in the -120/+67 promoter region were altered by oligonucleotide-directed mutagenesis. Several of these sequences were found to be required for proper promoter function; two of these are conserved in the promoter of the homologous gene isolated from the related species Drosophila virilis. Alteration of a conserved 9-bp sequence results in aberrant, low level expression in the body. Alteration of a separate 11-bp sequence, found in the promoter regions of several photoreceptor-specific genes of Drosophila, results in an approximately 15-fold reduction in promoter efficiency but without apparent alteration of tissue-specificity. A protein factor capable of interacting with this 11-bp sequence has been detected by DNaseI footprinting in embryonic nuclear extracts. Finally, we have further characterized two separable enhancer sequences previously shown to be required for normal levels of expression from this promoter. PMID:2521839

Molecular Variation of Adh and P6 Genes in an African Population of Drosophila Melanogaster and Its Relation to Chromosomal Inversions

PubMed Central

Benassi, V.; Aulard, S.; Mazeau, S.; Veuille, M.

1993-01-01

Four-cutter molecular polymorphism of Adh and P6, and chromosome inversion polymorphism of chromosome II were investigated in 95 isogenic lines of an Ivory Coast population of Drosophila melanogaster, a species assumed to have recently spread throughout the world from a West African origin. The P6 gene showed little linkage disequilibrium with the In(2L)t inversion, although it is located within this inversion. This suggests that the inversion and the P6 locus have extensively exchanged genetic information through either double crossover or gene conversion. Allozymic variation in ADH was in linkage disequilibrium with In(2L)t and In(2R)NS inversions. Evidence suggests either that inversion linkage with the Fast allele is selectively maintained, or that this allele only recently appeared. Molecular polymorphism at the Adh locus in the Ivory Coast is not higher than in North American populations. New haplotypes specific to the African population were found, some of them connect the ``Wa(s)-like'' haplotypes found at high frequencies in the United States to the other slow haplotypes. Their relation with In(2L)t supports the hypothesis that Wa(s) recently recombined away from an In(2L)t chromosome which may be the cause of its divergence from the other haplotypes. PMID:8349110

Adaptive evolution of newly emerged micro-RNA genes in Drosophila.

PubMed

Lu, Jian; Fu, Yonggui; Kumar, Supriya; Shen, Yang; Zeng, Kai; Xu, Anlong; Carthew, Richard; Wu, Chung-I

2008-05-01

How often micro-RNA (miRNA) genes emerged and how fast they evolved soon after their emergence are some of the central questions in the evolution of miRNAs. Because most known miRNA genes are ancient and highly conserved, these questions can be best answered by identifying newly emerged miRNA genes. Among the 78 miRNA genes in Drosophila reported before 2007, only 5 are confirmed to be newly emerged in the genus (although many more can be found in the newly reported data set; e.g., Ruby et al. 2007; Stark et al. 2007; Lu et al. 2008). These new miRNA genes have undergone numerous changes, even in the normally invariant mature sequences. Four of them (the miR-310/311/312/313 cluster, denoted miR-310s) were duplicated from other conserved miRNA genes. The fifth one (miR-303) appears to be a very young gene, originating de novo from a non-miRNA sequence recently. We sequenced these 5 miRNA genes and their neighboring regions from a worldwide collection of Drosophila melanogaster lines. The levels of divergence and polymorphism in these miRNA genes, vis-à-vis those of the neighboring DNA sequences, suggest that these 5 genes are evolving adaptively. Furthermore, the polymorphism pattern of miR-310s in D. melanogaster is indicative of hitchhiking under positive selection. Thus, a large number of adaptive changes over a long period of time may be essential for the evolution of newly emerged miRNA genes.

Histamine Promotes the Release of Interleukin-6 via the H1R/p38 and NF-κB Pathways in Nasal Fibroblasts.

PubMed

Park, Il-Ho; Um, Ji-Young; Cho, Jung-Sun; Lee, Seung Hoon; Lee, Sang Hag; Lee, Heung-Man

2014-11-01

Based on the close relationship between histamine and interleukin 6 (IL-6), we hypothesized that histamine may regulate the production of cytokines, such as IL-6, during allergic inflammation. Here, we examined the role of histamine in IL-6 production and histamine receptor activity in nasal fibroblasts, along with the mechanisms underlying these effects. Experiments were performed using nasal fibroblasts from 8 normal patients. RT-PCR was used to identify the major histamine receptors expressed in nasal fibroblasts. Fibroblasts were then treated with histamine with or without histamine-receptor antagonists, and monitored for IL-6 production using an ELISA. Four potential downstream signaling molecules, p38, extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and NF-κB, were evaluated by Western blot, and a luciferase reporter assay. Elevated expression was seen for all histamine receptors, with IL-6 protein levels increasing significantly following histamine stimulation. Among the histamine-receptor specific antagonists, only the H1R antagonist significantly decreased IL-6 production in histamine-stimulated nasal fibroblasts. Histamine increased the expression level of phosphorylated p38 (pp38), pERK, and pJNK, as well as NF-κB induction. The H1R antagonist actively suppressed pp38 and NF-κB expression in histamine-induced nasal fibroblasts, but not pERK and pJNK. The p38 inhibitor strongly attenuated IL-6 production in histamine-stimulated nasal fibroblasts. The data presented here suggest that antihistamines may be involved in the regulation of cytokines, such as IL-6, due to the role of histamine as an inflammatory mediator in nasal fibroblasts.

A Baculovirus Immediate-Early Gene, ie1, Promoter Drives Efficient Expression of a Transgene in Both Drosophila melanogaster and Bombyx mori

PubMed Central

Masumoto, Mika; Ohde, Takahiro; Shiomi, Kunihiro; Yaginuma, Toshinobu; Niimi, Teruyuki

2012-01-01

Many promoters have been used to drive expression of heterologous transgenes in insects. One major obstacle in the study of non-model insects is the dearth of useful promoters for analysis of gene function. Here, we investigated whether the promoter of the immediate-early gene, ie1, from the Bombyx mori nucleopolyhedrovirus (BmNPV) could be used to drive efficient transgene expression in a wide variety of insects. We used a piggyBac-based vector with a 3xP3-DsRed transformation marker to generate a reporter construct; this construct was used to determine the expression patterns driven by the BmNPV ie1 promoter; we performed a detailed investigation of the promoter in transgene expression pattern in Drosophila melanogaster and in B. mori. Drosophila and Bombyx belong to different insect orders (Diptera and Lepidoptera, respectively); however, and to our surprise, ie1 promoter-driven expression was evident in several tissues (e.g., prothoracic gland, midgut, and tracheole) in both insects. Furthermore, in both species, the ie1 promoter drove expression of the reporter gene from a relatively early embryonic stage, and strong ubiquitous ie1 promoter-driven expression continued throughout the larval, pupal, and adult stages by surface observation. Therefore, we suggest that the ie1 promoter can be used as an efficient expression driver in a diverse range of insect species. PMID:23152896

Strategies to re-express epigenetically silenced p15(INK4b) and p21(WAF1) genes in acute myeloid leukemia.

PubMed

Geyer, C Ronald

2010-01-01

p15(INK4B) and p21(WAF1) are TGF-β targets that are silenced in leukemia by epigenetic mechanisms involving DNA methylation and/or histone modifications. Mechanisms for establishing and maintaining epigenetic silencing of p15(INK4B) and p21(WAF1) are not well established. The reversible nature of epigenetic modifications has lead to the development of drugs that target DNA methyltransferases, histone deacetylases, and histone methyltransferases, which have been used to re-express aberrantly silenced genes in leukemia. Recently, non-coding RNA, referred to as natural antisense transcripts (NATs), have been implicated in the regulation of epigenetic modifications. Here, we review epigenetic mechanisms for silencing p15(INK4B) and p21(WAF1) and the role of NATs in this process. We also review epigenetic drugs and drug combinations used to re-express p15(INK4B) and p21(WAF1). Lastly, we discuss the potential use of NATs to target the activity of epigenetic drugs to specific genes and to permanently re-express epigenetically silenced genes.

Isolation and expression of scabrous, a gene regulating neurogenesis in Drosophila.

PubMed

Mlodzik, M; Baker, N E; Rubin, G M

1990-11-01

Mutations in the Drosophila scabrous (sca) gene affect eye and bristle development, leading to irregular spacing of ommatidia and bristle duplications in the adult fly. We have cloned the sca gene by P-element tagging. The sca transcription unit is 12 kb and consists of four exons that are joined in a 3.2-kb mRNA. In an enhancer trap screen we have isolated several P[lacZ] insertions close to the sca transcription start site. We have examined the expression pattern of sca by in situ hybridization to sca transcripts, by beta-galactosidase localization in the P[lacZ] lines, and by immunocytochemistry with an anti-sca antiserum. During embryogenesis, sca is expressed in a dynamic pattern associated with neural development. During imaginal development, sca is mainly expressed in the R8 photoreceptor precursor cells in the eye imaginal disc and in sensory organ precursor cells in other discs. In the wing disc, sca expression is coextensive with the anlagen for bristles and is controlled by genes of the achaete-scute complex. Based on its loss-of-function phenotype, expression pattern, and the predicted structure of its product, a secreted peptide with homology to the fibrinogen gene family, we propose that sca encodes a signal involved in lateral inhibition within individual domains of the developing nervous system.

RNF38 encodes a nuclear ubiquitin protein ligase that modifies p53

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sheren, Jamie E.; Kassenbrock, C. Kenneth, E-mail: ken.kassenbrock@ucdenver.edu; Department of Biology, Colorado State University, Fort Collins, CO 80523-1878

2013-11-01

Highlights: •RNF38 is shown to be a nuclear protein with a bipartite nuclear localization signal. •RNF38 protein is purified and shown to have ubiquitin protein ligase (E3) activity. •We show that RNF38 binds p53 and can ubiquitinate p53 in vitro. •Overexpression of RNF38 increases p53 ubiquitination in HEK293T cells. •Overexpression of RNF38 in HEK293T cells alters p53 localization. -- Abstract: The RNF38 gene encodes a RING finger protein of unknown function. Here we demonstrate that RNF38 is a functional ubiquitin protein ligase (E3). We show that RNF38 isoform 1 is localized to the nucleus by a bipartite nuclear localization sequencemore » (NLS). We confirm that RNF38 is a binding partner of p53 and demonstrate that RNF38 can ubiquitinate p53 in vitro and in vivo. Finally, we show that overexpression of RNF38 in HEK293T cells results in relocalization of p53 to discrete foci associated with PML nuclear bodies. These results suggest RNF38 is an E3 ubiquitin ligase that may play a role in regulating p53.« less

Variable sexually dimorphic gene expression in laboratory strains of Drosophila melanogaster.

PubMed

Baker, Dean A; Meadows, Lisa A; Wang, Jing; Dow, Julian At; Russell, Steven

2007-12-10

Wild-type laboratory strains of model organisms are typically kept in isolation for many years, with the action of genetic drift and selection on mutational variation causing lineages to diverge with time. Natural populations from which such strains are established, show that gender-specific interactions in particular drive many aspects of sequence level and transcriptional level variation. Here, our goal was to identify genes that display transcriptional variation between laboratory strains of Drosophila melanogaster, and to explore evidence of gender-biased interactions underlying that variability. Transcriptional variation among the laboratory genotypes studied occurs more frequently in males than in females. Qualitative differences are also apparent to suggest that genes within particular functional classes disproportionately display variation in gene expression. Our analysis indicates that genes with reproductive functions are most often divergent between genotypes in both sexes, however a large proportion of female variation can also be attributed to genes without expression in the ovaries. The present study clearly shows that transcriptional variation between common laboratory strains of Drosophila can differ dramatically due to sexual dimorphism. Much of this variation reflects sex-specific challenges associated with divergent physiological trade-offs, morphology and regulatory pathways operating within males and females.

Using FlyBase, a Database of Drosophila Genes & Genomes

PubMed Central

Marygold, Steven J.; Crosby, Madeline A.; Goodman, Joshua L.

2016-01-01

SUMMARY For nearly 25 years, FlyBase (flybase.org) has provided a freely available online database of biological information about Drosophila species, focusing on the model organism D. melanogaster. The need for a centralized, integrated view of Drosophila research has never been greater as advances in genomic, proteomic and high-throughput technologies add to the quantity and diversity of available data and resources. FlyBase has taken several approaches to respond to these changes in the research landscape. Novel report pages have been generated for new reagent types and physical interaction data; Drosophila models of human disease are now represented and showcased in dedicated Human Disease Model Reports; other integrated reports have been established that bring together related genes, datasets or reagents; Gene Reports have been revised to improve access to new data types and to highlight functional data; links to external sites have been organized and expanded; and new tools have been developed to display and interrogate all these data, including improved batch processing and bulk file availability. In addition, several new community initiatives have served to enhance interactions between researchers and FlyBase, resulting in direct user contributions and improved feedback. This chapter provides an overview of the data content, organization and available tools within FlyBase, focusing on recent improvements. We hope it serves as a guide for our diverse user base, enabling efficient and effective exploration of the database and thereby accelerating research discoveries. PMID:27730573

IL-17A causes depression-like symptoms via NFκB and p38MAPK signaling pathways in mice: Implications for psoriasis associated depression.

PubMed

Nadeem, Ahmed; Ahmad, Sheikh F; Al-Harbi, Naif O; Fardan, Ali S; El-Sherbeeny, Ahmed M; Ibrahim, Khalid E; Attia, Sabry M

2017-09-01

Psoriasis has been shown to be associated with an increased prevalence of comorbid major depression. IL-17A plays an important role in both depression and psoriasis. IL-17A has been shown to be elevated in systemic circulation of psoriatic patients. IL-17A released from different immune cells during psoriasis may be responsible for the development of neuropsychiatric symptoms associated with depression. Therefore, this study explored the association of systemic IL-17A with depression. The present study utilized imiquimod model of psoriatic inflammation as well as IL-17A administration in mice to investigate the effect of IL-17A on depression-like behavior. Psoriatic inflammation led to enhanced IL-17A expression in peripheral immune cells of both innate and adaptive origin. This was associated with increased NFκB/p38MAPK signaling and inflammatory mediators in different brain regions, and depression-like symptoms (as reflected by sucrose preference and tail suspension tests). The role of IL-17A was further confirmed by administering it alone for ten days, followed by assessment of the same parameters. IL-17A administration produced effects similar to psoriasis-like inflammation on neurobehavior and NFκB/p38MAPK pathways. Moreover, both NFκB and p38MAPK inhibitors led to attenuation in IL-17A associated with depression-like behavior via reduction in inflammatory mediators, such as MCP-1, iNOS, IL-6, and CXCL-2. Furthermore, anti-IL17A antibody also led to a reduction in imiquimod-induced depression-like symptoms, as well as NFκB/p38MAPK signaling. The present study shows that IL-17A plays an important role in comorbid depression associated with psoriatic inflammation, where both NFκB and p38MAPK pathways play significant roles via upregulation of inflammatory mediators in the brain. Copyright © 2017 Elsevier Ltd. All rights reserved.

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

PubMed

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 < 0.05. Significantly enriched GO terms were clustered using REVIGO and DAVID functional annotation clustering. Significance of overlap between transcript lists was determined with a Fisher's exact hypergeometric test. Treatment of cultured cells and adult flies with lithium and VPA induces transcriptional responses in genes with similar ontology, with as most prominent immune response, 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.

Glutamate-dependent transcriptional regulation in bergmann glia cells: involvement of p38 MAP kinase.

PubMed

Zepeda, Rossana C; Barrera, Iliana; Castelán, Francisco; Soto-Cid, Abraham; Hernández-Kelly, Luisa C; López-Bayghen, Esther; Ortega, Arturo

2008-07-01

Glutamate (Glu) is the major excitatory neurotransmitter in the Central Nervous System (CNS). Ionotropic and metabotropic glutamate receptors (GluRs) are present in neurons and glial cells and are involved in gene expression regulation. Mitogen-activated proteins kinases (MAPK) are critical for all the membrane to nuclei signaling pathways described so far. In cerebellar Bergmann glial cells, glutamate-dependent transcriptional regulation is partially dependent on p42/44 MAPK activity. Another member of this kinase family, p38 MAPK is activated by non-mitogenic stimuli through its Thr180/Tyr182 phosphorylation and phosphorylates cytoplasmic and nuclear protein targets involved in translational and transcriptional events. Taking into consideration that the role of p38MAPK in glial cells is not well understood, we demonstrate here that glutamate increases p38 MAPK phosphorylation in a time and dose dependent manner in cultured chick cerebellar Bergmann glial cells (BGC). Moreover, p38 MAPK is involved in the glutamate-induced transcriptional activation in these cells. Ionotropic as well as metabotropic glutamate receptors participate in p38 MAPK activation. The present findings demonstrate the involvement of p38 MAPK in glutamate-dependent gene expression regulation in glial cells.

Motor neurons in Drosophila flight control: could b1 be the one?

NASA Astrophysics Data System (ADS)

Whitehead, Samuel; Shirangi, Troy; Cohen, Itai

Similar to balancing a stick on one's fingertip, flapping flight is inherently unstable; maintaining stability is a delicate balancing act made possible only by near-constant, often-subtle corrective actions. For fruit flies, such corrective responses need not only be robust, but also fast: the Drosophila flight control reflex has a response latency time of ~5 ms, ranking it among the fastest reflexes in the animal kingdom. How is such rapid, robust control implemented physiologically? Here we present an analysis of a putatively crucial component of the Drosophila flight control circuit: the b1 motor neuron. Specifically, we apply mechanical perturbations to freely-flying Drosophila and analyze the differences in kinematics patterns between flies with manipulated and un-manipulated b1 motor neurons. Ultimately, we hope to identify the functional role of b1 in flight stabilization, with the aim of linking it to previously-proposed, reduced-order models for reflexive control.

The Drosophila gene collection: Identification of putative full-length cDNAs for 70 percent of D. melanogaster genes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Stapleton, Mark; Liao, Guochun; Brokstein, Peter

2002-08-12

Collections of full-length nonredundant cDNA clones are critical reagents for functional genomics. The first step toward these resources is the generation and single-pass sequencing of cDNA libraries that contain a high proportion of full-length clones. The first release of the Drosophila Gene Collection Release 1 (DGCr1) was produced from six libraries representing various tissues, developmental stages, and the cultured S2 cell line. Nearly 80,000 random 5prime expressed sequence tags (EST) from these libraries were collapsed into a nonredundant set of 5849 cDNAs, corresponding to {approx}40 percent of the 13,474 predicted genes in Drosophila. To obtain cDNA clones representing the remainingmore » genes, we have generated an additional 157,835 5prime ESTs from two previously existing and three new libraries. One new library is derived from adult testis, a tissue we previously did not exploit for gene discovery; two new cap-trapped normalized libraries are derived from 0-22hr embryos and adult heads. Taking advantage of the annotated D. melanogaster genome sequence, we clustered the ESTs by aligning them to the genome. Clusters that overlap genes not already represented by cDNA clones in the DGCr1 were analyzed further, and putative full-length clones were selected for inclusion in the new DGC. This second release of the DGC (DGCr2) contains 5061 additional clones, extending the collection to 10,910 cDNAs representing >70 percent of the predicted genes in Drosophila.« less

Temperature-dependent sex-reversal by a transformer-2 gene-edited mutation in the spotted wing drosophila, Drosophila suzukii

USDA-ARS?s Scientific Manuscript database

Female to male sex reversal was achieved in an emerging agricultural insect pest, Drosophila suzukii, by creating a temperature-sensitive point mutation in the sex-determination gene, transformer-2 (tra-2) using CRISPR/Cas9 (clustered regularly interspaced palindromic repeats/ CRISPR-associated) hom...

A chimeric cyclic interferon-α2b peptide induces apoptosis by sequential activation of phosphatidylinositol 3-kinase, protein kinase Cδ and p38 MAP kinase.

PubMed

Blank, V C; Bertucci, L; Furmento, V A; Peña, C; Marino, V J; Roguin, L P

2013-06-10

We have previously demonstrated that tyrosine phosphorylation of STAT1/3 and p38 mitogen-activated protein kinase (p38 MAPK) activation are involved in the apoptotic response triggered by a chimeric cyclic peptide of the interferon-α2b (IFN-α2b) in WISH cells. Since the peptide also induced serine phosphorylation of STAT proteins, in the present study we examined the kinase involved in serine STAT1 phosphorylation and the signaling effectors acting upstream such activation. We first found that p38 MAPK is involved in serine STAT1 phosphorylation, since a reduction of phophoserine-STAT1 levels was evident after incubating WISH cells with cyclic peptide in the presence of a p38 pharmacological inhibitor or a dominant-negative p38 mutant. Next, we demonstrated that the peptide induced activation of protein kinase Cδ (PKCδ). Based on this finding, the role of this kinase was then evaluated. After incubating WISH cells with a PKCδ inhibitor or after decreasing PKCδ expression levels by RNA interference, both peptide-induced serine STAT1 and p38 phosphorylation levels were significantly decreased, indicating that PKCδ functions as an upstream regulator of p38. We also showed that PKCδ and p38 activation stimulated by the peptide was inhibited by a specific pharmacological inhibitor of phosphatidylinositol 3-kinase (PI3K) or by a dominant-negative p85 PI3K-regulatory subunit, suggesting that PI3K is upstream in the signaling cascade. In addition, the role of PI3K and PKCδ in cyclic peptide-induced apoptosis was examined. Both signaling effectors were found to regulate the antiproliferative activity and the apoptotic response triggered by the cyclic peptide in WISH cells. In conclusion, we herein demonstrated that STAT1 serine phosphorylation is mediated by the sequential activation of PI3K, PKCδ and p38 MAPK. This signaling cascade contributes to the antitumor effect induced by the chimeric IFN-α2b cyclic peptide in WISH cells. Copyright © 2013 Elsevier Inc

Genomic cloning and chromosomal localization of HRY, the human homolog to the Drosophila segmentation gene, hairy

DOE Office of Scientific and Technical Information (OSTI.GOV)

Feder, J.N.; Jan, L.Y.; Jan, Y.N.

The Drosophila hairy gene encodes a basic helix- loop-helix protein that functions in at least two steps during Drosophila development: (1) during embryogenesis, when it partakes in the establishment of segments, and (2) during the larval stage, when it functions negatively in determining the pattern of sensory bristles on the adult fly. In the rat, a structurally homologous gene (RHL) behaves as an immediate-early gene in its response to growth factors and can, like that in Drosophila, suppress neuronal differentiation events. Here, the authors report the genomic cloning of the human hairy gene homolog (HRY). The coding region of themore » gene is contained within four exons. The predicted amino acid sequence reveals only four amino acid differences between the human and rat genes. Analysis of the DNA sequence 5[prime] to the coding region reveals a putatitve untranslated exon. To increase the value of the HRY gene as a genetic marker and to assess its potential involvement in genetic disorders, they sublocalized the locus to chromosome 3q28-q29 by fluorescence in situ hybridization. 34 refs., 4 figs., 1 tab.« less

The Drosophila melanogaster Muc68E Mucin Gene Influences Adult Size, Starvation Tolerance, and Cold Recovery.

PubMed

Reis, Micael; Silva, Ana C; Vieira, Cristina P; Vieira, Jorge

2016-07-07

Mucins have been implicated in many different biological processes, such as protection from mechanical damage, microorganisms, and toxic molecules, as well as providing a luminal scaffold during development. Nevertheless, it is conceivable that mucins have the potential to modulate food absorption as well, and thus contribute to the definition of several important phenotypic traits. Here we show that the Drosophila melanogaster Muc68E gene is 40- to 60-million-yr old, and is present in Drosophila species of the subgenus Sophophora only. The central repeat region of this gene is fast evolving, and shows evidence for repeated expansions/contractions. This and/or frequent gene conversion events lead to the homogenization of its repeats. The amino acid pattern P[ED][ED][ST][ST][ST] is found in the repeat region of Muc68E proteins from all Drosophila species studied, and can occur multiple times within a single conserved repeat block, and thus may have functional significance. Muc68E is a nonessential gene under laboratory conditions, but Muc68E mutant flies are smaller and lighter than controls at birth. However, at 4 d of age, Muc68E mutants are heavier, recover faster from chill-coma, and are more resistant to starvation than control flies, although they have the same percentage of lipids as controls. Mutant flies have enlarged abdominal size 1 d after chill-coma recovery, which is associated with higher lipid content. These results suggest that Muc68E has a role in metabolism modulation, food absorption, and/or feeding patterns in larvae and adults, and under normal and stress conditions. Such biological function is novel for mucin genes. Copyright © 2016 Reis et al.

Dynamic Repositioning of Dorsal to Two Different κB Motifs Controls Its Autoregulation during Immune Response in Drosophila

PubMed Central

Mrinal, Nirotpal; Nagaraju, Javaregowda

2010-01-01

Autoregulation is one of the mechanisms of imparting feedback control on gene expression. Positive autoregulatory feedback results in induction of a gene, and negative feedback leads to its suppression. Here, we report an interesting mechanism of autoregulation operating on Drosophila Rel gene dorsal that can activate as well as repress its expression. Using biochemical and genetic approaches, we show that upon immune challenge Dorsal regulates its activation as well as repression by dynamically binding to two different κB motifs, κBI (intronic κB) and κBP (promoter κB), present in the dorsal gene. Although the κBI motif functions as an enhancer, the κBP motif acts as a transcriptional repressor. Interestingly, Dorsal binding to these two motifs is dynamic; immediately upon immune challenge, Dorsal binds to the κBI leading to auto-activation, whereas at the terminal phase of the immune response, it is removed from the κBI and repositioned at the κBP, resulting in its repression. Furthermore, we show that repression of Dorsal as well as its binding to the κBP depends on the transcription factor AP1. Depletion of AP1 by RNA interference resulted in constitutive expression of Dorsal. In conclusion, this study suggests that during acute phase response dorsal is regulated by following two subcircuits: (i) Dl-κBI for activation and (ii) Dl-AP1-κBP for repression. These two subcircuits are temporally delineated and bring about overall regulation of dorsal during immune response. These results suggest the presence of a previously unknown mechanism of Dorsal autoregulation in immune-challenged Drosophila. PMID:20504768

Conditional inhibition of autophagy genes in adult Drosophila impairs immunity without compromising longevity.

