Drosophila melanogaster
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA116747 | ENA |
REPOSITORIES: ENA
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Project description:Primary piRNAs in Drosophila ovarian somatic cells arise from piRNA cluster transcripts and the 3′ UTRs of a subset of mRNAs, including Traffic jam (Tj) mRNA. However, it is unclear how these RNAs are determined as primary piRNA sources. Here, we identify a cis-acting 100-nt fragment in the Tj 3′ UTR that is sufficient for producing artificial piRNAs from unintegrated DNA. These artificial piRNAs were effective in endogenous gene transcriptional silencing. Yb, a core component of primary piRNA biogenesis center Yb bodies, directly bound the Tj-cis-element. Disruption of this interaction markedly reduced piRNA production. Thus, Yb is the trans-acting partner of the Tj-cis-element. Yb-CLIP revealed that Yb-binding correlated with somatic piRNA production but Tj-cis-element downstream sequences produced few artificial piRNAs. Thus, Yb determines primary piRNA sources by two modes of action; primary binding to cis-elements to specify substrates, and secondary binding to downstream regions to increase diversity in piRNA populations. HITS-CLIP of Yb in OSCs (Ovarian Somatic Cells) depleted for tj cis-element, and small RNA sequencing of Piwi-piRNAs in OSCs depleted for tj cis-element.
2015-06-05 | E-GEOD-69625 | biostudies-arrayexpress
Project description:Primary piRNAs in Drosophila ovarian somatic cells arise from piRNA cluster transcripts and the 3′ UTRs of a subset of mRNAs, including Traffic jam (Tj) mRNA. However, it is unclear how these RNAs are determined as primary piRNA sources. Here, we identify a cis-acting 100-nt fragment in the Tj 3′ UTR that is sufficient for producing artificial piRNAs from unintegrated DNA. These artificial piRNAs were effective in endogenous gene transcriptional silencing. Yb, a core component of primary piRNA biogenesis center Yb bodies, directly bound the Tj-cis-element. Disruption of this interaction markedly reduced piRNA production. Thus, Yb is the trans-acting partner of the Tj-cis-element. Yb-CLIP revealed that Yb-binding correlated with somatic piRNA production but Tj-cis-element downstream sequences produced few artificial piRNAs. Thus, Yb determines primary piRNA sources by two modes of action; primary binding to cis-elements to specify substrates, and secondary binding to downstream regions to increase diversity in piRNA populations.
2015-06-05 | GSE69625 | GEO
Project description:The PiwiM-bM-^@M-^SpiRNA complex (PiwiM-bM-^@M-^SpiRISC) in Drosophila ovarian somatic cells represses transposons transcriptionally to maintain genome integrity; however, the underlying mechanisms remain obscure. We performed mRNA-seq analysis from OSCs transfected with siRNAs against CG3893, the known piRNA pathway genes, Piwi, Maelstrom, HP1a and Armitage, and the control (EGFP), and PolII ChIP-seqanalysis from OSCs transfected with siRNAs against CG3893, Piwi, Mael and the control (EGFP). This result indicates that CG3893 is a novel factor for primary piRNA pathway in OSCs. RNA levels in wild-type (EGFP control knock-down) ovarian somatic cells (OSC) and RNAi knock-downs of Piwi, Armi, Mael, CG3893, and HP1a. RNA Polymerase II occupancy in wild-type (EGFP control knock-down) ovarian somatic cells (OSC) and RNAi knock-downs of Piwi, Mael, and CG3893.
2013-08-01 | E-GEOD-47006 | biostudies-arrayexpress
Project description:The Piwi–piRNA complex (Piwi–piRISC) in Drosophila ovarian somatic cells represses transposons transcriptionally to maintain genome integrity; however, the underlying mechanisms remain obscure. We performed mRNA-seq analysis from OSCs transfected with siRNAs against CG3893, the known piRNA pathway genes, Piwi, Maelstrom, HP1a and Armitage, and the control (EGFP), and PolII ChIP-seqanalysis from OSCs transfected with siRNAs against CG3893, Piwi, Mael and the control (EGFP). This result indicates that CG3893 is a novel factor for primary piRNA pathway in OSCs.
2013-08-01 | GSE47006 | GEO
Project description:Piwi-interacting RNAs (piRNAs) suppress transposon activity in animal germ cells. In the Drosophila ovary, primary Aubergine (Aub)-bound antisense piRNAs initiate the ping-pong cycle to produce secondary AGO3-bound sense piRNAs. This increases the number of secondary Aub-bound antisense piRNAs that can act to destroy transposon mRNAs. Here we show that Krimper (Krimp), a Tudor-domain protein, directly interacts with piRNA-free AGO3 to promote symmetrical dimethylarginine (sDMA) modification, ensuring sense piRNA-loading onto sDMA-modified AGO3. In aub mutant ovaries, AGO3 associates with ping-pong signature piRNAs, suggesting AGO3’s compatibility with primary piRNA loading. Krimp sequesters ectopically expressed AGO3 within Krimp bodies in cultured ovarian somatic cells (OSCs), in which only the primary piRNA pathway operates. Upon krimp-RNAi in OSCs, AGO3 loads with piRNAs, further showing the capacity of AGO3 for primary piRNA loading. We propose that Krimp enforces an antisense bias on piRNA pools by binding AGO3 and blocking its access to primary piRNAs. In order to investigate function of Krimp in piRNA pathway, sequencing of Piwi subfamily protein associated small RNAs was performed using adult Drosophila ovaries and Ovarian Somatic Cells (OSCs) depleted for Krimp or Aub.
