CENP-C unwraps the CENP-A nucleosome through the H2A C-terminal tail

Centromeres are defined epigenetically by nucleosomes containing the histone H3 variant CENPA, upon which the constitutive centromere-associated network of proteins (CCAN) is built. CENPC, is considered to be a central organizer of the CCAN. We provide new molecular insights into the structure of CENP-A nucleosomes, in isolation and in complex with the CENP-C central region (CENP-CCR), the main CENP-A binding module of CENP-C. We establish that the short αN-helix of CENP-A promotes DNA flexibility at the nucleosome ends, independently of the sequence it wraps. Furthermore, we show that, in vitro, two regions of CENP-C (CENP-CCR and CENP-Cmotif ) both bind exclusively to the CENP-A nucleosome. We find CENP-CCR to bind with high affinity due to an extended hydrophobic area made up of CENP-AV532 and CENP-AV533. Importantly, we identify two key conformational changes within the CENP-A nucleosome upon CENP-C binding. First, the loose DNA wrapping of CENP-A nucleosomes is further exacerbated, through destabilization of the H2A N-terminal tail. Second, CENP-CCR rigidifies the N-terminal tail of H4 in the conformation favoring H4K20 monomethylation, essential for a functional centromere.