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1.
Dokl Biochem Biophys ; 486(1): 187-191, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31367818

RESUMO

Using transgenic Drosophila model systems, we showed that four binding sites for the architectural protein dCTCF per se cannot form an effective insulator that blocks enhancers and protects against the Polycomb-dependent repression. These results suggest that, in the known Drosophila insulators, the dCTCF protein functions in cooperation with other architectural proteins.


Assuntos
Fator de Ligação a CCCTC/genética , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Animais , Animais Geneticamente Modificados
2.
Dokl Biochem Biophys ; 486(1): 224-228, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31367827

RESUMO

TRF2 protein (TBP-related factor 2) can substitute for TBP forming alternative transcription initiation complexes on TATA-less promoters, including the promoters of histone H1 and piRNA clusters required for transposon repression. The Drosophilatrf2 gene codes for two isoforms: a "short" and a "long" one, in which the same short TRF2 sequence is preceded by a long N-terminal domain. Here, we demonstrated that the long TFR2 isoform has a greater functional activity than the short isoform by expressing each of them at a reduced rate under the endogenous promoters. Expression of the long isoform alone affects neither the flies' viability nor the sex ratio. Expression of the short isoform alone leads to the phenotype described for the trf2 gene insufficiency and derepression of transposable elements, that is, decreased viability, disturbance of homologous chromosome pairing and segregation, and apparent female-biased sex ratio.


Assuntos
Drosophila melanogaster/metabolismo , Proteína 2 de Ligação a Repetições Teloméricas/química , Proteína 2 de Ligação a Repetições Teloméricas/metabolismo , Animais , Drosophila melanogaster/genética , Mutação , Isoformas de Proteínas/química , Isoformas de Proteínas/metabolismo , Proteína 2 de Ligação a Repetições Teloméricas/genética
4.
BMC Bioinformatics ; 20(1): 327, 2019 Jun 13.
Artigo em Inglês | MEDLINE | ID: mdl-31195954

RESUMO

BACKGROUND: The gap gene system controls the early cascade of the segmentation pathway in Drosophila melanogaster as well as other insects. Owing to its tractability and key role in embryo patterning, this system has been the focus for both computational modelers and experimentalists. The gap gene expression dynamics can be considered strictly as a one-dimensional process and modeled as a system of reaction-diffusion equations. While substantial progress has been made in modeling this phenomenon, there still remains a deficit of approaches to evaluate competing hypotheses. Most of the model development has happened in isolation and there has been little attempt to compare candidate models. RESULTS: The Bayesian framework offers a means of doing formal model evaluation. Here, we demonstrate how this framework can be used to compare different models of gene expression. We focus on the Papatsenko-Levine formalism, which exploits a fractional occupancy based approach to incorporate activation of the gap genes by the maternal genes and cross-regulation by the gap genes themselves. The Bayesian approach provides insight about relationship between system parameters. In the regulatory pathway of segmentation, the parameters for number of binding sites and binding affinity have a negative correlation. The model selection analysis supports a stronger binding affinity for Bicoid compared to other regulatory edges, as shown by a larger posterior mean. The procedure doesn't show support for activation of Kruppel by Bicoid. CONCLUSIONS: We provide an efficient solver for the general representation of the Papatsenko-Levine model. We also demonstrate the utility of Bayes factor for evaluating candidate models for spatial pattering models. In addition, by using the parallel tempering sampler, the convergence of Markov chains can be remarkably improved and robust estimates of Bayes factors obtained.


Assuntos
Drosophila melanogaster/genética , Redes Reguladoras de Genes , Animais , Teorema de Bayes , Proteínas de Drosophila/genética , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento , Funções Verossimilhança , Cadeias de Markov , Modelos Genéticos , Método de Monte Carlo
5.
Mol Biol (Mosk) ; 53(3): 476-484, 2019.
Artigo em Russo | MEDLINE | ID: mdl-31184613

