Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

The pausing zone and control of RNA polymerase II elongation by Spt5: implications for the pause-release model

ABSTRACT: The pause-release model of transcription proposes that pol II pauses 40-100 bases from the start site resulting in a pile-up that is relieved by subsequent release into productive elongation. Pause release is facilitated by PTEFb phosphorylation of the pol II elongation factor, Spt5. We mapped paused polymerases by eNETseq and found frequent pausing in zones that extend ~0.3-3kb into genes, even when PTEFb is inhibited. The fraction of paused polymerases or “pausiness” declines gradually over several kb, and not abruptly as predicted for a discrete pause release event. Spt5 depletion extends pausing zones suggesting that it promotes maturation of elongation complexes to a low-pausing state. Expression of mutants after Spt5 depletion showed that phosphomimetic substitutions in the Spt5 CTR1 domain diminished pausing throughout genes. In contrast mutants that prevent phosphorylation of the Spt5 RNA-binding domain strengthened pausing. Thus distinct Spt5 phospho-isoforms set the balance between pausing and elongation.

ORGANISM(S): Homo sapiens

PROVIDER: GSE202749 | GEO | 2022/10/01

REPOSITORIES: GEO

ACCESS DATA

Json Xml

Dataset's files

Source:

			Action	DRS
		Other

Items per page:

1 - 1 of 1

Similar Datasets

Stabilization of Pol II protein, orchestration of transcription cycles, and maintenance of enhancer landscape by general transcription regulator SPT5 [ATAC-seq]

Project description:Transcription machinery progression is governed by multitasking regulators including SPT5, an evolutionarily conserved factor implicated in virtually all transcriptional steps from enhancer activation to termination. Yet its mechanistic understanding in human cells remains incomplete. Here we utilize rapid degradation system and reveal crucial function of SPT5 in maintaining cellular and chromatin Pol II levels. Rapid SPT5 depletion causes a pronounced reduction of paused Pol II at promoters and enhancers, distinct from NELF degradation resulting in short-distance paused Pol II redistribution. Most of genes exhibit down- but not upregulation, accompanied by greatly impaired transcription activation and altered chromatin landscape at enhancers, and severe Pol II processivity defects at gene bodies. Phosphorylation of KOWx-4/5- linker potentiates pause release and is antagonized by Integrator-PP2A (INTAC) targeting SPT5 and Pol II. Our findings position SPT5 as an essential positive regulator of global transcription by controlling cellular Pol II levels, enhancer activation and landscape, paused Pol II stability, elongation processivity and termination in human.

2021-09-29 | GSE180844 | GEO

Stabilization of Pol II protein, orchestration of transcription cycles, and maintenance of enhancer landscape by general transcription regulator SPT5 [RNA-seq]

2021-09-29 | GSE180843 | GEO

Stabilization of Pol II protein, orchestration of transcription cycles, and maintenance of enhancer landscape by general transcription regulator SPT5 [ChIP-seq]

2021-09-29 | GSE180842 | GEO

Stabilization of Pol II protein, orchestration of transcription cycles, and maintenance of enhancer landscape by general transcription regulator SPT5 [TT-seq]

2021-09-29 | GSE183506 | GEO

Stabilization of Pol II protein, orchestration of transcription cycles, and maintenance of enhancer landscape by general transcription regulator SPT5 [PRO-seq]

2021-09-29 | GSE183505 | GEO

Transcriptional elongation requires DNA break-induced signaling

Project description:We have previously shown that RNA polymerase II (Pol II) pause release and transcriptional elongation involve phosphorylation of the factor TRIM28 by the DNA damage response (DDR) kinases ATM and DNA-PK. Here, we report a significant role for DNA breaks and DDR signaling in the mechanisms of transcriptional elongation in stimulus-inducible genes in humans. Our data show the enrichment of TRIM28 and Î³H2AX on serum-induced genes and the important function of DNA-PK for Pol II pause release and transcriptional activation-coupled DDR signaling on these genes. Î³H2AX accumulation decreases when P-TEFb is inhibited, confirming that DDR signaling results from transcriptional elongation. In addition, transcriptional elongation-coupled DDR signaling involves topoisomerase II because inhibiting this enzyme interferes with Pol II pause release and Î³H2AX accumulation. Our findings propose that DDR signaling is required for effective Pol II pause release and transcriptional elongation through a novel mechanism involving TRIM28, DNA-PK, and topoisomerase II 42 samples in total. IP targets were gammaH2ax, s2-pol-II, pol-II, pTRIM28, DNA-pk, topo-IIB. Experimental conditions included DMSO treatment (control), pTEFb, topoII-i, dnapk-i. Matched non-specific IP samples used for control in peak calling.

