Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Elongation/termination factor exchange mediated by PP1 phosphatase orchestrates trancription termination

ABSTRACT: Termination of RNA polymerase II (Pol II) transcription is a key step, that is important for 3’end formation of functional mRNA, mRNA release and Pol II recycling. Even so, this underlying termination mechanism is not yet understood. Here, we demonstrate that the conserved and essential termination factor Seb1 interacts with Pol II near the end of the RNA exit channel and the Rpb4/7 stalk. Furthermore, the Seb1 interaction surface with Pol II largely overlaps with that of the elongation factor Spt5. Notably, Seb1 co-transcriptional recruitment is dependent on Spt5 de-phosphorylation by the conserved PP1 phosphatase Dis2, which also de-phosphorylates threonine 4 within the Pol II heptad repeated C-terminal domain. We propose that Dis2 orchestrates the transition from elongation to termination phase during the transcription cycle by mediating elongation to termination factor exchange and de-phosphorylation of Pol II C-terminal domain.

ORGANISM(S): Schizosaccharomyces pombe

PROVIDER: GSE111326 | GEO | 2018/12/26

REPOSITORIES: GEO

ACCESS DATA

Json Xml

Similar Datasets

Control of RNA pol II speed by PNUTS-PP1 and Spt5 dephosphorylation facilitates termination by a “sitting duck torpedo” mechanism

Project description:Control of transcription speed, which influences many co-transcriptional processes, is poorly understood. We report that PNUTS-PP1 phosphatase is a negative regulator of RNA pol II elongation rate. The PNUTS W401A mutation, which disrupts PP1 binding, causes genome-wide acceleration of transcription associated with hyper-phosphorylation of the Spt5 elongation factor. Immediately downstream of poly(A) sites, pol II decelerates from >2kb/min to <1 kb/min, which correlates with Spt5 dephosphorylation. Pol II deceleration and Spt5 dephosphorylation require poly(A) site recognition and the PNUTS-PP1 complex, which is in turn necessary for transcription termination. These results lead to a new model for termination, the “sitting duck torpedo” mechanism, where poly(A) site-dependent deceleration caused by PNUTS-PP1 and Spt5 dephosphorylation is required to convert pol II into a viable target for the Xrn2 terminator exonuclease. Spt5 and its bacterial homologue NusG therefore have related functions controlling kinetic competition between RNA polymerases and the termination factors that pursue them.

2019-10-30 | GSE134198 | GEO

A Cdk9-PP1 switch regulates the elongation-termination transition of RNA polymerase II

Project description:The end of the RNA polymerase II (Pol II) transcription cycle is strictly regulated to ensure proper mRNA maturation and prevent interference between neighboring genes. Pol II slowing downstream of the cleavage and polyadenylation signal (CPS) leads to recruitment of cleavage and polyadenylation factors and termination, but how this chain of events is initiated remains unclear. In a chemical-genetic screen we identified protein phosphatase 1 (PP1) isoforms as substrates of human positive transcription elongation factor b (P-TEFb), the cyclin-dependent kinase 9 (Cdk9)-cyclin T1 complex. Here we show that Cdk9 and PP1 govern phosphorylation of the conserved transcription factor Spt5 in the fission yeast Schizosaccharomyces pombe. Cdk9 phosphorylates both Spt5 and a negative regulatory site on the PP1 isoform Dis2. Sites phosphorylated by Cdk9 in the Spt5 carboxy-terminal domain (CTD) are dephosphorylated by Dis2 in vitro, and Cdk9 inhibition in vivo leads to rapid Spt5 dephosphorylation that is retarded by concurrent Dis2 inactivation. Chromatin immunoprecipitation and sequencing (ChIP-seq) analysis indicates that Spt5 is dephosphorylated as transcription complexes traverse the CPS, prior to or concomitant with Pol II pausing. A Dis2-inactivating mutation stabilizes Spt5 phosphorylation (pSpt5) on chromatin, promotes transcription beyond the normal termination zone detected by precision run-on transcription and sequencing (PRO-seq), and is suppressed by ablation of Cdk9 target sites in Spt5. These results support a model whereby the transition of Pol II from elongation to termination is regulated by opposing activities of Cdk9 and Dis2 towards their common substrate Spt5—a bistable switch analogous to a Cdk1-PP1 module that controls exit from mitosis.

