Functional crosstalk between histone H2B ubiquitylation and H2A modifications and variants

Felix Wojcik,Leslie Y Beh,Raphael Hofmann,Galia T Debelouchina,Geoffrey P Dann,Tom W Muir

doi:10.1038/s41467-018-03895-5

Abstract

Ubiquitylation of histone H2B at lysine residue 120 (H2BK120ub) is a prominent histone posttranslational modification (PTM) associated with the actively transcribed genome. Although H2BK120ub triggers several critical downstream histone modification pathways and changes in chromatin structure, less is known about the regulation of the ubiquitylation reaction itself, in particular with respect to the modification status of the chromatin substrate. Here we employ an unbiased library screening approach to profile the impact of pre-existing chromatin modifications on de novo ubiquitylation of H2BK120 by the cognate human E2:E3 ligase pair, UBE2A:RNF20/40. Deposition of H2BK120ub is found to be highly sensitive to PTMs on the N-terminal tail of histone H2A, a crosstalk that extends to the common histone variant H2A.Z. Based on a series of biochemical and cell-based studies, we propose that this crosstalk contributes to the spatial organization of H2BK120ub on gene bodies, and is thus important for transcriptional regulation.

Highlights

Ubiquitylation of histone H2B at lysine residue 120 (H2BK120ub) is a prominent histone posttranslational modification (PTM) associated with the actively transcribed genome
We began our investigations by asking whether pre-existing modifications to the chromatin template affect the efficiency of H2BK120ub deposition by the UBE2A:RNF20/40 machinery
We introduced a high-throughput system for the quantitative analysis of chromatin biochemistry[17]. This technology is based on the use of a DNA-barcoded mononucleosome (MN) library containing a diverse set of histone PTMs, variants, and mutations

Summary

Introduction

Ubiquitylation of histone H2B at lysine residue 120 (H2BK120ub) is a prominent histone posttranslational modification (PTM) associated with the actively transcribed genome. Attachment of this 76-residue protein adds significant steric bulk to the nucleosome, increasing the available surface area by as much as ~ 4800 Å24,5 These attributes allow H2BK120ub to act as a signaling hub for several downstream biochemical processes associated with active transcriptional elongation, including lysine methylation events on histone H36,7, as well as binding of the histone chaperone complex, FACT8. The enzymatic machinery for installing H2BK120ub in humans is known and comprises the E2 ligase UBE2A/B (Rad[6] in yeast) and the hetero-dimeric RINGtype E3 ligase RNF20/40 (Bre[1] in yeast)[14] The activity of these enzymes is stimulated by synergistic interactions involving components of the Mediator transcription initiation complex ( the MED23 subunit)[15] and the polymeraseassociated factor transcription elongation complex[16]. These findings have led to the idea that efficient ubiquitylation of H2B is coupled to ongoing transcription[8,12,14]

Methods

Results

Conclusion