Yra2 regulates proteolysis of Cse4 to prevent its mislocalization to non-centromeric regions for chromosomal stability in budding yeast

Restricting the centromere-specific histone H3 variant Cse4 (CENP-A in human) to centromeric chromatin ensures faithful chromosome segregation. Mislocalization of overexpressed Cse4/CENP-A to non-centromeric regions contributes to chromosomal instability (CIN), an important hallmark of many cancers. Hence, defining mechanisms that prevent mislocalization of Cse4 is clinically significant. Here we report a role for YRA2 (Yeast RNA Annealing Protein 2) in ubiquitin mediated proteolysis of Cse4 to prevent its mislocalization for chromosomal stability. YRA2 was identified in a genome-wide screen for gene deletions that exhibit synthetic dosage lethality (SDL) upon overexpression of CSE4 (GALCSE4). We determined that yra2Δ strains exhibit increased Cse4 stability, enriched Cse4 chromatin association, reduced Cse4 ubiquitination, Cse4 mislocalization, and CIN. Defects in interaction of E3 ubiquitin ligase Psh1 with Cse4 contributes to stability of Cse4 in yra2Δ strains. Consistent with these results, overexpression of PSH1 suppresses GALCSE4 SDL in yra2Δ strain. Yra2 mediated proteolysis of Cse4 is separable from its RNA related functions as GALCSE4 SDL was not observed in yra2F96A,F108A strain with mutations in Yra2 RNA recognition motif. Furthermore, poly(A)+ RNA export mutants in YRA1 (yra1-2) and MEX67 (mex67-5), that interact with Yra2, do not exhibit GALCSE4 SDL and defects in RNA export are not observed in yra2Δ cells. We determined that evolutionarily conserved C-terminal ChTOP domain of Yra2 with an intact N-terminal RNA binding domain exhibits GALCSE4 SDL and defects in Cse4 proteolysis. In summary, we have defined a role for Yra2 in preventing mislocalization of Cse4 by facilitating its proteolysis for chromosomal stability.