The Effectcs of Histone H4 Acetylations in Nucleosome-Nucleosome Interactions and on Chromatin Folding and Fibre-Fibre Association

Abstract

We developed novel semi-synthetic methods for the preparation of precisely modified N-terminal histone tails and investigated the folding and self-association of recombinant nucleosome arrays using a range of biophysical methods. These arrays contained acetylations at positions 5, 8, 12, and 16 of histone H4 and the condensation of this chromatin system, induced by cations like Mg2+ was systematically studied. Additionally, the effects of the combination of acetylations and H4 tail methylations, linker histone H1 and nucleosome repate length is also studied. It was found that: i) The single clean and complete H4-K16 acetylation is sufficient to severely antagonize array folding, while acetylations at positions 5, 8 and 12 together only have a moderate effect that acts additively when combined with H4-K16Ac. ii) Inter-array self-association is unspecifically electrostatically controlled by the charge reduction effect of acetylation and follows polyelectrolyte behaviour. iii) The cation K+, at physiological concentrations, impede full nucleosome array folding almost to the same extent as H4-K16Ac. The observations suggests that nucleosome-nucleosome stacking, required for full folding, is mediated by H4K16 binding to a site on the H2B histone, which is disrupted by acetylation or the presence of K+ ions. The nature of the nucleosome-nucleosome interactions and role of tails and H4 acetylations were further investigated by studying solutions of mononucleosomes (NCP) with tail acetylations or alternatively, devoid of tails, using synchrotron X-ray scattering. Contrary to array folding, close nucleosome-nucleosome stacking is observed for NCPs with acetylated H4 histone tails, as shown by the formation of an hexagonal columnar phase of ordered mononuclesomes, induced by cations. Computer simulations with a detailed coarse grained model indicate that aggregation to stacked NCPs can be completely driven by electrostatic interactions.