RESUMO

It is known that long (200-300 nucleotides and longer) non-protein-coding RNAs (ncRNAs) tissue-specifically expressed from the regulatory regions of developmental genes can regulate the transcription of the mRNA of these genes. In this study, an attempt is made to identify differentially expressed ncRNAs in the extended promoter region of the fork head (fkh) gene of the fruit fly Drosophila melanogaster. We investigated four preparations of the total RNA: from embryos, from adult flies (separately from females and males), and from the S2 cell line of cultured Drosophila cells. In the total RNA preparations from embryos and adult flies, the levels of fkh expression differed substantially, whereas in S2 cells its expression is not detected at all (shown in this work). We perform classical Northern blot analysis of gel-separated RNAs hybridized to a series of radioactively labeled DNA fragments corresponding to the adjacent and partially overlapping regions of the promoter region of the fkh gene. Several previously unknown differentially expressed ncRNAs are detected, including those in the regions overlapping with the previously detected regulatory elements (TRE1 and salivary gland enhancer sgE) and the transcription start site of the fkh gene. The collected data complement and clarify the results of the previously conducted RNA-seq experiments, in particular, in terms of the length of the detected RNAs. These results may serve as a foundation for further studies of the mechanisms of tissue-specific regulation of the fkh gene expression.


Assuntos
Northern Blotting , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Fatores de Transcrição Forkhead/genética , Regulação da Expressão Gênica no Desenvolvimento , Regiões Promotoras Genéticas/genética , RNA Longo não Codificante/análise , RNA Longo não Codificante/genética , Animais , Linhagem Celular , Drosophila melanogaster/embriologia , Drosophila melanogaster/crescimento & desenvolvimento , Elementos Facilitadores Genéticos/genética , Feminino , Masculino , Especificidade de Órgãos
6.
Nat Cell Biol ; 21(6): 710-720, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31160709

RESUMO

The capacity of stem cells to self-renew or differentiate has been attributed to distinct metabolic states. A genetic screen targeting regulators of mitochondrial dynamics revealed that mitochondrial fusion is required for the maintenance of male germline stem cells (GSCs) in Drosophila melanogaster. Depletion of Mitofusin (dMfn) or Opa1 led to dysfunctional mitochondria, activation of Target of rapamycin (TOR) and a marked accumulation of lipid droplets. Enhancement of lipid utilization by the mitochondria attenuated TOR activation and rescued the loss of GSCs that was caused by inhibition of mitochondrial fusion. Moreover, constitutive activation of the TOR-pathway target and lipogenesis factor Sterol regulatory element binding protein (SREBP) also resulted in GSC loss, whereas inhibition of SREBP rescued GSC loss triggered by depletion of dMfn. Our findings highlight a critical role for mitochondrial fusion and lipid homeostasis in GSC maintenance, providing insight into the potential impact of mitochondrial and metabolic diseases on the function of stem and/or germ cells.


Assuntos
Proteínas de Drosophila/genética , Proteínas de Membrana/genética , Dinâmica Mitocondrial/genética , Células-Tronco/metabolismo , Proteínas de Ligação a Elemento Regulador de Esterol/genética , Animais , Diferenciação Celular/genética , Drosophila melanogaster/genética , Drosophila melanogaster/crescimento & desenvolvimento , Drosophila melanogaster/metabolismo , Homeostase , Metabolismo dos Lipídeos/genética , Masculino , Mitocôndrias/genética , Receptores Proteína Tirosina Quinases/genética , Transdução de Sinais/genética , Nicho de Células-Tronco/genética , Células-Tronco/citologia , Testículo/crescimento & desenvolvimento , Testículo/metabolismo
7.
BMC Evol Biol ; 19(1): 126, 2019 06 18.
Artigo em Inglês | MEDLINE | ID: mdl-31215418

