CryoEM structure of human CENP-A nucleosome alone and in complex with central domain of human CENP-C

Centromere is a chromosome locus necessary for genetic stability during cell division. Centromere is defined epigenetically, by nucleosome containing histone H3 variant, CENP-A, upon which constitutive centromere-associated network of proteins (CCAN) is built. CENP-C, a member of CCAN, is considered to be a blueprint of centromere. It interacts with several CCAN components and directly bridges CENP-A nucleosomes to kinetochore in mitosis. In this study, we provide new molecular details on structure of CENP-A nucleosome alone and in complex with CENP-C central region (CENP-CCR), main CENP-A binding module of CENP-C. We confirm high-flexibility of DNA ends as intrinsic features of CENP-A-containing nucleosomes independent on DNA sequence it wraps. We establish that, in vitro, two regions of CENP-C (CENP-CCR and CENP-Cmotif), both bind exclusively CENP-A nucleosome, albeit CENP-CCR with a higher affinity due to extended hydrophobic area made of CENP-AV532 and CENP-AV533. Finally, we identify conformational changes on CENP-A nucleosome that occur upon CENP-C binding. Pronounced DNA unwrapping is facilitated by destabilization of H2A N-terminal tail and N-terminal tail of H4 is stabilized in the upward conformation favored for CENP-A specific H4K20 monomethylation. Together, our findings provide foundation for understanding epigenetic basis for centromere formation and functioning.