Acetylation of histone H4 lysine 5 and 12 is required for CENP-A deposition into centromeres

Abstract

Centromeres are specified epigenetically through the deposition of the centromere-specific histone H3 variant CENP-A. However, how additional epigenetic features are involved in centromere specification is unknown. Here, we find that histone H4 Lys5 and Lys12 acetylation (H4K5ac and H4K12ac) primarily occur within the pre-nucleosomal CENP-A–H4–HJURP (CENP-A chaperone) complex, before centromere deposition. We show that H4K5ac and H4K12ac are mediated by the RbAp46/48–Hat1 complex and that RbAp48-deficient DT40 cells fail to recruit HJURP to centromeres and do not incorporate new CENP-A at centromeres. However, C-terminally-truncated HJURP, that does not bind CENP-A, does localize to centromeres in RbAp48-deficient cells. Acetylation-dead H4 mutations cause mis-localization of the CENP-A–H4 complex to non-centromeric chromatin. Crucially, CENP-A with acetylation-mimetic H4 was assembled specifically into centromeres even in RbAp48-deficient DT40 cells. We conclude that H4K5ac and H4K12ac, mediated by RbAp46/48, facilitates efficient CENP-A deposition into centromeres.

Posttranslational mechanisms controlling centromere function and assembly

“…Posttranslational modification of the histone H4 subunit is required for proper deposition of the CENP-A nucleosome. RbAp46 and RbAp48 subunits found in many chromatin assembly complexes are associated with pre-nucleosomal CENP-A/H4 and HJURP [29,35,36 •• ] (Figure 2b). Mis16, the S. pombe homolog of RbAp46/48, recognizes both CENP-A and histone H4 in the heterodimer [37].…”

“…In this respect, the modification of the preassembled CENP-A/H4 dimer is similar to histone H3/H4, which is also acetylated on the same residues of histone H4 [26]. Depletion of RbAp46/48/Mis16 in several organisms, or the mutation of histone H4 Lys5 or Lys12 residues to non-acetylated amino acids, reduces new CENP-A deposition [36 •• ,38,39]; implying that the modification of the H4 subunit of the pre-nucleosomal heterodimer is essential for CENP-A deposition.…”

Posttranslational modifications of CENP-A: marks of distinction

“…Modifications on histone H4 present in the prenucleosomal complex are also involved in CENP-A deposition. Histone H4 Lys-5 and Lys-12 acetylations occur in the pre-deposition complex, are removed after CENP-A nucleosome formation, and appear to be critical for deposition (Fukagawa 2017; Shang et al 2016). …”

Orchestrating the Specific Assembly of Centromeric Nucleosomes

“…Indeed, histone H4 is acetylated on K5ac and K12ac within the prenucleosomal complex, and these modifications are necessary for CENP-A deposition (Fig. 1) (Shang et al 2016). …”

“…A crystal structure of the Mis16-Scm3-CENP-A Cnp1 /H4 complex shows that Mis16 contacts both the Scm3 chaperone and histone H4 (An et al 2015). Depletion of RbAp proteins reduces HJURP recruitment and new CENP-A deposition (Dunleavy et al 2009; Shang et al 2016). K5 and K12 acetylation of the histone H4 bound to CENP-A within the prenucleosomal complex are dependent on RbAp48, and these modifications are required for CENP-A deposition in vivo (Fig.…”