RESUMO

BACKGROUND: L-ascorbate (Vitamin C) is an important antioxidant and co-factor in eukaryotic cells, and in mammals it is indispensable for brain development and cognitive function. Vertebrates usually become L-ascorbate auxothrophs when the last enzyme of the synthetic pathway, an L-gulonolactone oxidase (GULO), is lost. Since Protostomes were until recently thought not to have a GULO gene, they were considered to be auxothrophs for Vitamin C. RESULTS: By performing phylogenetic analyses with tens of non-Bilateria and Protostomian genomes, it is shown, that a GULO gene is present in the non-Bilateria Placozoa, Myxozoa (here reported for the first time) and Anthozoa groups, and in Protostomians, in the Araneae family, the Gastropoda class, the Acari subclass (here reported for the first time), and the Priapulida, Annelida (here reported for the first time) and Brachiopoda phyla lineages. GULO is an old gene that predates the separation of Animals and Fungi, although it could be much older. We also show that within Protostomes, GULO has been lost multiple times in large taxonomic groups, namely the Pancrustacea, Nematoda, Platyhelminthes and Bivalvia groups, a pattern similar to that reported for Vertebrate species. Nevertheless, we show that Drosophila melanogaster seems to be capable of synthesizing L-ascorbate, likely through an alternative pathway, as recently reported for Caenorhabditis elegans. CONCLUSIONS: Non-Bilaterian and Protostomians seem to be able to synthesize Vitamin C either through the conventional animal pathway or an alternative pathway, but in this animal group, not being able to synthesize L-ascorbate seems to be the exception rather than the rule.


Assuntos
Ácido Ascórbico/metabolismo , Eucariotos/enzimologia , Eucariotos/genética , Evolução Molecular , L-Gulonolactona Oxidase/genética , Animais , Drosophila melanogaster/genética , Eucariotos/classificação , Eucariotos/metabolismo , Genoma , L-Gulonolactona Oxidase/química , L-Gulonolactona Oxidase/metabolismo , Modelos Moleculares , Filogenia , Vertebrados/classificação , Vertebrados/genética
8.
Food Chem Toxicol ; 131: 110557, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31176925

RESUMO

The aim of the present study was to appraise the mutagenic and recombinogenic potential of bupropion hydrochloride (BHc) and trazodone hydrochloride (THc). We used standard (ST) and the high bioactivation (HB) crossings from Drosophila melanogaster in the Somatic Mutation and Recombination Test. We treated third-instar larvae from both crossings with different concentrations of BHc and THc (0.9375 to 7.5 mg/mL). BHc significantly increased the frequency of mutant spots in both crossings, except for the lowest concentration in the ST crossing. ST had also the mostly recombinogenic result, and in the HB, BHc was highly mutagenic. On the other hand, THc significantly increased the frequency of mutant spots in both the ST and HB crossings at all concentrations. The three initial concentrations were recombinogenic and the highest concentration was mutagenic for the THc. BHc and THc at high concentrations were toxic, even though their mutagenicity was not dose-related. THc significantly increased the frequency of mutant spots when metabolized, probably as a result of the production of 1-(3'-chlorophenyl) piperazine. BHc was essentially recombinogenic and when metabolized, it became mutagenic. THc was recombinogenic in both crossings. Further studies are needed to clarify the action mechanisms from BHc and THc.


Assuntos
Antidepressivos/toxicidade , Bupropiona/toxicidade , Drosophila melanogaster/efeitos dos fármacos , Mutagênicos/toxicidade , Recombinação Genética/efeitos dos fármacos , Trazodona/toxicidade , Animais , Drosophila melanogaster/genética , Feminino , Masculino , Testes de Mutagenicidade , Mutação , Asas de Animais/efeitos dos fármacos
9.
Nat Commun ; 10(1): 2891, 2019 06 28.
Artigo em Inglês | MEDLINE | ID: mdl-31253791

RESUMO

Our ability to manage acute myeloid leukemia (AML) is limited by our incomplete understanding of the epigenetic disruption central to leukemogenesis, including improper histone methylation. Here we examine 16 histone H3 genes in 434 primary AML samples and identify Q69H, A26P, R2Q, R8H and K27M/I mutations (1.6%), with higher incidence in secondary AML (9%). These mutations occur in pre-leukemic hematopoietic stem cells (HSCs) and exist in the major leukemic clones in patients. They increase the frequency of functional HSCs, alter differentiation, and amplify leukemic aggressiveness. These effects are dependent on the specific mutation. H3K27 mutation increases the expression of genes involved in erythrocyte and myeloid differentiation with altered H3K27 tri-methylation and K27 acetylation. The functional impact of histone mutations is independent of RUNX1 mutation, although they at times co-occur. This study establishes that H3 mutations are drivers of human pre-cancerous stem cell expansion and important early events in leukemogenesis.


