Structural plasticity of CENP-A regulated by H4 influences cellular levels and kinetochore assembly

The Histone variant CENP-ACse4 is a core component of the specialized nucleosome at the centromere in budding yeast. The level of Cse4 in cells is tightly regulated, primarily by ubiquitin-mediated proteolysis. However, the structural transitions in Cse4 that regulate centromere localization and interaction with regulatory components are poorly understood. Using time resolved fluorescence, NMR and molecular dynamics we show for the first time that soluble Cse4 can exist in a closed conformation, inaccessible to various regulatory components. We further determined that binding of its obligate partner H4, alters the inter-domain interaction within Cse4, ensuring an open state that will lend itself to proteolysis. This dynamic model allows kinetochore formation only in presence of H4, as the N-terminus, which is required for interaction with centromeric components will be unavailable in absence of H4. The specific requirement of H4 binding for the conformational regulation of Cse4 suggests a unique structure-based regulatory mechanism for Cse4 localization and prevention of premature kinetochore assembly.