Solution Scattering and FRET Studies on Nucleosomes Reveal DNA Unwrapping Effects of H3 and H4 Tail Removal

Kurt Andresen,Xiangyun Qiu,Isabel Jimenez-Useche,Steven C Howell,Chongli Yuan,Martin Fernandez-Zapico

doi:10.1371/journal.pone.0078587

Abstract

Using a combination of small-angle X-ray scattering (SAXS) and fluorescence resonance energy transfer (FRET) measurements we have determined the role of the H3 and H4 histone tails, independently, in stabilizing the nucleosome DNA terminal ends from unwrapping from the nucleosome core. We have performed solution scattering experiments on recombinant wild-type, H3 and H4 tail-removed mutants and fit all scattering data with predictions from PDB models and compared these experiments to complementary DNA-end FRET experiments. Based on these combined SAXS and FRET studies, we find that while all nucleosomes exhibited DNA unwrapping, the extent of this unwrapping is increased for nucleosomes with the H3 tails removed but, surprisingly, decreased in nucleosomes with the H4 tails removed. Studies of salt concentration effects show a minimum amount of DNA unwrapping for all complexes around 50-100mM of monovalent ions. These data exhibit opposite roles for the positively-charged nucleosome tails, with the ability to decrease access (in the case of the H3 histone) or increase access (in the case of the H4 histone) to the DNA surrounding the nucleosome. In the range of salt concentrations studied (0-200mM KCl), the data point to the H4 tail-removed mutant at physiological (50-100mM) monovalent salt concentration as the mononucleosome with the least amount of DNA unwrapping.

Highlights

In recent years, research investigating the problem of chromatin structure has increased dramatically
This led to the discovery that a change between these densely-packed and less densely-packed phases could be brought about by changes in ion composition or in composition of the histone tails with the eventual conclusion that the physics of this process could largely be attributed to packaging of the nucleosome core particle [3]
The combination of the two techniques used in this study can give us important complementary structural information

Summary

Introduction

Research investigating the problem of chromatin structure has increased dramatically. Chromatin studies concentrated on the varying levels of density of genomic material within the nucleus which creates hetero- and euchromatin This led to the discovery that a change between these densely-packed and less densely-packed phases could be brought about by changes in ion composition or in composition of the histone tails with the eventual conclusion that the physics of this process could largely be attributed to packaging of the nucleosome core particle [3]. The canonical nucleosome consists of 147 base pairs of DNA wrapped around an octameric histone core of four proteins (H2A, H2B, H3, and H4) These proteins contain positively-charged, flexible tails that protrude from the core either through (in the case of H2B and H3) or around (in the case of H2A and H4) the DNA [6,7]. Driven by increasingly refined structures of the nucleosome [7,8,9] and nucleosome arrays [10], the question of the packing of nucleosomes into the 30nm fiber (or nucleosome condensation) and other higher-order structures has attracted some of the most active research and debate [11]

Methods

Results

Conclusion