Role of Direct Interactions between the Histone H4 Tail and the H2A Core in Long Range Nucleosome Contacts

Divya Sinha,Michael A Shogren-Knaak

doi:10.1074/jbc.m109.091298

Abstract

In eukaryotic nuclei the majority of genomic DNA is believed to exist in higher order chromatin structures. Nonetheless, the nature of direct, long range nucleosome interactions that contribute to these structures is poorly understood. To determine whether these interactions are directly mediated by contacts between the histone H4 amino-terminal tail and the acidic patch of the H2A/H2B interface, as previously demonstrated for short range nucleosomal interactions, we have characterized the extent and effect of disulfide cross-linking between residues in histones contained in different strands of nucleosomal arrays. We show that in 208-12 5 S rDNA and 601-177-12 nucleosomal array systems, direct interactions between histones H4-V21C and H2A-E64C can be captured. This interaction depends on the extent of initial cross-strand association but does not require these specific residues, because interactions with residues flanking H4-V21C can also be captured. Additionally, we find that trapping H2A-H4 intra-array interactions antagonizes the ability of these arrays to undergo intermolecular self-association.

Highlights

The understanding of higher order chromatin structures has been significantly aided by in vitro studies of isolated and reconstituted nucleosomal array systems, where reversible short range intra-array and long range interarray nucleosome associations can be induced even in the absence of additional chromatin-associated proteins [5, 6]
Substitution of cysteines for histone residues H4-Val21 and H2A-Glu64 allowed them to trap interactions that occurred under conditions where nucleosomal arrays exhibit intramolecular but not intermolecular compaction. To adapt this technique to study the extent of interarray contacts between the H4 tail and H2A/H2B acidic patch, our strategy was to generate two different sets of arrays, where each array consisted of nucleosomes that included either H4-V21C histones or H2A-E64C histones
Role of Direct H4-H2A Interarray Interactions—In chromatin, the acid patch of the histone H2A/H2B dimer serves as a key protein interaction site

Summary

Introduction

The understanding of higher order chromatin structures has been significantly aided by in vitro studies of isolated and reconstituted nucleosomal array systems, where reversible short range intra-array and long range interarray nucleosome associations can be induced even in the absence of additional chromatin-associated proteins [5, 6]. Interarray Cross-linking in 208-12 Arrays via H4-V21C and H2A-E64C—Direct interactions between the histone H4 tail of one nucleosome and the H2A/H2B acidic patch of another have been previously demonstrated within the same nucleosomal array by Richmond and co-workers [18] using oxidative crosslinking. Significant sedimentation was not observed with similar treatment of arrays containing H2A-E64C histones (Fig. 1D), indicating that H2A-H2A cross-linking was not responsible for the interarray association.

Results

Conclusion