Characterization of AGO2 complexes

Abstract The AGO2 protein is part of a highly conserved family of RNA-binding protein called Argonaute. This protein family plays a key role in gene expression regulation in association with different classes of small non-coding RNA (miRNA, siRNA, piRNA). In addition to the post-transcriptional regulation in the cytoplasm, AGO2 protein acts also in the nucleus at the transcriptional level with a mechanism, in humans, not clearly identified. To elucidate the molecular functions of AGO2 in human cells, we performed an extensive analysis of both proteins and small RNAs associated to endogenous AGO2 protein by high-throughput technologies. We identified various AGO2-interactors not previously described. Interactors that can shed light on molecular functions exerted by AGO2 in novel as well in known biological processes. In particular, we provided several evidence regarding the mechanism of AGO2-mediated transcriptional regulation. Interestingly, we uncovered the interaction of AGO2 with the SWI/SNF chromatin remodeling complexes. The SWI/SNF is a multisubunit complex that alter the structure or the positioning of nucleosomes, thus modulating the access of regulatory proteins and transcription factors to DNA. Futhermore, we identified a new class of AGO2-associated small RNAs in the nucleus that showed a remarkable overlap with the binding sites of SWI/SNF complexes, regulatory regions in proximity of transcription start site and with chromatin marks associated with transcriptional active state. Taken together, our data suggests that nuclear AGO2 regulates the transcriptional initiation interacting with SWI/SNF complexes to ensure proper nucleosome positioning in proximity of promoter regions.