2018-06-07 | GSE102590 | GEO

mRNA decapping factors and the exonuclease Xrn2 function in widespread premature termination of RNA polymerase II transcription

Project description:We report a function of human mRNA decapping factors in control of transcription by RNA polymerase II. Decapping proteins Edc3, Dcp1a and Dcp2 and the termination factor TTF2 co-immunoprecipitate with Xrn2, the nuclear 5'-3' exonuclease torpedo that facilitates transcription termination at the 3' ends of genes. Dcp1a, Xrn2 and TTF2 localize near transcription start sites (TSSs) by ChIP-Seq. At genes with 5' peaks of paused pol II, knockdown of decapping or termination factors, Xrn2 and TTF2, shifted polymerase away from the TSS toward upstream and downstream distal positions. This re-distribution of pol II is similar in magnitude to that caused by depletion of the elongation factor Spt5. We propose that coupled decapping of nascent transcripts and premature termination by the torpedo mechanism is a widespread mechanism that limits bidirectional pol II elongation. Regulated co-transcriptional decapping near promoter-proximal pause sites followed by premature termination could control productive pol II elongation. RNA pol II (GSE30895: GSM766171), Xrn2, TTF2 and Dcp1a were localized by ChIP-Seq in HeLa cells. RNA pol II was localized in control HEK293 cells and cells infected with lentiviruses expressing a scrambled control shRNA (scr), and shRNAs targeting the following proteins: Xrn2, TTF2, Xrn2+TTF2, Edc3, Dcp1a, and Dcp2.

2012-04-04 | E-GEOD-36185 | biostudies-arrayexpress

Integrator Recruits Protein Phosphatase 2A to Prevent Pause Release and Enable Transcription Termination

Project description:Efficient release of promoter-proximally paused Pol II into productive elongation is essential for gene expression. Recently, we reported that the Integrator complex can bind paused Pol II and drive premature transcription termination, potently attenuating the activity of target genes. Premature termination requires RNA cleavage by the endonuclease subunit of Integrator, but the roles of other Integrator subunits in gene regulation have yet to be elucidated. Here, we report that Integrator subunit 8 (IntS8) is critical for transcription repression through its association with Protein Phosphatase 2A (PP2A). We find that Integrator-bound PP2A dephosphorylates the Pol II C-terminal domain and Spt5, and prevents the transition to productive elongation. Blocking PP2A association with Integrator thus stimulates pause release and gene activation. These results reveal a second catalytic function associated with Integrator-mediated transcription termination, and suggest a new model for the control of productive elongation involving active competition between transcriptional kinases and phosphatases.

2020-10-06 | GSE150844 | GEO

mRNA decapping factors and the exonuclease Xrn2 function in widespread premature termination of RNA polymerase II transcription

2012-04-05 | GSE36185 | GEO

Characterization of the SF3B1 interactome

Project description:CDK9 is a critical kinase required for the productive transcription of protein-coding genes by RNA polymerase II (pol II). As part of P-TEFb, CDK9 phosphorylates the carboxyl-terminal domain (CTD) of pol II and elongation factors, including SPT5, which allows pol II to elongate past the early elongation checkpoint (EEC) encountered soon after initiation. We show that, in addition to halting pol II at the EEC, loss of CDK9 activity causes premature termination of transcription across the last exon, loss of polyadenylation factors from chromatin, and loss of polyadenylation of nascent transcripts. Inhibition of the phosphatase PP2A abrogates the premature termination and loss of polyadenylation caused by CDK9 inhibition, indicating that this kinase/phosphatase pair regulates transcription elongation and RNA processing at the end of protein-coding genes. Our phosphoproteomic analyses after CDK9 inhibition, using either DRB or an analog-sensitive CDK9 cell line, confirm the splicing factor SF3B1 as an additional key target of this kinase. These results emphasize the important roles that CDK9 plays in coupling transcription elongation and termination to RNA maturation downstream of the EEC. As part of this project, we characterized the interactome of SF3B1 in HeLa cells in duplicate.

