Acetylation of Histone H3 at the Nucleosome Dyad Alters DNA-Histone Binding

Abstract

Histone post-translational modifications are essential for regulating and facilitating biological processes such as RNA transcription and DNA repair. Fifteen modifications are located in the DNA-histone dyad interface and include the acetylation of H3-K115 (H3-K115Ac) and H3-K122 (H3-K122Ac), but the functional consequences of these modifications are unknown. We have prepared semisynthetic histone H3 acetylated at Lys-115 and/or Lys-122 by expressed protein ligation and incorporated them into single nucleosomes. Competitive reconstitution analysis demonstrated that the acetylation of H3-K115 and H3-K122 reduces the free energy of histone octamer binding. Restriction enzyme kinetic analysis suggests that these histone modifications do not alter DNA accessibility near the sites of modification. However, acetylation of H3-K122 increases the rate of thermal repositioning. Remarkably, Lys 3 Gln substitution mutations, which are used to mimic Lys acetylation, do not fully duplicate the effects of the H3-K115Ac or H3-K122Ac modifications. Our results are consistent with the conclusion that acetylation in the dyad interface reduces DNA-histone interaction(s), which may facilitate nucleosome repositioning and/or assembly/disassembly.All eukaryotic genomes are organized into strings of nucleosomes, where 147 bp of DNA are tightly wrapped around a histone protein octamer (1). Many biological processes are dependent on DNA-protein interactions. However, access to DNA-binding sites is often restricted by the nucleosome structure. Alterations in nucleosome structure, dynamics, and positioning have been hypothesized to play a gatekeeper role in regulating biological processes such as DNA replication, repair, and transcription (2).The post-translational modification (PTM) 3 of core histones (3) plays a central role in regulating the biological processing of eukaryotic genomes. Until recently, known histone PTMs were almost exclusively located on the unstructured histone tail regions, which extend from the structured core of the nucleosomes. PTMs in the histone tails can function to directly alter nucleosome (4 -6) and/or chromatin structure and stability (7,8) and function as protein-binding sites (9) in the "histone code" model (10).During the past 5 years over 30 additional histone PTMs were identified within structured regions of the nucleosome (11-13). Many of these modifications are buried within the nucleosome core and thus are not readily accessible for protein binding. Fifteen of these histone PTMs are located in the DNAhistone interface, where the histone octamer contacts the phosphate backbone of the wrapped DNA (14). Studies have suggested that only mild structural perturbations occur in coremodified nucleosomes, which implies that modifications buried beneath the DNA are unlikely to provide a protein-binding site (15). This has led to two additional models for the function of nucleosome core PTMs.Modifications such as Lys acetylation that reduce the positive charge of the histone octamer surface may reduce the bindi...

Divergent Residues Within Histone H3 Dictate a Unique Chromatin Structure in Saccharomyces cerevisiae

“…This shift could be the result of altered interactions between histone H3 and the cellular machinery or direct effects on histone-DNA interactions. In support of the latter, it has been shown that neutralization of charge in this area by acetylation of H3 K122 destabilizes the nucleosome in vitro (Manohar et al 2009). To test between these two possibilities, we monitored the positioning of recombinant nucleosomes containing yeast H3 and yH3 a3h on DNA in vitro.…”

Acetylation-Deacetylation of the Transcription Factor Nrf2 (Nuclear Factor Erythroid 2-related Factor 2) Regulates Its Transcriptional Activity and Nucleocytoplasmic Localization

“…In this second set of mutations, we also made acetylation mimics (Lys 3 Gln) at the same lysine residues that were mutated to arginine residues. The Lys 3 Gln substitution is often used to mimic the acetylated form of the Lys residue (62)(63)(64)(65)(66). As shown in Fig.…”

“…For example, in their studies on DNA damage-induced acetylation of lysine 3016 of ataxiatelangiectasia mutated kinase, a modification that activates ataxia-telangiectasia mutated kinase, Sun et al (65) found that glutamine did not substitute for acetyl-lysine in ataxia-telangiectasia mutated kinase. In another example, Manohar et al (66) showed in studies on nucleosome dyad acetylation that the Lys 3 Gln substitution did not fully replicate the role of lysine acetylation; they found that the Lys 3 Gln substitution mimicked the change in charge but was a very poor mimic of the steric effect of acetylation. Thus, our findings may indicate that in addition to deacetylation of wild-type Nrf2, thereby inhibiting its transcriptional activity, SIRT1 expression may also affect Nrf2 activity by an additional mechanism when acetylatable lysines are mutated to glutamine.…”

“…This was rather surprising, given the fact that the K591Q and K588Q/K591Q sites cannot be deacetylated. Conceptually, the Lys 3 Gln substitution is understood to mimic the acetylated form of the Lys residue, whereas the Lys 3 Arg substitution only eliminates the ⑀-amino group of the acetylation site without a neutralization of positive charges (62)(63)(64)(65)(66). However, the Lys 3 Gln substitution does not always faithfully mimic the acetylated form of the Lys residue.…”

Histone Acetyltransferases

“…Although these data are highly valuable, a potential pitfall for the mutagenesis approach is that lysine to glutamine and arginine only moderately mimics the lysine in the acetylated and unacetylated form, respectively. 149,150 . Because the K274 in MOF or K327 in Tip60 resides near the head group of AcCoA, a chemical change to this residue may alter the active site microenvironment, thus compromising the catalytic activity of MYST proteins.…”