Assuntos
Epigenômica , Regulação Leucêmica da Expressão Gênica/fisiologia , Histonas/metabolismo , Leucemia Mieloide Aguda/metabolismo , Animais , Animais Geneticamente Modificados , Antineoplásicos/farmacologia , Sequência de Bases , Células da Medula Óssea , Diferenciação Celular , Transformação Celular Neoplásica , DNA/genética , Drosophila melanogaster/genética , Regulação Leucêmica da Expressão Gênica/efeitos dos fármacos , Hematopoese/fisiologia , Células-Tronco Hematopoéticas/metabolismo , Humanos , Camundongos , Mutação , Neoplasias Experimentais
10.
Behav Processes ; 164: 133-142, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31051219

RESUMO

Starting in late 1980's, Bill Timberlake and associates conducted a series of experiments on anticipatory contrast which showed that rats' feeding decisions were regulated by the nutritive value of currently ingested and anticipated food. The effects of nutrient sensing on feeding regulation have been studied intensively in rodents, and recently, in the fruit fly Drosophila melanogaster. In this study, we developed a new behavioral test to study rapid feeding decisions of tethered flies within 6-8 s of ingestion. Using a two-phase experimental design, we presented individual flies one of four serial combinations of a non-nutritive sugar, arabinose, or a nutritive sugar, sucrose. Feeding decisions of wildtype (Canton-S) flies are altered both by immediate effects of nutrient sensing and 1-hour delayed effects of nutrient-feeding, and the two effects act additively to yield a signature pattern of behavioral contrast based on nutritive contrast. Feeding phenotype of flies that carry a mutation of the dSLC5A11 (cupcake) gene varied with the mutant allele and genetic background. Fasted dSLC5A11 mutants showed an overeating phenotype and a defect in short-term feeding regulation irrespective of the nutritive value of sugar. Flies that carried the dSLC5A111 allele showed differential feeding for arabinose and sucrose. However, dSLC5A112 allele yielded a conspicuous deficit in delayed effects of nutrient ingestion, but only when it was expressed on a Canton-S background. Our results suggest that dSLC5A11 might function to integrate external stimulus properties and internal state for feeding regulation and action selection.


Assuntos
Tomada de Decisões , Proteínas de Drosophila/genética , Proteínas de Drosophila/fisiologia , Drosophila melanogaster/fisiologia , Comportamento Alimentar/fisiologia , Nutrientes/fisiologia , Proteínas de Transporte de Sódio-Glucose/genética , Proteínas de Transporte de Sódio-Glucose/fisiologia , Alelos , Animais , Arabinose , Drosophila melanogaster/genética , Mutação , Valor Nutritivo , Percepção/fisiologia , Fenótipo , Ratos , Sacarose , Fatores de Tempo
11.
Genes Dev ; 33(13-14): 844-856, 2019 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-31123065

RESUMO

The Piwi-interacting RNA (piRNA) pathway is a small RNA-based immune system that silences mobile genetic elements in animal germlines. piRNA biogenesis requires a specialized machinery that converts long single-stranded precursors into small RNAs of ∼25-nucleotides in length. This process involves factors that operate in two different subcellular compartments: the nuage/Yb body and mitochondria. How these two sites communicate to achieve accurate substrate selection and efficient processing remains unclear. Here, we investigate a previously uncharacterized piRNA biogenesis factor, Daedalus (Daed), that is located on the outer mitochondrial membrane. Daed is essential for Zucchini-mediated piRNA production and the correct localization of the indispensable piRNA biogenesis factor Armitage (Armi). We found that Gasz and Daed interact with each other and likely provide a mitochondrial "anchoring platform" to ensure that Armi is held in place, proximal to Zucchini, during piRNA processing. Our data suggest that Armi initially identifies piRNA precursors in nuage/Yb bodies in a manner that depends on Piwi and then moves to mitochondria to present precursors to the mitochondrial biogenesis machinery. These results represent a significant step in understanding a critical aspect of transposon silencing; namely, how RNAs are chosen to instruct the piRNA machinery in the nature of its silencing targets.