2022-08-22 | PXD033694 | Pride

Identification of CDK9 targets using DRB and quantitative phosphoproteomics

Project description:CDK9 is a critical kinase required for the productive transcription of protein-coding genes by RNA polymerase II (pol II) in higher eukaryotes. Phosphorylation of targets including the elongation factor SPT5 and the carboxyl-terminal domain (CTD) of RNA pol II allow the polymerase to pass an early elongation checkpoint (EEC), which is encountered soon after initiation. In addition to halting RNA polymerase II at the EEC, CDK9 inhibition also causes premature termination of transcription across the last exon, loss of polyadenylation factors from chromatin, and loss of polyadenylation of nascent transcripts. Inhibition of the phosphatase PP2A abrogates the premature termination and loss of polyadenylation caused by CDK9 inhibition, suggesting that CDK9 and PP2A, working together, regulate the coupling of elongation and transcription termination to RNA maturation. Our phosphoproteomic analyses, using either DRB or an analog-sensitive CDK9 cell line confirm the splicing factor SF3B1 as an additional key target of this kinase. CDK9 inhibition causes loss of interaction of splicing and export factors with SF3B1, suggesting that CDK9 also helps to co-ordinates coupling of splicing and export to transcription.

2022-08-22 | PXD026720 | Pride

Genome-wide analysis of RNA polymerase II termination at protein-coding genes

Project description:At the end of protein-coding genes, RNA polymerase (Pol) II undergoes a concerted transition that involves 3’-processing of the pre-mRNA and transcription termination. Here we conduct a genome-wide analysis of this 3’-transition in yeast. We find that the 3’-transition globally requires the Pol II elongation factor Spt5 and factors involved in the recognition of the poly-adenylation (pA) site and in endonucleolytic RNA cleavage. Pol II release from DNA occurs in a narrow termination window downstream of the pA site and generally requires the ‘torpedo’ exonuclease Rat1 (human XRN2). The Rat1-interacting factor Rai1 contributes to RNA degradation downstream of the pA site. Defects in the 3’-transition result in transcription interference at downstream genes, which is attenuated by a back-up termination mechanism.

2017-03-15 | GSE79222 | GEO

The conserved protein Seb1 drives transcription termination by binding RNA polymerase II and nascent RNA

Project description:Termination of Pol II transcription is an important step in the transcription cycle and is responsible for dislodgement of polymerase from DNA which leads to the release of a functional transcript. Recent studies have identified key players important for termination and showed a conserved domain that interacts with the phosphorylated C-terminus of Pol II (CTD-Interacting-Domain, CID) to constitute a common feature of these proteins. However, the mechanism by which transcription termination is achieved, is not understood. Using genome-wide methods, we demonstrate that the fission yeast CID protein Seb1 is essential for termination of protein-coding and non-coding genes through interacting with S2-phosphorylated Pol II and nascent RNA. Furthermore, we present the crystal structures of the Seb1 CTD- and RNA-binding modules. Unexpectedly, the latter reveals a novel intertwined two-domain arrangement of a canonical RRM and a second domain. These results provide important insights into the mechanism underlying eukaryotic transcription termination.

2017-02-05 | GSE89914 | GEO

Stabilization of Pol II protein, orchestration of transcription cycles, and maintenance of enhancer landscape by general transcription regulator SPT5 [TT-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 | GSE183506 | GEO

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data