Investigation into the cohesin loading complex of Caenorhabditis elegans

Abstract

Sister chromatid cohesion is important to ensure faithful chromosome segregation occurs during mitosis and is mediated by the multisubunit cohesin complex. In Saccharomyces cereviside the cohesin complex is loaded onto chromosomes by a protein complex composed of Scc2 and Scc4. Caenorhabdifis elegans PQN-85 is readily identified as the orthologue of Scc2 and the sequence similarity between PQN-85 and its orthologues from all organisms suggests that their function in cohesin loading is conserved throughout evolution. Data presented in this thesis shows that in accordance with its suspected role in cohesin loading RNA interference against pqn-85 results in chromosome segregation defects in C elegans embryos similar to those observed when the cohesin subunit scc-3 is depleted. Although it has been problematic to identify homologues of Scc4 in metazoans, it has recently been proven that C elegans MAU-2 is distantly related to Scc4. Co-depleting scc-3 with either pqn-85 or mau-2 result in exacerbation of the chromosome segregation defects with very similar phenotypes suggesting that mau-2 as well as pqn-85 functions in chromosome segregation. Data is also presented demonstrating that MAU-2 physically interacts with the N-terminus of the PQN-85 protein in a directed yeast two-hybrid assay consistent with the idea that PQN-85 and MAU-2 form a complex. Microscopic examination of mau-2:: GFP embryos in this thesis demonstratesth at MAU-2:: GFP localises to the nucleus in accordancew ith a role in cohesin loading, whilst depletion of pqn-85 by RNAi in mau-2:: GFP embryos resulted in the reduction of MAU-2 :: GFP within the nucleus and concurrent accumulation within the cytoplasm. Together these results suggest that the cohesin loading complex is conserved in C elegans whilst the extensive homology between PQN-85, MAU-2 and their metazoan orthologues suggests that it is also likely that the cohesin complex is conserved amongst all metazoans.