Assuntos
Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Mitocôndrias/metabolismo , RNA Helicases/metabolismo , RNA Interferente Pequeno/biossíntese , Animais , Linhagem Celular , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Técnicas de Silenciamento de Genes , Ligação Proteica , Transporte Proteico , RNA Interferente Pequeno/metabolismo
12.
Nature ; 570(7759): 122-126, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31092928

RESUMO

Transcriptional cofactors (COFs) communicate regulatory cues from enhancers to promoters and are central effectors of transcription activation and gene expression1. Although some COFs have been shown to prefer certain promoter types2-5 over others (for example, see refs 6,7), the extent to which different COFs display intrinsic specificities for distinct promoters is unclear. Here we use a high-throughput promoter-activity assay in Drosophila melanogaster S2 cells to screen 23 COFs for their ability to activate 72,000 candidate core promoters (CPs). We observe differential activation of CPs, indicating distinct regulatory preferences or 'compatibilities'8,9 between COFs and specific types of CPs. These functionally distinct CP types are differentially enriched for known sequence elements2,4, such as the TATA box, downstream promoter element (DPE) or TCT motif, and display distinct chromatin properties at endogenous loci. Notably, the CP types differ in their relative abundance of H3K4me3 and H3K4me1 marks (see also refs 10-12), suggesting that these histone modifications might distinguish trans-regulatory factors rather than promoter- versus enhancer-type cis-regulatory elements. We confirm the existence of distinct COF-CP compatibilities in two additional Drosophila cell lines and in human cells, for which we find COFs that prefer TATA-box or CpG-island promoters, respectively. Distinct compatibilities between COFs and promoters can explain how different enhancers specifically activate distinct sets of genes9, alternative promoters within the same genes, and distinct transcription start sites within the same promoter13. Thus, COF-promoter compatibilities may underlie distinct transcriptional programs in species as divergent as flies and humans.


Assuntos
Regiões Promotoras Genéticas/genética , Fatores de Transcrição/metabolismo , Transcrição Genética , Ativação Transcricional , Animais , Linhagem Celular , Cromatina/genética , Ilhas de CpG/genética , Drosophila melanogaster/citologia , Drosophila melanogaster/genética , Elementos Facilitadores Genéticos/genética , Histonas/metabolismo , Humanos , Especificidade por Substrato , TATA Box/genética , Sítio de Iniciação de Transcrição
13.
Environ Sci Pollut Res Int ; 26(19): 19560-19574, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31079296

RESUMO

The current study checks the effect of various concentrations of dietary graphene oxide (GO) nano-sheets on the development of Drosophila melanogaster. GO was synthesized and characterized by XRD, FTIR, FESEM, and TEM analytical techniques. Various concentrations of GO were mixed with the fly food and flies were transferred to the vial. Various behavioral and morphological as well as genetic defects were checked on the different developmental stages of the offspring. In the larval stage of development, the crawling speed and trailing path change significantly than the control. GO induces the generation of oxygen radicals within the larval hemolymph as evidenced by nitroblue tetrazolium assay. GO induces DNA damage within the gut cell, which was detected by Hoechst staining and within hemolymph by comet assay. Adult flies hatched after GO treatment show defective phototaxis and geotaxis behavior. Besides behavior, phenotypic defects were observed in the wing, eye, thorax bristles, and mouth parts. At 300 mg/L concentration, wing spots were observed. Altogether, the current study finds oral administration of GO which acts as a mutagen and causes various behavioral and developmental defects in the offspring. Here for the first time, we are reporting GO, which acts as a teratogen in Drosophila, besides its extensive medical applications.


Assuntos
Drosophila melanogaster/efeitos dos fármacos , Grafite/toxicidade , Mutagênicos/toxicidade , Nanoestruturas/toxicidade , Estresse Oxidativo/efeitos dos fármacos , Teratogênios/toxicidade , Administração Oral , Animais , Comportamento Animal/efeitos dos fármacos , Dano ao DNA , Drosophila melanogaster/genética , Drosophila melanogaster/crescimento & desenvolvimento , Larva/efeitos dos fármacos , Larva/genética , Larva/crescimento & desenvolvimento
14.
Mol Biol (Mosk) ; 53(2): 225-239, 2019.
Artigo em Russo | MEDLINE | ID: mdl-31099773

