Molecular basis for Cdk1-regulated timing of Mis18 complex assembly and CENP-A deposition

The centromere, a chromosomal locus that acts as a microtubule attachment site, is epigenetically specified by the enrichment of CENP-A nucleosomes. Centromere maintenance during the cell cycle requires HJURP-mediated CENP-A deposition, a process regulated by the Mis18 complex (Mis18 alpha/Mis18 beta/Mis18BP1). Spatial and temporal regulation of Mis18 complex assembly is crucial for its centromere association and function. Here, we provide the molecular basis for the assembly and regulation of the Mis18 complex. We show that the N-terminal region of Mis18BP1 spanning amino acid residues 20-130 directly interacts with Mis18 alpha/beta to form the Mis18 complex. Within Mis18 alpha/beta, the Mis18 alpha MeDiY domain can directly interact with Mis18BP1. Mis18 alpha/beta forms a hetero-hexamer with 4 Mis18 alpha and 2 Mis18 beta. However, only two copies of Mis18BP1 interact with Mis18 alpha/beta to form a hetero-octameric assembly, highlighting the role of Mis18 oligomerization in limiting the number of Mis18BP1 within the Mis18 complex. Furthermore, we demonstrate the involvement of consensus Cdk1 phosphorylation sites on Mis18 complex assembly and thus provide a rationale for cell cycle-regulated timing of Mis18 assembly and CENP-A deposition.