Stable centromere association of the yeast histone variant Cse4 requires its essential N-terminal domain
Chromosome segregation relies on kinetochores that assemble on specialized centromeric chromatin containing a histone H3 variant. In budding yeast, a single centromeric nucleosome containing Cse4 assembles at a sequence-defined 125 bp centromere. Yeast centromeric sequences are poor templates for nucleosome formation in vitro, suggesting the existence of mechanisms that specifically stabilize Cse4 nucleosomes in vivo. The extended Cse4 N-terminal tail binds to the chaperone Scm3, and a short essential region called END within the N-terminal tail binds the inner kinetochore complex OA. To address the roles of these interactions, we utilized single molecule fluorescence assays to monitor Cse4 during kinetochore assembly. We found that OA and Scm3 independently stabilize Cse4 at centromeres via their END interaction. Scm3 binding to the Cse4 END is enhanced by Ipl1/Aurora B phosphorylation, identifying a previously unknown role for Ipl1 in ensuring Cse4 stability. Strikingly, an Ipl1 phosphomimetic mutation in the Cse4 END enhances Scm3 binding and can restore Cse4 recruitment in mutants defective in OA binding. Together, these data suggest that a key function of the essential Cse4 N-terminus is to ensure Cse4 localization at centromeres.