RESUMO

The ensemble of gap genes is one of the best studied and most conserved gene regulatory networks (GRNs). Gap genes, such as hunchback (hb), Krüppel (Kr), pou-domain (pdm; pdm1 and pdm2), and castor (cas) genes belong to the well-known families Ikaros (IKZF1/hb), Krüppel-like factor (KLF/Kr), POU domain (BRN1/pdm-1, BRN2/pdm-2), and Castor homologs (CASZ1/cas), which are present in all vertebrate genomes and code for site-specific transcription factors. Gap genes form a core of an embryonic segmentation control subnetwork and define the temporal identity of neuroblasts in Drosophila embryos. The key gene regulatory mechanisms whereby the gap genes govern segmentation and neurogenesis are similar. Moreover, the gap genes are evolutionarily conserved in terms of their function as a core of the temporal specification GRN during neurogenesis in vertebrates, including humans. A problem of special interest is to understand the extent of conservation for the molecular mechanisms involved in the regulatory functions of the gap genes. The problem is especially important because human orthologs of the gap gens are crucial for many pathophysiological processes, including tumor growth suppression.


Assuntos
Drosophila melanogaster/citologia , Drosophila melanogaster/genética , Evolução Molecular , Redes Reguladoras de Genes , Neurônios/metabolismo , Animais , Drosophila melanogaster/embriologia , Regulação da Expressão Gênica no Desenvolvimento , Fatores de Tempo , Fatores de Transcrição/metabolismo
15.
J Insect Sci ; 19(3)2019 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-31115476

RESUMO

Alkaline ceramidase (Dacer) in Drosophila melanogaster was demonstrated to be resistant to paraquat-induced oxidative stress. However, the underlying mechanism for this resistance remained unclear. Here, we showed that sphingosine feeding triggered the accumulation of hydrogen peroxide (H2O2). Dacer-deficient D. melanogaster (Dacer mutant) has higher catalase (CAT) activity and CAT transcription level, leading to higher resistance to oxidative stress induced by paraquat. By performing a quantitative proteomic analysis, we identified 79 differentially expressed proteins in comparing Dacer mutant to wild type. Three oxidoreductases, including two cytochrome P450 (CG3050, CG9438) and an oxoglutarate/iron-dependent dioxygenase (CG17807), were most significantly upregulated in Dacer mutant. We presumed that altered antioxidative activity in Dacer mutant might be responsible for increased oxidative stress resistance. Our work provides a novel insight into the oxidative antistress response in D. melanogaster.


Assuntos
Ceramidase Alcalina/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/enzimologia , Estresse Oxidativo , Esfingosina/administração & dosagem , Ceramidase Alcalina/efeitos dos fármacos , Ceramidase Alcalina/genética , Animais , Catalase/metabolismo , Proteínas de Drosophila/efeitos dos fármacos , Proteínas de Drosophila/genética , Drosophila melanogaster/efeitos dos fármacos , Drosophila melanogaster/genética , Peróxido de Hidrogênio/metabolismo , Paraquat , Proteoma
16.
Genes Cells ; 24(7): 496-510, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31124270

RESUMO

In the Drosophila brain, neurons form genetically specified synaptic connections with defined neuronal targets. It is proposed that each central nervous system neuron expresses specific cell surface proteins, which act as identification tags. Through an RNAi screen of cell surface molecules in the Drosophila visual system, we found that the cell adhesion molecule Klingon (Klg) plays an important role in repressing the ectopic formation of extended axons, preventing the formation of excessive synapses. Cell-specific manipulation of klg showed that Klg is required in both photoreceptors and the glia, suggesting that the balanced homophilic interaction between photoreceptor axons and the glia is required for normal synapse formation. Previous studies suggested that Klg binds to cDIP and our genetic analyses indicate that cDIP is required in glia for ectopic synaptic repression. These data suggest that Klg play a critical role together with cDIP in refining synaptic specificity and preventing unnecessary connections in the brain.