PubMed

Ren, Chunli; Finkel, Steven E; Tower, John

2009-03-01

Immune function declines with age in Drosophila and humans, and autophagy is implicated in immune function. In addition, autophagy genes are required for life span extension caused by reduced insulin/IGF1-like signaling and dietary restriction in Caenorhabditiselegans. To test if the autophagy pathway might be limiting for immunity and/or life span in adult Drosophila, the Geneswitch system was used to cause conditional inactivation of the autophagy genes Atg5, Atg7 and Atg12 by RNAi. Conditional inhibition of Atg genes in adult flies reduced lysotracker staining of adult tissues, and reduced resistance to injected Escherichia coli, as evidenced by increased bacterial titers and reduced fly survival. However, survival of uninjected flies was unaffected by Atg gene inactivation. The data indicate that Atg gene activity is required for normal immune function in adult flies, and suggest that neither autophagy nor immune function are limiting for adult life span under typical laboratory conditions.

Decreased microRNA-125a-3p contributes to upregulation of p38 MAPK in rat trigeminal ganglions with orofacial inflammatory pain.

PubMed

Dong, Yingchun; Li, Pengfei; Ni, Yanhong; Zhao, Junjie; Liu, Zhiqiang

2014-01-01

Orofacial inflammatory pain is a difficult clinical problem, and the specific molecular mechanisms for this pain remain largely unexplained. The present study aimed to determine the differential expression of microRNAs (miRNAs) and disclose the underlying role of miR-125a-3p in orofacial inflammatory pain induced by complete Freund's adjuvant (CFA). Thirty-two differentially expressed miRNAs were first screened using a microarray chip in ipsilateral trigeminal ganglions (TGs) following CFA injection into the orofacial skin innervated by trigeminal nerve, and a portion of them, including miR-23a*, -24-2*, -26a, -92a, -125a-3p, -183 and -299 were subsequently selected and validated by qPCR. The target genes were predicted based on the miRWalk website and were further analyzed by gene ontology (GO). Further studies revealed miR-125a-3p expression was down-regulated, whereas both the expression of p38 MAPK (mitogen-activated protein kinase) alpha and CGRP (calcitonin gene-related peptide) were up-regulated in ipsilateral TGs at different time points after CFA injection compared with control. Furthermore, mechanistic study revealed that miR-125a-3p negatively regulates p38 alpha gene expression and is positively correlated with the head withdrawal threshold reflecting pain. Luciferase assay showed that binding of miR-125a-3p to the 3'UTR of p38 alpha gene suppressed the transcriptional activity, and overexpression of miR-125a-3p significantly inhibited the p38 alpha mRNA level in ND8/34 cells. Taken together, our results show that miR-125a-3p is negatively correlated with the development and maintenance of orofacial inflammatory pain via regulating p38 MAPK.

Flamenco, a gene controlling the gypsy retrovirus of drosophila melanogaster

DOE Office of Scientific and Technical Information (OSTI.GOV)

Prud`homme, N.; Gans, M.; Masson, M.

1995-02-01

Gypsy is an endogenous retrovirus of Drosophila melanogaster. It is table and does not transpose with detectable frequencies in most Drosophila strains. However, we have characterized unstable strains, known as MG, in which it transposes at high frequency. These stocks contain more copies of gypsy than usual stocks. Transposition results in mutations in several genes such as ovo and cut. They are stable and are due to gypsy insertions. Integrations into the ovo{sup D1} female sterile-dominant mutation result in a null allele of the gene and occurrence of fertile females. This phenomenon, known as the ovo{sup D1} reversion assay, canmore » be used to quantitate gypsy activity. We have shown that the properties of MG strains result from mutation of a host gene that we called flamenco (flam). It has a strict maternal effect on gypsy mobilization: transposition occurs at high frequency only in the germ line of the progeny of females homozygous for mutations of the gene. It is located at position 65.9 (20A1-3) on the X chromosome. The mutant allele present in MG strains is essentially recessive. Flamenco seems to control the infective properties of gypsy. 40 refs., 10 figs., 6 tabs.« less

The evolution of courtship behaviors through the origination of a new gene in Drosophila

PubMed Central

Dai, Hongzheng; Chen, Ying; Chen, Sidi; Mao, Qiyan; Kennedy, David; Landback, Patrick; Eyre-Walker, Adam; Du, Wei; Long, Manyuan

2008-01-01

New genes can originate by the combination of sequences from unrelated genes or their duplicates to form a chimeric structure. These chimeric genes often evolve rapidly, suggesting that they undergo adaptive evolution and may therefore be involved in novel phenotypes. Their functions, however, are rarely known. Here, we describe the phenotypic effects of a chimeric gene, sphinx, that has recently evolved in Drosophila melanogaster. We show that a knockout of this gene leads to increased male–male courtship in D. melanogaster, although it leaves other aspects of mating behavior unchanged. Comparative studies of courtship behavior in other closely related Drosophila species suggest that this mutant phenotype of male–male courtship is the ancestral condition because these related species show much higher levels of male–male courtship than D. melanogaster. D. melanogaster therefore seems to have evolved in its courtship behaviors by the recruitment of a new chimeric gene. PMID:18508971

Methods to identify and analyze gene products involved in neuronal intracellular transport using Drosophila

PubMed Central

Neisch, Amanda L.; Avery, Adam W.; Machame, James B.; Li, Min-gang; Hays, Thomas S.

2017-01-01

Proper neuronal function critically depends on efficient intracellular transport and disruption of transport leads to neurodegeneration. Molecular pathways that support or regulate neuronal transport are not fully understood. A greater understanding of these pathways will help reveal the pathological mechanisms underlying disease. Drosophila melanogaster is the premier model system for performing large-scale genetic functional screens. Here we describe methods to carry out primary and secondary genetic screens in Drosophila aimed at identifying novel gene products and pathways that impact neuronal intracellular transport. These screens are performed using whole animal or live cell imaging of intact neural tissue to ensure integrity of neurons and their cellular environment. The primary screen is used to identify gross defects in neuronal function indicative of a disruption in microtubule-based transport. The secondary screens, conducted in both motoneurons and dendritic arborization neurons, will confirm the function of candidate gene products in intracellular transport. Together, the methodologies described here will support labs interested in identifying and characterizing gene products that alter intracellular transport in Drosophila. PMID:26794520

A gain-of-function screen for genes that influence axon guidance identifies the NF-kappaB protein dorsal and reveals a requirement for the kinase Pelle in Drosophila photoreceptor axon targeting.

PubMed

Mindorff, Elizabeth N; O'Keefe, David D; Labbé, Alain; Yang, Jennie Ping; Ou, Yimiao; Yoshikawa, Shingo; van Meyel, Donald J

2007-08-01

To identify novel regulators of nervous system development, we used the GAL4-UAS misexpression system in Drosophila to screen for genes that influence axon guidance in developing embryos. We mobilized the Gene Search (GS) P element and identified 42 lines with insertions in unique loci, including leak/roundabout2, which encodes an axon guidance receptor and confirms the utility of our screen. The genes we identified encode proteins of diverse classes, some acting near the cell surface and others in the cytoplasm or nucleus. We found that one GS line drove misexpression of the NF-kappaB transcription factor Dorsal, causing motor axons to bypass their correct termination sites. In the developing visual system, Dorsal misexpression also caused photoreceptor axons to reach incorrect positions within the optic lobe. This mistargeting occurred without observable changes of cell fate and correlated with localization of ectopic Dorsal in distal axons. We found that Dorsal and its inhibitor Cactus are expressed in photoreceptors, though neither was required for axon targeting. However, mutation analyses of genes known to act upstream of Dorsal revealed a requirement for the interleukin receptor-associated kinase family kinase Pelle for layer-specific targeting of photoreceptor axons, validating our screen as a means to identify new molecular determinants of nervous system development in vivo.

Sex-specific control of CNS autoimmunity by p38 MAPK signaling in myeloid cells

PubMed Central

Krementsov, Dimitry N.; Noubade, Rajkumar; Dragon, Julie A.; Otsu, Kinya; Rincon, Mercedes; Teuscher, Cory

2013-01-01

Objective Multiple sclerosis (MS) is a chronic inflammatory demyelinating disease of the central nervous system (CNS), characterized by a global increasing incidence driven by relapsing-remitting disease in females. p38 MAP kinase (MAPK) has been described as a key regulator of inflammatory responses in autoimmunity, but its role in the sexual dimorphism in MS or MS models remains unexplored. Methods Toward this end, we used experimental autoimmune encephalomyelitis (EAE), the principal animal model of MS, combined with pharmacologic and genetic inhibition of p38 MAPK activity and transcriptomic analyses. Results Pharmacologic inhibition of p38 MAPK selectively ameliorated EAE in female mice. Conditional deletion studies demonstrated that p38α signaling in macrophages/myeloid cells, but not T cells or dendritic cells, recapitulated this sexual dimorphism. Analysis of CNS inflammatory infiltrates showed that female, but not male mice lacking p38α in myeloid cells exhibited reduced immune cell activation compared with controls, while peripheral T cell priming was unaffected in both sexes. Transcriptomic analyses of myeloid cells revealed differences in p38α-controlled transcripts comprising female- and male-specific gene modules, with greater p38α dependence of pro-inflammatory gene expression in females. Interpretation Our findings demonstrate a key role for p38α in myeloid cells in CNS autoimmunity and uncover important molecular mechanisms underlying sex differences in disease pathogenesis. Taken together, our results suggest that the p38 MAPK signaling pathway represents a novel target for much needed disease modifying therapies for MS. PMID:24027119

Characterization of the intergenic RNA profile at abdominal-A and Abdominal-B in the Drosophila bithorax complex

PubMed Central

Bae, Esther; Calhoun, Vincent C.; Levine, Michael; Lewis, Edward B.; Drewell, Robert A.

2002-01-01

The correct spatial expression of two Drosophila bithorax complex (BX-C) genes, abdominal-A (abdA) and Abdominal-B (AbdB), is dependent on the 100-kb intergenic infraabdominal (iab) region. The iab region is known to contain a number of different domains (iab2 through iab8) that harbor cis-regulatory elements responsible for directing expression of abdA and AbdB in the second through eighth abdominal segments. Here, we use in situ hybridization to perform high-resolution mapping of the transcriptional activity in the iab control regions. We show that transcription of the control regions themselves is abundant and precedes activation of the abdA and AbdB genes. As with the homeotic genes of the BX-C, the transcription patterns of the RNAs from the iab control regions demonstrate colinearity with the sequence of the iab regions along the chromosome and the domains in the embryo under the control of the specific iab regions. These observations suggest that the intergenic RNAs may play a role in initiating cis regulation at the BX-C early in development. PMID:12481037

The gene for PAX7, a member of the paired-box-containing genes, is localized on human chromosome arm 1p36.

PubMed

Shapiro, D N; Sublett, J E; Li, B; Valentine, M B; Morris, S W; Noll, M

1993-09-01

The murine Pax-7 gene and the cognate human gene, formerly designated HuP1, are members of the multigene paired-box-containing class of developmental regulatory genes first identified in Drosophila. By analysis of somatic cell hybrids segregating human chromosomes, the gene encoding PAX7 was localized to human chromosome 1. Fluorescence in situ hybridization confirmed this assignment and allowed mapping of the gene to the terminal region of the short arm (1p36) of the chromosome. Additionally, these results confirm the extensive homology between human chromosome 1p and the distal segment of mouse chromosome 4, extending from bands C5 through E2.

p38 MAPK inhibition suppresses the TLR-hypersensitive phenotype in FANCC- and FANCA-deficient mononuclear phagocytes

PubMed Central

Anur, Praveen; Yates, Jane; Garbati, Michael R.; Vanderwerf, Scott; Keeble, Winifred; Rathbun, Keaney; Hays, Laura E.; Tyner, Jeffrey W.; Svahn, Johanna; Cappelli, Enrico; Dufour, Carlo

2012-01-01

Fanconi anemia, complementation group C (FANCC)–deficient hematopoietic stem and progenitor cells are hypersensitive to a variety of inhibitory cytokines, one of which, TNFα, can induce BM failure and clonal evolution in Fancc-deficient mice. FANCC-deficient macrophages are also hypersensitive to TLR activation and produce TNFα in an unrestrained fashion. Reasoning that suppression of inhibitory cytokine production might enhance hematopoiesis, we screened small molecules using TLR agonist–stimulated FANCC- and Fanconi anemia, complementation group A (FANCA)–deficient macrophages containing an NF-κB/AP-1–responsive reporter gene (SEAP). Of the 75 small molecules screened, the p38 MAPK inhibitor BIRB 796 and dasatinib potently suppressed TLR8-dependent expression of the reporter gene. Fanconi anemia (FA) macrophages were hypersensitive to the TLR7/8 activator R848, overproducing SEAP and TNFα in response to all doses of the agonist. Low doses (50nM) of both agents inhibited p38 MAPK–dependent activation of MAPKAPK2 (MK2) and suppressed MK2-dependent TNFα production without substantially influencing TNFα gene transcription. Overproduction of TNFα by primary FA cells was likewise suppressed by these agents and involved inhibition of MK2 activation. Because MK2 is also known to influence production and/or sensitivity to 2 other suppressive factors (MIP-1α and IFNγ) to which FA hematopoietic progenitor cells are uniquely vulnerable, targeting of p38 MAPK in FA hematopoietic cells is a rational objective for preclinical evaluation. PMID:22234699

p38 MAPK inhibition suppresses the TLR-hypersensitive phenotype in FANCC- and FANCA-deficient mononuclear phagocytes.

PubMed

Anur, Praveen; Yates, Jane; Garbati, Michael R; Vanderwerf, Scott; Keeble, Winifred; Rathbun, Keaney; Hays, Laura E; Tyner, Jeffrey W; Svahn, Johanna; Cappelli, Enrico; Dufour, Carlo; Bagby, Grover C

2012-03-01

Fanconi anemia, complementation group C (FANCC)-deficient hematopoietic stem and progenitor cells are hypersensitive to a variety of inhibitory cytokines, one of which, TNFα, can induce BM failure and clonal evolution in Fancc-deficient mice. FANCC-deficient macrophages are also hypersensitive to TLR activation and produce TNFα in an unrestrained fashion. Reasoning that suppression of inhibitory cytokine production might enhance hematopoiesis, we screened small molecules using TLR agonist-stimulated FANCC- and Fanconi anemia, complementation group A (FANCA)-deficient macrophages containing an NF-κB/AP-1-responsive reporter gene (SEAP). Of the 75 small molecules screened, the p38 MAPK inhibitor BIRB 796 and dasatinib potently suppressed TLR8-dependent expression of the reporter gene. Fanconi anemia (FA) macrophages were hypersensitive to the TLR7/8 activator R848, overproducing SEAP and TNFα in response to all doses of the agonist. Low doses (50nM) of both agents inhibited p38 MAPK-dependent activation of MAPKAPK2 (MK2) and suppressed MK2-dependent TNFα production without substantially influencing TNFα gene transcription. Overproduction of TNFα by primary FA cells was likewise suppressed by these agents and involved inhibition of MK2 activation. Because MK2 is also known to influence production and/or sensitivity to 2 other suppressive factors (MIP-1α and IFNγ) to which FA hematopoietic progenitor cells are uniquely vulnerable, targeting of p38 MAPK in FA hematopoietic cells is a rational objective for preclinical evaluation.

STAT, Wingless, and Nurf-38 determine the accuracy of regeneration after radiation damage in Drosophila.

PubMed

Verghese, Shilpi; Su, Tin Tin

2017-10-01

We report here a study of regeneration in Drosophila larval wing imaginal discs after damage by ionizing radiation. We detected faithful regeneration that restored a wing disc and abnormal regeneration that produced an extra wing disc. We describe a sequence of changes in cell number, location and fate that occur to produce an ectopic disc. We identified a group of cells that not only participate in ectopic disc formation but also recruit others to do so. STAT92E (Drosophila STAT3/5) and Nurf-38, which encodes a member of the Nucleosome Remodeling Factor complex, oppose each other in these cells to modulate the frequency of ectopic disc growth. The picture that emerges is one in which activities like STAT increase after radiation damage and fulfill essential roles in rebuilding the tissue. But such activities must be kept in check so that one and only one wing disc is regenerated.

Two rapidly evolving genes contribute to male fitness in Drosophila

PubMed Central

Reinhardt, Josephine A; Jones, Corbin D

2013-01-01

Purifying selection often results in conservation of gene sequence and function. The most functionally conserved genes are also thought to be among the most biologically essential. These observations have led to the use of sequence conservation as a proxy for functional conservation. Here we describe two genes that are exceptions to this pattern. We show that lack of sequence conservation among orthologs of CG15460 and CG15323 – herein named jean-baptiste (jb) and karr respectively – does not necessarily predict lack of functional conservation. These two Drosophila melanogaster genes are among the most rapidly evolving protein-coding genes in this species, being nearly as diverged from their D. yakuba orthologs as random sequences are. jb and karr are both expressed at an elevated level in larval males and adult testes, but they are not accessory gland proteins and their loss does not affect male fertility. Instead, knockdown of these genes in D. melanogaster via RNA interference caused male-biased viability defects. These viability effects occur prior to the third instar for jb and during late pupation for karr. We show that putative orthologs to jb and karr are also expressed strongly in the testes of other Drosophila species and have similar gene structure across species despite low levels of sequence conservation. While standard molecular evolution tests could not reject neutrality, other data hint at a role for natural selection. Together these data provide a clear case where a lack of sequence conservation does not imply a lack of conservation of expression or function. PMID:24221639

Genes for Drosophila small heat shock proteins are regulated differently by ecdysterone

DOE Office of Scientific and Technical Information (OSTI.GOV)

Amin, J.; Voellmy, R.; Mestril, R.

Genes for small heat shock proteins (hsp27 to hsp22) are activated in late third-instar larvae of Drosophila melanogaster in the absence of heat stress. This regulation has been stimulated in cultured Drosophila cells in which the genes are activated by the addition of ecdysterone. Sequence elements (HERE) involved in ecdysterone regulation of the hsp27 and hsp23 genes have been defined by transfection studies and have recently been identified as binding sites for ecdysterone receptor. The authors report here that the shp27 and hsp23 genes are regulated differently by ecdysterone. The hsp27 gene is activated rapidly by ecdysterone, even in themore » absence of protein synthesis. In contrast, high-level expression of the hsp23 gene begins only after a lag of about 6 h, is dependent on the continuous presence of ecdysterone, and is sensitive to low concentrations of protein synthesis inhibitors. Transfection experiments with reported constructs show that this difference in regulation is at the transcriptional level. Synthetic hsp27 or hsp23 HERE sequences confer hsp27- or hsp23-type ecdysterone regulation on a basal promoter. These findings indicate that the hsp27 gene is primary, and the hsp23 gene is mainly a secondary, hormone-responsive gene. Ecdysterone receptor is implied to play a role in the regulation of both genes.« less

The Foldback-like element Galileo belongs to the P superfamily of DNA transposons and is widespread within the Drosophila genus.

PubMed

Marzo, Mar; Puig, Marta; Ruiz, Alfredo

2008-02-26

Galileo is the only transposable element (TE) known to have generated natural chromosomal inversions in the genus Drosophila. It was discovered in Drosophila buzzatii and classified as a Foldback-like element because of its long, internally repetitive, terminal inverted repeats (TIRs) and lack of coding capacity. Here, we characterized a seemingly complete copy of Galileo from the D. buzzatii genome. It is 5,406 bp long, possesses 1,229-bp TIRs, and encodes a 912-aa transposase similar to those of the Drosophila melanogaster 1360 (Hoppel) and P elements. We also searched the recently available genome sequences of 12 Drosophila species for elements similar to Dbuz\\Galileo by using bioinformatic tools. Galileo was found in six species (ananassae, willistoni, peudoobscura, persimilis, virilis, and mojavensis) from the two main lineages within the Drosophila genus. Our observations place Galileo within the P superfamily of cut-and-paste transposons and extend considerably its phylogenetic distribution. The interspecific distribution of Galileo indicates an ancient presence in the genus, but the phylogenetic tree built with the transposase amino acid sequences contrasts significantly with that of the species, indicating lineage sorting and/or horizontal transfer events. Our results also suggest that Foldback-like elements such as Galileo may evolve from DNA-based transposon ancestors by loss of the transposase gene and disproportionate elongation of TIRs.

The Foldback-like element Galileo belongs to the P superfamily of DNA transposons and is widespread within the Drosophila genus

PubMed Central

Marzo, Mar; Puig, Marta; Ruiz, Alfredo

2008-01-01

Galileo is the only transposable element (TE) known to have generated natural chromosomal inversions in the genus Drosophila. It was discovered in Drosophila buzzatii and classified as a Foldback-like element because of its long, internally repetitive, terminal inverted repeats (TIRs) and lack of coding capacity. Here, we characterized a seemingly complete copy of Galileo from the D. buzzatii genome. It is 5,406 bp long, possesses 1,229-bp TIRs, and encodes a 912-aa transposase similar to those of the Drosophila melanogaster 1360 (Hoppel) and P elements. We also searched the recently available genome sequences of 12 Drosophila species for elements similar to Dbuz\\Galileo by using bioinformatic tools. Galileo was found in six species (ananassae, willistoni, peudoobscura, persimilis, virilis, and mojavensis) from the two main lineages within the Drosophila genus. Our observations place Galileo within the P superfamily of cut-and-paste transposons and extend considerably its phylogenetic distribution. The interspecific distribution of Galileo indicates an ancient presence in the genus, but the phylogenetic tree built with the transposase amino acid sequences contrasts significantly with that of the species, indicating lineage sorting and/or horizontal transfer events. Our results also suggest that Foldback-like elements such as Galileo may evolve from DNA-based transposon ancestors by loss of the transposase gene and disproportionate elongation of TIRs. PMID:18287066

Characterization of new regulatory elements within the Drosophila bithorax complex.

PubMed

Pérez-Lluch, Sílvia; Cuartero, Sergi; Azorín, Fernando; Espinàs, M Lluïsa

2008-12-01

The homeotic Abdominal-B (Abd-B) gene expression depends on a modular cis-regulatory region divided into discrete functional domains (iab) that control the expression of the gene in a particular segment of the fly. These domains contain regulatory elements implicated in both initiation and maintenance of homeotic gene expression and elements that separate the different domains. In this paper we have performed an extensive analysis of the iab-6 regulatory region, which regulates Abd-B expression at abdominal segment A6 (PS11), and we have characterized two new polycomb response elements (PREs) within this domain. We report that PREs at Abd-B cis-regulatory domains present a particular chromatin structure which is nuclease accessible all along Drosophila development and both in active and repressed states. We also show that one of these regions contains a dCTCF and CP190 dependent activity in transgenic enhancer-blocking assays, suggesting that it corresponds to the Fab-6 boundary element of the Drosophila bithorax complex.

Repression of the interleukin 6 gene promoter by p53 and the retinoblastoma susceptibility gene product

DOE Office of Scientific and Technical Information (OSTI.GOV)

Santhanam, U.; Ray, A.; Sehgal, P.B.

1991-09-01

The aberrant overexpression of interleukin 6 (IL-6) is implicated as an autocrine mechanism in the enhanced proliferation of the neoplastic cell elements in various B- and T-cell malignancies and in some carcinomas and sarcomas; many of these neoplasms have been shown to be associated with a mutated p53 gene. The possibility that wild-type (wt) p53, a nuclear tumor-suppressor protein, but not its transforming mutants might serve to repress IL-6 gene expression was investigated in HeLa cells. The authors transiently cotransfected these cells with constitutive cytomegalovirus (CMV) enhancer/promoter expression plasmids overproducing wt or mutant human or murine p53 and with appropriatemore » chloramphenicol acetyltransferase (CAT) reporter plasmids containing the promoter elements of human IL-6, c-fos, or {beta}-actin genes or of porcine major histocompatibility complex (MHC) class I gene in pN-38 to evaluate the effect of the various p53 species on these promoters. These observations identify transcriptional repression as a property of p53 and suggest that p53 and RB may be involved as transcriptional repressors in modulating IL-6 gene expression during cellular differentiation and oncogenesis.« less

P element excision in drosophila melanogaster and related drosophilids

USDA-ARS?s Scientific Manuscript database

The frequency of P element excision and the structure of the resulting excision products were determined in three drosophilid species, Drosophila melanogaster, D. virilis, and Chymomyza procnemis. A transient P element mobility assay was conducted in the cells of developing insect embryos, but unlik...

[Expression of the Drosophila melanogaster limk1 gene 3'-UTRs mRNA in Yeast Saccharomyces cerevisiae].

PubMed

Rumyantsev, A M; Zakharov, G A; Zhuravlev, A V; Padkina, M V; Savvateeva-Popova, E V; Sambuk, E V

2014-06-01

The stability of mRNA and its translation efficacy in higher eukaryotes are influenced by the interaction of 3'-untranscribed regions (3'-UTRs) with microRNAs and RNA-binding proteins. Since Saccharomyces cerevisiae lack microRNAs, it is possible to evaluate the contribution of only 3'-UTRs' and RNA-binding proteins' interaction in post-transcriptional regulation. For this, the post-transcriptional regulation of Drosophila limk1 gene encoding for the key enzyme of actin remodeling was studied in yeast. Analysis of limkl mRNA 3'-UTRs revealed the potential sites of yeast transcriptional termination. Computer remodeling demonstrated the possibility of secondary structure formation in limkl mRNA 3'-UTRs. For an evaluation of the functional activity of Drosophila 3'-UTRs in yeast, the reporter gene PHO5 encoding for yeast acid phosphatase (AP) fused to different variants of Drosophila limk1 mRNA 3'-UTRs (513, 1075, 1554 bp) was used. Assessments of AP activity and RT-PCR demonstrated that Drosophila limkl gene 3'-UTRs were functionally active and recognized in yeast. Therefore, yeast might be used as an appropriate model system for studies of 3'-UTR's role in post-transcriptional regulation.

