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

Abstract

AbstractCentromeres are defined epigenetically by nucleosomes containing the histone H3 variant CENP-A, upon which the constitutive centromere-associated network of proteins (CCAN) is built. CENP-C, 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-CCRand CENP-Cmotif) both bind exclusively to the CENP-A nucleosome. We find CENP-CCRto bind with high affinity due to an extended hydrophobic area made up of CENP-AV532and 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-CCRrigidifies the N-terminal tail of H4 in the conformation favoring H4K20monomethylation, essential for a functional centromere.SynopsisCENP-A nucleosomes have a short αN helix incompatible with complete DNA wrapping, independently of DNA sequence. CENP-C binds exclusively to CENP-A nucleosomes and this binding induces conformational changes that further differentiate CENP-A-containing from canonical nucleosomes.CENP-C binds CENP-A nucleosomes specificallyDNA ends of the CENP-A nucleosome are further unwrapped in the CENP-A/CENP-C complex, due to flexible H2A C-terminal tailsThe N-terminal tail of H4 adopts a conformation favored for centromere specific H4K20monomethylation when CENP-C is bound