Determining the Mechanism of CSR-1 RNAi Pathway in Organizing Holocentromere in Caenorhabditis elegans

Abstract

CSR-1 RNAi pathway is required for the holocentromere organization and proper chromosome segregation in Caenorhabditis elegans. It remains obscure how this pathway functionally links to these two events. A previous study showed that germline genes, which are targets of CSR-1 RNAi pathway, are inversely correlated to HCP-3 occupancy on chromatin. We propose that CSR-1 RNAi pathway may restrict HCP-3 localization on the centromere. To study how the CSR-1 RNAi pathway affects the centromere function, we have closely examined the centromeric protein localization and expression followed by RNAi knockdown of the Argonaute CSR-1. By live imaging, we found that upon csr-1 knockdown, embryos have an altered localization of the centromeric histone H3 protein variant, HCP-3. Our imaging and biochemical data also show that csr-1 knockdown elevated both centromeric and cellular level of HCP-3, respectively, but it did not affect the bipolar arrangement of HCP-3 on centromere. We proposed that the increase in HCP-3 level, which is comparable to CENP-A overexpression in other organisms, may lead to merotelic kinetochore attachment and aneuploidy. We will also test if HCP-3 occupancy is reduced after tethering CSR-1 onto an HCP-3-enriched region by ChIP-qPCR. This study will provide insights on how RNAi and non-coding RNA transcripts may function in centromere organization and implicate how centromere are defined.