The three-dimensional structure of MAP kinase p38[beta]: different features of the ATP-binding site in p38[beta] compared with p38[alpha

DOE Office of Scientific and Technical Information (OSTI.GOV)

Patel, Sangita B.; Cameron, Patricia M.; O'Keefe, Stephen J.

2010-10-18

The p38 mitogen-activated protein kinases are activated in response to environmental stress and cytokines and play a significant role in transcriptional regulation and inflammatory responses. Of the four p38 isoforms known to date, two (p38{alpha} and p38{beta}) have been identified as targets for cytokine-suppressive anti-inflammatory drugs. Recently, it was reported that specific inhibition of the p38{alpha} isoform is necessary and sufficient for anti-inflammatory efficacy in vivo, while further inhibition of p38{beta} may not provide any additional benefit. In order to aid the development of p38{alpha}-selective compounds, the three-dimensional structure of p38{beta} was determined. To do so, the C162S and C119S,C162Smore » mutants of human MAP kinase p38{beta} were cloned, expressed in Escherichia coli and purified. Initial screening hits in crystallization trials in the presence of an inhibitor led upon optimization to crystals that diffracted to 2.05 {angstrom} resolution and allowed structure determination (PDB codes 3gc8 and 3gc9 for the single and double mutant, respectively). The structure of the p38{alpha} C162S mutant in complex with the same inhibitor is also reported (PDB code 3gc7). A comparison between the structures of the two kinases showed that they are highly similar overall but that there are differences in the relative orientation of the N- and C-terminal domains that causes a reduction in the size of the ATP-binding pocket in p38{beta}. This difference in size between the two pockets could be exploited in order to achieve selectivity.« less

Hybrid sterility and evolution in Hawaiian Drosophila: differential gene and allele-specific expression analysis of backcross males.

PubMed

Brill, E; Kang, L; Michalak, K; Michalak, P; Price, D K

2016-08-01

The Hawaiian Drosophila are an iconic example of sequential colonization, adaptive radiation and speciation on islands. Genetic and phenotypic analysis of closely related species pairs that exhibit incomplete reproductive isolation can provide insights into the mechanisms of speciation. Drosophila silvestris from Hawai'i Island and Drosophila planitibia from Maui are two closely related allopatric Hawaiian picture-winged Drosophila that produce sterile F1 males but fertile F1 females, a pattern consistent with Haldane's rule. Backcrossing F1 hybrid females between these two species to parental species gives rise to recombinant males with three distinct sperm phenotypes despite a similar genomic background: motile sperm, no sperm (sterile), and immotile sperm. We found that these three reproductive morphologies of backcross hybrid males produce divergent gene expression profiles in testes, as measured with RNA sequencing. There were a total of 71 genes significantly differentially expressed between backcross males with no sperm compared with those backcross males with motile sperm and immotile sperm, but no significant differential gene expression between backcross males with motile sperm and backcross males with immotile sperm. All of these genes were underexpressed in males with no sperm, including a number of genes with previously known activities in adult testis. An allele-specific expression analysis showed overwhelmingly more cis-divergent than trans-divergent genes, with no significant difference in the ratio of cis- and trans-divergent genes among the sperm phenotypes. Overall, the results indicate that the regulation of gene expression involved in sperm production likely diverged relatively rapidly between these two closely related species.

Molecular Population Genetics of Sex Determination Genes: The Transformer Gene of Drosophila Melanogaster

PubMed Central

Walthour, C. S.; Schaeffer, S. W.

1994-01-01

The transformer locus (tra) produces an RNA processing protein that alternatively splices the doublesex pre-mRNA in the sex determination hierarchy of Drosophila melanogaster. Comparisons of the tra coding region among Drosophila species have revealed an unusually high degree of divergence in synonymous and nonsynonymous sites. In this study, we tested the hypothesis that the tra gene will be polymorphic in synonymous and nonsynonymous sites within species by investigating nucleotide sequence variation in eleven tra alleles within D. melanogaster. Of the 1063 nucleotides examined, two synonymous sites were polymorphic and no amino acid variation was detected. Three statistical tests were used to detect departures from an equilibrium neutral model. Two tests failed to reject a neutral model of molecular evolution because of low statisitical power associated with low levels of genetic variation (Tajima/Fu and Li). The Hudson, Kreitman, and Aguade test rejected a neutral model when the tra region was compared to the 5'-flanking region of alcohol dehydrogenase (Adh). The lack of variability in the tra gene is consistent with a recent selective sweep of a beneficial allele in or near the tra locus. PMID:8013913

Cloning and characterization of the Drosophila homolog of the xeroderma pigmentosum complementation-group B correcting gene, ERCC3.

PubMed Central

Koken, M H; Vreeken, C; Bol, S A; Cheng, N C; Jaspers-Dekker, I; Hoeijmakers, J H; Eeken, J C; Weeda, G; Pastink, A

1992-01-01

Previously the human nucleotide excision repair gene ERCC3 was shown to be responsible for a rare combination of the autosomal recessive DNA repair disorders xeroderma pigmentosum (complementation group B) and Cockayne's syndrome (complementation group C). The human and mouse ERCC3 proteins contain several sequence motifs suggesting that it is a nucleic acid or chromatin binding helicase. To study the significance of these domains and the overall evolutionary conservation of the gene, the homolog from Drosophila melanogaster was isolated by low stringency hybridizations using two flanking probes of the human ERCC3 cDNA. The flanking probe strategy selects for long stretches of nucleotide sequence homology, and avoids isolation of small regions with fortuitous homology. In situ hybridization localized the gene onto chromosome III 67E3/4, a region devoid of known D.melanogaster mutagen sensitive mutants. Northern blot analysis showed that the gene is continuously expressed in all stages of fly development. A slight increase (2-3 times) of ERCC3Dm transcript was observed in the later stages. Two almost full length cDNAs were isolated, which have different 5' untranslated regions (UTR). The SD4 cDNA harbours only one long open reading frame (ORF) coding for ERCC3Dm. Another clone (SD2), however, has the potential to encode two proteins: a 170 amino acids polypeptide starting at the optimal first ATG has no detectable homology with any other proteins currently in the data bases, and another ORF beginning at the suboptimal second startcodon which is identical to that of SD4. Comparison of the encoded ERCC3Dm protein with the homologous proteins of mouse and man shows a strong amino acid conservation (71% identity), especially in the postulated DNA binding region and seven 'helicase' domains. The ERCC3Dm sequence is fully consistent with the presumed functions and the high conservation of these regions strengthens their functional significance. Microinjection and DNA

Regulation of chromatin organization and inducible gene expression by a Drosophila insulator

PubMed Central

Wood, Ashley M.; Van Bortle, Kevin; Ramos, Edward; Takenaka, Naomi; Rohrbaugh, Margaret; Jones, Brian C.; Jones, Keith C.; Corces, Victor G.

2011-01-01

SUMMARY Insulators are multi-protein-DNA complexes thought to affect gene expression by mediating inter- and intra-chromosomal interactions. Drosophila insulators contain specific DNA binding proteins plus common components, such as CP190, that facilitate these interactions. Here we examine changes in the distribution of Drosophila insulator proteins during the heat-shock and ecdysone responses. We find that CP190 recruitment to insulator sites is the main regulatable step in controlling insulator function during heat shock. In contrast, both CP190 and DNA binding protein recruitment are regulated during the ecdysone response. CP190 is necessary to stabilize specific chromatin loops and for proper activation of transcription of genes regulated by this hormone. These findings suggest that cells may regulate recruitment of insulator proteins to the DNA in order to activate insulator activity at specific sites and create distinct patterns of nuclear organization that are necessary to achieve proper gene expression in response to different stimuli. PMID:21981916

Regulation of chromatin organization and inducible gene expression by a Drosophila insulator.

PubMed

Wood, Ashley M; Van Bortle, Kevin; Ramos, Edward; Takenaka, Naomi; Rohrbaugh, Margaret; Jones, Brian C; Jones, Keith C; Corces, Victor G

2011-10-07

Insulators are multiprotein-DNA complexes thought to affect gene expression by mediating inter- and intrachromosomal interactions. Drosophila insulators contain specific DNA-binding proteins plus common components, such as CP190, that facilitate these interactions. Here, we examine changes in the distribution of Drosophila insulator proteins during the heat-shock and ecdysone responses. We find that CP190 recruitment to insulator sites is the main regulatable step in controlling insulator function during heat shock. In contrast, both CP190 and DNA-binding protein recruitment are regulated during the ecdysone response. CP190 is necessary to stabilize specific chromatin loops and for proper activation of transcription of genes regulated by this hormone. These findings suggest that cells may regulate recruitment of insulator proteins to DNA to activate insulator activity at specific sites and create distinct patterns of nuclear organization that are necessary to achieve proper gene expression in response to different stimuli. Copyright © 2011 Elsevier Inc. All rights reserved.

Discovery of biaryl-4-carbonitriles as antihyperglycemic agents that may act through AMPK-p38 MAPK pathway.

PubMed

Goel, Atul; Nag, Pankaj; Rahuja, Neha; Srivastava, Rohit; Chaurasia, Sumit; Gautam, Sudeep; Chandra, Sharat; Siddiqi, Mohammad Imran; Srivastava, Arvind K

2014-08-25

A series of functionalized biaryl-4-carbonitriles was synthesized in three steps and evaluated for PTP-1B inhibitory activity. Among the synthesized compounds, four biaryls 6a-d showed inhibition (IC50 58-75 μM) against in vitro PTP-1B assay possibly due to interaction with amino acid residues Lys120, Tyr46 through hydrogen bonding and aromatic-aromatic interactions, respectively. Two biaryl-4-carbonitriles 6b and 6c showed improved glucose tolerance, fasting as well as postprandial blood glucose, serum total triglycerides, and increased high-density lipoprotein-cholesterol in SLM, STZ, STZ-S and C57BL/KsJ-db/db animal models. The bioanalysis of 4'-bromo-2,3-dimethyl-5-(piperidin-1-yl)biphenyl-4-carbonitrile (6b) revealed that like insulin, it increased 2-deoxyglucose uptake in skeletal muscle cells (L6 and C2C12 myotubes). The compound 6b significantly up-regulated the genes related to the insulin signaling pathways like AMPK, MAPK including glucose transporter-4 (GLUT-4) gene in muscle tissue of C57BL/KsJ-db/db mice. Furthermore, it was observed that the compound 6b up-regulated PPARα, UCP2 and HNF4α, which are key regulator of glucose, lipid, and fatty acid metabolism. Western blot analysis of the compound 6b showed that it significantly increased the phosphorylation of AMPK and p38 MAPK and ameliorated glucose uptake in C57BL/KsJ-db/db mice through the AMPK-p38 MAPK pathway. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

Abdominal-B and caudal inhibit the formation of specific neuroblasts in the Drosophila tail region

PubMed Central

Birkholz, Oliver; Vef, Olaf; Rogulja-Ortmann, Ana; Berger, Christian; Technau, Gerhard M.

2013-01-01

The central nervous system of Drosophila melanogaster consists of fused segmental units (neuromeres), each generated by a characteristic number of neural stem cells (neuroblasts). In the embryo, thoracic and anterior abdominal neuromeres are almost equally sized and formed by repetitive sets of neuroblasts, whereas the terminal abdominal neuromeres are generated by significantly smaller populations of progenitor cells. Here we investigated the role of the Hox gene Abdominal-B in shaping the terminal neuromeres. We show that the regulatory isoform of Abdominal-B (Abd-B.r) not only confers abdominal fate to specific neuroblasts (e.g. NB6-4) and regulates programmed cell death of several progeny cells within certain neuroblast lineages (e.g. NB3-3) in parasegment 14, but also inhibits the formation of a specific set of neuroblasts in parasegment 15 (including NB7-3). We further show that Abd-B.r requires cooperation of the ParaHox gene caudal to unfold its full competence concerning neuroblast inhibition and specification. Thus, our findings demonstrate that combined action of Abdominal-B and caudal contributes to the size and composition of the terminal neuromeres by regulating both the number and lineages of specific neuroblasts. PMID:23903193

Using FlyBase, a Database of Drosophila Genes and Genomes.

PubMed

Marygold, Steven J; Crosby, Madeline A; Goodman, Joshua L

2016-01-01

For nearly 25 years, FlyBase (flybase.org) has provided a freely available online database of biological information about Drosophila species, focusing on the model organism D. melanogaster. The need for a centralized, integrated view of Drosophila research has never been greater as advances in genomic, proteomic, and high-throughput technologies add to the quantity and diversity of available data and resources.FlyBase has taken several approaches to respond to these changes in the research landscape. Novel report pages have been generated for new reagent types and physical interaction data; Drosophila models of human disease are now represented and showcased in dedicated Human Disease Model Reports; other integrated reports have been established that bring together related genes, datasets, or reagents; Gene Reports have been revised to improve access to new data types and to highlight functional data; links to external sites have been organized and expanded; and new tools have been developed to display and interrogate all these data, including improved batch processing and bulk file availability. In addition, several new community initiatives have served to enhance interactions between researchers and FlyBase, resulting in direct user contributions and improved feedback.This chapter provides an overview of the data content, organization, and available tools within FlyBase, focusing on recent improvements. We hope it serves as a guide for our diverse user base, enabling efficient and effective exploration of the database and thereby accelerating research discoveries.

p53 genes function to restrain mobile elements

PubMed Central

Wylie, Annika; Jones, Amanda E.; D'Brot, Alejandro; Lu, Wan-Jin; Kurtz, Paula; Moran, John V.; Rakheja, Dinesh; Chen, Kenneth S.; Hammer, Robert E.; Comerford, Sarah A.; Amatruda, James F.; Abrams, John M.

2016-01-01

Throughout the animal kingdom, p53 genes govern stress response networks by specifying adaptive transcriptional responses. The human member of this gene family is mutated in most cancers, but precisely how p53 functions to mediate tumor suppression is not well understood. Using Drosophila and zebrafish models, we show that p53 restricts retrotransposon activity and genetically interacts with components of the piRNA (piwi-interacting RNA) pathway. Furthermore, transposon eruptions occurring in the p53− germline were incited by meiotic recombination, and transcripts produced from these mobile elements accumulated in the germ plasm. In gene complementation studies, normal human p53 alleles suppressed transposons, but mutant p53 alleles from cancer patients could not. Consistent with these observations, we also found patterns of unrestrained retrotransposons in p53-driven mouse and human cancers. Furthermore, p53 status correlated with repressive chromatin marks in the 5′ sequence of a synthetic LINE-1 element. Together, these observations indicate that ancestral functions of p53 operate through conserved mechanisms to contain retrotransposons. Since human p53 mutants are disabled for this activity, our findings raise the possibility that p53 mitigates oncogenic disease in part by restricting transposon mobility. PMID:26701264

Cardioprotective role of P38 MAPK during myocardial infarction via parallel activation of α-crystallin B and Nrf2.

PubMed

Mitra, Arkadeep; Ray, Aramita; Datta, Ritwik; Sengupta, Shantanu; Sarkar, Sagartirtha

2014-09-01

Myocardial infarction (MI) is defined as cardiac cell death due to prolonged ischemia. Although necrotic cell death was considered to be solely responsible for myocyte death during MI, it was recently revealed that apoptosis also plays its part in this death process. Our laboratory has recently shown that endoplasmic reticulum (ER) stress-induced apoptosis is the predominant route for apoptosis during MI and the conventional mitochondrial pathway is bypassed by activation of a small heat shock protein α-crystallin B (CRYAB). Since CRYAB is a direct target of P38 mitogen-activated protein kinase (MAPK) cascade, we were prompted to check the role of P38 MAPK in 20-week-old male Wister rats immediately after infarct formation. Interestingly, parallel activation of mitochondrial apoptotic pathway with an increase in ER stress-induced apoptotic load was observed along with decreased activation of CRYAB and Nrf2 (a pro-survival protein activated in response to ER stress) in MI rats treated with SB203580, a specific inhibitor of P38α and P38β compared to the MI alone. As a cumulative effect, this inhibitor treatment also resulted in significant increase in the levels of caspase3 activity and TUNEL positivity, the end point apoptotic markers. Furthermore, SB203580-treated hypoxic adult cardiomyocytes showed formation of desmin aggregates which were previously associated with impaired cardiac function. Thus, this study shows for the first time the precise mechanism by which P38 MAPK plays a pro-survival role and confers protection of cardiomyocytes, during infarct formation. © 2014 Wiley Periodicals, Inc.

The SDF-1-CXCR4 Axis Functions Through p38-MAPK Signaling to Drive Breast Cancer Progression and Metastasis

DTIC Science & Technology

2009-09-01

line) could induce proliferation and lead to hormone independent tumors in vivo. Upon analysis of these tumors by real-time PCR, it was found that the... analysis we have shown that overexpression of CXCR4 leads to increased levels of ER-mediated gene expression; specifically we found increased levels of...SDF-1 and, the classic ER-mediated gene, Progesterone receptor (PgR). 1.B Determine if CXCR4 activates p38. Western blot analysis of breast

Inter- and intra-species variation in genome-wide gene expression of Drosophila in response to parasitoid wasp attack.

PubMed

Salazar-Jaramillo, Laura; Jalvingh, Kirsten M; de Haan, Ammerins; Kraaijeveld, Ken; Buermans, Henk; Wertheim, Bregje

2017-04-27

Parasitoid resistance in Drosophila varies considerably, among and within species. An immune response, lamellocyte-mediated encapsulation, evolved in a subclade of Drosophila and was subsequently lost in at least one species within this subclade. While the mechanisms of resistance are fairly well documented in D. melanogaster, much less is known for closely related species. Here, we studied the inter- and intra-species variation in gene expression after parasitoid attack in Drosophila. We used RNA-seq after parasitization of four closely related Drosophila species of the melanogaster subgroup and replicated lines of D. melanogaster experimentally selected for increased resistance to gain insights into short- and long-term evolutionary changes. We found a core set of genes that are consistently up-regulated after parasitoid attack in the species and lines tested, regardless of their level of resistance. Another set of genes showed no up-regulation or expression in D. sechellia, the species unable to raise an immune response against parasitoids. This set consists largely of genes that are lineage-restricted to the melanogaster subgroup. Artificially selected lines did not show significant differences in gene expression with respect to non-selected lines in their responses to parasitoid attack, but several genes showed differential exon usage. We showed substantial similarities, but also notable differences, in the transcriptional responses to parasitoid attack among four closely related Drosophila species. In contrast, within D. melanogaster, the responses were remarkably similar. We confirmed that in the short-term, selection does not act on a pre-activation of the immune response. Instead it may target alternative mechanisms such as differential exon usage. In the long-term, we found support for the hypothesis that the ability to immunologically resist parasitoid attack is contingent on new genes that are restricted to the melanogaster subgroup.

Telomeric P elements associated with cytotype regulation of the P transposon family in Drosophila melanogaster.

PubMed Central

Stuart, Jeremy R; Haley, Kevin J; Swedzinski, Douglas; Lockner, Samuel; Kocian, Paul E; Merriman, Peter J; Simmons, Michael J

2002-01-01

P elements inserted at the left end of the Drosophila X chromosome were isolated genetically from wild-type P strains. Stocks carrying these elements were tested for repression of P-strain-induced gonadal dysgenesis in females and for repression of transposase-catalyzed P-element excision in males and females. Both traits were repressed by stocks carrying either complete or incomplete P elements inserted near the telomere of the X chromosome in cytological region 1A, but not by stocks carrying only nontelomeric X-linked P elements. All three of the telomeric P elements that were analyzed at the molecular level were inserted in one of the 1.8-kb telomere-associated sequence (TAS) repeats near the end of the X chromosome. Stocks with these telomeric P elements strongly repressed P-element excision induced in the male germline by a P strain or by the transposase-producing transgenes H(hsp/CP)2, H(hsp/CP)3, a combination of these two transgenes, and P(ry(+), delta2-3)99B. For H(hsp/CP)2 and P(ry(+), delta2-3)99B, the repression was also effective when the flies were subjected to heat-shock treatments. However, these stocks did not repress the somatic transposase activity of P(ry(+), delta2-3)99B. Repression of transposase activity in the germline required maternal transmission of the telomeric P elements themselves. Paternal transmission of these elements, or maternal transmission of the cytoplasm from carriers, both were insufficient to repress transposase activity. Collectively, these findings indicate that the regulatory abilities of telomeric P elements are similar to those of the P cytotype. PMID:12524339

Mutations in CG8878, a Novel Putative Protein Kinase, Enhance P Element Dependent Silencing (PDS) and Position Effect Variegation (PEV) in Drosophila melanogaster

PubMed Central

McCracken, Allen; Locke, John

2014-01-01

Genes in multicellular organisms are expressed as part of a developmental program that is largely dependent on self-perpetuating higher-order chromatin states. The mechanism of establishing and maintaining these epigenetic events is well studied in Drosophila. The first known example of an epigenetic effect was that of (PEV) in Drosophila, which has been shown to be due to gene silencing via heterochromatin formation. We are investigating a process similar to Position Effect Variegation (PEV) using a mini-w transgene, called Pci, inserted in the upstream regulatory region of ci. The mini-white + transgene in Pci is expressed throughout the adult eye; however, when other P or KP elements are present, a variegated eye phenotype results indicating random w + silencing during development. This P element dependent silencing (PDS) can be modified by the haplo-suppressors/triplo-enhancers, Su(var)205 and Su(var)3–7, indicating that these heterochromatic modifiers also act dose dependently in PDS. Here we use a spontaneous derivative mutation of Pci called PciE1 (E1) that variegates like PDS in the absence of P elements, presumably due to an adjacent gypsy element insertion, to screen for second-site modifier mutations that enhance variable silencing of white + in E1. We isolated 7 mutations in CG8878, an essential gene, that enhance the E1 variegated phenotype. CG8878, a previously uncharacterized gene, potentially encodes a serine/threonine kinase whose closest Drosophila paralogue, ballchen (nhk-1), phosphorylates histones. These mutant alleles enhance both PDS at E1 and Position Effect Variegation (PEV) at wm4, indicating a previously unknown common silencing mechanism between the two. PMID:24614804

Adaptive response due to changes in gene regulation: a study with Drosophila.

PubMed Central

McDonald, J F; Chambers, G K; David, J; Ayala, F J

1977-01-01

In spite of the critical role of the process of adaptation in evolution, there are few detailed studies of the genotypic and molecular basis of the process. Drosophila melanogaster flies selected for increased tolerance to ethanol exhibited higher levels of alcohol dehydrogenase (alcohol:NAD+ oxidoreductase; EC 1.1.1.1) activity than unselected controls. A series of tests (electrophoresis, product inhibition, temperature stability, pH optima, substrate specificity, and Michaelis constants) gave no evidence of structural differences in the enzyme of the selected and the control flies. However, quantitative immunological assays showed that the selected flies contained significantly higher amounts of alcohol dehydrogenase. Adaptation of the selected flies to higher alcohol tolerance has most likely taken place by changes not in the structural gene locus coding for the enzyme, but by regulatory changes affecting the amount of gene product. Images PMID:412190

Molecular evolution of the insect chemoreceptor gene superfamily in Drosophila melanogaster.

PubMed

Robertson, Hugh M; Warr, Coral G; Carlson, John R

2003-11-25

The insect chemoreceptor superfamily in Drosophila melanogaster is predicted to consist of 62 odorant receptor (Or) and 68 gustatory receptor (Gr) proteins, encoded by families of 60 Or and 60 Gr genes through alternative splicing. We include two previously undescribed Or genes and two previously undescribed Gr genes; two previously predicted Or genes are shown to be alternative splice forms. Three polymorphic pseudogenes and one highly defective pseudogene are recognized. Phylogenetic analysis reveals deep branches connecting multiple highly divergent clades within the Gr family, and the Or family appears to be a single highly expanded lineage within the superfamily. The genes are spread throughout the Drosophila genome, with some relatively recently diverged genes still clustered in the genome. The Gr5a gene on the X chromosome, which encodes a receptor for the sugar trehalose, has transposed from one such tandem cluster of six genes at cytological location 64, as has Gr61a, and all eight of these receptors might bind sugars. Analysis of intron evolution suggests that the common ancestor consisted of a long N-terminal exon encoding transmembrane domains 1-5 followed by three exons encoding transmembrane domains 6-7. As many as 57 additional introns have been acquired idiosyncratically during the evolution of the superfamily, whereas the ancestral introns and some of the older idiosyncratic introns have been lost at least 48 times independently. Altogether, these patterns of molecular evolution suggest that this is an ancient superfamily of chemoreceptors, probably dating back at least to the origin of the arthropods.

Molecular evolution of the insect chemoreceptor gene superfamily in Drosophila melanogaster

PubMed Central

Robertson, Hugh M.; Warr, Coral G.; Carlson, John R.

2003-01-01

The insect chemoreceptor superfamily in Drosophila melanogaster is predicted to consist of 62 odorant receptor (Or) and 68 gustatory receptor (Gr) proteins, encoded by families of 60 Or and 60 Gr genes through alternative splicing. We include two previously undescribed Or genes and two previously undescribed Gr genes; two previously predicted Or genes are shown to be alternative splice forms. Three polymorphic pseudogenes and one highly defective pseudogene are recognized. Phylogenetic analysis reveals deep branches connecting multiple highly divergent clades within the Gr family, and the Or family appears to be a single highly expanded lineage within the superfamily. The genes are spread throughout the Drosophila genome, with some relatively recently diverged genes still clustered in the genome. The Gr5a gene on the X chromosome, which encodes a receptor for the sugar trehalose, has transposed from one such tandem cluster of six genes at cytological location 64, as has Gr61a, and all eight of these receptors might bind sugars. Analysis of intron evolution suggests that the common ancestor consisted of a long N-terminal exon encoding transmembrane domains 1-5 followed by three exons encoding transmembrane domains 6-7. As many as 57 additional introns have been acquired idiosyncratically during the evolution of the superfamily, whereas the ancestral introns and some of the older idiosyncratic introns have been lost at least 48 times independently. Altogether, these patterns of molecular evolution suggest that this is an ancient superfamily of chemoreceptors, probably dating back at least to the origin of the arthropods. PMID:14608037

Signal Integration by the IκB Protein Pickle Shapes Drosophila Innate Host Defense.

