Structural Basis for Mis18 Complex Assembly: Implications for Centromere Maintenance

Abstract

The centromere, defined by the enrichment of CENP-A (a Histone H3 variant) containing nucleosomes, is a specialised chromosomal locus that acts as a microtubule attachment site. To preserve centromere identity, CENP-A levels must be maintained through active CENP-A loading during the cell cycle. A central player mediating this process is the Mis18 complex (Mis18α, Mis18ý and Mis18BP1), which recruits the CENP-A specific chaperone HJURP to centromeres for CENP-A deposition. Here, using a multi-pronged approach, we characterise the structure of the Mis18 complex and show that multiple hetero- and homo-oligomeric interfaces facilitate the hetero-octameric Mis18 complex assembly composed of 4 Mis18α, 2 Mis18ý and 2 Mis18BP1. Evaluation of structure-guided/separation-of-function mutants reveals structural determinants essential for Mis18 complex assembly and centromere maintenance. Our results provide new mechanistic insights on centromere maintenance, highlighting that while Mis18α can associate with centromeres and deposit CENP-A independently of Mis18ý, the latter is indispensable for the optimal level of CENP-A loading required for preserving the centromere identity.

Direct recruitment of Mis18 to interphase spindle poles promotes CENP-A chromatin assembly

“…Association of the conserved Yippee domain of hMis18α with hM18BP1 is required for hMis18α recruitment to human centromeres 25,26,30,72 . As our structural analyses predict that the Yippee domain of S. pombe Mis18 mediates its interaction with the Sad1 N-terminal region (Figure 4C), we conclude that Sad1 performs the equivalent function of human M18BP1 in mediating S. pombe Mis18C recruitment 26,27,30,72 .…”

“…Association of the conserved Yippee domain of hMis18α with hM18BP1 is required for hMis18α recruitment to human centromeres 25,26,30,72 . As our structural analyses predict that the Yippee domain of S. pombe Mis18 mediates its interaction with the Sad1 N-terminal region (Figure 4C), we conclude that Sad1 performs the equivalent function of human M18BP1 in mediating S. pombe Mis18C recruitment 26,27,30,72 . Our finding that neither the N-nor C-terminal domains of Mis18 were sufficient to bind the Sad1 nucleoplasmic region in vitro (Figure S4A), and that the short Mis18 C-terminal tail was required for robust binding (Figures S4B-S4D), suggests additional binding interactions between Mis18 and Sad1 may exist.…”