Assuntos
Moléculas de Adesão Celular/metabolismo , Proteínas de Drosophila/metabolismo , Drosophila melanogaster/fisiologia , Proteínas do Olho/metabolismo , Neuroglia/fisiologia , Células Fotorreceptoras de Invertebrados/fisiologia , Sinapses/fisiologia , Vias Visuais , Animais , Animais Geneticamente Modificados/genética , Animais Geneticamente Modificados/fisiologia , Axônios/fisiologia , Moléculas de Adesão Celular/genética , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Proteínas do Olho/genética , Feminino
17.
BMC Bioinformatics ; 20(1): 174, 2019 Apr 05.
Artigo em Inglês | MEDLINE | ID: mdl-30953451

RESUMO

BACKGROUND: Identifying transcriptional enhancers and other cis-regulatory modules (CRMs) is an important goal of post-sequencing genome annotation. Computational approaches provide a useful complement to empirical methods for CRM discovery, but it is critical that we develop effective means to evaluate their performance in terms of estimating their sensitivity and specificity. RESULTS: We introduce here pCRMeval, a pipeline for in silico evaluation of any enhancer prediction tools that are flexible enough to be applied to the Drosophila melanogaster genome. pCRMeval compares the result of predictions with the extensive existing knowledge of experimentally-validated Drosophila CRMs in order to estimate the precision and relative sensitivity of the prediction method. In the case of supervised prediction methods-when training data composed of validated CRMs are used-pCRMeval can also assess the sensitivity of specific training sets. We demonstrate the utility of pCRMeval through evaluation of our SCRMshaw CRM prediction method and training data. By measuring the impact of different parameters on SCRMshaw performance, as assessed by pCRMeval, we develop a more robust version of SCRMshaw, SCRMshaw_HD, that improves the number of predictions while maintaining sensitivity and specificity. Our analysis also demonstrates that SCRMshaw_HD, when applied to increasingly less well-assembled genomes, maintains its strong predictive power with only a minor drop-off in performance. CONCLUSION: Our pCRMeval pipeline provides a general framework for evaluation that can be applied to any CRM prediction method, particularly a supervised method. While we make use of it here primarily to test and improve a particular method for CRM prediction, SCRMshaw, pCRMeval should provide a valuable platform to the research community not only for evaluating individual methods, but also for comparing between competing methods.


Assuntos
Biologia Computacional , Drosophila melanogaster/genética , Elementos Facilitadores Genéticos , Animais , Sequência de Bases , Mapeamento Cromossômico , Drosophila melanogaster/metabolismo , Feminino , Genoma de Inseto , Masculino , Especificidade da Espécie , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismo
18.
Nat Commun ; 10(1): 1626, 2019 04 09.
Artigo em Inglês | MEDLINE | ID: mdl-30967537

RESUMO

MicroRNAs (miRNAs) are key mediators of post-transcriptional gene expression silencing. So far, no comprehensive experimental annotation of functional miRNA target sites exists in Drosophila. Here, we generated a transcriptome-wide in vivo map of miRNA-mRNA interactions in Drosophila melanogaster, making use of single nucleotide resolution in Argonaute1 (AGO1) crosslinking and immunoprecipitation (CLIP) data. Absolute quantification of cellular miRNA levels presents the miRNA pool in Drosophila cell lines to be more diverse than previously reported. Benchmarking two CLIP approaches, we identify a similar predictive potential to unambiguously assign thousands of miRNA-mRNA pairs from AGO1 interaction data at unprecedented depth, achieving higher signal-to-noise ratios than with computational methods alone. Quantitative RNA-seq and sub-codon resolution ribosomal footprinting data upon AGO1 depletion enabled the determination of miRNA-mediated effects on target expression and translation. We thus provide the first comprehensive resource of miRNA target sites and their quantitative functional impact in Drosophila.