PubMed

Morris, Otto; Liu, Xi; Domingues, Celia; Runchel, Christopher; Chai, Andrea; Basith, Shaherin; Tenev, Tencho; Chen, Haiyang; Choi, Sangdun; Pennetta, Giuseppa; Buchon, Nicolas; Meier, Pascal

2016-09-14

Pattern recognition receptors are activated following infection and trigger transcriptional programs important for host defense. Tight regulation of NF-κB activation is critical to avoid detrimental and misbalanced responses. We describe Pickle, a Drosophila nuclear IκB that integrates signaling inputs from both the Imd and Toll pathways by skewing the transcriptional output of the NF-κB dimer repertoire. Pickle interacts with the NF-κB protein Relish and the histone deacetylase dHDAC1, selectively repressing Relish homodimers while leaving other NF-κB dimer combinations unscathed. Pickle's ability to selectively inhibit Relish homodimer activity contributes to proper host immunity and organismal health. Although loss of pickle results in hyper-induction of Relish target genes and improved host resistance to pathogenic bacteria in the short term, chronic inactivation of pickle causes loss of immune tolerance and shortened lifespan. Pickle therefore allows balanced immune responses that protect from pathogenic microbes while permitting the establishment of beneficial commensal host-microbe relationships. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

Characterization of the Drosophila Methoprene -tolerant gene product. Juvenile hormone binding and ligand-dependent gene regulation.

PubMed

Miura, Ken; Oda, Masahito; Makita, Sumiko; Chinzei, Yasuo

2005-03-01

Juvenile hormones (JHs) of insects are sesquiterpenoids that regulate a great diversity of processes in development and reproduction. As yet the molecular modes of action of JH are poorly understood. The Methoprene-tolerant (Met) gene of Drosophila melanogaster has been found to be responsible for resistance to a JH analogue (JHA) insecticide, methoprene. Previous studies on Met have implicated its involvement in JH signaling, although direct evidence is lacking. We have now examined the product of Met (MET) in terms of its binding to JH and ligand-dependent gene regulation. In vitro synthesized MET directly bound to JH III with high affinity (Kd = 5.3 +/- 1.5 nm, mean +/- SD), consistent with the physiological JH concentration. In transient transfection assays using Drosophila S2 cells the yeast GAL4-DNA binding domain fused to MET exerted JH- or JHA-dependent activation of a reporter gene. Activation of the reporter gene was highly JH- or JHA-specific with the order of effectiveness: JH III > JH II > JH I > methoprene; compounds which are only structurally related to JH or JHA did not induce any activation. Localization of MET in the S2 cells was nuclear irrespective of the presence or absence of JH. These results suggest that MET may function as a JH-dependent transcription factor.

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

PubMed Central

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

Mapping Linked Genes in "Drosophila Melanogaster" Using Data from the F2 Generation of a Dihybrid Cross

ERIC Educational Resources Information Center

Marshall, Pamela A.

2008-01-01

"Drosophila melanogaster" is a commonly utilized organism for testing hypotheses about inheritance of traits. Students in both high school and university labs study the genetics of inheritance by analyzing offspring of appropriate "Drosophila" crosses to determine inheritance patterns, including gene linkage. However, most genetics investigations…

Identification and partial characterization of the enzyme of omega: one of five putative DPP IV genes in Drosophila melanogaster

PubMed Central

Chihara, Carol J.; Song, Chunyan; LaMonte, Greg; Fetalvero, Kristina; Hinchman, Kristy; Phan, Helen; Pineda, Mario; Robinson, Kelly; Schneider, Gregory P.

2005-01-01

The omega (ome) gene product is a modifier of larval cuticle protein 5 and its alleles (and duplicates) in the third instar of Drosophila melanogaster. Using deletion mapping the locus mapped to 70F-71A on the left arm of chromosome 3. A homozygote null mutant (ome 1) shows a pleiotropic phenotype that affected the size, developmental time of the flies, and the fertility (or perhaps the behavior) of homozygous mutant males. The omega gene was verified as producing a dipeptidyl peptidase IV (DPPIV) by genetic analysis, substrate specificity and pH optimum. The identity of the gene was confirmed as CG32145 (cytology 70F4) in the Celera Database (Berkeley Drosophila Genome Project), which is consistent with its deletion map position. The genomic structure of the gene is described and the decrease in DPPIV activity in the mutant ome1 is shown to be due to the gene CG32145 (omega). The D. melanogaster omega DPPIV enzyme was partially purified and characterized. The exons of the ome1 mutant were sequenced and a base substitution mutation in exon 4 was identified that would yield a truncated protein caused by a stop codon. A preliminary study of the compartmentalization of the omega DPPIV enzyme in several organs is also reported. Abbreviations: DPPIV dipeptidyl peptidase IV LCP5 & LCP6 third instar larval cuticle proteins 5 & 6 ome & ome1 omega locus name (CG32145) and mutant allele in D. melanogaster pNA paranotroanilide PMID:17119608

A Metagenome-Wide Association Study and Arrayed Mutant Library Confirm Acetobacter Lipopolysaccharide Genes Are Necessary for Association with Drosophila melanogaster.

PubMed

White, K Makay; Matthews, Melinda K; Hughes, Rachel C; Sommer, Andrew J; Griffitts, Joel S; Newell, Peter D; Chaston, John M

2018-03-28

A metagenome wide association (MGWA) study of bacterial host association determinants in Drosophila predicted that LPS biosynthesis genes are significantly associated with host colonization. We were unable to create site-directed mutants for each of the predicted genes in Acetobacter , so we created an arrayed transposon insertion library using Acetobacter fabarum DsW_054 isolated from Drosophila Creation of the A. fabarum DsW_054 gene knock-out library was performed by combinatorial mapping and Illumina sequencing of random transposon insertion mutants. Transposon insertion locations for 6,418 mutants were successfully mapped, including hits within 63% of annotated genes in the A. fabarum DsW_054 genome. For 45/45 members of the library, insertion sites were verified by arbitrary PCR and Sanger sequencing. Mutants with insertions in four different LPS biosynthesis genes were selected from the library to validate the MGWA predictions. Insertion mutations in two genes biosynthetically upstream of Lipid-A formation, lpxC and lpxB , show significant differences in host association, whereas mutations in two genes encoding LPS biosynthesis functions downstream of Lipid-A biosynthesis had no effect. These results suggest an impact of bacterial cell surface molecules on the bacterial capacity for host association. Also, the transposon insertion mutant library will be a useful resource for ongoing research on the genetic basis for Acetobacter traits. Copyright © 2018 White et al.

Depletion of a Drosophila homolog of yeast Sup35p disrupts spindle assembly, chromosome segregation, and cytokinesis during male meiosis.

PubMed

Basu, J; Williams, B C; Li, Z; Williams, E V; Goldberg, M L

1998-01-01

In the course of a genetic screen for male-sterile mutations in Drosophila affecting chromosome segregation during the meiotic divisions in spermatocytes, we identified the mutation dsup35(63D). Examination of mutant testes showed that chromosome misbehavior was a consequence of major disruptions in meiotic spindle assembly. These perturbations included problems in aster formation, separation, and migration around the nuclear envelope; aberrations in spindle organization and integrity; and disappearance of the ana/telophase central spindle, which in turn disrupts cytokinesis. The dsup35(63D) mutation is caused by a P element insertion that affects, specifically in the testis, the expression of a gene (dsup35) encoding the Drosophila homolog of the yeast Sup35p and Xenopus eRF3 proteins. These proteins are involved in the termination of polypeptide synthesis on ribosomes, but previous studies have suggested that Sup35p and closely related proteins of the same family also interact directly with microtubules. An affinity-purified antibody directed against the product of the dsup35 gene was prepared; interestingly, this antibody specifically labels primary spermatocytes in one or two discrete foci of unknown structure within the nucleoplasm. We discuss how depletion of the dsup35 gene product in spermatocytes might lead to the global disruptions in meiotic spindle assembly seen in mutant spermatocytes.

Genetic Resistance to Viral Infection: The Molecular Cloning of a Drosophila Gene That Restricts Infection by the Rhabdovirus Sigma

PubMed Central

Contamine, D.; Petitjean, A. M.; Ashburner, M.

1989-01-01

The ref(2)P gene of Drosophila melanogaster has two common alleles, ref(2)P(o) which permits the infection of flies by the rhabdovirus sigma (σ), and ref(2)P(p) which is restrictive for σ infection. This gene has been cloned by P element tagging and shown to code for two RNAs in adult flies. These RNAs are expressed in both males and females, but only the larger is expressed in ovaries. Both transcripts are shorter, by about 50 nucleotides, in flies carrying the ref(2)P(p) allele than in those carrying ref(2)P(o). The dominance relationships of these two alleles, and the fact that ref(2)P(null) alleles are permissive to σ infection, suggest that the ref(2)P(o) product is antimorphic to that of the ref(2)P(p) allele. PMID:2557263

Genome-wide identification of 52 cytochrome P450 (CYP) genes in the copepod Tigriopus japonicus and their B[α]P-induced expression patterns.

PubMed

Han, Jeonghoon; Kim, Duck-Hyun; Kim, Hui-Su; Nelson, David R; Lee, Jae-Seong

2017-09-01

Cytochrome P450s (CYPs) are enzymes with a heme-binding domain that are found in all living organisms. CYP enzymes have important roles associated with detoxification of xenobiotics and endogenous compounds (e.g. steroids, fatty acids, and hormones). Although CYP enzymes have been reported in several invertebrates, including insects, little is known about copepod CYPs. Here, we identified the entire repertoire of CYP genes (n=52) from whole genome and transcriptome sequences of the benthic copepod Tigriopus japonicus, including a tandem duplication (CYP3026A3, CYP3026A4, CYP3026A5), and examined patterns of gene expression over various developmental stages and in response to benzo[α]pyrene (B[α]P) exposure. Through phylogenetic analysis, the 52 T. japonicus CYP genes were assigned to five distinct clans: CYP2 (22 genes), CYP3 (19 genes), CYP4 (two genes), CYP20 (one gene), and mitochondrial (eight genes). Developmental stage and gender-specific expression patterns of the 52 T. japonicus CYPs were analyzed. CYP3022A1 was constitutively expressed during all developmental stages. CYP genes in clans 2 and 3 were induced in response to B[α]P, suggesting that these differentially modulated CYP transcripts are likely involved in defense against exposure to B[α]P and other pollutants. This study enhances our understanding of the repertoire of CYP genes in copepods and of their potential role in development and detoxification in copepods. Copyright © 2017 Elsevier Inc. All rights reserved.

The myosin ID pathway and left-right asymmetry in Drosophila.

PubMed

Géminard, Charles; González-Morales, Nicanor; Coutelis, Jean-Baptiste; Noselli, Stéphane

2014-06-01

Drosophila is a classical model to study body patterning, however left-right (L/R) asymmetry had remained unexplored, until recently. The discovery of the conserved myosin ID gene as a major determinant of L/R asymmetry has revealed a novel L/R pathway involving the actin cytoskeleton and the adherens junction. In this process, the HOX gene Abdominal-B plays a major role through the control of myosin ID expression and therefore symmetry breaking. In this review, we present organs and markers showing L/R asymmetry in Drosophila and discuss our current understanding of the underlying molecular genetic mechanisms. Drosophila represents a valuable model system revealing novel strategies to establish L/R asymmetry in invertebrates and providing an evolutionary perspective to the problem of laterality in bilateria. © 2014 Wiley Periodicals, Inc.

Adhesive pad differentiation in Drosophila melanogaster depends on the Polycomb group gene Su(z)2.

PubMed

Hüsken, Mirko; Hufnagel, Kim; Mende, Katharina; Appel, Esther; Meyer, Heiko; Peisker, Henrik; Tögel, Markus; Wang, Shuoshuo; Wolff, Jonas; Gorb, Stanislav N; Paululat, Achim

2015-04-15

The ability of many insects to walk on vertical smooth surfaces such as glass or even on the ceiling has fascinated biologists for a long time, and has led to the discovery of highly specialized adhesive organs located at the distal end of the animals' legs. So far, research has primarily focused on structural and ultrastructural investigations leading to a deeper understanding of adhesive organ functionality and to the development of new bioinspired materials. Genetic approaches, e.g. the analysis of mutants, to achieve a better understanding of adhesive organ differentiation have not been used so far. Here, we describe the first Drosophila melanogaster mutant that develops malformed adhesive organs, resulting in a complete loss of climbing ability on vertical smooth surfaces. Interestingly, these mutants fail to make close contact between the setal tips and the smooth surface, a crucial condition for wet adhesion mediated by capillary forces. Instead, these flies walk solely on their claws. Moreover, we were able to show that the mutation is caused by a P-element insertion into the Su(z)2 gene locus. Remobilization of the P-element restores climbing ability. Furthermore, we provide evidence that the P-element insertion results in an artificial Su(z)2 transcript, which most likely causes a gain-of-function mutation. We presume that this transcript causes deregulation of yet unknown target genes involved in pulvilli differentiation. Our results nicely demonstrate that the genetically treatable model organism Drosophila is highly suitable for future investigations on adhesive organ differentiation. © 2015. Published by The Company of Biologists Ltd.

Retinal determination genes coordinate neuroepithelial specification and neurogenesis modes in the Drosophila optic lobe

PubMed Central

Apitz, Holger

2016-01-01

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

Ubiquitin over-expression phenotypes and ubiquitin gene molecular misreading during aging in Drosophila melanogaster

PubMed Central

Hoe, Nicholas; Huang, Chung M.; Landis, Gary; Verhage, Marian; Ford, Daniel; Yang, Junsheng; van Leeuwen, Fred W.; Tower, John

2011-01-01

Molecular Misreading (MM) is the inaccurate conversion of genomic information into aberrant proteins. For example, when RNA polymerase II transcribes a GAGAG motif it synthesizes at low frequency RNA with a two-base deletion. If the deletion occurs in a coding region, translation will result in production of misframed proteins. During mammalian aging, misframed versions of human amyloid precursor protein (hApp) and ubiquitin (hUbb) accumulate in the aggregates characteristic of neurodegenerative diseases, suggesting dysfunctional degradation or clearance. Here cDNA clones encoding wild-type hUbb and the frame-shifted version hUbb+1 were expressed in transgenic Drosophila using the doxycycline-regulated system. Misframed proteins were abundantly produced, both from the transgenes and from endogenous Drosophila ubiquitin-encoding genes, and their abundance increased during aging in whole-fly extracts. Over-expression of wild-type hUbb, but not hUbb+1, was toxic during fly development. In contrast, when over-expressed specifically in adult flies, hUbb+1 caused small decreases in life span, whereas hUbb was associated with small increases, preferentially in males. The data suggest that MM occurs in Drosophila and that the resultant misframed proteins accumulate with age. MM of the ubiquitin gene can produce alternative ubiquitin gene products with different and sometimes opposing phenotypic effects. PMID:21415465

Isolation and characterization of polygalacturonase genes (pecA and pecB) from Aspergillus flavus.

PubMed Central

Whitehead, M P; Shieh, M T; Cleveland, T E; Cary, J W; Dean, R A

1995-01-01

Two genes, pecA and pecB, encoding endopolyglacturonases were cloned from a highly aggressive strain of Aspergillus flavus. The pecA gene consisted of 1,228 bp encoding a protein of 363 amino acids with a predicted molecular mass of 37.6 kDa, interrupted by two introns of 58 and 81 bp in length. Accumulation of pecA mRNA in both pectin- or glucose-grown mycelia in the highly aggressive strain matched the activity profile of a pectinase previously identified as P2c. Transformants of a weakly aggressive strain containing a functional copy of the pecA gene produced P2c in vitro, confirming that pecA encodes P2c. The coding region of pecB was determined to be 1,217 bp in length interrupted by two introns of 65 and 54 bp in length. The predicted protein of 366 amino acids had an estimated molecular mass of 38 kDa. Transcripts of this gene accumulated in mycelia grown in medium containing pectin alone, never in mycelia grown in glucose-containing medium, for both highly and weakly aggressive strains. Thus, pecB encodes the activity previously identified as P1 or P3. pecA and pecB share a high degree of sequence identity with polygalacturonase genes from Aspergillus parasiticus and Aspergillus oryzae, further establishing the close relationships between members of the A. flavus group. Conservation of intron positions in these genes also indicates that they share a common ancestor with genes encoding endopolyglacturonases of Aspergillus niger. PMID:7574642

Control of sigma virus multiplication by the ref(2)P gene of Drosophila melanogaster: an in vivo study of the PB1 domain of Ref(2)P.

PubMed

Carré-Mlouka, A; Gaumer, S; Gay, P; Petitjean, A M; Coulondre, C; Dru, P; Bras, F; Dezélée, S; Contamine, D

2007-05-01

Ref(2)P has been described as one of the Drosophila proteins that interacts with the sigma virus cycle. We generated alleles to identify critical residues involved in the restrictive (inhibiting viral multiplication) or permissive (allowing viral multiplication) character of Ref(2)P. We demonstrate that permissive alleles increase the ability of the sigma virus to infect Drosophila when compared to null alleles and we confirm that restrictive alleles decrease this capacity. Moreover, we have created alleles unfunctional in viral cycling while functional for Ref(2)P fly functions. This type of allele had never been observed before and shows that fly- and virus-related activities of Ref(2)P are separable. The viral status of Ref(2)P variants is determined by the amino-terminal PB1 domain polymorphism. In addition, an isolated PB1 domain mimics virus-related functions even if it is similar to a loss of function toward fly-related activities. The evolutionary tree of the Ref(2)P PB1 domain that we could build on the basis of the natural allele sequences is in agreement with an evolution of PB1 domain due to successive transient selection waves.

Effects of P Element Insertions on Quantitative Traits in Drosophila Melanogaster

PubMed Central

Mackay, TFC.; Lyman, R. F.; Jackson, M. S.

1992-01-01

P element mutagenesis was used to construct 94 third chromosome lines of Drosophila melanogaster which contained on average 3.1 stable P element inserts, in an inbred host strain background previously free of P elements. The homozygous and heterozygous effects of the inserts on viability and abdominal and sternopleural bristle number were ascertained by comparing the chromosome lines with inserts to insert-free control lines of the inbred host strain. P elements reduced average homozygous viability by 12.2% per insert and average heterozygous viability by 5.5% per insert, and induced recessive lethal mutations at a rate of 3.8% per insert. Mutational variation for the bristle traits averaged over both sexes was 0.03V(e) per homozygous P insert and 0.003V(e) per heterozygous P insert, where V(e) is the environmental variance. Mutational variation was greater for the sexes considered separately because inserts had large pleiotropic effects on sex dimorphism of bristle characters. The distributions of homozygous effects of inserts on the bristle traits were asymmetrical, with the largest effects in the direction of reducing bristle number; and highly leptokurtic, with most of the increase in variance contributed by a few lines with large effects. The inserts had partially recessive effects on the bristle traits. Insert lines with extreme bristle effects had on average greatly reduced viability. PMID:1311697

Large clusters of co-expressed genes in the Drosophila genome.

PubMed

Boutanaev, Alexander M; Kalmykova, Alla I; Shevelyov, Yuri Y; Nurminsky, Dmitry I

2002-12-12

Clustering of co-expressed, non-homologous genes on chromosomes implies their co-regulation. In lower eukaryotes, co-expressed genes are often found in pairs. Clustering of genes that share aspects of transcriptional regulation has also been reported in higher eukaryotes. To advance our understanding of the mode of coordinated gene regulation in multicellular organisms, we performed a genome-wide analysis of the chromosomal distribution of co-expressed genes in Drosophila. We identified a total of 1,661 testes-specific genes, one-third of which are clustered on chromosomes. The number of clusters of three or more genes is much higher than expected by chance. We observed a similar trend for genes upregulated in the embryo and in the adult head, although the expression pattern of individual genes cannot be predicted on the basis of chromosomal position alone. Our data suggest that the prevalent mechanism of transcriptional co-regulation in higher eukaryotes operates with extensive chromatin domains that comprise multiple genes.

Attenuation of p38α MAPK stress response signaling delays the in vivo aging of skeletal muscle myofibers and progenitor cells.

PubMed

Papaconstantinou, John; Wang, Chen Z; Zhang, Min; Yang, San; Deford, James; Bulavin, Dmitry V; Ansari, Naseem H

2015-09-01

Functional competence and self-renewal of mammalian skeletal muscle myofibers and progenitor cells declines with age. Progression of the muscle aging phenotype involves the decline of juvenile protective factorsi.e., proteins whose beneficial functions translate directly to the quality of life, and self-renewal of progenitor cells. These characteristics occur simultaneously with the age-associated increase of p38α stress response signaling. This suggests that the maintenance of low levels of p38α activity of juvenile tissues may delay or attenuate aging. We used the dominant negative haploinsufficient p38α mouse (DN-p38α(AF/+)) to demonstrate that in vivo attenuation of p38α activity in the gastrocnemius of the aged mutant delays age-associated processes that include: a) the decline of the juvenile protective factors, BubR1, aldehyde dehydrogenase 1A (ALDH1A1), and aldehyde dehydrogenase 2 (ALDH2); b) attenuated expression of p16(Ink4a) and p19(Arf) tumor suppressor genes of the Cdkn2a locus; c) decreased levels of hydroxynonenal protein adducts, expression of COX2 and iNOS; d) decline of the senescent progenitor cell pool level and d) the loss of gastrocnemius muscle mass. We propose that elevated P-p38α activity promotes skeletal muscle aging and that the homeostasis of p38α impacts the maintenance of a beneficial healthspan.

Administration of chlorogenic acid alleviates spinal cord injury via TLR4/NF‑κB and p38 signaling pathway anti‑inflammatory activity.

PubMed

Chen, Dayong; Pan, Dan; Tang, Shaolong; Tan, Zhihong; Zhang, Yanan; Fu, Yunfeng; Lü, Guohua; Huang, Qinghua

2018-01-01

Chlorogenic acid, as a secondary metabolite of plants, exhibits a variety of effects including free radical scavenging, antiseptic, anti‑inflammatory and anti‑viral, in addition to its ability to reduce blood glucose, protect the liver and act as an anti‑hyperlipidemic agent and cholagogue. The present study demonstrated that administration of chlorogenic acid alleviated spinal cord injury (SCI) via anti‑inflammatory activity mediated by nuclear factor (NF)‑κB and p38 signaling pathways. Wistar rats were used to structure a SCI model rat to explore the effects of administration of chlorogenic acid on SCI. The Basso, Beattie and Bresnahan test was executed for assessment of neuronal functional recovery and then spinal cord tissue wet/dry weight ratio was recorded. The present study demonstrated that chlorogenic acid increased SCI‑inhibition of BBB scores and decreased SCI‑induction of spinal cord wet/dry weight ratio in rats. In addition, chlorogenic acid suppressed SCI‑induced inflammatory activity, inducible nitric oxide synthase activity and cyclooxygenase‑2 protein expression in the SCI rat. Furthermore, chlorogenic acid suppressed Toll like receptor (TLR)‑4/myeloid differentiation primary response 88 (MyD88)/NF‑κB/IκB signaling pathways and downregulated p38 mitogen activated protein kinase protein expression in SCI rats. The findings suggest that administration of chlorogenic acid alleviates SCI via anti‑inflammatory activity mediated by TLR4/MyD88/NF‑κB and p38 signaling pathways.

Adenosine 5'-monophosphate-induced hypothermia inhibits the activation of ERK1/2, JNK, p38 and NF-κB in endotoxemic rats.

PubMed

Wang, Yunlong; Zhang, Aihua; Lu, Shulai; Pan, Xinting; Jia, Dongmei; Yu, Wenjuan; Jiang, Yanxia; Li, Xinde; Wang, Xuefeng; Zhang, Jidong; Hou, Lin; Sun, Yunbo

2014-11-01

Many studies have shown that LPS mainly activates four signal transduction pathways to induce inflammation, namely the p38, ERK1/2, JNK and IKK/NF-κB pathways. Studies have demonstrated that 5'-AMP-induced hypothermia (AIH) exhibits high anti-inflammatory capabilities. In this study, we explore that how AIH inhibits the inflammatory response. Wistar rats were divided into five groups: a control group, an LPS group, a 5'-AMP pre-treatment group, a 5'-AMP post-treatment group and a 5'-AMP group. For each group, plasma and lung were collected from the rats at 6h and 12h after LPS injection. ELISA assays were used to detect plasma levels of CD14, CRP and MCP-1. Inflammatory pathway activation and TLR4 expression were assayed separately by Western blot analysis and immunohistochemistry. Our results showed that rats treated with AIH either before or after an LPS-challenge had a significant decrease in plasma levels of CD14, CRP and TLR4 compared with rats that received LPS only. Western blot analysis showed that AIH inhibited the activation of extracellular signal-regulated kinases (ERK) 1/2, p38, c-Jun N-terminal kinase (JNK) and NF-κB in inflammatory rats. Our study concluded that AIH attenuated LPS-induced inflammation mainly by inhibiting activation on the ERK1/2, p38, JNK and NF-κB signaling pathways. Copyright © 2014 Elsevier B.V. All rights reserved.

Dopa decarboxylase (Ddc) affects variation in Drosophila longevity.

