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. 2014 Dec;20(12):1977–1986. doi: 10.1261/rna.046300.114

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© 2014 Post et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society

This article is distributed exclusively by the RNA Society for the first 12 months after the full-issue publication date (see http://rnajournal.cshlp.org/site/misc/terms.xhtml). After 12 months, it is available under a Creative Commons License (Attribution-NonCommercial 4.0 International), as described at http://creativecommons.org/licenses/by-nc/4.0/.

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FIGURE 4. — PIWI-mediated TGS operates through chromatin changes. (A) Chromatin immunoprecipitation (ChIP) assays measuring RNA Polymerase II (RNA Pol II) and Histone H3 lysine 9 tri-methylation (H3K9me3) enrichment at OSS cell genomic loci and the flamenco piRNA reporter. Blood is an endogenous TE that is epigenetically silenced, Actin5C is a constitutively expressed housekeeping gene locus, Mec2 is a locus with a de novo 297 TE and under PIWI-mediated silencing in OSS cells (Sytnikova et al. 2014), and the flam element 3′-UTR reporter constructs were transiently transfected plasmids. Error bars represent standard error of mean of biological triplicates. Wilcoxon rank-sum tests for certain comparisons of siPIWI versus siGFP conditions are represented by (***) P < 0.05, (**) P < 0.1, and (*) P < 0.2. The inset for the Blood ChIP is a different scale of the main figure to show the RNA Pol II levels. (B) A model that suggests the dominant mechanism for PIWI directed gene silencing is TGS and this mechanism requires a bulk of piRNAs interacting with the nascent transcript. Most protein-coding transcripts are unaffected by less-frequent PIWI/piRNA interactions.