Assuntos
Proteínas Argonauta/genética , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Regulação da Expressão Gênica , MicroRNAs/metabolismo , RNA Mensageiro/metabolismo , Animais , MicroRNAs/genética , MicroRNAs/isolamento & purificação , RNA Mensageiro/genética , RNA Mensageiro/isolamento & purificação , Análise de Sequência de RNA , Transcriptoma/genética
19.
Mol Genet Genomics ; 294(4): 1073-1083, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31006039

RESUMO

Nopp140, often called the nucleolar and Cajal body phosphoprotein (NOLC1), is an evolutionarily conserved chaperone for the transcription and processing of rRNA during ribosome subunit assembly. Metazoan Nopp140 contains an amino terminal LisH dimerization domain and a highly conserved carboxyl domain. A large central domain consists of alternating basic and acidic motifs of low sequence complexity. Orthologous versions of Nopp140 contain variable numbers of repeating basic-acidic units. While vertebrate Nopp140 genes use multiple exons to encode the central domain, the Nopp140 gene in Drosophila uses exclusively exon 2 to encode the central domain. Here, we define three overlapping repeat sequence patterns (P, P', and P″) within the central domain of D. melanogaster Nopp140. These repeat patterns are poorly conserved in other Drosophila species. We also describe a length polymorphism in exon 2 that pertains specifically to the P' pattern in D. melanogaster. The pattern displays either two or three 96 base pair repeats, respectively, referred to as Nopp140-Short and Nopp140-Long. Fly lines homozygous for one or the other allele, or heterozygous for both alleles, show no discernible phenotypes. PCR characterization of the long and short alleles shows a poorly defined, artifactual bias toward amplifying the long allele over the short allele. The significance of this polymorphism will be in discerning the largely unknown properties of Nopp140's large central domain in rDNA transcription and ribosome biogenesis.


Assuntos
Proteínas de Drosophila/química , Proteínas de Drosophila/genética , Drosophila melanogaster/metabolismo , Proteínas Nucleares/química , Proteínas Nucleares/genética , Polimorfismo Genético , Animais , Drosophila melanogaster/genética , Éxons , Fenótipo , Domínios Proteicos , RNA Ribossômico/genética , Sequências Repetitivas de Ácido Nucleico
20.
BMC Genomics ; 20(1): 305, 2019 Apr 23.
Artigo em Inglês | MEDLINE | ID: mdl-31014230

RESUMO

BACKGROUND: Evolutionary theory indicates that the dynamics of aging in the soma and reproductive tissues may be distinct. This difference arises from the fact that only the germline lineage establishes future generations. In the soma, changes in the landscape of heterochromatin have been proposed to have an important role in aging. This is because redistribution of heterochromatin during aging has been linked to the derepression of transposable elements and an overall loss of somatic gene regulation. A role for changes in the chromatin landscape in the aging of reproductive tissues is less well established. Whether or not epigenetic factors, such as heterochromatin marks, are perturbed in aging reproductive tissues is of interest because, in special cases, epigenetic variation may be heritable. Using mRNA sequencing data from late-stage egg chambers in Drosophila melanogaster, we characterized the landscape of altered gene and transposable element expression in aged reproductive tissues. This allowed us to test the hypothesis that reproductive tissues may differ from somatic tissues in their response to aging. RESULTS: We show that age-related expression changes in late-stage egg chambers tend to occur in genes residing in heterochromatin, particularly on the largely heterochromatic 4th chromosome. However, these expression differences are seen as both decreases and increases during aging, inconsistent with a general loss of heterochromatic silencing. We also identify an increase in expression of the piRNA machinery, suggesting an age-related increased investment in the maintenance of genome stability. We further identify a strong age-related reduction in the expression of mitochondrial transcripts. However, we find no evidence for global TE derepression in reproductive tissues. Rather, the observed effects of aging on TEs are primarily strain and family specific. CONCLUSIONS: These results identify unique responses in somatic versus reproductive tissue with regards to aging. As in somatic tissues, female reproductive tissues show reduced expression of mitochondrial genes. In contrast, the piRNA machinery shows increased expression during aging. Overall, these results also indicate that global loss of TE control observed in other studies may be unique to the soma and sensitive to genetic background and TE family.


Assuntos
Envelhecimento/genética , Elementos de DNA Transponíveis/genética , Drosophila melanogaster/fisiologia , Perfilação da Expressão Gênica , Mitocôndrias/genética , Ovário/fisiologia , RNA Interferente Pequeno/genética , Animais , Drosophila melanogaster/genética , Feminino , Genoma Mitocondrial/genética , Heterocromatina/genética , Óvulo/metabolismo , RNA Mensageiro/genética
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