Control of RNA polymerase II promoter-proximal pausing by DNA supercoiling [RNA-seq]
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ABSTRACT:
PROVIDER: PRJNA594746 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR10662926.fastq.gz | Fastqsanger.gz | |||
| SRR10662927.fastq.gz | Fastqsanger.gz | |||
| SRR10662928.fastq.gz | Fastqsanger.gz | |||
| SRR10662929.fastq.gz | Fastqsanger.gz | |||
| SRR10662930.fastq.gz | Fastqsanger.gz |
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Similar Datasets
Project description:Control of RNA polymerase II promoter-proximal pausing by DNA supercoiling
| PRJNA594745 | ENA
Project description:During transcription, DNA supercoiling generated by the advance of RNA polymerase II (Pol II) is resolved by DNA topoisomerases, enzymes that bind chromatin and produce transient breaks to relax DNA. Recently, this idea of mere facilitators of transcription progression is changing, as topoisomerases are being assigned new functions in regulating the expression of specific genes. In fact, mammalian type II topoisomerases, both the [Symbol] (TOP2A) and [Symbol] (TOP2B) paralogs, are enriched at promoter regions, where they have been proposed to trigger transcription through the generation of DNA double-strand breaks (DSBs). However, this is difficult to reconcile with the intrinsic catalytic properties of TOP2 and the high risk of genome instability that continuous production and repair of DSBs implies. Here, we show that TOP2A enforces promoter-proximal pausing of Pol II by removing transcription-associated negative DNA supercoiling. Interestingly, this topological balance and its disruption is essential for the transcriptional control of Immediate Early Genes (IEGs) and their typical bursting behaviour in response to stimulus. We therefore uncover a novel layer of transcriptional regulation that relies on canonical functions of TOP2A that are independent of aberrant DSB formation, providing a topological framework for the control of promoter-proximal pausing and the tight regulation of IEGs.
2021-04-14 | GSE141799 | GEO
Project description:During transcription, DNA supercoiling generated by the advance of RNA polymerase II (Pol II) is resolved by DNA topoisomerases, enzymes that bind chromatin and produce transient breaks to relax DNA. Recently, this idea of mere facilitators of transcription progression is changing, as topoisomerases are being assigned new functions in regulating the expression of specific genes. In fact, mammalian type II topoisomerases, both the [Symbol] (TOP2A) and [Symbol] (TOP2B) paralogs, are enriched at promoter regions, where they have been proposed to trigger transcription through the generation of DNA double-strand breaks (DSBs). However, this is difficult to reconcile with the intrinsic catalytic properties of TOP2 and the high risk of genome instability that continuous production and repair of DSBs implies. Here, we show that TOP2A enforces promoter-proximal pausing of Pol II by removing transcription-associated negative DNA supercoiling. Interestingly, this topological balance and its disruption is essential for the transcriptional control of Immediate Early Genes (IEGs) and their typical bursting behaviour in response to stimulus. We therefore uncover a novel layer of transcriptional regulation that relies on canonical functions of TOP2A that are independent of aberrant DSB formation, providing a topological framework for the control of promoter-proximal pausing and the tight regulation of IEGs.
2021-04-14 | GSE141798 | GEO
Project description:Control of RNA polymerase II promoter-proximal pausing by DNA supercoiling [ChIP-seq]
| PRJNA594743 | ENA
Project description:TFIID enables RNA polymerase II promoter-proximal pausing
| PRJNA548938 | ENA
Project description:RNA polymerase II (pol II) transcribes all protein-coding and many non-coding RNAs in the human genome. Pol II transcription initiation is governed by the Pre-Initiation Complex (PIC), which contains TFIIA, TFIIB, TFIID, TFIIE, TFIIF, TFIIH, pol II, and Mediator. After initiation, pol II enzymes typically pause after transcribing less than 100 bases, and paused polymerases represent a common regulatory intermediate. Accordingly, paused pol II has been implicated in enhancer function, development and homeostasis, and diseases ranging from cancer to viral pathogenesis. Precisely how pol II promoter-proximal pausing is enforced and regulated remains unclear; however, protein complexes such as NELF and DSIF increase pausing whereas the activity of CDK9 (P-TEFb complex) correlates with pause release. To address specific mechanistic questions about pol II pausing and its regulation, we reconstituted human pol II promoter-proximal pausing in vitro, entirely with purified factors (no extracts). As expected, NELF and DSIF increased pol II pausing in vitro, whereas P-TEFb promoted pause release. Unexpectedly, the PIC alone was sufficient to reconstitute pol II pausing, suggesting that pausing is an inherent property of the PIC. In agreement, pol II pausing was lost upon replacement of the TFIID complex with TATA-binding protein (TBP); moreover, pausing was dependent upon TFIID subunits TAF1 and TAF2. TAF1/2 bind genomic DNA downstream of the pol II initiation site, invoking a “complex interaction” model for pausing. Consistent with this model, PRO-Seq experiments revealed increased transcription upon acute depletion (t=60 min) of TAF1 and TAF2 in human cells, and pol II pausing was disrupted at thousands of genes. Similar results were obtained in TAF1-depleted Drosophila S2 cells. Collectively, these data establish the general transcription factor TFIID as a genome-wide regulator of pol II promoter-proximal pausing.