PubMed

De Luca, Maria; Roshina, Nataliya V; Geiger-Thornsberry, Gretchen L; Lyman, Richard F; Pasyukova, Elena G; Mackay, Trudy F C

2003-08-01

Mutational analyses in model organisms have shown that genes affecting metabolism and stress resistance regulate life span, but the genes responsible for variation in longevity in natural populations are largely unidentified. Previously, we mapped quantitative trait loci (QTLs) affecting variation in longevity between two Drosophila melanogaster strains. Here, we show that the longevity QTL in the 36E;38B cytogenetic interval on chromosome 2 contains multiple closely linked QTLs, including the Dopa decarboxylase (Ddc) locus. Complementation tests to mutations show that Ddc is a positional candidate gene for life span in these strains. Linkage disequilibrium (LD) mapping in a sample of 173 alleles from a single population shows that three common molecular polymorphisms in Ddc account for 15.5% of the genetic contribution to variance in life span from chromosome 2. The polymorphisms are in strong LD, and the effects of the haplotypes on longevity suggest that the polymorphisms are maintained by balancing selection. DDC catalyzes the final step in the synthesis of the neurotransmitters, dopamine and serotonin. Thus, these data implicate variation in the synthesis of bioamines as a factor contributing to natural variation in individual life span.

Behavioral Teratogenesis in Drosophila melanogaster.

PubMed

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.

Bitter receptor gene (TAS2R38) P49A genotypes and their associations with aversion to vegetables and sweet/fat foods in Malaysian subjects.

PubMed

Ooi, Shee-Xuen; Lee, Pui-Leng; Law, Huey-Yi; Say, Yee-How

2010-01-01

Recently, the bitter receptor gene (TAS2R38) was identified to be responsible for phenylthiocarbamide (PTC) bitter sensitivity. Its two predominant haplotypes at three Single Nucleotide Polymorphisms (SNPs) are found to be definitive for the PTC status, which the ProAlaVal and AlaValIle haplotypes are associated with tasters and non-tasters, respectively. TAS2R38 haplotypes have been reported to influence food preferences (like cruciferous vegetables and fat foods) and cardiovascular disease risk factors. We examined, in 215 Malaysian subjects (100 males, 115 females), the association of the P49A SNP of TAS2R38 with anthropometric measurements and aversion to a list of 36 vegetables, 4 soy products, green tea and 37 sweet/fat foods. The subjects were successfully genotyped as 110 PA, 81 PP and 24 AA (with the A49 allelic frequency of 0.37), by Polymerase Chain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP). Ethnicity (Malay, Chinese or Indian), but not gender, was associated with the P49A TAS2R38 genotypes (p<0.001). However, no significant differences in terms of Body Mass Index, Total Body Fat, waist circumference and Waist-Hip Ratio were found between the genotypes (p<0.05). Only aversions to green tea, mayonnaise and whipped cream, but not soy products, vegetables, and other sweet/fat foods, were associated with the P49A genotypes (p<0.05). Therefore, the P49A SNP of the bitter receptor gene TAS2R38 could not serve as a predictor of anthropometric measurements and aversion to vegetables or sweet/fat foods in the sampled Malaysian subjects, and this suggests the existence of other possible factors influencing food selection among Malaysians.

The Goddard and Saturn Genes Are Essential for Drosophila Male Fertility and May Have Arisen De Novo

PubMed Central

Gubala, Anna M.; Schmitz, Jonathan F.; Kearns, Michael J.; Vinh, Tery T.; Bornberg-Bauer, Erich; Wolfner, Mariana F.

2017-01-01

New genes arise through a variety of mechanisms, including the duplication of existing genes and the de novo birth of genes from noncoding DNA sequences. While there are numerous examples of duplicated genes with important functional roles, the functions of de novo genes remain largely unexplored. Many newly evolved genes are expressed in the male reproductive tract, suggesting that these evolutionary innovations may provide advantages to males experiencing sexual selection. Using testis-specific RNA interference, we screened 11 putative de novo genes in Drosophila melanogaster for effects on male fertility and identified two, goddard and saturn, that are essential for spermatogenesis and sperm function. Goddard knockdown (KD) males fail to produce mature sperm, while saturn KD males produce few sperm, and these function inefficiently once transferred to females. Consistent with a de novo origin, both genes are identifiable only in Drosophila and are predicted to encode proteins with no sequence similarity to any annotated protein. However, since high levels of divergence prevented the unambiguous identification of the noncoding sequences from which each gene arose, we consider goddard and saturn to be putative de novo genes. Within Drosophila, both genes have been lost in certain lineages, but show conserved, male-specific patterns of expression in the species in which they are found. Goddard is consistently found in single-copy and evolves under purifying selection. In contrast, saturn has diversified through gene duplication and positive selection. These data suggest that de novo genes can acquire essential roles in male reproduction. PMID:28104747

Geraniol ameliorates TNBS-induced colitis: Involvement of Wnt/β-catenin, p38MAPK, NFκB, and PPARγ signaling pathways.

PubMed

Soubh, Ayman A; Abdallah, Dalaal M; El-Abhar, Hanan S

2015-09-01

Geraniol, a natural component of plant essential oils, exhibits potent chemopreventive effects in the colon; however, its possible role/mechanisms in experimental colitis have not been elucidated, which is the aim of this study. To fulfill this goal, rats were treated for 11days with geraniol and/or sulfasalazine using a TNBS-induced colitis model. Geraniol significantly hindered the colitis-clinical signs (weight loss, colon edema,ulcerative area, colon/spleen mass indices) and opposed the altered oxidative/nitrosative stress. It restored the depleted total antioxidant capacity and lessened the elevated levels of nitric oxide and lipid peroxide. TNBS induced apoptosis and inflammatory cell infiltration, whereas geraniol curtailed these effects by diminishing the levels of caspase-3, intercellular adhesion molecule-1, and myeloperoxidase. The anti-inflammatory effect was documented by inhibiting the colon contents of prostaglandin E2 and interleukin-1β. In order to delve into the anti-colitic signaling pathways, geraniol inhibited the content/expression of glycogen synthase kinase (GSK)-3β, β-catenin, p38 mitogen activated protein kinase (p38MAPK), and nuclear factor kappa B (NFκB), but upregulated that of peroxisome proliferator activated receptor γ (PPARγ). These effects were comparable to those of sulfasalazine, the standard drug, whereas its combination with geraniol mediated effects that surpassed either treatment alone. Geraniol in the current study improved experimental colitis partly via its antioxidant, anti-inflammatory, and immunosuppressive potentials, possibly by modulating the Wnt/GSK-3β/β-catenin, p38MAPK, NFκB, and PPARγ signaling pathways. The study also revealed that geraniol represents a valuable asset against colitis alone or in combination with the conventional anti-colitic therapies. Copyright © 2015 Elsevier Inc. All rights reserved.

Regulation of P-element transposase activity in Drosophila melanogaster by hobo transgenes that contain KP elements.

PubMed Central

Simmons, Michael J; Haley, Kevin J; Grimes, Craig D; Raymond, John D; Fong, Joseph C L

2002-01-01

Fusions between the Drosophila hsp70 promoter and three different incomplete P elements, KP, SP, and BP1, were inserted into the Drosophila genome by means of hobo transformation vectors and the resulting transgenic stocks were tested for repression of P-element transposase activity. Only the H(hsp/KP) transgenes repressed transposase activity, and the degree of repression was comparable to that of a naturally occurring KP element. The KP transgenes repressed transposase activity both with and without heat-shock treatments. Both the KP element and H(hsp/KP) transgenes repressed the transposase activity encoded by the modified P element in the P(ry(+), Delta2-3)99B transgene more effectively than that encoded by the complete P element in the H(hsp/CP)2 transgene even though the P(ry(+), Delta2-3)99B transgene was the stronger transposase source. Repression of both transposase sources appeared to be due to a zygotic effect of the KP element or transgene. There was no evidence for repression by a strictly maternal effect; nor was there any evidence for enhancement of KP repression by the joint maternal transmission of H(hsp/KP) and H(hsp/CP) transgenes. These results are consistent with the idea that KP-mediated repression of P-element activity involves a KP-repressor polypeptide that is not maternally transmitted and that KP-mediated repression is not strengthened by the 66-kD repressor produced by complete P elements through alternate splicing of their RNA. PMID:12019235

Doxycycline Suppresses Microglial Activation by Inhibiting the p38 MAPK and NF-kB Signaling Pathways.

PubMed

Santa-Cecília, Flávia V; Socias, Benjamin; Ouidja, Mohand O; Sepulveda-Diaz, Julia E; Acuña, Leonardo; Silva, Rangel L; Michel, Patrick P; Del-Bel, Elaine; Cunha, Thiago M; Raisman-Vozari, Rita

2016-05-01

In neurodegenerative diseases, the inflammatory response is mediated by activated glial cells, mainly microglia, which are the resident immune cells of the central nervous system. Activated microglial cells release proinflammatory mediators and neurotoxic factors that are suspected to cause or exacerbate these diseases. We recently demonstrated that doxycycline protects substantia nigra dopaminergic neurons in an animal model of Parkinson's disease. This effect was associated with a reduction of microglial cell activation, which suggests that doxycycline may operate primarily as an anti-inflammatory drug. In the present study, we assessed the anti-inflammatory potential of doxycycline using lipopolysaccharide (LPS)-activated primary microglial cells in culture as a model of neuroinflammation. Doxycycline attenuated the expression of key activation markers in LPS-treated microglial cultures in a concentration-dependent manner. More specifically, doxycycline treatment lowered the expression of the microglial activation marker IBA-1 as well as the production of ROS, NO, and proinflammatory cytokines (TNF-α and IL-1β). In primary microglial cells, we also found that doxycycline inhibits LPS-induced p38 MAP kinase phosphorylation and NF-kB nuclear translocation. The present results indicate that the effect of doxycycline on LPS-induced microglial activation probably occurs via the modulation of p38 MAP kinase and NF-kB signaling pathways. These results support the idea that doxycycline may be useful in preventing or slowing the progression of PD and other neurodegenerative diseases that exhibit altered glia function.

TES inhibits colorectal cancer progression through activation of p38.

PubMed

Li, Huili; Huang, Kun; Gao, Lu; Wang, Lixia; Niu, Yanfeng; Liu, Hongli; Wang, Zheng; Wang, Lin; Wang, Guobin; Wang, Jiliang

2016-07-19

The human TESTIN (TES) gene has been identified as a candidate tumor suppressor based on its location at a common fragile site - a region where loss of heterozygosity has been detected in numerous types of tumors. To investigate its role in colorectal cancer (CRC), we examined TES protein levels in CRC tissue samples and cell lines. We observed that TES was markedly reduced in both CRC tissue and cell lines. Additionally, overexpression of TES significantly inhibited cell proliferation, migration, and invasion, while increasing cell apoptosis in colon cancer cells. By contrast, shRNA-mediated TES knockdown elicited the opposite effects. TES inhibited the progression of CRC by up-regulating pro-apoptotic proteins, down-regulating anti-apoptotic proteins, and simultaneously activating p38 mitogen-activated protein kinase (MAPK) signaling pathways. Collectively, these data indicate that TES functions as a necessary suppressor of CRC progression by activating p38-MAPK signaling pathways. This suggests that TES may have a potential application in CRC diagnosis and targeted gene therapy.

Gene expression variations during Drosophila metamorphosis in real and simulated gravity

NASA Astrophysics Data System (ADS)

Marco, R.; Leandro-García, L. J.; Benguría, A.; Herranz, R.; Zeballos, A.; Gassert, G.; van Loon, J. J.; Medina, F. J.

Establishing the extent and significance of the effects of the exposure to microgravity of complex living organisms is a critical piece of information if the long-term exploration of near-by planets involving human beings is going to take place in the Future As a first step in this direction we have started to look into the patterns of gene expression during Drosophila development in real and simulated microgravity using microarray analysis of mRNA isolated from samples exposed to different environmental conditions In these experiments we used Affymetrix chips version 1 0 containing probes for more than 14 000 genes almost the complete Drosophila genome 55 of which are tagged with some molecular or functional designation while 45 are still waiting to be identified in functional terms The real microgravity exposure was imposed on the samples during the crew exchanging Soyuz 8 Mission to the ISS in October 2003 when after 11 days in Microgravity the Spanish-born astronaut Pedro Duque returned in the Soyuz 7 capsule carrying the experiments prepared by our Team Due to the constraints in the current ISS experiments in these Missions we limited the stages explored in our experiment to the developmental processes occurring during Drosophila metamorphosis As the experimental conditions at the launch site Baikonour were fairly limited we prepared the experiment in Madrid Toulouse and transp o rted the samples at 15 C in a temperature controlled container to slow down the developmental process a

Characterization of p38 MAPK isoforms for drug resistance study using systems biology approach.

PubMed

Peng, Huiming; Peng, Tao; Wen, Jianguo; Engler, David A; Matsunami, Risë K; Su, Jing; Zhang, Le; Chang, Chung-Che Jeff; Zhou, Xiaobo

2014-07-01

p38 mitogen-activated protein kinase activation plays an important role in resistance to chemotherapeutic cytotoxic drugs in treating multiple myeloma (MM). However, how the p38 mitogen-activated protein kinase signaling pathway is involved in drug resistance, in particular the roles that the various p38 isoforms play, remains largely unknown. To explore the underlying mechanisms, we developed a novel systems biology approach by integrating liquid chromatography-mass spectrometry and reverse phase protein array data from human MM cell lines with computational pathway models in which the unknown parameters were inferred using a proposed novel algorithm called modularized factor graph. New mechanisms predicted by our models suggest that combined activation of various p38 isoforms may result in drug resistance in MM via regulating the related pathways including extracellular signal-regulated kinase (ERK) pathway and NFкB pathway. ERK pathway regulating cell growth is synergistically regulated by p38δ isoform, whereas nuclear factor kappa B (NFкB) pathway regulating cell apoptosis is synergistically regulated by p38α isoform. This finding that p38δ isoform promotes the phosphorylation of ERK1/2 in MM cells treated with bortezomib was validated by western blotting. Based on the predicted mechanisms, we further screened drug combinations in silico and found that a promising drug combination targeting ERK1/2 and NFκB might reduce the effects of drug resistance in MM cells. This study provides a framework of a systems biology approach to studying drug resistance and drug combination selection. RPPA experimental Data and Matlab source codes of modularized factor graph for parameter estimation are freely available online at http://ctsb.is.wfubmc.edu/publications/modularized-factor-graph.php. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Mental Retardation Genes in Drosophila: New Approaches to Understanding and Treating Developmental Brain Disorders

ERIC Educational Resources Information Center

Restifo, Linda L.

2005-01-01

"Drosophila melanogaster" is emerging as a valuable genetic model system for the study of mental retardation (MR). MR genes are remarkably similar between humans and fruit flies. Cognitive behavioral assays can detect reductions in learning and memory in flies with mutations in MR genes. Neuroanatomical methods, including some at single-neuron…

Evolution under monogamy feminizes gene expression in Drosophila melanogaster.

PubMed

Hollis, Brian; Houle, David; Yan, Zheng; Kawecki, Tadeusz J; Keller, Laurent

2014-03-18

Many genes have evolved sexually dimorphic expression as a consequence of divergent selection on males and females. However, because the sexes share a genome, the extent to which evolution can shape gene expression independently in each sex is controversial. Here, we use experimental evolution to reveal suboptimal sex-specific expression for much of the genome. By enforcing a monogamous mating system in populations of Drosophila melanogaster for over 100 generations, we eliminated major components of selection on males: female choice and male-male competition. If gene expression is subject to sexually antagonistic selection, relaxed selection on males should cause evolution towards female optima. Monogamous males and females show this pattern of feminization in both the whole-body and head transcriptomes. Genes with male-biased expression patterns evolved decreased expression under monogamy, while genes with female-biased expression evolved increased expression, relative to polygamous populations. Our results demonstrate persistent and widespread evolutionary tension between male and female adaptation.

The white gene of Drosophila melanogaster encodes a protein with a role in courtship behavior.

PubMed

Anaka, Matthew; MacDonald, C Danielle; Barkova, Eva; Simon, Karl; Rostom, Reem; Godoy, Ruth A; Haigh, Andrew J; Meinertzhagen, Ian A; Lloyd, Vett

2008-01-01

The white gene of Drosophila melanogaster has been extensively studied, yet it is still not understood how its ectopic overexpression induces male-male courtship. To investigate the cellular basis of this behavior, we examined the sexual behavior of several classes of mutants. We find that male-male courtship is seen not only in flies overexpressing the white gene, but also in mutants expected to have mislocalized White protein. This finding confirms that mislocalizing White transporter in the cells in which it is normally expressed will produce male-male courtship behaviors; the courtship behavior is not an indirect consequence of aberrant physiological changes elsewhere in the body. Male-male courtship is also seen in some mutants with altered monoamine metabolism and deficits in learning and memory, but can be distinguished from that produced by White mislocalization by its reduced intensity and locomotor activity. Double mutants overexpressing white and with mutations in genes for serotonergic neurons suggest that male-male courtship produced by mislocalizing White may not be mediated exclusively by serotonergic neurons. We also find decreased olfactory learning in white mutants and in individuals with mutations in the genes for White's binding partners, brown and scarlet. Finally, in cultured Drosophila and mammalian cells, the White transporter is found in the endosomal compartment. The additional genes identified here as being involved in male-male courtship increase the repertoire of mutations available to study sexual behavior in Drosophila.

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

PubMed

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.

Polydatin ameliorates Staphylococcus aureus-induced mastitis in mice via inhibiting TLR2-mediated activation of the p38 MAPK/NF-κB pathway.

PubMed

Jiang, Kang-Feng; Zhao, Gan; Deng, Gan-Zhen; Wu, Hai-Chong; Yin, Nan-Nan; Chen, Xiu-Ying; Qiu, Chang-Wei; Peng, Xiu-Li

2017-02-01

Recent studies show that Polydatin (PD) extracted from the roots of Polygonum cuspidatum Sieb, a widely used traditional Chinese remedies, possesses anti-inflammatory activity in several experimental models. In this study, we investigated the anti-inflammatory effects of PD on Staphylococcus aureus-induced mastitis in mice and elucidated the potential mechanisms. In mice with S aureus-induced mastitis, administration of PD (15, 30, 45 mg/kg, ip) or dexamethasone (Dex, 5 mg/kg, ip) significantly suppressed the infiltration of inflammatory cells, ameliorated the mammary structural damage, and inhibited the activity of myeloperoxidase, a biomarker of neutrophils accumulation. Furthermore, PD treatment dose-dependently decreased the levels of TNF-α, IL-1β, IL-6 and IL-8 in the mammary gland tissues. PD treatment also dose-dependently decreased the expression of TLR2, MyD88, IRAK1, IRAK4 and TRAF6 as well as the phosphorylation of TAK1, MKK3/6, p38 MAPK, IκB-α and NF-κB in the mammary gland tissues. In mouse mammary epithelial cells (mMECs) infected by S aureus in vitro, pretreatment with PD dose-dependently suppressed the upregulated pro-inflammatory cytokines and signaling proteins, and the nuclear translocation of NF-κB p65 and AP-1. A TLR2-neutralizing antibody mimicked PD in its suppression on S aureus-induced upregulation of MyD88, p-p38 and p-p65 levels in mMECs. PD (50, 100 μg/mL) affected neither the growth of S aureus in vitro, nor the viability of mMECs. In conclusion, PD does not exhibit antibacterial activity against S aureus, its therapeutic effects in mouse S aureus-induced mastitis depend on its ability to down-regulate pro-inflammatory cytokine levels via inhibiting TLR2-mediated activation of the p38 MAPK/NF-κB signaling pathway.

Balancing selection on immunity genes: review of the current literature and new analysis in Drosophila melanogaster.

PubMed

Croze, Myriam; Živković, Daniel; Stephan, Wolfgang; Hutter, Stephan

2016-08-01

Balancing selection has been widely assumed to be an important evolutionary force, yet even today little is known about its abundance and its impact on the patterns of genetic diversity. Several studies have shown examples of balancing selection in humans, plants or parasites, and many genes under balancing selection are involved in immunity. It has been proposed that host-parasite coevolution is one of the main forces driving immune genes to evolve under balancing selection. In this paper, we review the literature on balancing selection on immunity genes in several organisms, including Drosophila. Furthermore, we performed a genome scan for balancing selection in an African population of Drosophila melanogaster using coalescent simulations of a demographic model with and without selection. We find very few genes under balancing selection and only one novel candidate gene related to immunity. Finally, we discuss the possible causes of the low number of genes under balancing selection. Copyright © 2016 The Authors. Published by Elsevier GmbH.. All rights reserved.

Targeted sequencing identifies 91 neurodevelopmental disorder risk genes with autism and developmental disability biases

PubMed Central

Stessman, Holly A. F.; Xiong, Bo; Coe, Bradley P.; Wang, Tianyun; Hoekzema, Kendra; Fenckova, Michaela; Kvarnung, Malin; Gerdts, Jennifer; Trinh, Sandy; Cosemans, Nele; Vives, Laura; Lin, Janice; Turner, Tychele N.; Santen, Gijs; Ruivenkamp, Claudia; Kriek, Marjolein; van Haeringen, Arie; Aten, Emmelien; Friend, Kathryn; Liebelt, Jan; Barnett, Christopher; Haan, Eric; Shaw, Marie; Gecz, Jozef; Anderlid, Britt-Marie; Nordgren, Ann; Lindstrand, Anna; Schwartz, Charles; Kooy, R. Frank; Vandeweyer, Geert; Helsmoortel, Celine; Romano, Corrado; Alberti, Antonino; Vinci, Mirella; Avola, Emanuela; Giusto, Stefania; Courchesne, Eric; Pramparo, Tiziano; Pierce, Karen; Nalabolu, Srinivasa; Amaral, David; Scheffer, Ingrid E.; Delatycki, Martin B.; Lockhart, Paul J.; Hormozdiari, Fereydoun; Harich, Benjamin; Castells-Nobau, Anna; Xia, Kun; Peeters, Hilde; Nordenskjöld, Magnus; Schenck, Annette; Bernier, Raphael A.; Eichler, Evan E.

2017-01-01

Gene-disruptive mutations contribute to the biology of neurodevelopmental disorders (NDDs), but most pathogenic genes are not known. We sequenced 208 candidate genes from >11,730 patients and >2,867 controls. We report 91 genes with an excess of de novo mutations or private disruptive mutations in 5.7% of patients, including 38 novel NDD genes. Drosophila functional assays of a subset bolster their involvement in NDDs. We identify 25 genes that show a bias for autism versus intellectual disability and highlight a network associated with high-functioning autism (FSIQ>100). Clinical follow-up for NAA15, KMT5B, and ASH1L reveals novel syndromic and non-syndromic forms of disease. PMID:28191889

p75 Neurotrophin Receptor Signaling Activates Sterol Regulatory Element-binding Protein-2 in Hepatocyte Cells via p38 Mitogen-activated Protein Kinase and Caspase-3.

PubMed

Pham, Dan Duc; Do, Hai Thi; Bruelle, Céline; Kukkonen, Jyrki P; Eriksson, Ove; Mogollón, Isabel; Korhonen, Laura T; Arumäe, Urmas; Lindholm, Dan

2016-05-13

Nerve growth factor (NGF) influences the survival and differentiation of a specific population of neurons during development, but its role in non-neuronal cells has been less studied. We observed here that NGF and its pro-form, pro-NGF, are elevated in fatty livers from leptin-deficient mice compared with controls, concomitant with an increase in low density lipoprotein receptors (LDLRs). Stimulation of mouse primary hepatocytes with NGF or pro-NGF increased LDLR expression through the p75 neurotrophin receptor (p75NTR). Studies using Huh7 human hepatocyte cells showed that the neurotrophins activate the sterol regulatory element-binding protein-2 (SREBP2) that regulates genes involved in lipid metabolism. The mechanisms for this were related to stimulation of p38 mitogen-activated protein kinase (p38 MAPK) and activation of caspase-3 and SREBP2 cleavage following NGF and pro-NGF stimulations. Cell fractionation experiments showed that caspase-3 activity was increased particularly in the membrane fraction that harbors SREBP2 and caspase-2. Experiments showed further that caspase-2 interacts with pro-caspase-3 and that p38 MAPK reduced this interaction and caused caspase-3 activation. Because of the increased caspase-3 activity, the cells did not undergo cell death following p75NTR stimulation, possibly due to concomitant activation of nuclear factor-κB (NF-κB) pathway by the neurotrophins. These results identify a novel signaling pathway triggered by ligand-activated p75NTR that via p38 MAPK and caspase-3 mediate the activation of SREBP2. This pathway may regulate LDLRs and lipid uptake particularly after injury or during tissue inflammation accompanied by an increased production of growth factors, including NGF and pro-NGF. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

The Genetic and Molecular Organization of the Dopa Decarboxylase Gene Cluster of Drosophila Melanogaster

PubMed Central

Stathakis, D. G.; Pentz, E. S.; Freeman, M. E.; Kullman, J.; Hankins, G. R.; Pearlson, N. J.; Wright, TRF.

1995-01-01

We report the complete molecular organization of the Dopa decarboxylase gene cluster. Mutagenesis screens recovered 77 new Df(2L)TW130 recessive lethal mutations. These new alleles combined with 263 previously isolated mutations in the cluster to define 18 essential genes. In addition, seven new deficiencies were isolated and characterized. Deficiency mapping, restriction fragment length polymorphism (RFLP) analysis and P-element-mediated germline transformation experiments determined the gene order for all 18 loci. Genomic and cDNA restriction endonuclease mapping, Northern blot analysis and DNA sequencing provided information on exact gene location, mRNA size and transcriptional direction for most of these loci. In addition, this analysis identified two transcription units that had not previously been identified by extensive mutagenesis screening. Most of the loci are contained within two dense subclusters. We discuss the effectiveness of mutagens and strategies used in our screens, the variable mutability of loci within the genome of Drosophila melanogaster, the cytological and molecular organization of the Ddc gene cluster, the validity of the one band-one gene hypothesis and a possible purpose for the clustering of genes in the Ddc region. PMID:8647399

The maternal genes Ci-p53/p73-a and Ci-p53/p73-b regulate zygotic ZicL expression and notochord differentiation in Ciona intestinalis embryos.

