DmGTSF1 is necessary for Piwi–piRISC-mediated transcriptional transposon silencing in the <i>Drosophila</i> ovary

Ohtani, Hitoshi; Iwasaki, Yuka W.; Shibuya, Aoi; Siomi, Haruhiko; Siomi, Mikiko C.; Saito, Kuniaki

doi:10.1101/gad.221515.113

Cited by 136 publications

(199 citation statements)

References 39 publications

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“…6A,C; Supplemental Fig. S5D; Ohtani et al 2013). Consistent with this, HP1a is present at piRNA-targeted TE insertions as well as at the flanking genomic regions in a Piwi-dependent manner (Fig.…”

Section: Cg9754/silencio Is Required For H3k9 Methylation and Transcrsupporting

confidence: 66%

“…However, neither of these factors is conserved in animals. In turn, two proteins with largely unknown functions that are required for piRNA-guided transcriptional silencing in animals (Gtsf1/Asterix and Maelstrom) (Sienski et al 2012;Donertas et al 2013;Muerdter et al 2013;Ohtani et al 2013) are absent in S. pombe, highlighting significant molecular differences in these two systems. Moreover, the Piwi-piRNA complex fails to mediate transcriptional silencing if recruited artificially (not involving a piRNAtarget interaction) to a reporter RNA in a Drosophila cell culture model that expresses a fully functional piRNA pathway centered on nuclear Piwi (Post et al 2014).…”

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confidence: 99%

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Silencio/CG9754 connects the Piwi–piRNA complex to the cellular heterochromatin machinery

et al. 2015

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The repression of transposable elements in eukaryotes often involves their transcriptional silencing via targeted chromatin modifications. In animal gonads, nuclear Argonaute proteins of the PIWI clade complexed with small guide RNAs (piRNAs) serve as sequence specificity determinants in this process. How binding of nuclear PIWIpiRNA complexes to nascent transcripts orchestrates heterochromatin formation and transcriptional silencing is unknown. Here, we characterize CG9754/Silencio as an essential piRNA pathway factor that is required for Piwimediated transcriptional silencing in Drosophila. Ectopic targeting of Silencio to RNA or DNA is sufficient to elicit silencing independently of Piwi and known piRNA pathway factors. Instead, Silencio requires the H3K9 methyltransferase Eggless/SetDB1 for its silencing ability. In agreement with this, SetDB1, but not Su(var)3-9, is required for Piwi-mediated transcriptional silencing genome-wide. Due to its interaction with the target-engaged PiwipiRNA complex, we suggest that Silencio acts as linker between the sequence specificity factor Piwi and the cellular heterochromatin machinery.[Keywords: H3K9 methylation; Piwi; transposon silencing; heterochromatin formation; piRNA pathway; transcriptional silencing] Supplemental material is available for this article. In fungi, plants, and animals, nuclear Argonaute proteins target nascent transcripts-RNAs that are still attached to their originating DNA locus via the transcribing RNA polymerase. This provides an opportunity for small RNA silencing pathways to connect to the various cellular chromatin-modifying activities for the sequence-specific formation of heterochromatin, typically at transposable element (TE) insertions (Castel and Martienssen 2013).Pioneering work in fission yeast has identified methylation of histone H3 at Lys9 (H3K9me2/3) and histone deacetylation as two major hallmarks that are required for small RNA-guided silencing and heterochromatin formation (Nakayama et al. 2001;Hall et al. 2002;Volpe et al. 2002;Yamada et al. 2005). A sequential order of events downstream from the recruitment of the Schizosaccharomyces pombe Ago1/siRNA complex to nascent RNA of centromeric repeats has been described. According to this, the Ago1/siRNA complex recruits the H3K9 methyltransferase Clr4 to chromatin, which results in H3K9 methylation (Noma et al. 2004;Verdel et al. 2004). This is believed to be a binding platform for the chromodomain-containing protein Swi6/HP1, which in turn recruits the major histone deacetylase and remodeling complex SHREC (Sugiyama et al. 2007). The final outcome of these events is the establishment of a nucleosome-dense region with low histone turnover, which effectively prevents the recruitment of RNA polymerase II (Pol II) to transcription initiation sites and hence transcription as such (e.g., Aygun et al. 2013).H3K9 methylation is also a hallmark of small RNAguided heterochromatin formation in plants, ciliates, and multicellular animals. In animal gonads, many TE insertions are methylated at...

