CSR-1 is an argonaute of a RNA interference pathway that is important for chromosome segregation in C. elegans. Live-cell imaging revealed that CSR-1 depletion slows down spindle pole separation in a kinetochore-dependent manner. In csr-1(RNAi) embryos, the kinetochores may be misattached to the microtubules and chromosome segregation is disrupted. On the holocentromeres, there are increased levels of some kinetochore proteins, including the centromeric epigenetic mark, CENP-A or HCP-3. Without affecting HCP-3 expression level, HCP-3 density is higher on stretched chromatin fibers in CSR-1-depleted embryos. The increased HCP-3 deposition on chromatin after CSR-1 depletion is at least partially independent of HCP-3 loading factors, KNL-2 and LIN-53, suggesting a non-classical, improper HCP-3 loading pathway. Negative regulation of HCP-3 holocentromere loading by CSR-1 required its slicer activity and the b isoform. CSR-1 acts as a HCP-3 repressor for its chromosomal occupancy, shedding light on the role of RNAi pathways in specifying the localization of centromere proteins.
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