PubMed

Noda, Takeshi

2011-12-01

I isolated a Ciona intestinalis homolog of p53, Ci-p53/p73-a, in a microarray screen of rapidly degraded maternal mRNA by comparing the transcriptomes of unfertilized eggs and 32-cell stage embryos. Higher expression of the gene in eggs and lower expression in later embryonic stages were confirmed by whole-mount in situ hybridization (WISH) and quantitative reverse transcription-PCR (qRT-PCR); expression was ubiquitous in eggs and early embryos. Knockdown of Ci-p53/p73-a by injection of antisense morpholino oligonucleotides (MOs) severely perturbed gastrulation cell movements and expression of notochord marker genes. A key regulator of notochord differentiation in Ciona embryos is Brachyury (Ci-Bra), which is directly activated by a zic-like gene (Ci-ZicL). The expression of Ci-ZicL and Ci-Bra in A-line notochord precursors was downregulated in Ci-p53/p73-a knockdown embryos. Maternal expression of Ci-p53/p73-b, a homolog of Ci-p53/p73-a, was also detected. In Ci-p53/p73-b knockdown embryos, gastrulation cell movements, expression of Ci-ZicL and Ci-Bra in A-line notochord precursors, and expression of notochord marker gene at later stages were perturbed. The upstream region of Ci-ZicL contains putative p53-binding sites. Cis-regulatory analysis of Ci-ZicL showed that these sites are involved in expression of Ci-ZicL in A-line notochord precursors at the 32-cell and early gastrula stages. These results suggest that p53 genes are maternal factors that play a crucial role in A-line notochord differentiation in C. intestinalis embryos by regulating Ci-ZicL expression. Copyright © 2011 Elsevier Inc. All rights reserved.

Baicalin plays an anti-inflammatory role through reducing nuclear factor-κB and p38 phosphorylation in S. aureus-induced mastitis.

PubMed

Guo, Mengyao; Zhang, Naisheng; Li, Depeng; Liang, Dejie; Liu, Zhicheng; Li, Fenyang; Fu, Yunhe; Cao, Yongguo; Deng, Xuming; Yang, Zhengtao

2013-06-01

Mastitis is an inflammatory disease caused by microbial infection. Staphylococcus aureus is the major etiological microorganism responsible for both clinical and subclinical mastitis in dairy cows. A mouse model of S. aureus mastitis is available. Baicalin is isolated from Scutellaria and is known to have anti-inflammatory properties. This study was designed to evaluate the effects of baicalin in S. aureus mastitis. In the present study, the mouse model was infected with S. aureus to cause mammary gland inflammation. Baicalin treatment was administered from 6h until 24h after infection. Baicalin significantly attenuated inflammatory cell infiltration and decreased levels of TNF-α, IL-β, and IL-6. Further studies revealed that baicalin downregulated phosphorylation of NF-κB and p38 in the mammary gland with S. aureus mastitis. Our results demonstrated that baicalin reduced the expression of the proinflammatory cytokines TNF-α, IL-β, and IL-6 by inhibiting NF-κB and p38 phosphorylation and mRNA expression. Copyright © 2013 Elsevier B.V. All rights reserved.

The Goddard and Saturn Genes Are Essential for Drosophila Male Fertility and May Have Arisen De Novo.

PubMed

Gubala, Anna M; Schmitz, Jonathan F; Kearns, Michael J; Vinh, Tery T; Bornberg-Bauer, Erich; Wolfner, Mariana F; Findlay, Geoffrey D

2017-05-01

New genes arise through a variety of mechanisms, including the duplication of existing genes and the de novo birth of genes from noncoding DNA sequences. While there are numerous examples of duplicated genes with important functional roles, the functions of de novo genes remain largely unexplored. Many newly evolved genes are expressed in the male reproductive tract, suggesting that these evolutionary innovations may provide advantages to males experiencing sexual selection. Using testis-specific RNA interference, we screened 11 putative de novo genes in Drosophila melanogaster for effects on male fertility and identified two, goddard and saturn, that are essential for spermatogenesis and sperm function. Goddard knockdown (KD) males fail to produce mature sperm, while saturn KD males produce few sperm, and these function inefficiently once transferred to females. Consistent with a de novo origin, both genes are identifiable only in Drosophila and are predicted to encode proteins with no sequence similarity to any annotated protein. However, since high levels of divergence prevented the unambiguous identification of the noncoding sequences from which each gene arose, we consider goddard and saturn to be putative de novo genes. Within Drosophila, both genes have been lost in certain lineages, but show conserved, male-specific patterns of expression in the species in which they are found. Goddard is consistently found in single-copy and evolves under purifying selection. In contrast, saturn has diversified through gene duplication and positive selection. These data suggest that de novo genes can acquire essential roles in male reproduction. © The Author 2017. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Drosophila mitochondrial DNA: a novel gene order.

PubMed Central

Clary, D O; Goddard, J M; Martin, S C; Fauron, C M; Wolstenholme, D R

1982-01-01

Part of the replication origin-containing A+T-rich region of the Drosophila yakuba mtDNA molecule and segments on either side of this region have been sequenced, and the genes within them identified. The data confirm that the small and large rRNA genes lie in tandem adjacent to that side of the A+T-rich region which is replicated first, and establish that a tRNAval gene lies between the two rRNA genes and that URF1 follows the large rRNA gene. The data further establish that the genes for tRNAile, tRNAgln, tRNAf-met and URF2 lie in the order given, on the opposite side of the A+T-rich region to the rRNA genes and, except for tRNAgln, are contained in the opposite strand to the rRNA, tRNAval and URF1 genes. This is in contrast to mammalian mtDNAs where all of these genes are located on the side of the replication origin which is replicated last, within the order tRNAphe, small (12S) rRNA, tRNAval, large (16S) rRNA, tRNAleu, URF1, tRNAile, tRNAgln, tRNAf-met and URF2, and, except tRNAgln, are all contained in the same (H) strand. In D. yakuba URF1 and URF2, the triplet AGA appears to specify an amino acid, which is again different from the situation found in mammalian mtDNAs, where AGA is used only as a rare termination codon. PMID:6294611

The RB-related gene Rb2/p130 in neuroblastoma differentiation and in B-myb promoter down-regulation.

PubMed

Raschellà, G; Tanno, B; Bonetto, F; Negroni, A; Claudio, P P; Baldi, A; Amendola, R; Calabretta, B; Giordano, A; Paggi, M G

1998-05-01

The retinoblastoma family of nuclear factors is composed of RB, the prototype of the tumour suppressor genes and of the strictly related genes p107 and Rb2/p130. The three genes code for proteins, namely pRb, p107 and pRb2/p130, that share similar structures and functions. These proteins are expressed, often simultaneously, in many cell types and are involved in the regulation of proliferation and differentiation. We determined the expression and the phosphorylation of the RB family gene products during the DMSO-induced differentiation of the N1E-115 murine neuroblastoma cells. In this system, pRb2/p130 was strongly up-regulated during mid-late differentiation stages, while, on the contrary, pRb and p107 resulted markedly decreased at late stages. Differentiating N1E-115 cells also showed a progressive decrease in B-myb levels, a proliferation-related protein whose constitutive expression inhibits neuronal differentiation. Transfection of each of the RB family genes in these cells was able, at different degrees, to induce neuronal differentiation, to inhibit [3H]thymidine incorporation and to down-regulate the activity of the B-myb promoter.

A regulatory circuit for piwi by the large Maf gene traffic jam in Drosophila.

PubMed

Saito, Kuniaki; Inagaki, Sachi; Mituyama, Toutai; Kawamura, Yoshinori; Ono, Yukiteru; Sakota, Eri; Kotani, Hazuki; Asai, Kiyoshi; Siomi, Haruhiko; Siomi, Mikiko C

2009-10-29

PIWI-interacting RNAs (piRNAs) silence retrotransposons in Drosophila germ lines by associating with the PIWI proteins Argonaute 3 (AGO3), Aubergine (Aub) and Piwi. piRNAs in Drosophila are produced from intergenic repetitive genes and piRNA clusters by two systems: the primary processing pathway and the amplification loop. The amplification loop occurs in a Dicer-independent, PIWI-Slicer-dependent manner. However, primary piRNA processing remains elusive. Here we analysed piRNA processing in a Drosophila ovarian somatic cell line where Piwi, but not Aub or AGO3, is expressed; thus, only the primary piRNAs exist. In addition to flamenco, a Piwi-specific piRNA cluster, traffic jam (tj), a large Maf gene, was determined as a new piRNA cluster. piRNAs arising from tj correspond to the untranslated regions of tj messenger RNA and are sense-oriented. piRNA loading on to Piwi may occur in the cytoplasm. zucchini, a gene encoding a putative cytoplasmic nuclease, is required for tj-derived piRNA production. In tj and piwi mutant ovaries, somatic cells fail to intermingle with germ cells and Fasciclin III is overexpressed. Loss of tj abolishes Piwi expression in gonadal somatic cells. Thus, in gonadal somatic cells, tj gives rise simultaneously to two different molecules: the TJ protein, which activates Piwi expression, and piRNAs, which define the Piwi targets for silencing.

Gene Model Annotations for Drosophila melanogaster: The Rule-Benders

PubMed Central

Crosby, Madeline A.; Gramates, L. Sian; dos Santos, Gilberto; Matthews, Beverley B.; St. Pierre, Susan E.; Zhou, Pinglei; Schroeder, Andrew J.; Falls, Kathleen; Emmert, David B.; Russo, Susan M.; Gelbart, William M.

2015-01-01

In the context of the FlyBase annotated gene models in Drosophila melanogaster, we describe the many exceptional cases we have curated from the literature or identified in the course of FlyBase analysis. These range from atypical but common examples such as dicistronic and polycistronic transcripts, noncanonical splices, trans-spliced transcripts, noncanonical translation starts, and stop-codon readthroughs, to single exceptional cases such as ribosomal frameshifting and HAC1-type intron processing. In FlyBase, exceptional genes and transcripts are flagged with Sequence Ontology terms and/or standardized comments. Because some of the rule-benders create problems for handlers of high-throughput data, we discuss plans for flagging these cases in bulk data downloads. PMID:26109356

The MAP kinase pathway is involved in odontoblast stimulation via p38 phosphorylation.

PubMed

Simon, Stephane; Smith, Anthony J; Berdal, Ariane; Lumley, Philip J; Cooper, Paul R

2010-02-01

We have previously shown that the p38 gene is highly expressed in odontoblasts during active primary dentinogenesis, but is drastically down-regulated as cells become quiescent in secondary dentinogenesis. Based on these observations, we hypothesized that p38 expression might be upregulated, and the protein activated by phosphorylation, when odontoblasts are stimulated such as during tertiary reactionary dentinogenesis. We stimulated immortalized, odontoblast-like MDPC-23 cells, alone or in combination, with heat-inactivated Streptococcus mutans, EDTA-extracted dentine matrix proteins (DMPs), or growth factors, including transforming growth factor (TGF)-beta1, tumor necrosis factor-alpha (TNF-alpha), and adrenomedullin (ADM). We used ELISA to measure the resulting phosphorylation of the p38 protein, as well as its degree of nuclear translocation. Our results suggest that the p38-MAPKinase pathway is activated during odontoblast stimulation in tertiary dentinogenesis by both p38 phosphorylation and enhanced nuclear translocation. Data indicate that odontoblast behaviour therefore potentially recapitulates that during active primary dentinogenesis. Copyright 2010 American Association of Endodontists. All rights reserved.

The impact of trisomy 12, retinoblastoma gene and P53 in prognosis of B-cell chronic lymphocytic leukemia.

PubMed

AbdelSalam, M; El Sissy, A; Samra, M A; Ibrahim, S; El Markaby, D; Gadallah, F

2008-06-01

Routine cytogenetic analysis frequently fails to identify an abnormal clone in B-cell lymphocytic leukaemia (B-CLL) due to poor response to mitogen stimulation. Fluorescence in situ hybridization (FISH) suggest that chromosomal abnormalities occur more frequently, most commonly trisomy 12, retinoblastoma gene deletion (Rb1 gene) and P53 gene deletion. 30 patients with B-CLL were enrolled in the trial from two centers in Cairo, Egypt during the period May 2000 to January 2002. Karyotyping and FISH assessment for possible chromosomal abnormalities (trisomy 12, Rb1 gene and P53 gene) were done at initial diagnosis. Results of cytogenetic abnormalities were correlated with clinical picture and survival. The median age was 57.4 years (range 40-75). Karyotyping technique showed that no metaphase could be detected in 30%, metaphase with normal karyotyping was observed in 63% and cytogenetic abnormalities were detected in two cases (one trisomy 12 and one deletion in chromosome 13). FISH examination of interphase and metaphase nuclei revealed cytogenetic abnormalities in 15 cases (50%), trisomy 12 in 9 (30%), Rb1 gene deletion in 5 (17%) and P53 gene deletion in 3. At diagnosis, patients with trisomy 12 were significantly associated with advanced stage and absolute lymphocyte count of >or=30,000/mm(3). Univariate analysis showed that absolute lymphocyte count >or=30,000/mm(3) (p=0.004) and trisomy 12 (p=0.024) were associated with poor progression free survival. Interphase and metaphase FISH studies improve the cytogenetic diagnosis of chromosomal abnormalities in B-CLL. Lymphocytosis and trisomy 12 may be a good indicator of poor prognosis.

The slowpoke gene is necessary for rapid ethanol tolerance in Drosophila.

PubMed

Cowmeadow, R B; Krishnan, H R; Atkinson, N S

2005-10-01

Ethanol is one of the most commonly used drugs in the world. We are interested in the compensatory mechanisms used by the nervous system to counter the effects of ethanol intoxication. Recently, the slowpoke BK-type calcium-activated potassium channel gene has been shown to be involved in ethanol sensitivity in Caenorhabditis elegans and in rapid tolerance to the anesthetic benzyl alcohol in Drosophila. We used Drosophila mutants to investigate the role of slowpoke in rapid tolerance to sedation with ethanol vapor. Rapid tolerance was defined as a reduction in the sedative phase caused by a single previous sedation. The ethanol and water contents of flies were measured to determine if pharmacodynamic changes could account for tolerance. A saturated ethanol air stream caused sedation in <20 min and resulted in rapid tolerance that was apparent 4 hr after sedation. Two independently isolated null mutations in the slowpoke gene eliminated the capacity for tolerance. In addition, a third mutation that blocked expression specifically in the nervous system also blocked rapid tolerance. Water measurements showed that both ethanol and mock sedation caused equivalent dehydration. Furthermore, a single prior exposure to ethanol did not cause a change in the ethanol clearance rate. Rapid tolerance, measured as a reduction in the duration of sedation, is a pharmacokinetic response to ethanol that does not occur without slowpoke expression in the nervous system in Drosophila. The slowpoke channel must be involved in triggering or producing a homeostatic mechanism that opposes the sedative effects of ethanol.

ISA virus regulates the generation of reactive oxygen species and p47phox expression in a p38 MAPK-dependent manner in Salmo salar.

PubMed

Olavarría, Víctor H; Valdivia, Sharin; Salas, Boris; Villalba, Melina; Sandoval, Rodrigo; Oliva, Harold; Valdebenito, Samuel; Yañez, Alejandro

2015-02-01

Several viruses, including Orthomyxovirus, utilize cellular reactive oxygen species (ROS) for viral genomic replication and survival within host cells. However, the role of ROS in early events of viral entry and signal induction has not been elucidated. Here, we show that ISA virus (ISAV) induces ROS production very early during infection of CHSE-214 and SHK-1Ycells, and that production is sustained over the observed 24h post-infection. The mitogen-activated protein kinase (MAPK) family is responsible for important signaling pathways. In this study, we report that ISAV activates ERK and p38 in Salmo salar. In salmonid macrophages, while ERK was required for SOD, GLURED, p47phox expression, p38 regulated the ROS production by the NADPH oxidase complex activation. These results, together with the presence of several consensus target motifs for p38 MAPK in the promoter of the S. salar p47phox gene, suggest that p38 MAPK regulates p47phox gene expression in fish through the activation of this key transcription factor. Copyright © 2014 Elsevier Ltd. All rights reserved.

Expression of Genes Involved in Drosophila Wing Morphogenesis and Vein Patterning Are Altered by Spaceflight

NASA Technical Reports Server (NTRS)

Parsons-Wingerter, Patricia A.; Hosamani, Ravikumar; Bhattacharya, Sharmila

2015-01-01

Imaginal wing discs of Drosophila melanogaster (fruit fly) defined during embryogenesis ultimately result in mature wings of stereotyped (specific) venation patterning. Major regulators of wing disc development are the epidermal growth factor receptor (EGF), Notch, Hedgehog (Hh), Wingless (Wg), and Dpp signaling pathways. Highly stereotyped vascular patterning is also characteristic of tissues in other organisms flown in space such as the mouse retina and leaves of Arabidopsis thaliana. Genetic and other adaptations of vascular patterning to space environmental factors have not yet been systematically quantified, despite widespread recognition of their critical importance for terrestrial and microgravity applications. Here we report changes in gene expression with space flight related to Drosophila wing morphogenesis and vein patterning. In addition, genetically modified phenotypes of increasingly abnormal ectopic wing venation in the Drosophila wing1 were analyzed by NASA's VESsel GENeration Analysis (VESGEN) software2. Our goal is to further develop insightful vascular mappings associated with bioinformatic dimensions of genetic or other molecular phenotypes for correlation with genetic and other molecular profiling relevant to NASA's GeneLab and other Space Biology exploration initiatives.

A genomewide survey of basic helix–loop–helix factors in Drosophila

PubMed Central

Moore, Adrian W.; Barbel, Sandra; Jan, Lily Yeh; Jan, Yuh Nung

2000-01-01

The basic helix–loop–helix (bHLH) transcription factors play important roles in the specification of tissue type during the development of animals. We have used the information contained in the recently published genomic sequence of Drosophila melanogaster to identify 12 additional bHLH proteins. By sequence analysis we have assigned these proteins to families defined by Atonal, Hairy-Enhancer of Split, Hand, p48, Mesp, MYC/USF, and the bHLH-Per, Arnt, Sim (PAS) domain. In addition, one single protein represents a unique family of bHLH proteins. mRNA in situ analysis demonstrates that the genes encoding these proteins are expressed in several tissue types but are particularly concentrated in the developing nervous system and mesoderm. PMID:10973473

Transcriptome analysis of a long-lived natural Drosophila variant: a prominent role of stress- and reproduction-genes in lifespan extension

PubMed Central

2012-01-01

Background While studying long-lived mutants has advanced our understanding of the processes involved in ageing, the mechanisms underlying natural variation in lifespan and ageing rate remain largely unknown. Here, we characterise genome-wide expression patterns of a long-lived, natural variant of Drosophila melanogaster resulting from selection for starvation resistance (SR) and compare it with normal-lived control flies (C). We do this at two time points representing middle age (90% survival) and old age (10% survival) respectively, in three adult diets (malnutrition, optimal food, and overfeeding). Results We found profound differences between Drosophila lines in their age-related expression. Most of the age-associated changes in normal-lived flies were abrogated in long-lived Drosophila. The stress-related genes, including those involved in proteolysis and cytochrome P450, were generally higher expressed in SR flies and showed a smaller increase in expression with age compared to C flies. The genes involved in reproduction showed a lower expression in middle-aged SR than in C flies and, unlike C flies, a lack of their downregulation with age. Further, we found that malnutrition strongly affected age-associated transcript patterns overriding the differences between the lines. However, under less stressful dietary conditions, line and diet affected age-dependent expression similarly. Finally, we present lists of candidate markers of ageing and lifespan extension. Conclusions Our study unveils transcriptional changes associated with lifespan extension in SR Drosophila. The results suggest that natural genetic variation for SR and lifespan can operate through similar transcriptional mechanisms as those of dietary restriction and life-extending mutations. PMID:22559237

New Molecular Bridge between RelA/p65 and NF-κB Target Genes via Histone Acetyltransferase TIP60 Cofactor*

PubMed Central

Kim, Jung-Woong; Jang, Sang-Min; Kim, Chul-Hong; An, Joo-Hee; Kang, Eun-Jin; Choi, Kyung-Hee

2012-01-01

The nuclear factor-κB (NF-κB) family is involved in the expressions of numerous genes, in development, apoptosis, inflammatory responses, and oncogenesis. In this study we identified four NF-κB target genes that are modulated by TIP60. We also found that TIP60 interacts with the NF-κB RelA/p65 subunit and increases its transcriptional activity through protein-protein interaction. Although TIP60 binds with RelA/p65 using its histone acetyltransferase domain, TIP60 does not directly acetylate RelA/p65. However, TIP60 maintained acetylated Lys-310 RelA/p65 levels in the TNF-α-dependent NF-κB signaling pathway. In chromatin immunoprecipitation assay, TIP60 was primarily recruited to the IL-6, IL-8, C-IAP1, and XIAP promoters in TNF-α stimulation followed by acetylation of histones H3 and H4. Chromatin remodeling by TIP60 involved the sequential recruitment of acetyl-Lys-310 RelA/p65 to its target gene promoters. Furthermore, we showed that up-regulated TIP60 expression was correlated with acetyl-Lys-310 RelA/p65 expressions in hepatocarcinoma tissues. Taken together these results suggest that TIP60 is involved in the NF-κB pathway through protein interaction with RelA/p65 and that it modulates the transcriptional activity of RelA/p65 in NF-κB-dependent gene expression. PMID:22249179

P38/TRHr-Dependent Regulation of TPO in Thyroid Cells Contributes to the Hypothyroidism of Triclosan-Treated Rats.

PubMed

Zhang, Pei; Yang, Min; Zeng, Li; Liu, Changjiang

2018-01-01

Triclosan, as an antimicrobial agent and a potential endocrine disruptor, has been used extensively in diverse products, resulting in widespread human exposure. In recent years, studies suggest that triclosan could disturb thyroid functions and decline thyroid hormones (THs). To verify our hypothesis that the MAPK pathway may function significantly in triclosan-induced hypothyroidism, Sprague-Dawley rats were gavaged with triclosan for 31 consecutive days; Nthy-ori 3-1 cells were treated with triclosan in the presence/absence of NAC, inhibitors (SB203580 and SB202474), or TRHr siRNA. Tissues and/or cells were analyzed by several techniques including transmission electron microscopy, confocal laser scanning microscopy, gene silencing, western blot, and real-time PCR. Triclosan led to histopathologic changes in the thyroid and decreases in triiodothyronine (T3) and thyroxine (T4). Triclosan stimulated ROS production and oxidative stress occurrence, thereby activating the p38 pathway in vivo and in vitro. Thyrotropin releasing hormone receptor (TRHr) was induced when the p38 pathway was activated, and was suppressed when that pathway was inhibited. Moreover, thyroid peroxidase (TPO) was restrained and modulated by the p38/TRHr pathway after triclosan treatment. Furthermore, deiodinase 3 (D3) and hepatic enzymes (Ugt2b1, CYP1a1, CYP1a2, CYP2b1, CYP3a1, and Sult1e1) were also induced by triclosan. Taken together, p38/TRHr-dependent regulation of TPO in thyroid cells contributes to the hypothyroidism of triclosan-treated rats. © 2018 The Author(s). Published by S. Karger AG, Basel.

TES inhibits colorectal cancer progression through activation of p38

PubMed Central

Gao, Lu; Wang, Lixia; Niu, Yanfeng; Liu, Hongli; Wang, Zheng; Wang, Lin; Wang, Guobin; Wang, Jiliang

2016-01-01

The human TESTIN (TES) gene has been identified as a candidate tumor suppressor based on its location at a common fragile site – a region where loss of heterozygosity has been detected in numerous types of tumors. To investigate its role in colorectal cancer (CRC), we examined TES protein levels in CRC tissue samples and cell lines. We observed that TES was markedly reduced in both CRC tissue and cell lines. Additionally, overexpression of TES significantly inhibited cell proliferation, migration, and invasion, while increasing cell apoptosis in colon cancer cells. By contrast, shRNA-mediated TES knockdown elicited the opposite effects. TES inhibited the progression of CRC by up-regulating pro-apoptotic proteins, down-regulating anti-apoptotic proteins, and simultaneously activating p38 mitogen-activated protein kinase (MAPK) signaling pathways. Collectively, these data indicate that TES functions as a necessary suppressor of CRC progression by activating p38-MAPK signaling pathways. This suggests that TES may have a potential application in CRC diagnosis and targeted gene therapy. PMID:27323777

7 CFR 15b.38 - Health care facilities.

Code of Federal Regulations, 2012 CFR

2012-01-01

... 7 Agriculture 1 2012-01-01 2012-01-01 false Health care facilities. 15b.38 Section 15b.38... ACTIVITIES RECEIVING FEDERAL FINANCIAL ASSISTANCE Other Aid, Benefits, or Services § 15b.38 Health care... impaired. A recipient hospital that provides health services or benefits shall establish a procedure for...

7 CFR 15b.38 - Health care facilities.

Code of Federal Regulations, 2010 CFR

2010-01-01

... 7 Agriculture 1 2010-01-01 2010-01-01 false Health care facilities. 15b.38 Section 15b.38... ACTIVITIES RECEIVING FEDERAL FINANCIAL ASSISTANCE Other Aid, Benefits, or Services § 15b.38 Health care... impaired. A recipient hospital that provides health services or benefits shall establish a procedure for...

7 CFR 15b.38 - Health care facilities.

Code of Federal Regulations, 2011 CFR

2011-01-01

... 7 Agriculture 1 2011-01-01 2011-01-01 false Health care facilities. 15b.38 Section 15b.38... ACTIVITIES RECEIVING FEDERAL FINANCIAL ASSISTANCE Other Aid, Benefits, or Services § 15b.38 Health care... impaired. A recipient hospital that provides health services or benefits shall establish a procedure for...