2015-05-28 | E-GEOD-69293 | biostudies-arrayexpress
Project description:piRNAs function in silencing retrotransposons by associating with the PIWI proteins, AGO3, Aub, and Piwi, in Drosophila germlines. Bioinformatics analyses of piRNAs in Drosophila ovaries suggested that piRNAs are produced by two systems, the primary processing pathway and the amplification loop, from repetitive genes and piRNA clusters in the genome. The amplification loop occurs in a Dicer-independent, PIWI-Slicer-dependent manner. However, the primary processing pathway remains largely conceptual. Here we show that in ovarian somatic cells, which lack Aub and AGO3 but express Piwi, the primary processing pathway for piRNAs indeed exists. Keywords: Small RNA profiling by high throughput sequencing Piwi-associated small RNAs were extracted from Drosophila ovarian somatic cells and their deep sequencing was carried out.
2009-10-01 | E-GEOD-15137 | biostudies-arrayexpress
Project description:The animal piRNA pathway is a small RNA silencing system that acts in gonads and protects the genome against the deleterious influence of transposons. A major bottleneck in the field is the lack of comprehensive knowledge of the factors and molecular processes that constitute this pathway. We conducted an RNAi screen in Drosophila and identified ~50 genes that strongly impact the ovarian somatic piRNA pathway. Many identified genes fall into functional categories that indicate essential roles for mitochondrial metabolism, RNA export, the nuclear pore, transcription elongation and chromatin regulation in the pathway. Follow-up studies on two factors demonstrate the identification of components acting at distinct hierarchical levels of the pathway. Finally, we define CG2183/Gasz as a novel primary piRNA biogenesis factor in somatic and germline cells. Based on the similarities between insect and vertebrate piRNA pathways our results have far-reaching implications for the understanding of this conserved genome defense system. Steady-state RNA levels in wild-type ovarian somatic cells (OSC) and RNAi knock-downs of the piRNA pathway components.
2013-05-10 | E-GEOD-45894 | biostudies-arrayexpress
Project description:RNAi-related silencing mechanisms concern as diverse biological processes as gene regulation, mostly via the miRNA pathway, and defense against molecular parasites, mainly controlled by the siRNA and the piRNA pathways. In Drosophila somatic ovarian cells, transposable elements (TEs) are repressed by chromatin changes induced by Piwi-interacting RNAs (piRNAs). We show here that a functional miRNA pathway is required for this piRNA-mediated TE transcriptional silencing to operate in this tissue. A general miRNA depletion, caused by tissue- and stage-specific knock-down of either drosha (involved in miRNA biogenesis) or AG01 and gawky (both responsible for miRNA activity) resulted in chromatin-mediated TE transcriptional desilencing and piRNA loss. For unknown reasons, the amount of piRNA produced by the traffic jam 3' UTR was apparently unaffected in miRNA-defective somatic ovarian cells. Although weaker, similar phenotypes could also be observed upon individual titration (by expression of the complementary miR-sponge) of at least three miRNAs, miR-14, miR-34 and miR-989. This work adds the maintenance of genome stability, via the piRNA-mediated TE repression, to the list of the already reported miRNA-controlled biological functions. Analyses of RNA present in 3 different genetic backgounds with or without Drosha-IP: a control, a second one expressing the wild-type Drosha protein and a third one expressing the trans-dominant negative Drosha protein in the somatic cells.
2015-04-08 | E-GEOD-60973 | biostudies-arrayexpress
Project description:RNAi-related silencing mechanisms concern as diverse biological processes as gene regulation, mostly via the miRNA pathway, and defense against molecular parasites, mainly controlled by the siRNA and the piRNA pathways. In Drosophila somatic ovarian cells, transposable elements (TEs) are repressed by chromatin changes induced by Piwi-interacting RNAs (piRNAs). We show here that a functional miRNA pathway is required for this piRNA-mediated TE transcriptional silencing to operate in this tissue. A general miRNA depletion, caused by tissue- and stage-specific knock-down of either drosha (involved in miRNA biogenesis) or AG01 and gawky (both responsible for miRNA activity) resulted in chromatin-mediated TE transcriptional desilencing and piRNA loss. For unknown reasons, the amount of piRNA produced by the traffic jam 3' UTR was apparently unaffected in miRNA-defective somatic ovarian cells. Although weaker, similar phenotypes could also be observed upon individual titration (by expression of the complementary miR-sponge) of at least three miRNAs, miR-14, miR-34 and miR-989. This work adds the maintenance of genome stability, via the piRNA-mediated TE repression, to the list of the already reported miRNA-controlled biological functions. Study of small regulatory RNA populations present in ovaries expressing either wild-type Drosha protein or the trans-dominant negative Drosha protein coupled or not with the Tub-GAL80[ts]GAL4 inhibitor.
2015-04-08 | E-GEOD-60972 | biostudies-arrayexpress
Project description:Here, we analyzed nine Drosophila piRNA pathway mutants for their impacts on both small RNA populations and the sub-cellular localization patterns of Piwi proteins. We find that distinct piRNA pathways with differing components function in ovarian germ and somatic cells. Keywords: Epigenetics
2009-04-30 | GSE15186 | GEO