Dissecting the Molecular Roles of Histone Chaperones in Histone Acetylation by Type B Histone Acetyltransferases (HAT-B)

Abstract

The HAT-B enzyme complex is responsible for acetylating newly synthesized histone H4 on lysines K5 and K12. HAT-B is a multisubunit complex composed of the histone acetyltransferase 1 (Hat1) catalytic subunit and the Hat2 (rbap46) histone chaperone. Hat1 is predominantly localized in the nucleus as a member of a trimeric NuB4 complex containing Hat1, Hat2, and a histone H3-H4 specific histone chaperone called Hif1 (NASP). In addition to Hif1 and Hat2, Hat1 interacts with Asf1 (antisilencing function 1), a histone chaperone that has been reported to be involved in both replication-dependent and -independent chromatin assembly. To elucidate the molecular roles of the Hif1 and Asf1 histone chaperones in HAT-B histone binding and acetyltransferase activity, we have characterized the stoichiometry and binding mode of Hif1 and Asf1 to HAT-B and the effect of this binding on the enzymatic activity of HAT-B. We find that Hif1 and Asf1 bind through different modes and independently to HAT-B, whereby Hif1 binds directly to Hat2, and Asf1 is only capable of interactions with HAT-B through contacts with histones H3-H4. We also demonstrate that HAT-B is significantly more active against an intact H3-H4 heterodimer over a histone H4 peptide, independent of either Hif1 or Asf1 binding. Mutational studies further demonstrate that HAT-B binding to the histone tail regions is not sufficient for this enhanced activity. Based on these data, we propose a model for HAT-B/histone chaperone assembly and acetylation of H3-H4 complexes.Histone acetylation has been linked to diverse functions including transcriptional activation, gene silencing, DNA repair, cell cycle progression, chromatin maturation and dynamics, and nucleosome assembly (1, 2). There are two main classes of enzymes responsible for acetylation of histones: type A and type B histone acetyltransferases. Type A histone acetyltransferases are nuclear enzymes that acetylate histones in the context of chromatin, whereas type B acetyltransferases are responsible for the acetylation of newly synthesized histones H3 and H4. Although histone H3 is only moderately acetylated upon its synthesis (ϳ30%) (3, 4), newly synthesized histone H4 is highly acetylated on lysine residues K5 (H4K5) and K12 (H4K12) (3, 5, 6). These modifications are transient, and once transported across the nucleus, the histones are deacetylated during the course of chromatin maturation (7). Although acetylation of H4K5/12 is highly conserved throughout evolution, it has been shown to be nonessential through mutagenic analysis, in which replacing these residues with arginine (mimicking the constitutively unacetylated state) has no effect on cell growth in yeast (8). Thus the evolutionarily conserved function of H4K5/ K12 acetylation remains unknown.The HAT-B 2 enzyme complex is responsible for acetylating newly synthesized histone H4 on lysines K5 and K12 (9 -13). HAT-B is a multisubunit complex composed of histone acetyltransferase 1 (Hat1), the catalytic subunit, and Hat2 (rbap46), a histone chaperone that in...

Functional Analysis of Hif1 Histone Chaperone in Saccharomyces cerevisiae

“…Hif1 has previously been shown to physically interact with the Hat1/Hat2-complex via Hat2 ( Haigney et al 2015 ). To further characterize the Hif1 interaction with the Hat1/Hat2-complex, we extended our Co-IP analysis and mapped the Hat2 interaction region on Hif1 ( Figure 2C, D ).…”

“…Asf1 and Hif1 interaction has been extensively reported where Asf1 is thought to function downstream of Hif1 in the H3/H4 transport pathway ( Campos et al 2010 ). In vitro studies have suggested that the interaction between Hif1 and Asf1 is likely mediated by histones H3/H4 ( Haigney et al 2015 ; Bowman et al 2017 ). To examine what region of Hif1 is required for an interaction with Asf1 in vivo , we employed our described Co-IP strategy.…”

Histone chaperone networks shaping chromatin function

“…Highlighting its central role in chaperoning H3–H4, ASF1 forms a large number of co-chaperone complexes with H3.1, H3.2, H3.3 and one or more of the chaperones Vps75, sNASP, RBAP46, RBAP48, MCM2 and the reader and chaperone TONSL 51,76,84,85,87–91 (FIG. 4).…”

“…4). Curiously, sNASP, RBAP46 and ASF1 seem to be capable of engaging histones H3–H4 in a single co-chaperone complex 87,89–91 . However, ASF1 and sNASP may have overlapping binding sites at the H3–H4 tetramerization interface 22,23,28 .…”

“…4). ASF1 can bind H3–H4 within the RBAP46–HAT1–H3–H4–sNASP co-chaperone complex 91 , which may provide ASF1 with access to both the cytoplasmic and nuclear stores of H3–H4. Most vertebrates have two ASF1 paralogues, ASF1A and ASF1B, which differ primarily in their amino- and carboxy-terminal regions outside the core histone-binding domain 95,119 .…”