2020-05-28 | GSE132764 | GEO
Project description:The control of promoter-proximal pausing and the release of RNA polymerase II (RNA Pol II) is a widely used mechanism for regulating gene expression in metazoans, especially for genes that respond to environmental and developmental cues. Here, we identify Pol II associated Factor 1 (PAF1) as a major regulator of promoter-proximal pausing. Knockdown of PAF1 leads to increased release of paused Pol II into gene bodies at thousands of genes. Genes with the highest levels of paused Pol II exhibit the largest redistribution of Pol II from the promoter-proximal region into the gene body in the absence of PAF1. PAF1 depletion results in increased nascent transcription and increased levels of phosphorylation of Pol II’s c-terminal domain on serine 2 (Ser2P). These changes can be explained by the recruitment of the Ser2P kinase Super Elongation Complex (SEC) effecting increased release of paused Pol II into productive elongation, thus establishing a novel function for PAF1 as a major regulator of pausing in metazoans. ChIP-seq of Pol II of different forms, SEC subunits, PAFc subunits and H2Bub in human cell lines targeted by PAF1 or scramble shRNA. ChIP-seq of total Pol II in HCT116 cells targeted by BRE1A or scramble shRNA. ChIP-seq of total Pol II in S2 cells targeted by Paf1 or LacZ RNAi. Total RNA-seq, nascent RNA-seq and GRO-seq in HCT116 cells targeted by PAF1 or scramble shRNA.
2015-08-13 | E-GEOD-70408 | biostudies-arrayexpress
Project description:The control of promoter-proximal pausing and the release of RNA polymerase II (RNA Pol II) is a widely used mechanism for regulating gene expression in metazoans, especially for genes that respond to environmental and developmental cues. Here, we identify Pol II associated Factor 1 (PAF1) as a major regulator of promoter-proximal pausing. Knockdown of PAF1 leads to increased release of paused Pol II into gene bodies at thousands of genes. Genes with the highest levels of paused Pol II exhibit the largest redistribution of Pol II from the promoter-proximal region into the gene body in the absence of PAF1. PAF1 depletion results in increased nascent transcription and increased levels of phosphorylation of Pol II’s c-terminal domain on serine 2 (Ser2P). These changes can be explained by the recruitment of the Ser2P kinase Super Elongation Complex (SEC) effecting increased release of paused Pol II into productive elongation, thus establishing a novel function for PAF1 as a major regulator of pausing in metazoans.
2015-08-13 | GSE70408 | GEO
Project description:A Role for FACT in RNA Polymerase II Promoter-Proximal Pausing
| PRJNA530563 | ENA
Project description:RNA polymerase II (RNAPII) transcription is governed by the pre-initiation complex (PIC), which contains TFIIA, TFIIB, TFIID, TFIIE, TFIIF, TFIIH, RNAPII, and Mediator. After initiation, RNAPII enzymes pause after transcribing less than 100 bases; precisely how RNAPII pausing is enforced and regulated remains unclear. To address specific mechanistic questions, we reconstituted human RNAPII promoter-proximal pausing in vitro, entirely with purified factors (no extracts). As expected, NELF and DSIF increased pausing, and P-TEFb promoted pause release. Unexpectedly, the PIC alone was sufficient to reconstitute pausing, suggesting RNAPII pausing is an inherent PIC function. In agreement, pausing was lost upon replacement of the TFIID complex with TATA-binding protein (TBP), and PRO-seq experiments revealed widespread disruption of RNAPII pausing upon acute depletion (t = 60 min) of TFIID subunits in human or Drosophila cells. These results establish a TFIID requirement for RNAPII pausing and suggest pause regulatory factors may function directly or indirectly through TFIID.
| S-EPMC7245555 | biostudies-literature