7 CFR 15b.38 - Health care facilities.

Code of Federal Regulations, 2013 CFR

2013-01-01

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Apoptotic activity and gene responses in Drosophila melanogaster S2 cells, induced by azadirachtin A.

PubMed

Xu, Lin; Li, Sheng; Ran, Xueqin; Liu, Chang; Lin, Rutao; Wang, Jiafu

2016-09-01

Azadirachtin has been used as an antifeedant and growth disruption agent for many insect species. Previous investigations have reported the apoptotic effects of azadirachtin on some insect cells, but the molecular mechanisms are still not clear. This study investigated the underlying molecular mechanisms for the apoptotic effects induced by azadirachtin on Drosophila melanogaster S2 cells in vitro. The results of the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide assay demonstrated that azadirachtin exhibited significant cytotoxicity to S2 cells in a time- and dose-dependent manner. The changes in cellular morphology and the DNA fragmentation demonstrated that azadirachtin induced remarkable apoptosis of S2 cells. Expression levels of 276 genes were found to be significantly changed in S2 cells after exposure to azadirachtin, as detected by Drosophila genome array. Among these genes, calmodulin (CaM) was the most highly upregulated gene. Azadirachtin was further demonstrated to trigger intracellular Ca(2+) release in S2 cells. The genes related to the apoptosis pathway, determined from chip data, were validated by the real-time quantitative polymerase chain reaction method. The results showed that azadirachtin-mediated intracellular Ca(2+) release was the primary event that triggered apoptosis in Drosophila S2 cells through both pathways of the Ca(2+) -CaM and EcR/Usp signalling cascade. © 2015 Society of Chemical Industry. © 2015 Society of Chemical Industry.

Loss of the starvation-induced gene Rack1 leads to glycogen deficiency and impaired autophagic responses in Drosophila.

PubMed

Erdi, Balázs; Nagy, Péter; Zvara, Agnes; Varga, Agnes; Pircs, Karolina; Ménesi, Dalma; Puskás, László G; Juhász, Gábor

2012-07-01

Autophagy delivers cytoplasmic material for lysosomal degradation in eukaryotic cells. Starvation induces high levels of autophagy to promote survival in the lack of nutrients. We compared genome-wide transcriptional profiles of fed and starved control, autophagy-deficient Atg7 and Atg1 null mutant Drosophila larvae to search for novel regulators of autophagy. Genes involved in catabolic processes including autophagy were transcriptionally upregulated in all cases. We also detected repression of genes involved in DNA replication in autophagy mutants compared with control animals. The expression of Rack1 (receptor of activated protein kinase C 1) increased 4.1- to 5.5-fold during nutrient deprivation in all three genotypes. The scaffold protein Rack1 plays a role in a wide range of processes including translation, cell adhesion and migration, cell survival and cancer. Loss of Rack1 led to attenuated autophagic response to starvation, and glycogen stores were decreased 11.8-fold in Rack1 mutant cells. Endogenous Rack1 partially colocalized with GFP-Atg8a and early autophagic structures on the ultrastructural level, suggesting its involvement in autophagosome formation. Endogenous Rack1 also showed a high degree of colocalization with glycogen particles in the larval fat body, and with Shaggy, the Drosophila homolog of glycogen synthase kinase 3B (GSK-3B). Our results, for the first time, demonstrated the fundamental role of Rack1 in autophagy and glycogen synthesis.

Isolation of new polar granule components in Drosophila reveals P body and ER associated proteins

PubMed Central

Thomson, Travis; Liu, Niankun; Arkov, Alexey; Lehmann, Ruth; Lasko, Paul

2008-01-01

Germ plasm, a specialized cytoplasm present at the posterior of the early Drosophila embryo, is necessary and sufficient for germ cell formation. Germ plasm is rich in mitochondria and contains electron dense structures called polar granules. To identify novel polar granule components we isolated proteins that associate in early embryos with Vasa (VAS) and Tudor (TUD), two known polar granule associated molecules. We identified Maternal expression at 31B (ME31B), eIF4A, Aubergine (AUB) and Transitional Endoplasmic Reticulum 94 (TER94) as components of both VAS and TUD complexes and confirmed their localization to polar granules by immuno-electron microscopy. ME31B, eIF4A and AUB are also present in processing (P) bodies, suggesting that polar granules, which are necessary for germ line formation, might be related to P bodies. Our recovery of ER associated proteins TER94 and ME31B confirms that polar granules are closely linked to the translational machinery and to mRNP assembly. PMID:18590813

An essential cell cycle regulation gene causes hybrid inviability in Drosophila

PubMed Central

Phadnis, Nitin; Baker, EmilyClare P.; Cooper, Jacob C.; Frizzell, Kimberly A.; Hsieh, Emily; de la Cruz, Aida Flor A.; Shendure, Jay; Kitzman, Jacob O.; Malik, Harmit S.

2015-01-01

Speciation, the process by which new biological species arise, involves the evolution of reproductive barriers such as hybrid sterility or inviability between populations. However, identifying hybrid incompatibility genes remains a key obstacle in understanding the molecular basis of reproductive isolation. We devised a genomic screen, which identified a cell cycle regulation gene as the cause of male inviability in hybrids between Drosophila melanogaster and D. simulans. Ablation of the D. simulans allele of this gene is sufficient to rescue the adult viability of hybrid males. This dominantly acting cell cycle regulator causes mitotic arrest and, thereby, inviability of male hybrid larvae. Our genomic method provides a facile means to accelerate the identification of hybrid incompatibility genes in other model and non-model systems. PMID:26680200

Identification of the Drosophila eIF4A gene as a target of the DREF transcription factor

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ida, Hiroyuki; Insect Biomedical Research Center, Kyoto Institute of Technology, Matsugasaki, Sakyo-ku, Kyoto 606-8585; Yoshida, Hideki

2007-12-10

The DNA replication-related element-binding factor (DREF) regulates cell proliferation-related gene expression in Drosophila. We have carried out a genetic screening, taking advantage of the rough eye phenotype of transgenic flies that express full-length DREF in the eye imaginal discs and identified the eukaryotic initiation factor 4A (eIF4A) gene as a dominant suppressor of the DREF-induced rough eye phenotype. The eIF4A gene was here found to carry three DRE sequences, DRE1 (- 40 to - 47), DRE2 (- 48 to - 55), and DRE3 (- 267 to - 274) in its promoter region, these all being important for the eIF4A genemore » promoter activity in cultured Drosophila Kc cells and in living flies. Knockdown of DREF in Drosophila S2 cells decreased the eIF4A mRNA level and the eIF4A gene promoter activity. Furthermore, specific binding of DREF to genomic regions containing DRE sequences was demonstrated by chromatin immunoprecipitation assays using anti-DREF antibodies. Band mobility shift assays using Kc cell nuclear extracts revealed that DREF could bind to DRE1 and DRE3 sequences in the eIF4A gene promoter in vitro, but not to the DRE2 sequence. The results suggest that the eIF4A gene is under the control of the DREF pathway and DREF is therefore involved in the regulation of protein synthesis.« less

p38 MAPK-dependent small HSP27 and αB-crystallin phosphorylation in regulation of myocardial function following cardioplegic arrest.

PubMed

Clements, Richard T; Feng, Jun; Cordeiro, Brenda; Bianchi, Cesario; Sellke, Frank W

2011-05-01

We previously demonstrated that myocardial p38 mitogen-activated protein kinase (MAPK) and heat shock protein 27 (HSP27) are phosphorylated following cardioplegic arrest in patients undergoing cardiac surgery and correlate with reduced cardiac function. The following studies were performed to determine whether inhibition of p38 MAPK and/or overexpression of nonphosphorylatable HSP27 improves cardiac function following cardioplegic arrest. Langendorff-perfused isolated rat hearts were subjected to 2 h of intermittent cold cardioplegia followed by 30 min of reperfusion. Hearts were treated with (CP+SB) or without (CP) the p38 MAPK inhibitor SB-203580 (5 μM) supplied in the cardioplegia. Sham-treated hearts served as controls. In separate experiments, isolated rat ventricular myocytes infected with either green fluorescent protein (GFP) or a nonphosphorylatable HSP27 mutant (3A-HSP27) were subjected to 3 h of cold hypoxic cardioplegia and simulated reperfusion (CP) followed by video microscopy and length change measurements. Baseline parameters of cardiac function were similar between groups [left ventricular developed pressure (LVDP), 119 ± 4.9 mmHg; positive and negative first derivatives of LV pressure (± dP/dt), 3,139 ± 245 and 2, 314 ± 110 mmHg/s]. CP resulted in reduced cardiac function (LVDP, 72.2 ± 5.8 mmHg; ± dP/dt, 2,076 ± 231 and -1,317 ± 156 mmHg/s) compared with baseline. Treatment with 5 μM SB-203580 significantly improved CP-induced cardiac function (LVDP, 101.9 ± 0 mmHg; ± dP/dt, 2,836 ± 163 and -2,108 ± 120 mmHg/s; P = 0.03, 0.01, and 0.04, CP+SB vs. CP). Inhibition of p38 MAPK significantly lowered CP-induced p38 MAPK, HSP27, and αB-crystallin (cryAB) phosphorylation. In vitro CP decreased myocyte length changes from 10.3 ± 1.5% (GFP) to 5.7 ± 0.8% (GFP+CP). Infection with 3A-HSP27 completely rescued CP-induced decreased myocyte contraction (11.1 ± 1.0%). However, infection with 3A-HSP27 did not block the endogenous HSP27 response

6-Gingerol protects intestinal barrier from ischemia/reperfusion-induced damage via inhibition of p38 MAPK to NF-κB signalling.

PubMed

Li, Yanli; Xu, Bin; Xu, Ming; Chen, Dapeng; Xiong, Yongjian; Lian, Mengqiao; Sun, Yuchao; Tang, Zeyao; Wang, Li; Jiang, Chunling; Lin, Yuan

2017-05-01

Intestinal ischemia reperfusion (I/R) injury caused by severe trauma, intestinal obstruction, and operation is one of the tough challenges in clinic. 6-Gingerol (6G), a main active ingredient of ginger, is found to have anti-microbial, anti-inflammatory, anti-oxidative, and anti-cancer activities. The present study was designed to characterize the potential protective effects of 6G on rat intestinal I/R injury and reveal the correlated mechanisms. Rat intestinal I/R model was established with clamping the superior mesenteric artery (SMA) and 6G was intragastrically administered for three consecutive days before I/R injury. Caco-2 and IEC-6 cells were incubated under hypoxia/reoxygenation (H/R) conditions to simulate I/R injury in vitro. The results showed that 6G significantly alleviated intestinal injury in I/R injured rats by reducing the generation of oxidative stress and inhibiting p38 MAPK signaling pathway. 6G significantly reduced MDA level and increased the levels of SOD, GSH, and GSH-Px in I/R injured intestinal tissues. 6G significantly decreased the production of proinflammatory cytokines including TNF-α, IL-1β, and IL-6, and inhibited the expression of inflammatory mediators iNOS/NO in I/R injured intestinal tissues. The impaired intestinal barrier function was restored by using 6G in I/R injured rats and in both Caco-2 and IEC-6 cells characterized by inhibiting p38 MAPK phosphorylation, nuclear translocation of NF-κB, and expression of myosin light chain kinase (MLCK) protein. 6G also reduced the generation of reactive oxygen species (ROS) in both Caco-2 and IEC-6 cells. In vitro transfection of p38 MAPK siRNA mitigated the impact of 6G on NF-κB and MLCK expression, and the results further corroborated the protective effects of 6G on intestinal I/R injury by repressing p38 MAPK signaling. In conclusion, the present study suggests that 6G exerts protective effects against I/R-induced intestinal mucosa injury by inhibiting the formation of ROS and p

p62/Sequestosome-1, Autophagy-related Gene 8, and Autophagy in Drosophila Are Regulated by Nuclear Factor Erythroid 2-related Factor 2 (NRF2), Independent of Transcription Factor TFEB.

PubMed

Jain, Ashish; Rusten, Tor Erik; Katheder, Nadja; Elvenes, Julianne; Bruun, Jack-Ansgar; Sjøttem, Eva; Lamark, Trond; Johansen, Terje

2015-06-12

The selective autophagy receptor p62/sequestosome 1 (SQSTM1) interacts directly with LC3 and is involved in oxidative stress signaling in two ways in mammals. First, p62 is transcriptionally induced upon oxidative stress by the NF-E2-related factor 2 (NRF2) by direct binding to an antioxidant response element in the p62 promoter. Second, p62 accumulation, occurring when autophagy is impaired, leads to increased p62 binding to the NRF2 inhibitor KEAP1, resulting in reduced proteasomal turnover of NRF2. This gives chronic oxidative stress signaling through a feed forward loop. Here, we show that the Drosophila p62/SQSTM1 orthologue, Ref(2)P, interacts directly with DmAtg8a via an LC3-interacting region motif, supporting a role for Ref(2)P in selective autophagy. The ref(2)P promoter also contains a functional antioxidant response element that is directly bound by the NRF2 orthologue, CncC, which can induce ref(2)P expression along with the oxidative stress-associated gene gstD1. However, distinct from the situation in mammals, Ref(2)P does not interact directly with DmKeap1 via a KEAP1-interacting region motif; nor does ectopically expressed Ref(2)P or autophagy deficiency activate the oxidative stress response. Instead, DmAtg8a interacts directly with DmKeap1, and DmKeap1 is removed upon programmed autophagy in Drosophila gut cells. Strikingly, CncC induced increased Atg8a levels and autophagy independent of TFEB/MitF in fat body and larval gut tissues. Thus, these results extend the intimate relationship between oxidative stress-sensing NRF2/CncC transcription factors and autophagy and suggest that NRF2/CncC may regulate autophagic activity in other organisms too. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

Topological Constraints on Transvection between White Genes within the Transposing Element Te35b in Drosophila Melanogaster

PubMed Central

Gubb, D.; Roote, J.; Trenear, J.; Coulson, D.; Ashburner, M.

1997-01-01

The transposable element TE35B carries two copies of the white (w) gene at 35B1.2 on the second chromosome. These w genes are suppressed in a zeste-1 (z(1)) mutant background in a synapsis-dependent manner. Single-copy derivatives of the original TE35B stock give red eyes when heterozygous, but zeste eyes when homozygous. TE35B derivatives carrying single, double or triple copies of w were crossed to generate flies carrying from two to five ectopic w genes. Within this range, z(1)-mediated suppression is insensitive to copynumber and does not distinguish between w genes that are in cis or in trans. Suppression does not require the juxtaposition of even numbers of w genes, but is extremely sensitive to chromosomal topology. When arranged in a tight cluster, in triple-copy TE derivatives, w genes are nonsuppressible. Breakpoints falling within TE35B and separating two functional w genes act as partial suppressors of z(1). Similarly, breakpoints immediately proximal or distal to both w genes give partial suppression. This transvection-dependent downregulation of w genes may result from mis-activation of the X-chromosome dosage compensation mechanism. PMID:9215897

Single Nucleotide Polymorphism Markers for Genetic Mapping in Drosophila melanogaster

PubMed Central

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 span the genome. Most of these markers are single nucleotide polymorphisms and sequences for these variants are provided in an accessible format. The average density of the new markers is one per 225 kb on the autosomes and one per megabase on the X chromosome. We include in this survey a set of P-element strains that provide additional use for high-resolution mapping. We show one application of the new markers in a simple set of crosses to map a mutation in the hedgehog gene to an interval of <1 Mb. This new map resource significantly increases the efficiency and resolution of recombination mapping and will be of immediate value to the Drosophila research community. PMID:11381036

Single nucleotide polymorphism markers for genetic mapping in Drosophila melanogaster

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hoskins, Roger A.; Phan, Alexander C.; Naeemuddin, Mohammed

2001-04-16

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 have recently revolutionized human, mouse and plant genetics. Here, we describe the systematic characterization of a dense set of molecular markers in Drosophila using an STS-based physical map of the genome. We identify 474 biallelic markers in standard laboratory strains of Drosophila that the genome. The majority of these markers are single nucleotide polymorphisms (SNPs) and sequences for these variants are provided in an accessible format. The average density of the new markersmore » is 1 marker per 225 kb on the autosomes and 1 marker per 1 Mb on the X chromosome. We include in this survey a set of P-element strains that provide additional utility for high-resolution mapping. We demonstrate one application of the new markers in a simple set of crosses to map a mutation in the hedgehog gene to an interval of <1 Mb. This new map resource significantly increases the efficiency and resolution of recombination mapping and will be of immediate value to the Drosophila research community.« less

Flexible exportation mechanisms of arthrofactin in Pseudomonas sp. MIS38.

PubMed

Lim, S P; Roongsawang, N; Washio, K; Morikawa, M

2009-07-01

To obtain further insights into transportation mechanisms of a most effective biosurfactant, arthrofactin in Pseudomonas sp. MIS38. A cluster genes arfA/B/C encodes an arthrofactin synthetase complex (ArfA/B/C). Downstream of the arfA/B/C lie genes encoding a putative periplasmic protein (ArfD, 362 aa) and a putative ATP-binding cassette transporter (ArfE, 651 aa), namely arfD and arfE, respectively. The arfA/B/C, arfD, and arfE form an operon suggesting their functional connection. Gene knockout mutants ArfD:Km, ArfE:Km, ArfD:Tc/ArfE:Km, and gene overexpression strains MIS38(pME6032_arfD/E) and ArfE:Km(pME6032_arfD/E) were prepared and analysed for arthrofactin production profiles. It was found that the production levels of arthrofactin were temporally reduced in the mutants or increased in the gene overexpression strains, but they eventually became similar level to that of MIS38. Addition of ABC transporter inhibitors, glibenclamide and sodium ortho-vanadate dramatically reduced the production levels of arthrofactin. This excludes a possibility that arthrofactin is exported by diffusion with the aid of its own high surfactant activity. ArfD/E is not an exclusive but a primary exporter of arthrofactin during early growth stage. Reduction in the arthrofactin productivity of arfD and arfE knockout mutants was eventually rescued by another ABC transporter system. Effects of arfD and arfE overexpression were evident only for 1-day cultivation. Multiple ATP dependent active transporter systems are responsible for the production of arthrofactin. Pseudomonas bacteria are characterized to be endued with multiple exporter and efflux systems for secondary metabolites including antibiotics, plant toxins, and biosurfactants. The present work demonstrates exceptionally flexible and highly controlled transportation mechanisms of a most effective lipopeptide biosurfactant, arthrofactin in Pseudomonas sp. MIS38. Because lipopeptide biosurfactants are known to enhance efficacy of

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

PubMed Central

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

2016-01-01

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. PMID:26757981

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

PubMed

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.

S100A8 and S100A9 Promotes Invasion and Migration through p38 Mitogen-Activated Protein Kinase-Dependent NF-κB Activation in Gastric Cancer Cells

PubMed Central

Kwon, Chae Hwa; Moon, Hyun Jung; Park, Hye Ji; Choi, Jin Hwa; Park, Do Youn

2013-01-01

S100A8 and S100A9 (S100A8/A9) are low-molecular weight members of the S100 family of calcium-binding proteins. Recent studies have reported S100A8/A9 promote tumorigenesis. We have previously reported that S100A8/A9 is mostly expressed in stromal cells and inflammatory cells between gastric tumor cells. However, the role of environmental S100A8/A9 in gastric cancer has not been defined. We observed in the present study the effect of S100A8/A9 on migration and invasion of gastric cancer cells. S100A8/A9 treatment increased migration and invasionat lower concentrations that did not affect cell proliferation and cell viability. S100A8/A9 caused activation of p38 mitogen-activated protein kinase (MAPK) and nuclear factor-κB (NF-κB). The phosphorylation of p38 MAPK was not affected by the NF-κB inhibitor Bay whereas activation of NF-κB was blocked by p38 MAPK inhibitor SB203580, indicating that S100A8/A9-induced NF-κB activation is mediated by phosphorylation of p38 MAPK. S100A8/A9-induced cell migration and invasion was inhibited by SB203580 and Bay, suggesting that activation of p38 MAPK and NF-κB is involved in the S100A8/A9 induced cell migration and invasion. S100A8/A9 caused an increase in matrix metalloproteinase 2 (MMP2) and MMP12 expression, which were inhibited by SB203580 and Bay. S100A8/A9-induced cell migration and invasion was inhibited by MMP2 siRNA and MMP12 siRNA, indicating that MMP2 and MMP12 is related to the S100A8/A9 induced cell migration and invasion. Taken together, these results suggest that S100A8/A9 promotes cell migration and invasion through p38 MAPK-dependent NF-κB activation leading to an increase of MMP2 and MMP12 in gastric cancer. PMID:23456298

Reverse engineering the gap gene network of Drosophila melanogaster.

PubMed

Perkins, Theodore J; Jaeger, Johannes; Reinitz, John; Glass, Leon

2006-05-01

A fundamental problem in functional genomics is to determine the structure and dynamics of genetic networks based on expression data. We describe a new strategy for solving this problem and apply it to recently published data on early Drosophila melanogaster development. Our method is orders of magnitude faster than current fitting methods and allows us to fit different types of rules for expressing regulatory relationships. Specifically, we use our approach to fit models using a smooth nonlinear formalism for modeling gene regulation (gene circuits) as well as models using logical rules based on activation and repression thresholds for transcription factors. Our technique also allows us to infer regulatory relationships de novo or to test network structures suggested by the literature. We fit a series of models to test several outstanding questions about gap gene regulation, including regulation of and by hunchback and the role of autoactivation. Based on our modeling results and validation against the experimental literature, we propose a revised network structure for the gap gene system. Interestingly, some relationships in standard textbook models of gap gene regulation appear to be unnecessary for or even inconsistent with the details of gap gene expression during wild-type development.

Emodin Inhibition of Influenza A Virus Replication and Influenza Viral Pneumonia via the Nrf2, TLR4, p38/JNK and NF-kappaB Pathways.

PubMed

Dai, Jian-Ping; Wang, Qian-Wen; Su, Yun; Gu, Li-Ming; Zhao, Ying; Chen, Xiao-Xua; Chen, Cheng; Li, Wei-Zhong; Wang, Ge-Fei; Li, Kang-Sheng

2017-10-18

Lasting activations of toll-like receptors (TLRs), MAPK and NF-κB pathways can support influenza A virus (IAV) infection and promote pneumonia. In this study, we have investigated the effect and mechanism of action of emodin on IAV infection using qRT-PCR, western blotting, ELISA, Nrf2 luciferase reporter, siRNA and plaque inhibition assays. The results showed that emodin could significantly inhibit IAV (ST169, H1N1) replication, reduce IAV-induced expressions of TLR2/3/4/7, MyD88 and TRAF6, decrease IAV-induced phosphorylations of p38/JNK MAPK and nuclear translocation of NF-κB p65. Emodin also activated the Nrf2 pathway, decreased ROS levels, increased GSH levelss and GSH/GSSG ratio, and upregulated the activities of SOD, GR, CAT and GSH-Px after IAV infection. Suppression of Nrf2 via siRNA markedly blocked the inhibitory effects of emodin on IAV-induced activations of TLR4, p38/JNK, and NF-κB pathways and on IAV-induced production of IL-1β, IL-6 and expression of IAV M2 protein. Emodin also dramatically increased the survival rate of mice, reduced lung edema, pulmonary viral titer and inflammatory cytokines, and improved lung histopathological changes. In conclusion, emodin can inhibit IAV replication and influenza viral pneumonia, at least in part, by activating Nrf2 signaling and inhibiting IAV-induced activations of the TLR4, p38/JNK MAPK and NF-κB pathways.

Effect of spaceflight on the circadian rhythm, lifespan and gene expression of Drosophila melanogaster.

PubMed

Ma, Lingling; Ma, Jun; Xu, Kanyan

2015-01-01

Space travelers are reported to experience circadian rhythm disruption during spaceflight. However, how the space environment affects circadian rhythm is yet to be determined. The major focus of this study was to investigate the effect of spaceflight on the Drosophila circadian clock at both the behavioral and molecular level. We used China's Shenzhou-9 spaceship to carry Drosophila. After 13 days of spaceflight, behavior tests showed that the flies maintained normal locomotor activity rhythm and sleep pattern. The expression level and rhythm of major clock genes were also unaffected. However, expression profiling showed differentially regulated output genes of the circadian clock system between space flown and control flies, suggesting that spaceflight affected the circadian output pathway. We also investigated other physiological effects of spaceflight such as lipid metabolism and lifespan, and searched genes significantly affected by spaceflight using microarray analysis. These results provide new information on the effects of spaceflight on circadian rhythm, lipid metabolism and lifespan. Furthermore, we showed that studying the effect of spaceflight on gene expression using samples collected at different Zeitgeber time could obtain different results, suggesting the importance of appropriate sampling procedures in studies on the effects of spaceflight.

Effect of Spaceflight on the Circadian Rhythm, Lifespan and Gene Expression of Drosophila melanogaster

PubMed Central

Xu, Kanyan

2015-01-01

Space travelers are reported to experience circadian rhythm disruption during spaceflight. However, how the space environment affects circadian rhythm is yet to be determined. The major focus of this study was to investigate the effect of spaceflight on the Drosophila circadian clock at both the behavioral and molecular level. We used China’s Shenzhou-9 spaceship to carry Drosophila. After 13 days of spaceflight, behavior tests showed that the flies maintained normal locomotor activity rhythm and sleep pattern. The expression level and rhythm of major clock genes were also unaffected. However, expression profiling showed differentially regulated output genes of the circadian clock system between space flown and control flies, suggesting that spaceflight affected the circadian output pathway. We also investigated other physiological effects of spaceflight such as lipid metabolism and lifespan, and searched genes significantly affected by spaceflight using microarray analysis. These results provide new information on the effects of spaceflight on circadian rhythm, lipid metabolism and lifespan. Furthermore, we showed that studying the effect of spaceflight on gene expression using samples collected at different Zeitgeber time could obtain different results, suggesting the importance of appropriate sampling procedures in studies on the effects of spaceflight. PMID:25798821

Metabolomic Studies in Drosophila.

