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Comprehensive analysis of promoter-proximal RNA polymerase II pausing across mammalian cell types.

ABSTRACT: BACKGROUND:For 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. RESULTS:Here, 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. CONCLUSIONS:Our results provide new insights into the contribution of RNA polymerase II pausing in mammalian gene regulation and chromatin structure.

SUBMITTER: Day DS

PROVIDER: S-EPMC4893286 | biostudies-literature | 2016 Jun

REPOSITORIES: biostudies-literature

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Comprehensive analysis of promoter-proximal RNA polymerase II pausing across mammalian cell types.

Day Daniel S DS Zhang Bing B Stevens Sean M SM Ferrari Francesco F Larschan Erica N EN Park Peter J PJ Pu William T WT

Genome biology 20160603 1

<h4>Background</h4>For 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.<h4>Results</h4>Here, we analyze 85 RNA polymerase II chromatin immunoprecipitation (ChIP)-sequencing experiments from 35 different mur ...[more]

PMID: 27259512

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Project description:Herpes simplex virus 1 (HSV-1) genes are transcribed by cellular RNA polymerase II (Pol II). Expression of viral immediate early (?) genes is followed sequentially by early (?), late (?1), and true late (?2) genes. We used precision nuclear run-on with deep sequencing to map and to quantify Pol II on the HSV-1(F) genome with single-nucleotide resolution. Approximately 30% of total Pol II relocated to viral genomes within 3?h postinfection (hpi), when it occupied genes of all temporal classes. At that time, Pol II on ? genes accumulated most heavily at promoter-proximal pause (PPP) sites located ?60?nucleotides downstream of the transcriptional start site, while ? genes bore Pol II more evenly across gene bodies. At 6 hpi, Pol II increased on ?1 and ?2 genes while Pol II pausing remained prominent on ? genes. At that time, average cytoplasmic mRNA expression from ? and ? genes decreased, relative to levels at 3 hpi, while ?1 relative expression increased slightly and ?2 expression increased more substantially. Cycloheximide treatment during the first 3?h reduced the amount of Pol II associated with the viral genome and confined most of the remaining Pol II to ? gene PPP sites. Inhibition of both cyclin-dependent kinase 9 activity and viral DNA replication reduced Pol II on the viral genome and restricted much of the remaining Pol II to PPP sites.IMPORTANCE These data suggest that viral transcription is regulated not only by Pol II recruitment to viral genes but also by control of elongation into viral gene bodies. We provide a detailed map of Pol II occupancy on the HSV-1 genome that clarifies features of the viral transcriptome, including the first identification of Pol II PPP sites. The data indicate that Pol II is recruited to late genes early in infection. Comparing ? and ? gene occupancy at PPP sites and gene bodies suggests that Pol II is released more efficiently into the bodies of ? genes than ? genes at 3 hpi and that repression of ? gene expression late in infection is mediated by prolonged promoter-proximal pausing. In addition, DNA replication is required to maintain full Pol II occupancy on viral DNA and to promote elongation on late genes later in infection.

Dataset Information

Comprehensive analysis of promoter-proximal RNA polymerase II pausing across mammalian cell types.

Publications

Comprehensive analysis of promoter-proximal RNA polymerase II pausing across mammalian cell types.

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