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Section: Cg9754/silencio Is Required For H3k9 Methylation and Transcrsupporting

confidence: 66%

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confidence: 99%

Silencio/CG9754 connects the Piwi–piRNA complex to the cellular heterochromatin machinery

et al. 2015

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“…Several recent genome-wide studies examining mutants and genetic perturbations of the Piwi pathway in flies have confirmed the transcriptional silencing mechanism of TEs by PIWI via a transcription-repressive chromatin state (Sienski et al 2012;Huang et al 2013;Le Thomas et al 2013;Ohtani et al 2013;Rozhkov et al 2013). Here, we present a reporter assay platform that shows a dependence on PIWI and piRNA complementarity for triggering robust gene silencing, and the silencing capacity appeared to be present in both TE-directed piRNAs as well as genic piRNAs.…”

Section: Discussionmentioning

confidence: 95%

The capacity of target silencing by Drosophila PIWI and piRNAs

Post

Clark

Sytnikova

et al. 2014

RNA

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Although Piwi proteins and Piwi-interacting RNAs (piRNAs) genetically repress transposable elements (TEs), it is unclear how the highly diverse piRNA populations direct Piwi proteins to silence TE targets without silencing the entire transcriptome. To determine the capacity of piRNA-mediated silencing, we introduced reporter genes into Drosophila OSS cells, which express microRNAs (miRNAs) and piRNAs, and compared the Piwi pathway to the Argonaute pathway in gene regulation. Reporter constructs containing several target sites that were robustly silenced by miRNAs were not silenced to the same degrees by piRNAs. However, another set of reporters we designed to enable a large number of both TE-directed and genic piRNAs to bind were robustly silenced by the PIWI/piRNA complex in OSS cells. These reporters show that a bulk of piRNAs are required to pair to the reporter's transcripts and not the reporter's DNA sequence to engage PIWI-mediated silencing. Following our genome-wide study of PIWI-regulated targets in OSS cells, we assessed candidate gene elements with our reporter platform. These results suggest TE sequences are the most direct of PIWI regulatory targets while coding genes are less directly affected by PIWI targeting. Finally, our study suggests that the PIWI transcriptional silencing mechanism triggers robust chromatin changes on targets with sufficient piRNA binding, and preferentially regulates TE transcripts because proteincoding transcripts lack a threshold of targeting by piRNA populations. This reporter platform will facilitate future dissections of the PIWI-targeting mechanism.

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“…Although miRNAs and endo-siRNAs are expressed in both animals and plants, piRNA expression is largely restricted to animal gonads, where they primarily mediate TE silencing for genome integrity and germ cell development (Thomson and Lin 2009;Siomi et al 2011). piRNAs act as guides for the cleavage of TE transcripts by PIWI protein Slicer activity in the cytoplasm, which often couples with the ping-pong cycle, or for Slicer-independent heterochromatin formation at TE loci in the nucleus (Sienski et al 2012;Dönertas et al 2013;Huang et al 2013;Le Thomas et al 2013;Muerdter et al 2013;Ohtani et al 2013).…”

Section: Introductionmentioning

confidence: 99%

Small RNA profiling and characterization of piRNA clusters in the adult testes of the common marmoset, a model primate

Hirano

Iwasaki

Lin

et al. 2014

RNA

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Small RNAs mediate gene silencing by binding Argonaute/Piwi proteins to regulate target RNAs. Here, we describe small RNA profiling of the adult testes of Callithrix jacchus, the common marmoset. The most abundant class of small RNAs in the adult testis was piRNAs, although 353 novel miRNAs but few endo-siRNAs were also identified. MARWI, a marmoset homolog of mouse MIWI and a very abundant PIWI in adult testes, associates with piRNAs that show characteristics of mouse pachytene piRNAs. As in other mammals, most marmoset piRNAs are derived from conserved clustered regions in the genome, which are annotated as intergenic regions. However, unlike in mice, marmoset piRNA clusters are also found on the X chromosome, suggesting escape from meiotic sex chromosome inactivation by the X-linked clusters. Some of the piRNA clusters identified contain antisenseorientated pseudogenes, suggesting the possibility that pseudogene-derived piRNAs may regulate parental functional proteincoding genes. More piRNAs map to transposable element (TE) subfamilies when they have copies in piRNA clusters. In addition, the strand bias observed for piRNAs mapped to each TE subfamily correlates with the polarity of copies inserted in clusters. These findings suggest that pachytene piRNA clusters determine the abundance and strand-bias of TE-derived piRNAs, may regulate protein-coding genes via pseudogene-derived piRNAs, and may even play roles in meiosis in the adult marmoset testis.

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DmGTSF1 is necessary for Piwi–piRISC-mediated transcriptional transposon silencing in the Drosophila ovary

Cited by 136 publications

References 39 publications

Silencio/CG9754 connects the Piwi–piRNA complex to the cellular heterochromatin machinery

Silencio/CG9754 connects the Piwi–piRNA complex to the cellular heterochromatin machinery

The capacity of target silencing by Drosophila PIWI and piRNAs

Small RNA profiling and characterization of piRNA clusters in the adult testes of the common marmoset, a model primate

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