Drosophila melanogaster
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA286011 | 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:In Drosophila, PIWI proteins and bound PIWI interacting RNAs (piRNAs) form the core of a small RNA mediated defense system against selfish genetic elements. Within germline cells piRNAs are processed from piRNA clusters and transposons to be loaded into Piwi/Aubergine/AGO3 and a subset of piRNAs undergoes target dependent amplification. In contrast, gonadal somatic support cells express only Piwi, lack signs of piRNA amplification and exhibit primary piRNA biogenesis from piRNA clusters. Neither piRNA processing/loading nor Piwi mediated target silencing is understood at the genetic, cellular or molecular level. We developed an in vivo RNAi assay for the somatic piRNA pathway and identified the RNA helicase Armitage, the Tudor domain containing RNA helicase Yb and the putative nuclease Zucchini as essential factors for primary piRNA biogenesis. Lack of any of these proteins leads to transposon de-silencing, to a collapse in piRNA levels and to a failure in Piwi nuclear accumulation. We show that Armitage and Yb interact physically and co-localize in cytoplasmic Yb-bodies, which flank P-bodies. Loss of Zucchini leads to an accumulation of Piwi and Armitage in Yb-bodies indicating that Yb-bodies are sites of primary piRNA biogenesis. small RNA libraries were prepared from Piwi immuno-precipitates of five different genotypes
2010-10-03 | E-GEOD-23560 | biostudies-arrayexpress
Project description:In Drosophila, PIWI proteins and bound PIWI interacting RNAs (piRNAs) form the core of a small RNA mediated defense system against selfish genetic elements. Within germline cells piRNAs are processed from piRNA clusters and transposons to be loaded into Piwi/Aubergine/AGO3 and a subset of piRNAs undergoes target dependent amplification. In contrast, gonadal somatic support cells express only Piwi, lack signs of piRNA amplification and exhibit primary piRNA biogenesis from piRNA clusters. Neither piRNA processing/loading nor Piwi mediated target silencing is understood at the genetic, cellular or molecular level. We developed an in vivo RNAi assay for the somatic piRNA pathway and identified the RNA helicase Armitage, the Tudor domain containing RNA helicase Yb and the putative nuclease Zucchini as essential factors for primary piRNA biogenesis. Lack of any of these proteins leads to transposon de-silencing, to a collapse in piRNA levels and to a failure in Piwi nuclear accumulation. We show that Armitage and Yb interact physically and co-localize in cytoplasmic Yb-bodies, which flank P-bodies. Loss of Zucchini leads to an accumulation of Piwi and Armitage in Yb-bodies indicating that Yb-bodies are sites of primary piRNA biogenesis.
2010-10-04 | GSE23560 | GEO
Project description:piRNAs direct Piwi to repress transposons to maintain genome integrity in Drosophila ovarian somatic cells. piRNA maturation and association with Piwi occur at perinuclear Yb bodies, the centers of piRNA biogenesis. Here, we show that piRNA intermediates arising from the piRNA cluster flamenco (flam) concentrate into perinuclear foci adjacent to Yb bodies, termed Flam bodies. Although flam expression is not required for Yb body formation, Yb, the core component of Yb bodies, is required for Flam body formation. Abolishment of the RNA-binding activity of Yb disrupts both Yb bodies and Flam bodies. Loss of Zucchini, an endoribonuclease necessary for piRNA maturation, enlarges Flam bodies, which now superimpose with Yb bodies. Yb directly binds flam, but not neighboring protein-coding gene, transcripts. Thus, Yb integrates piRNA processing factors and piRNA intermediates into Yb bodies and Flam bodies, respectively, through direct binding to enhance piRNA biogenesis and formation of piRNA-inducing silencing complexes. HITS-CLIP was performed using OSC (Ovarian Somatic Cells). The antibody for Drosophila Yb, which was generated in this study, was used. Obtained CLIP tags were analyzed using illumina HiSeq200.
2014-06-24 | E-GEOD-54875 | biostudies-arrayexpress
Project description:piRNAs direct Piwi to repress transposons to maintain genome integrity in Drosophila ovarian somatic cells. piRNA maturation and association with Piwi occur at perinuclear Yb bodies, the centers of piRNA biogenesis. Here, we show that piRNA intermediates arising from the piRNA cluster flamenco (flam) concentrate into perinuclear foci adjacent to Yb bodies, termed Flam bodies. Although flam expression is not required for Yb body formation, Yb, the core component of Yb bodies, is required for Flam body formation. Abolishment of the RNA-binding activity of Yb disrupts both Yb bodies and Flam bodies. Loss of Zucchini, an endoribonuclease necessary for piRNA maturation, enlarges Flam bodies, which now superimpose with Yb bodies. Yb directly binds flam, but not neighboring protein-coding gene, transcripts. Thus, Yb integrates piRNA processing factors and piRNA intermediates into Yb bodies and Flam bodies, respectively, through direct binding to enhance piRNA biogenesis and formation of piRNA-inducing silencing complexes.
