| description |
RNA interference (RNAi) is a degradative cell defense mechanism that uses double-stranded RNA (dsRNA) to regulate the intracellular levels of target RNA with homologous sequences. Although RNAi-mediated post-transcriptional silencing (PTGS) is well documented, RNAi-mediated transcriptional gene silencing (TGS) and in particular the role of RNAi in chromatin are still elusive. Here we investigate RNAi-mediated TGS in Drosophila. We show that RNAi components Dicer-2 (Dcr2) and Argonaute-2 (AGO2) associate with chromatin, with strong preference for euchromatic, transcriptionally active loci and that loss of Dcr2 and AGO2 affects the basal level of expression of various genes including hsp70 and hsp68 heat shock loci. We find that Dcr2 and AGO2 interact with the transcription machinery as they associate with RNA polymerase II (Pol II) and negative elongation factor NELF. Notably Dcr2 and AGO2 loss of function show that transcriptional defects are accompanied by perturbation of Pol II positioning on promoters. Further, both Dcr2 and Ago2 null mutations as well as missense mutations compromising the RNAi activity impair Pol II dynamics upon heat shock, suggesting a global role for chromatin-associated RNAi components in the transcriptional stress response. Finally, AGO2 RIP-seq experiments performed in normal and heat shock conditions reveal that, particularly upon stress, AGO2 is strongly enriched in small-RNAs encompassing promoter as well as other parts of heat shock and other gene loci on both sense and antisense, with a strong bias for antisense, particularly after heat shock. Taken together our results reveal a new scenario for RNAi function in Drosophila chromatin in which Dcr2 and AGO2 are globally associated with transcriptionally active loci and may play a pivotal role in shaping the transcriptome by controlling RNA Pol II processivity in the context of pervasive sense/antisense transcription none provided |