Abstract
BackgroundFor many genes, RNA polymerase II stably pauses before transitioning to productive elongation. Although polymerase II pausing has been shown to be a mechanism for regulating transcriptional activation, the extent to which it is involved in control of mammalian gene expression and its relationship to chromatin structure remain poorly understood.ResultsHere, we analyze 85 RNA polymerase II chromatin immunoprecipitation (ChIP)-sequencing experiments from 35 different murine and human samples, as well as related genome-wide datasets, to gain new insights into the relationship between polymerase II pausing and gene regulation. Across cell and tissue types, paused genes (pausing index > 2) comprise approximately 60 % of expressed genes and are repeatedly associated with specific biological functions. Paused genes also have lower cell-to-cell expression variability. Increased pausing has a non-linear effect on gene expression levels, with moderately paused genes being expressed more highly than other paused genes. The highest gene expression levels are often achieved through a novel pause-release mechanism driven by high polymerase II initiation. In three datasets examining the impact of extracellular signals, genes responsive to stimulus have slightly lower pausing index on average than non-responsive genes, and rapid gene activation is linked to conditional pause-release. Both chromatin structure and local sequence composition near the transcription start site influence pausing, with divergent features between mammals and Drosophila. Most notably, in mammals pausing is positively correlated with histone H2A.Z occupancy at promoters.ConclusionsOur results provide new insights into the contribution of RNA polymerase II pausing in mammalian gene regulation and chromatin structure.Electronic supplementary materialThe online version of this article (doi:10.1186/s13059-016-0984-2) contains supplementary material, which is available to authorized users.
Highlights
For many genes, RNA polymerase II stably pauses before transitioning to productive elongation
Characterization of RNA polymerase II (RNAP2) pausing across multiple cell types We analyzed RNAP2 pausing at each gene based on its “Pausing Index” (PI; referred to as “Traveling Ratio”) [1, 7, 8, 13, 14]
RNAP2 chromatin immunoprecipitation (ChIP)-seq is less sensitive than other techniques designed for measuring paused RNAP2, such as GRO-seq [15] or PRO-seq [3], a comparison between GRO-seq and RNAP2 ChIP-seq data suggested that most signals observed in RNAP2 ChIP-seq data come from transcriptionally engaged RNAP2 [9], supporting their use for measuring differences in RNAP2 pausing
Summary
RNA polymerase II stably pauses before transitioning to productive elongation. Polymerase II pausing has been shown to be a mechanism for regulating transcriptional activation, the extent to which it is involved in control of mammalian gene expression and its relationship to chromatin structure remain poorly understood. The relationship between RNAP2 pausing and the regulation of chromatin structure and gene expression is poorly understood [1]. Several studies have suggested that RNAP2 pausing-release mediates rapid gene expression changes in response to external stimuli [1, 4, 5]. Recent reports suggest that stably paused RNAP2 shapes the chromatin structure of promoters [9,10,11], but the extent that RNAP2 pausing regulates chromatin remodeling across mammalian cell types has not been carefully examined. With advances in profiling various histone modifications, chromatin-bound proteins, and genomewide expression levels in an unbiased manner, a broad, in-depth analysis of such data would better highlight and refine the role RNAP2 pausing has on regulating gene expression and chromatin structure in mammals
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