2014-06-24 | GSE54875 | GEO
Project description:The piRNA pathway is a small RNA-based immune system that silences mobile genetic elements in animal germlines. In Drosophila ovaries, piRNAs are produced from discrete genomic loci, called piRNA clusters, which are composed of inactive transposon copies and fragments and thus constitute a genetically encoded memory of past transposon challenges. Two types of piRNA clusters exist in flies: dual-strand clusters, expressed only in the germline via a highly specialised machinery, and uni-strand cluster, which are predominantly expressed in the somatic follicle cells. Flamenco (flam) is the major uni-strand piRNA cluster in Drosophila, giving rise to the majority of somatic piRNAs. Flam resembles a canonical RNA polymerase II transcriptional unit, nonetheless it can be specifically recognised by the piRNA pathway and directed to the biogenesis machinery. Recent work has implicated the RNA helicase Yb in the licensing of somatic piRNA production, however a detailed understanding of the molecular mechanisms underlying flam export and specification is still lacking. Here, we show that flam export triggers the assembly of peri-nuclear condensates of Yb and provide evidence that piRNA production from flam specifically requires subunits of the Nuclear Pore Complex (NPC). In the absence of some NPC subunits, transposons become de-silenced and piRNA biogenesis is compromised exclusively from flam. We also show that Yb transiently associates with the NPC to promote flam export. Taken together, our data shed light on how the export of uni-strand cluster transcripts is achieved and suggest the evolution of a specialised machinery that couples transcription, nuclear export and piRNA production.
2021-04-15 | PXD019670 | Pride
Project description:The piRNA pathway is a small RNA-based immune system that silences mobile genetic elements in animal germlines. In Drosophila ovaries, piRNAs are produced from discrete genomic loci, called piRNA clusters, which are composed of inactive transposon copies and fragments and thus constitute a genetically encoded memory of past transposon challenges. Two types of piRNA clusters exist in flies: dual-strand clusters, expressed only in the germline via a highly specialised machinery, and uni-strand cluster, which are predominantly expressed in the somatic follicle cells. Flamenco (flam) is the major uni-strand piRNA cluster in Drosophila, giving rise to the majority of somatic piRNAs. Flam resembles a canonical RNA polymerase II transcriptional unit, nonetheless it can be specifically recognised by the piRNA pathway and directed to the biogenesis machinery. Recent work has implicated the RNA helicase Yb in the licensing of somatic piRNA production, however a detailed understanding of the molecular mechanisms underlying flam export and specification is still lacking. Here, we show that flam export triggers the assembly of peri-nuclear condensates of Yb and provide evidence that piRNA production from flam specifically requires subunits of the Nuclear Pore Complex (NPC). In the absence of some NPC subunits, transposons become de-silenced and piRNA biogenesis is compromised exclusively from flam. We also show that Yb transiently associates with the NPC to promote flam export. Taken together, our data shed light on how the export of uni-strand cluster transcripts is achieved and suggest the evolution of a specialised machinery that couples transcription, nuclear export and piRNA production.
2021-04-16 | PXD019671 | Pride
Project description:The piRNA pathway is a small RNA-based immune system that silences mobile genetic elements in animal germlines. In Drosophila ovaries, piRNAs are produced from discrete genomic loci, called piRNA clusters, which are composed of inactive transposon copies and fragments and thus constitute a genetically encoded memory of past transposon challenges. Two types of piRNA clusters exist in flies: dual-strand clusters, expressed only in the germline via a highly specialised machinery, and uni-strand cluster, which are predominantly expressed in the somatic follicle cells. Flamenco (flam) is the major uni-strand piRNA cluster in Drosophila, giving rise to the majority of somatic piRNAs. Flam resembles a canonical RNA polymerase II transcriptional unit, nonetheless it can be specifically recognised by the piRNA pathway and directed to the biogenesis machinery. Recent work has implicated the RNA helicase Yb in the licensing of somatic piRNA production, however a detailed understanding of the molecular mechanisms underlying flam export and specification is still lacking. Here, we show that flam export triggers the assembly of peri-nuclear condensates of Yb and provide evidence that piRNA production from flam specifically requires subunits of the Nuclear Pore Complex (NPC). In the absence of some NPC subunits, transposons become de-silenced and piRNA biogenesis is compromised exclusively from flam. We also show that Yb transiently associates with the NPC to promote flam export. Taken together, our data shed light on how the export of uni-strand cluster transcripts is achieved and suggest the evolution of a specialised machinery that couples transcription, nuclear export and piRNA production.
2021-07-02 | PXD019674 | Pride
Project description:Despite exciting progress in understanding the Piwi-associ-ated RNA (piRNA) pathway in the germ line, less is known about this pathway in somatic cells. We showed previously that Piwi, a key component of the piRNA pathway in Drosophila, is regulated in somatic cells by Yb, a novel protein containing an RNA helicase-like motif and a Tudor-like domain. Yb is specifically expressed in gonadal somatic cells and regulates piwi in somatic niche cells to control germ line and somatic stem cell self-renewal. However, the molecular basis of the regulation remains elusive. Here, we report that Yb recruits Armitage (Armi), a putative RNA helicase involved in the piRNA pathway, to the Yb body, a cytoplasmic sphere to which Yb is exclusively localized. Moreover, co-immunoprecipitation experiments show that Yb forms a complex with Armi. In Yb mu- tants, Armi is dispersed throughout the cytoplasm, and Piwi fails to enter the nucleus and is rarely detectable in the cytoplasm. Furthermore, somatic piRNAs are drastically diminished, and soma-expressing transposons are desilenced. These observations indicate a crucial role of Yb and the Yb body in piRNA biogenesis, possibly by regulating the activity of Armi that controls the entry of Piwi into the nucleus for its function. Finally, we discovered putative endo-siRNAs in the flamenco locus and the Yb dependence of their expression. These observations further implicate a role for Yb in transposon silencing via both the piRNA and endo-siRNA pathways. Examination of the effect of Yb on piRNA pathway
2011-02-10 | E-GEOD-26407 | biostudies-arrayexpress