Small regulatory RNAs inhibit RNA polymerase II during the elongation phase of transcription

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Abstract
Small RNAs function in both the cytoplasm, inhibiting expression from messenger RNAs, and in the nucleus, to silence heterochromatin and prevent genome rearrangement. In this study, Guang et al. characterize a new protein involved in RNA interference in the nucleus. NRDE-2 associates with the Argonaute protein NRDE-3 and siRNAs on nascent transcripts. This association prevents elongation by RNA polymerase II, thereby making this a co-transcriptional form of gene silencing. Small regulatory RNAs function both in the cytoplasm, inhibiting expression from messenger RNAs, and in the nucleus, silencing heterochromatin and preventing genome rearrangement. Now a new protein involved in RNA interference in the nucleus has been characterized. This protein, NRDE-2, associates with NRDE-3 and short interfering RNAs on nascent transcripts. This association prevents elongation of the transcripts by RNA polymerase II, making this a co-transcriptional form of gene silencing. Eukaryotic cells express a wide variety of endogenous small regulatory RNAs that regulate heterochromatin formation, developmental timing, defence against parasitic nucleic acids and genome rearrangement. Many small regulatory RNAs are thought to function in nuclei1,2. For instance, in plants and fungi, short interfering RNA (siRNAs) associate with nascent transcripts and direct chromatin and/or DNA modifications1,2. To understand further the biological roles of small regulatory RNAs, we conducted a genetic screen to identify factors required for RNA interference (RNAi) in Caenorhabditis elegans nuclei3. Here we show that the gene nuclear RNAi defective-2 (nrde-2) encodes an evolutionarily conserved protein that is required for siRNA-mediated silencing in nuclei. NRDE-2 associates with the Argonaute protein NRDE-3 within nuclei and is recruited by NRDE-3/siRNA complexes to nascent transcripts that have been targeted by RNAi. We find that nuclear-localized siRNAs direct an NRDE-2-dependent silencing of pre-messenger RNAs (pre-mRNAs) 3′ to sites of RNAi, an NRDE-2-dependent accumulation of RNA polymerase (RNAP) II at genomic loci targeted by RNAi, and NRDE-2-dependent decreases in RNAP II occupancy and RNAP II transcriptional activity 3′ to sites of RNAi. These results define NRDE-2 as a component of the nuclear RNAi machinery and demonstrate that metazoan siRNAs can silence nuclear-localized RNAs co-transcriptionally. In addition, these results establish a novel mode of RNAP II regulation: siRNA-directed recruitment of NRDE factors that inhibit RNAP II during the elongation phase of transcription.