Proteolysis contributes to the exclusive centromere localization of the yeast Cse4/CENP-A histone H3 variant

Kinetochores are the specialized protein structures that form on centromeric DNA and direct chromosome segregation. It is critical that all chromosomes assemble a single kinetochore every cell cycle. One hallmark of all eukaryotic kinetochores is CENP-A, an essential centromeric histone H3 (CenH3) variant. Overexpression of CENP-A causes mislocalization to euchromatin, which could lead to deleterious consequences because CENP-A overexpression is associated with colorectal cancer [1, 2]. Although CENP-A protein levels are important for genomic stability, little is known about the mechanisms of CenH3 regulation. Here, we show that the levels of the budding yeast CenH3, Cse4, are regulated by ubiquitin-proteasome-mediated proteolysis. Because mutation of all Cse4 lysine residues did not completely stabilize the protein, we isolated a dominant lethal mutant, CSE4-351, that was stable. The Cse4-351 protein localized to euchromatin, suggesting that proteolysis prevents CenH3 euchromatic localization. When wild-type Cse4 was fused to a degron signal, the soluble Cse4 protein was rapidly degraded, but the centromere bound Cse4 was stable, indicating that centromere localization protects Cse4 from degradation. Taken together, these data identify proteolysis as one mechanism that contributes to the restricted centromere localization of the yeast CenH3.