Mechanism of centromere recruitment of the CENP-A chaperone HJURP and its implications for centromere licensing

Nucleosomes containing the histone H3 variant CENP-A are the epigenetic mark of centromeres, the kinetochore assembly sites required for chromosome segregation. HJURP is the CENP-A chaperone, which associates with Mis18 alpha, Mis18 beta, and M18BP1 to target centromeres and deposit new CENP-A. How these proteins interact to promote CENP-A deposition remains poorly understood. Here we show that two repeats in human HJURP proposed to be functionally distinct are in fact interchangeable and bind concomitantly to the 4: 2: 2 Mis18 alpha:Mis18 beta:M18BP1 complex without dissociating it. HJURP binds CENP-A: H4 dimers, and therefore assembly of CENP-A: H4 tetramers must be performed by two Mis18 alpha beta:M18BP1:HJURP complexes, or by the same complex in consecutive rounds. The Mis18 alpha N-terminal tails blockade two identical HJURP-repeat binding sites near the Mis18 alpha beta C-terminal helices. These were identified by photo-cross-linking experiments and mutated to separate Mis18 from HJURP centromere recruitment. Our results identify molecular underpinnings of eukaryotic chromosome inheritance and shed light on how centromeres license CENP-A deposition.