PubMed

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.

Recurrent Innovation at Genes Required for Telomere Integrity in Drosophila.

PubMed

Lee, Yuh Chwen G; Leek, Courtney; Levine, Mia T

2017-02-01

Telomeres are nucleoprotein complexes at the ends of linear chromosomes. These specialized structures ensure genome integrity and faithful chromosome inheritance. Recurrent addition of repetitive, telomere-specific DNA elements to chromosome ends combats end-attrition, while specialized telomere-associated proteins protect naked, double-stranded chromosome ends from promiscuous repair into end-to-end fusions. Although telomere length homeostasis and end-protection are ubiquitous across eukaryotes, there is sporadic but building evidence that the molecular machinery supporting these essential processes evolves rapidly. Nevertheless, no global analysis of the evolutionary forces that shape these fast-evolving proteins has been performed on any eukaryote. The abundant population and comparative genomic resources of Drosophila melanogaster and its close relatives offer us a unique opportunity to fill this gap. Here we leverage population genetics, molecular evolution, and phylogenomics to define the scope and evolutionary mechanisms driving fast evolution of genes required for telomere integrity. We uncover evidence of pervasive positive selection across multiple evolutionary timescales. We also document prolific expansion, turnover, and expression evolution in gene families founded by telomeric proteins. Motivated by the mutant phenotypes and molecular roles of these fast-evolving genes, we put forward four alternative, but not mutually exclusive, models of intra-genomic conflict that may play out at very termini of eukaryotic chromosomes. Our findings set the stage for investigating both the genetic causes and functional consequences of telomere protein evolution in Drosophila and beyond. © The Author 2016. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

Detection of Volatile Indicators of Illicit Substances by the Olfactory Receptors of Drosophila melanogaster

PubMed Central

Marshall, Brenton; Warr, Coral G.

2010-01-01

Insects can detect a large range of odors with a numerically simple olfactory system that delivers high sensitivity and accurate discrimination. Therefore, insect olfactory receptors hold great promise as biosensors for detection of volatile organic chemicals in a range of applications. The array of olfactory receptor neurons of Drosophila melanogaster is rapidly becoming the best-characterized natural nose. We have investigated the suitability of Drosophila receptors as detectors for volatiles with applications in law enforcement, emergency response, and security. We first characterized responses of the majority of olfactory neuron types to a set of diagnostic odorants. Being thus able to correctly identify neurons, we then screened for responses from 38 different types of neurons to 35 agents. We identified 13 neuron types with responses to 13 agents. As individual Drosophila receptor genes have been mapped to neuron types, we can infer which genes confer responsiveness to the neurons. The responses were confirmed for one receptor by expressing it in a nonresponsive neuron. The fly olfactory system is mainly adapted to detect volatiles from fermenting fruits. However, our findings establish that volatiles associated with illicit substances, many of which are of nonnatural origin, are also detected by Drosophila receptors. PMID:20530374

Relationship between p53 dysfunction, CD38 expression, and IgV(H) mutation in chronic lymphocytic leukemia.

PubMed

Lin, Ke; Sherrington, Paul D; Dennis, Michael; Matrai, Zoltan; Cawley, John C; Pettitt, Andrew R

2002-08-15

Established adverse prognostic factors in chronic lymphocytic leukemia (CLL) include CD38 expression, relative lack of IgV(H) mutation, and defects of the TP53 gene. However, disruption of the p53 pathway can occur through mechanisms other than TP53 mutation, and we have recently developed a simple screening test that detects p53 dysfunction due to mutation of the genes encoding either p53 or ATM, a kinase that regulates p53. The present study was conducted to examine the predictive value of this test and to establish the relationship between p53 dysfunction, CD38 expression, and IgV(H) mutation. CLL cells from 71 patients were examined for IgV(H) mutation, CD38 expression, and p53 dysfunction (detected as an impaired p53/p21 response to ionizing radiation). Survival data obtained from 69 patients were analyzed according to each of these parameters. Relative lack of IgV(H) mutation (less than 5%; n = 45), CD38 positivity (antigen expressed on more than 20% of malignant cells; n = 19), and p53 dysfunction (n = 19) were independently confirmed as adverse prognostic factors. Intriguingly, all p53-dysfunctional patients and all but one of the CD38(+) patients had less [corrected] than 5% IgV(H) mutation. Moreover, patients with p53 dysfunction and/or CD38 positivity (n = 31) accounted for the short survival of the less mutated group. These findings indicate that the poor outcome associated with having less than 5% IgV(H) mutation may be due to the overrepresentation of high-risk patients with p53 dysfunction and/or CD38 positivity within this group, and that CD38(-) patients with functionally intact p53 may have a prolonged survival regardless of the extent of IgV(H) mutation.

Introgression in the Drosophila subobscura--D. Madeirensis sister species: evidence of gene flow in nuclear genes despite mitochondrial differentiation.

PubMed

Herrig, Danielle K; Modrick, Alec J; Brud, Evgeny; Llopart, Ana

2014-03-01

Species hybridization, and thus the potential for gene flow, was once viewed as reproductive mistake. However, recent analysis based on large datasets and newly developed models suggest that gene exchange is not as rare as originally suspected. To investigate the history and speciation of the closely related species Drosophila subobscura, D. madeirensis, and D. guanche, we obtained polymorphism and divergence data for 26 regions throughout the genome, including the Y chromosome and mitochondrial DNA. We found that the D. subobscura X/autosome ratio of silent nucleotide diversity is significantly smaller than the 0.75 expected under neutrality. This pattern, if held genomewide, may reflect a faster accumulation of beneficial mutations on the X chromosome than on autosomes. We also detected evidence of gene flow in autosomal regions, while sex chromosomes remain distinct. This is consistent with the large X effect on hybrid male sterility seen in this system and the presence of two X chromosome inversions fixed between species. Overall, our data conform to chromosomal speciation models in which rearrangements are proposed to serve as gene flow barriers. Contrary to other observations in Drosophila, the mitochondrial genome appears resilient to gene flow in the presence of nuclear exchange. © 2013 The Authors. Evolution published by Wiley Periodicals, Inc. on behalf of The Society for the Study of Evolution.

The glutamate dehydrogenase GENE of Drosophila melanogaster: molecular analysis and expression.

PubMed

Papadopoulou, D; Louis, C

2000-09-01

Glutamate dehydrogenase is an enzyme that, in addition to its role in the energy metabolism in mitochondria, is involved in neuromuscular transmission. Here we present the structure and sequence of the Gdh gene of Drosophila melanogaster, as well as the analysis of its spatial and temporal pattern of expression. Unlike all other organisms analyzed so far, two forms of the enzyme, differing by the inclusion of 13 extra amino acids, are found in the fruitfly. We show the presence of Gdh mRNA in several tissues of the developing embryo, including the central nervous system, muscles and the alimentary tract. Moreover, we detect the localization of the Gdh protein in specific areas of the muscles, a fact that is consistent both with an involvement in energy metabolism and the role of glutamate as the major neuromuscular transmitter in Drosophila.

Structure and expression of the rat CYP3A1 gene: isolation of the gene (P450/6betaB) and characterization of the recombinant protein.

PubMed

Nagata, K; Ogino, M; Shimada, M; Miyata, M; Gonzalez, F J; Yamazoe, Y

1999-02-15

A P450 gene (P450/6betaB) of the CYP3A subfamily was isolated from a rat genomic library. Nucleotide sequencing of the exons revealed a high similarity with P450PCN1 cDNA (Gonzalez et al. (1985), J. Biol. Chem. 260, 7345-7441), but differed in 41 nucleotides, resulting in 11 changes and 2 deletions of amino acid residues. The P450/6betaB spanned about 30 kbp and consisted of 13 exons, and was in exon number and size identical with CYP3A2 gene except in the 6th exon, which was shorter than that of CYP3A2. 6beta-B mRNA, which may be transcribed from P450/6betaB, was detected on Northern blotting and by reverse transcription-polymerase chain reaction (RT-PCR). Profiles of the developmental change and induction by a treatment with several chemicals were very similar to those of P450PCN1 mRNA reported previously. P450PCN1 mRNA and gene, however, were not detected by PCR in rats. To determine whether P450/6betaB encodes an active protein, a cDNA was isolated and expressed. Expression of 6beta-B cDNA in COS-1 cells was carried out and revealed that the recombinant protein comigrated with purified P4506beta-4 previously identified as CYP3A1. The recombinant 6beta-B protein showed similar turnover rate and regioselectivity for testosterone with purified P4506beta-4 by the simultaneous addition of NADPH-cytochrome P450 reductase and cytochrome b5. These data suggest that P450/6betaB encodes an active P450 form corresponding to CYP3A1 and P450PCN1 reported previously does not exist in rats. Copyright 1999 Academic Press.

Intratracheal administration of p38α short-hairpin RNA plasmid ameliorates lung ischemia-reperfusion injury in rats.

PubMed

Lv, Xiangqi; Tan, Jing; Liu, Dongdong; Wu, Ping; Cui, Xiaoguang

2012-06-01

Lung ischemia-reperfusion injury (LIRI) remains a significant problem after lung transplantation. A crucial signaling enzyme involved in inflammation and apoptosis during LIRI is p38 mitogen-activated protein kinase (MAPK). Gene silencing of p38α by short hairpin RNA (shRNA) can downregulate p38α expression. The lungs have an extremely large surface area, which makes the absorption of shRNA highly effective. Therefore, we evaluated the therapeutic efficacy of p38α shRNA plasmids in a rat model of lung transplantation. The delivery of p38α shRNA plasmid was performed by intratracheal administration 48 hours before transplantation into donor rats. Control animals received scrambled shRNA plasmids. Reverse-transcription polymerase chain reaction and Western blots were used to assess gene silencing efficacy. The therapeutic effects of shRNA plasmids were evaluated by lung function tests. We determined the levels of inflammatory cytokines, the level of intercellular adhesion molecule 1 (ICAM-1), c-Myc mRNA expression, and ICAM-1 protein expression, and the presence of cell apoptosis. Rats administered p38α shRNA plasmids showed a significant downregulation in lung expression of p38α transcripts and protein levels. Compared with the control group, the p38α shRNA group showed a higher pulmonary vein oxygen level, lower wet weight-to-dry weight ratio, lower lung injury score, and lower serum levels of tumor necrosis factor-α, interleukin-6, and interleukin-8. Messenger RNA levels of ICAM-1 and c-Myc in the p38α shRNA group were dramatically lower than in the control group. Levels of ICAM-1 protein expression exhibited a similar trend. Cell apoptosis decreased in the p38α shRNA group vs the control group. Intratracheal administration of p38α shRNA plasmids provided therapeutic effects in a rat model of lung transplantation. Crown Copyright © 2012. Published by Elsevier Inc. All rights reserved.

Netrin-1 induces the migration of Schwann cells via p38 MAPK and PI3K-Akt signaling pathway mediated by the UNC5B receptor

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lv, Jianwei; Tianjin Institute of Orthopedics in Traditional Chinese and Western Medicine, No. 155, Munan Road, Tianjin 300050; Sun, Xiaolei

2015-08-14

Schwann cells (SCs) play an essentially supportive role in the regeneration of injured peripheral nerve system (PNS). As Netrin-1 is crucial for the normal development of nervous system (NS) and can direct the process of damaged PNS regeneration, our study was designed to determine the role of Netrin-1 in RSC96 Schwann cells (an immortalized rat Schwann cell line) proliferation and migration. Our studies demonstrated that Netrin-1 had no effect on RSC96 cells proliferation, while significantly promoted RSC96 cells migration. The Netrin-1-induced RSC96 cells migration was significantly attenuated by inhibition of p38 and PI3K through pretreatment with SB203580 and LY294002 respectively,more » but not inhibition of MEK1/2 and JNK by U0126-EtOH and SP600125 individually. Treatment with Netrin-1 enhanced the phosphorylation of p38 and Akt. QRT-PCR indicated that Netrin-1 and only its receptors Unc5a, Unc5b and Neogenin were expressed in RSC96 cells, among which Unc5b expressed the most. And UNC5B protein was significantly increased after stimulated by Netrin-1. In conclusion, we show here that Netrin-1-enhanced SCs migration is mediated by activating p38 MAPK and PI3K-Akt signal cascades via receptor UNC5B, which suggests that Netrin-1 could serve as a new therapeutic strategy and has potential application value for PNS regeneration. - Highlights: • Netrin-1 attracts RSC96 Schwann cells migration in a dose dependent manner. • Netrin-1 induced Schwann cells migration is p38 and PI3K-Akt signaling dependent. • UNC5B may be dominant receptor mediating Netrin-1′ effect on RSC96 cells motility. • Netrin-1 may promote peripheral nerve repair by enhancing Schwann cells motility.« less

The Transposable Element Mariner Mediates Germline Transformation in Drosophila Melanogaster

PubMed Central

Lidholm, D. A.; Lohe, A. R.; Hartl, D. L.

1993-01-01

A vector for germline transformation in Drosophila melanogaster was constructed using the transposable element mariner. The vector, denoted pMlwB, contains a mariner element disrupted by an insertion containing the wild-type white gene from D. melanogaster, the β-galactosidase gene from Escherichia coli and sequences that enable plasmid replication and selection in E. coli. The white gene is controlled by the promoter of the D. melanogaster gene for heat-shock protein 70, and the β-galactosidase gene is flanked upstream by the promoter of the transposable element P as well as that of mariner. The MlwB element was introduced into the germline of D. melanogaster by co-injection into embryos with an active mariner element, Mos1, which codes for a functional transposase and serves as a helper. Two independent germline insertions were isolated and characterized. The results show that the MlwB element inserted into the genome in a mariner-dependent manner with the termini of the inverted repeats inserted at a TA dinucleotide. Both insertions exhibit an unexpected degree of germline and somatic stability, even in the presence of an active mariner element in the genetic background. These results demonstrate that the mariner transposable element, which is small (1286 bp) and relatively homogeneous in size among different copies, is nevertheless capable of promoting the insertion of the large (13.2 kb) MlwB element. Because of the widespread phylogenetic distribution of mariner among insects, these results suggest that mariner might provide a wide hostrange transformation vector for insects. PMID:8394264

Molecular and Genetic Characterization of the Drosophila Melanogaster 87e Actin Gene Region

PubMed Central

Manseau, L. J.; Ganetzky, B.; Craig, E. A.

1988-01-01

A combined molecular and genetic analysis of the 87E actin gene (Act87E) in Drosophila melanogaster was undertaken. A clone of Act87E was isolated and characterized. The Act87E transcription unit is 1.57 kb and includes a 556-base intervening sequence in the 5' leader of the gene. The protein-coding region is contiguous and encodes a protein that is >93% identical to the other Drosophila actins. By in situ hybridization with a series of deficiencies that break in 87E, Act87E was localized to a region encompassing one to three faint, polytene chromosome bands. The region between the deficiency endpoints that flank the actin gene was isolated and measures approximately 24-30 kb. The closest proximal deficiency endpoint lies 8-10 kb 5' to the actin gene; the closest distal deficiency endpoint lies 16-20 kb 3' to the actin gene. A single, recessive lethal complementation group lies between the deficiency endpoints that flank the actin gene. An EMS mutagenesis screen produced four additional members of this recessive lethal complementation group. Molecular analysis of the members of this complementation group indicated that two of the newly induced mutations have deletions of approximately 1 kb in a transcribed region 4-5 kb 3' (distal) to the actin gene. This result suggests that the recessive lethal complementation group represents a gene separate from and distal to the actin gene. The mutagenesis screen failed to identify additional recessive lethal complementation groups in the actin gene-containing region. The implications of the failure to identify recessive lethal mutations in the actin gene are discussed in reference to studies of other conserved multigene families and other muscle protein mutations. PMID:2840338

FBI-1 enhances transcription of the nuclear factor-kappaB (NF-kappaB)-responsive E-selectin gene by nuclear localization of the p65 subunit of NF-kappaB.

PubMed

Lee, Dong-Kee; Kang, Jae-Eun; Park, Hye-Jin; Kim, Myung-Hwa; Yim, Tae-Hee; Kim, Jung-Min; Heo, Min-Kyu; Kim, Kyu-Yeun; Kwon, Ho Jeong; Hur, Man-Wook

2005-07-29

The POZ domain is a highly conserved protein-protein interaction motif found in many regulatory proteins. Nuclear factor-kappaB (NF-kappaB) plays a key role in the expression of a variety of genes in response to infection, inflammation, and stressful conditions. We found that the POZ domain of FBI-1 (factor that binds to the inducer of short transcripts of human immunodeficiency virus-1) interacted with the Rel homology domain of the p65 subunit of NF-kappaB in both in vivo and in vitro protein-protein interaction assays. FBI-1 enhanced NF-kappaB-mediated transcription of E-selectin genes in HeLa cells upon phorbol 12-myristate 13-acetate stimulation and overcame gene repression by IkappaB alpha or IkappaB beta. In contrast, the POZ domain of FBI-1, which is a dominant-negative form of FBI-1, repressed NF-kappaB-mediated transcription, and the repression was cooperative with IkappaB alpha or IkappaB beta. In contrast, the POZ domain tagged with a nuclear localization sequence polypeptide of FBI-1 enhanced NF-kappaB-responsive gene transcription, suggesting that the molecular interaction between the POZ domain and the Rel homology domain of p65 and the nuclear localization by the nuclear localization sequence are important in the transcription enhancement mediated by FBI-1. Confocal microscopy showed that FBI-1 increased NF-kappaB movement into the nucleus and increased the stability of NF-kappaB in the nucleus, which enhanced NF-kappaB-mediated transcription of the E-selectin gene. FBI-1 also interacted with IkappaB alpha and IkappaB beta.

Identification of Drosophila melanogaster yellow-f and yellow-f2 proteins as dopachrome-conversion enzymes.

PubMed Central

Han, Qian; Fang, Jianmin; Ding, Haizhen; Johnson, Jody K; Christensen, Bruce M; Li, Jianyong

2002-01-01

This study describes the identification of Drosophila yellow-f and yellow-f2 as dopachrome-conversion enzymes responsible for catalysing the conversion of dopachrome into 5,6-dihydroxyindole in the melanization pathway. Drosophila yellow -y gene and yellow -b, -c, -f and -f2 genes were expressed in an insect cell/baculovirus expression system and their corresponding recombinant proteins were screened for dopachrome-conversion enzyme activity. Among the yellow and yellow -related genes, the yellow -f and yellow -f2 genes were identified as the genes coding for Drosophila dopachrome-conversion enzyme based on the high activity of their recombinant proteins in catalysing the production of 5,6-dihydroxyindole from dopachrome. Both yellow-f and yellow-f2 are capable of mediating a decarboxylative structural rearrangement of dopachrome, as well as an isomerization/tautomerization of dopamine chrome and dopa methyl ester chrome. Northern hybridization revealed the transcription of yellow -f in larvae and pupae, but a high abundance of mRNA was observed in later larval and early pupal stages. In contrast, yellow-f2 transcripts were present at all stages, but high abundance of its mRNA was observed in later-stage pupae and adults. These data indicate that yellow-f and yellow-f2 complement each other during Drosophila development and that the yellow-f is involved in larval and pupal melanization, and yellow-f2 plays a major role in melanization reactions in Drosophila during later pupal and adult development. Results from this study provide the groundwork towards a better understanding of the physiological roles of the Drosophila yellow gene family. PMID:12164780

Mobile phone electromagnetic radiation activates MAPK signaling and regulates viability in Drosophila.

PubMed

Lee, Kyu-Sun; Choi, Jong-Soon; Hong, Sae-Yong; Son, Tae-Ho; Yu, Kweon

2008-07-01

Mobile phones are widely used in the modern world. However, biological effects of electromagnetic radiation produced by mobile phones are largely unknown. In this report, we show biological effects of the mobile phone 835 MHz electromagnetic field (EMF) in the Drosophila model system. When flies were exposed to the specific absorption rate (SAR) 1.6 W/kg, which is the proposed exposure limit by the American National Standards Institute (ANSI), more than 90% of the flies were viable even after the 30 h exposure. However, in the SAR 4.0 W/kg strong EMF exposure, viability dropped from the 12 h exposure. These EMF exposures triggered stress response and increased the production of reactive oxygen species. The EMF exposures also activated extracellular signal regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) signaling, but not p38 kinase signaling. Interestingly, SAR 1.6 W/kg activated mainly ERK signaling and expression of an anti-apoptotic gene, whereas SAR 4.0 W/kg strongly activated JNK signaling and expression of apoptotic genes. In addition, SAR 4.0 W/kg amplified the number of apoptotic cells in the fly brain. These findings demonstrate that the exposure limit on electromagnetic radiation proposed by ANSI triggered ERK-survival signaling but the strong electromagnetic radiation activated JNK-apoptotic signaling in Drosophila.

FlpStop, a tool for conditional gene control in Drosophila

PubMed Central

Fisher, Yvette E; Yang, Helen H; Isaacman-Beck, Jesse; Xie, Marjorie; Gohl, Daryl M; Clandinin, Thomas R

2017-01-01

Manipulating gene function cell type-specifically is a common experimental goal in Drosophila research and has been central to studies of neural development, circuit computation, and behavior. However, current cell type-specific gene disruption techniques in flies often reduce gene activity incompletely or rely on cell division. Here we describe FlpStop, a generalizable tool for conditional gene disruption and rescue in post-mitotic cells. In proof-of-principle experiments, we manipulated apterous, a regulator of wing development. Next, we produced conditional null alleles of Glutamic acid decarboxylase 1 (Gad1) and Resistant to dieldrin (Rdl), genes vital for GABAergic neurotransmission, as well as cacophony (cac) and paralytic (para), voltage-gated ion channels central to neuronal excitability. To demonstrate the utility of this approach, we manipulated cac in a specific visual interneuron type and discovered differential regulation of calcium signals across subcellular compartments. Thus, FlpStop will facilitate investigations into the interactions between genes, circuits, and computation. DOI: http://dx.doi.org/10.7554/eLife.22279.001 PMID:28211790

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

PubMed

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.

Automated update, revision, and quality control of the maize genome annotations using MAKER-P improves the B73 RefGen_v3 gene models and identifies new genes.

PubMed

Law, MeiYee; Childs, Kevin L; Campbell, Michael S; Stein, Joshua C; Olson, Andrew J; Holt, Carson; Panchy, Nicholas; Lei, Jikai; Jiao, Dian; Andorf, Carson M; Lawrence, Carolyn J; Ware, Doreen; Shiu, Shin-Han; Sun, Yanni; Jiang, Ning; Yandell, Mark

2015-01-01

The large size and relative complexity of many plant genomes make creation, quality control, and dissemination of high-quality gene structure annotations challenging. In response, we have developed MAKER-P, a fast and easy-to-use genome annotation engine for plants. Here, we report the use of MAKER-P to update and revise the maize (Zea mays) B73 RefGen_v3 annotation build (5b+) in less than 3 h using the iPlant Cyberinfrastructure. MAKER-P identified and annotated 4,466 additional, well-supported protein-coding genes not present in the 5b+ annotation build, added additional untranslated regions to 1,393 5b+ gene models, identified 2,647 5b+ gene models that lack any supporting evidence (despite the use of large and diverse evidence data sets), identified 104,215 pseudogene fragments, and created an additional 2,522 noncoding gene annotations. We also describe a method for de novo training of MAKER-P for the annotation of newly sequenced grass genomes. Collectively, these results lead to the 6a maize genome annotation and demonstrate the utility of MAKER-P for rapid annotation, management, and quality control of grasses and other difficult-to-annotate plant genomes. © 2015 American Society of Plant Biologists. All Rights Reserved.

The protective effect of lidocaine on lipopolysaccharide-induced acute lung injury in rats through NF-κB and p38 MAPK signaling pathway and excessive inflammatory responses.

PubMed

Chen, L-J; Ding, Y-B; Ma, P-L; Jiang, S-H; Li, K-Z; Li, A-Z; Li, M-C; Shi, C-X; Du, J; Zhou, H-D

2018-04-01

Acute lung injury is a severe disease with a high rate of mortality, leading to more important illness. We aimed at exploring the protective role and potential mechanisms of lidocaine on lipopolysaccharide (LPS)-induced acute lung injury (ALI). Sprague Dawley (SD) rats were randomly assigned to control group receiving 0.9% saline solution, LPS group treated with 4 mg/kg LPS i.p., LPS + lidocaine(treated with 4 mg/kg LPS i.p. followed by giving 1 mg/kg, 3 mg/kg, 5 mg/kg of lidocaine i.v.). Lung specimens and the bronchoalveolar lavage fluid (BALF) were collected for histopathological examination and biochemical analyze 12 h after LPS induction. The cytokines expression of TNF-α, IL-6 and MCP-1 was measured by ELISA. In addition, the malondialdehyde (MDA) content, the activities of total antioxidant capacity (T-AOC) and superoxide dismutase (SOD) in lung tissues were also detected using ELISA. The protein expressions of p38, p-p38, p65, p-p65 and IκB were analyzed by Western blot. The results indicated that after lidocaine treatment was able to decrease significantly wet-to-dry (W/D) ratio and ameliorate the histopathologic damage. Additionally, total protein content and the number of leukocytes in BALF significantly decreased. ELISA result indicated that the levels of TNF-α, IL-6 and MCP-1 in BALF were markedly suppressed. Meanwhile, the activities of T-AOC and SOD in lung tissues significantly increased, while the content of MDA significantly decreased after treatment with lidocaine. Moreover, Western blot suggested that lidocaine inhibited phosphorylation of NF-κB p65 and p38 MAPK. Therefore, lidocaine could ameliorate the LPS-induced lung injury via NF-κB/p38 MAPK signaling and excessive inflammatory responses, providing a potential for becoming the anti-inflammatory agent against lung injury.