2015-11-20 | E-GEOD-75170 | biostudies-arrayexpress

Stable pausing by RNA polymerase II provides an opportunity to target and integrate regulatory signals

Project description:Metazoan gene expression is often regulated after the recruitment of RNA polymerase II (Pol II) to promoters, through the controlled release of promoter-proximally paused Pol II into productive RNA synthesis. Despite the prevalence of paused Pol II, very little is known about the dynamics of these early elongation complexes or the fate of short transcription start site-associated (tss) RNAs they produce. Here, we demonstrate that paused elongation complexes can be remarkably stable, with half-lives exceeding 15 minutes at genes with inefficient pause release. Promoter-proximal termination by Pol II is infrequent and released tssRNAs are targeted for rapid degradation. Further, we provide evidence that the predominant tssRNA species observed are nascent RNAs held within early elongation complexes. We propose that stable pausing of polymerase provides a temporal window of opportunity for recruitment of factors to modulate gene expression and that the nascent tssRNA represents an appealing target for these interactions. This submission includes 13 raw data files. Four samples (chromatin, soluble, mock-treated, and Rrp40-depleted) are represented by two biological replicate raw data files, while five samples (cells, DMSO, FP, mock-treated + FP, and Rrp-40depleted + FP) are represented by single raw data files.

2013-10-31 | E-GEOD-49078 | biostudies-arrayexpress

Functional association of Gdown1 with RNA Polymerase II in Human

Project description:Most human genes are loaded with promoter proximally paused RNA polymerase II (RNAP II) molecules that are poised for release into productive elongation by P-TEFb. Gdown1, a protein that renders RNAP II responsive to mediator, is involved in RNAP II elongation control. During in vitro transcription assays Gdown1 specifically blocked elongation stimulation by TFIIF, inhibited the termination activity of TTF2, and influenced pausing factors NELF and DSIF, but did not affect the function of TFIIS or the mRNA capping enzyme. Without P-TEFb, Gdown1 led to the production of stably paused polymerases in the presence of nuclear extract. ChIP-Seq data demonstrated that Gdown1 mapped over essentially all poised polymerases across the human genome. Gdown1 increases the stability of poised polymerases while maintaining their responsiveness to P-TEFb, and mediator overcomes a Gdown1-mediated block of initiation by allowing TFIIF function. 7 ChIP-Seq data for Pol II and Gdown1 in human HeLa cell. See associated publication.

2012-01-13 | E-GEOD-32442 | biostudies-arrayexpress

RNA Polymerase II pausing temporally coordinates cell cycle progression and erythroid differentiation [RNA-seq]

Project description:The controlled release of promoter-proximal RNA polymerase II (Pol II) pausing into productive elongation is a major step in gene regulation. Yet, functionally analyzing Pol II pausing is difficult because the factors that regulate pause release are generally essential. Here, we identified heterozygous loss-of-function mutations in SUPT5H, encoding SPT5, in individuals with β-thalassemia trait unlinked to the β-globin locus. Recapitulating the pathogenic mutations in human erythroid cells or acute treatment with transcription inhibitors replicates the patient phenotype of altered globin gene expression. Furthermore, in SUPT5H-edited cells, Pol II pause release was globally disrupted. We observed dynamic patterns of pausing at cell cycle and erythroid differentiation genes at the transition from progenitors to precursors during erythropoiesis. At this same transition, in SUPT5H-edited cells, we observed delayed differentiation and altered cell cycle kinetics, as well as a delay in the onset of gene expression programs. Therefore, hindering pause release perturbs cell cycle and delays differentiation at key stages of erythropoiesis, revealing a role for Pol II pausing in the temporal coordination between the cell cycle and differentiation.

2023-07-21 | GSE214558 | GEO