Cdk9 and H2Bub1 signal to Clr6-CII/Rpd3S to suppress aberrant antisense transcription.

Jürg Bähler,Pabitra K Parua,Sarah Mackinnon,Patricia Garcia,Viviane Pagé,Robert P Fisher,Danny A Bitton,Miriam Sansó,Daniel Pinto,Elena Hidalgo,Jason C Tanny,J Peter Svensson

doi:10.1093/nar/gkaa474

Abstract

Mono-ubiquitylation of histone H2B (H2Bub1) and phosphorylation of elongation factor Spt5 by cyclin-dependent kinase 9 (Cdk9) occur during transcription by RNA polymerase II (RNAPII), and are mutually dependent in fission yeast. It remained unclear whether Cdk9 and H2Bub1 cooperate to regulate the expression of individual genes. Here, we show that Cdk9 inhibition or H2Bub1 loss induces intragenic antisense transcription of ∼10% of fission yeast genes, with each perturbation affecting largely distinct subsets; ablation of both pathways de-represses antisense transcription of over half the genome. H2Bub1 and phospho-Spt5 have similar genome-wide distributions; both modifications are enriched, and directly proportional to each other, in coding regions, and decrease abruptly around the cleavage and polyadenylation signal (CPS). Cdk9-dependence of antisense suppression at specific genes correlates with high H2Bub1 occupancy, and with promoter-proximal RNAPII pausing. Genetic interactions link Cdk9, H2Bub1 and the histone deacetylase Clr6-CII, while combined Cdk9 inhibition and H2Bub1 loss impair Clr6-CII recruitment to chromatin and lead to decreased occupancy and increased acetylation of histones within gene coding regions. These results uncover novel interactions between co-transcriptional histone modification pathways, which link regulation of RNAPII transcription elongation to suppression of aberrant initiation.

Highlights

The elongation phase of transcription by RNA polymerase II (RNAPII) is subject to stringent regulation [1]
Enzymes involved in RNA processing and chromatin modification are recruited directly to the RNAPII elongation complex, ensuring that these events are coupled to RNA synthesis [2]
Perturbations of chromatin structure due to passage of the transcription complex carry an inherent risk of inappropriate access by initiation factors

Summary

Introduction

The elongation phase of transcription by RNA polymerase II (RNAPII) is subject to stringent regulation [1]. H2Bub positively regulates methylation of histone H3 at Lys (H3K4me) and Lys (H3K79me)––marks likewise associated with transcribed chromatin [7,8]––and functions independently of methylation [9,10], for example, to regulate nucleosome stability and positioning within coding regions [11,12,13]. The mechanisms underlying these methylation-independent effects and their consequences for gene expression are not known. Despite the presence of H2Bub at most or all transcribed genes, its loss affects steady-state levels of only a small fraction of mRNAs, suggesting redundant or compensatory homeostatic mechanisms [4,9,11,14]

Methods

Results

Conclusion