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Genome-associated RNA Polymerase II includes the dissociable Rpb4/7 subcomplex

ABSTRACT: Yeast RNA polymerase (Pol) II consists of a ten-subunit core enzyme and the Rpb4/7 subcomplex, which is dispensable for catalytic activity and dissociates in vitro. To investigate whether Rpb4/7 is an integral part of DNA-associated Pol II in vivo, we used chromatin immuno-precipitation coupled to high-resolution tiling microarray analysis. We show that the genome-wide occupancy profiles for Rpb7 and the core subunit Rpb3 are essentially identical. Thus, the complete Pol II associates with DNA in vivo, consistent with functional roles of Rpb4/7 throughout the transcription cycle. Keywords: ChIP-chip

ORGANISM(S): Saccharomyces cerevisiae

PROVIDER: GSE12060 | GEO | 2008/07/30

SECONDARY ACCESSION(S): PRJNA113393

REPOSITORIES: GEO

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Genome-associated RNA Polymerase II includes the dissociable Rpb4/7 subcomplex

Project description:Yeast RNA polymerase (Pol) II consists of a ten-subunit core enzyme and the Rpb4/7 subcomplex, which is dispensable for catalytic activity and dissociates in vitro. To investigate whether Rpb4/7 is an integral part of DNA-associated Pol II in vivo, we used chromatin immuno-precipitation coupled to high-resolution tiling microarray analysis. We show that the genome-wide occupancy profiles for Rpb7 and the core subunit Rpb3 are essentially identical. Thus, the complete Pol II associates with DNA in vivo, consistent with functional roles of Rpb4/7 throughout the transcription cycle. Keywords: ChIP-chip Comparison of Rpb3 vs. Rpb4/7 genome-wide occupancy profiles. Data obtained from independent ChIP-chip experiments on two yeast strains: S288C Rpb3TAP vs. Rpb7-TAP strain and W303 Rpb3-TAP vs. wild type strain (IP with the monoclonal antibody for Rpb4/7). Biological replicates W303 strains: 1 x Rpb3-TAP, 1 x wt with Rpb4/7 antibody, independently grown and harvested. Biological replicates S288C strains: 3 x Rpb3-TAP, 2 x Rpb7-TAP, independently grown and harvested. One replicate per array.

2008-07-29 | E-GEOD-12060 | biostudies-arrayexpress

Rpb4 functions mainly in mRNA synthesis by RNA polymerase II

Project description:RNA polymerase II (Pol II) is the central enzyme that carries out eukaryotic mRNA transcription and consists of a 10-subunit catalytic core and a heterodimeric subcomplex of subunits Rpb4 and Rpb7 (Rpb4/7). Rpb4/7 has been proposed to shuttle from the nucleus to the cytoplasm, and to function there in mRNA translation and degradation. Here we provide evidence that Rpb4 mainly functions in nuclear mRNA synthesis by Pol II, and evidence arguing against an important cytoplasmic role. We used metabolic RNA labeling and comparative Dynamic Transcriptome Analysis (cDTA) to show that Rpb4 deletion in Saccharomyces cerevisiae causes a drastic defect in mRNA synthesis that is compensated by down-regulation of mRNA degradation, resulting in mRNA level buffering. Deletion of Rpb4 can be rescued by covalent fusion of Rpb4 to the Pol II core subunit Rpb2, which largely restores mRNA synthesis and degradation defects caused by Rpb4 deletion. Thus Rpb4 is a bona fide Pol II core subunit which functions mainly in mRNA synthesis.

2014-05-09 | E-MTAB-2539 | biostudies-arrayexpress

Whole genome recruitment of Rpb3 and Rpb4 in S.cerevesiae

Project description:ChIP on chip microarray experiment to decipher Rpb3 and Rpb4 recruitment in S.cerevesiae genome. Keywords: ChIP-chip

2007-11-15 | GSE9265 | GEO

Numerous post-translational modifications of RNA polymerase II subunit Rpb4 link transcription to post-transcriptional mechanisms

Project description:Rpb4/7 binds RNA Polymerase II (Pol II) transcripts co-transcriptionally and accompanies them throughout their lives. By virtue of its capacity to interact with key regulators (e.g., Pol II, eIF3, Pat1) both temporarily and spatially, Rpb4/7 regulates the major stages of the mRNA lifecycle. Here we show that Rpb4/7 can undergo over 100 combinations of post-translational modifications (PTMs). Remarkably, the Rpb4/7 PTMs repertoire changes as the mRNA/Rpb4/7 complex progresses from one stage to the next. A mutagenesis approach in residues that undergo PTMs suggests that temporal Rpb4 PTMs regulate its interactions with key regulators of gene expression that control transcriptional and post-transcriptional stages. Moreover, one mutant type specifically affects mRNA synthesis despite its normal association with Pol II, whereas the other affects both mRNA synthesis and decay; both types disrupt the balance between mRNA synthesis and decay (‘mRNA buffering’) and the cell’s capacity to respond to the environment. Taken together, we propose that temporal Rpb4/7 PTMs are involved in cross talks among the various stages of the mRNA lifecycle.

2020-09-25 | PXD020511 | Pride

Potential modulation of Puf3 chromatin association mediated by Rpb4

2022-11-09 | GSE152093 | GEO

Rpb4 and Puf3 imprint and post-transcriptionally control stability of a common set of mRNAs in yeast

Project description:Gene expression involving RNA polymerase II is regulated by the concerted interplay between mRNA synthesis and degradation, a crosstalk where mRNA decay machinery impacts transcription and transcription machinery influences mRNA stability. Rpb4, and likely the dimer Rpb4/7 seem to be the central components of the RNA pol II governing these processes. In this work we unravel more precisely the molecular mechanisms participated by Rpb4 that mediates the posttranscriptional events regulating mRNA imprinting and stability. We analysed genome-wide, by RIP-seq, the association of Rpb4 with mRNAs and demonstrated that it targets a broad population of about 1400 transcripts. A group of mRNAs are also targets of the RPB, Puf3. We demonstrated that Rpb4 and Puf3, physically, genetically and functionally interact and cooperate to imprint and regulate stability of this group of mRNA. We also demonstrated, by the first time, that Puf3 associates with the chromatin, in an Rpb4-dependent manner. Our data also point to Rpb4 as the key element of the RNA pol II that interplay mRNA synthesis, imprinting and stability, in cooperation with RBPs.

2022-11-09 | GSE152094 | GEO

Genome-wide nucleosome, histone and Pol II maps for wild type Saccharomyces cerevisiae induced with 3-aminotriazole (3AT)

2014-10-28 | GSE54524 | GEO

Genome-associated RNA polymerase II includes the dissociable Rpb4/7 subcomplex.

Project description:Yeast RNA polymerase (Pol) II consists of a 10-subunit core enzyme and the Rpb4/7 subcomplex, which is dispensable for catalytic activity and dissociates in vitro. To investigate whether Rpb4/7 is an integral part of DNA-associated Pol II in vivo, we used chromatin immunoprecipitation coupled to high resolution tiling microarray analysis. We show that the genome-wide occupancy profiles for Rpb7 and the core subunit Rpb3 are essentially identical. Thus, the complete Pol II associates with DNA in vivo, consistent with functional roles of Rpb4/7 throughout the transcription cycle.

| S-EPMC3258907 | biostudies-literature

Genome-wide nucleosome, histone and Pol II maps for wild type Saccharomyces cerevisiae induced with 3-aminotriazole (3AT)

Project description:Paired-end sequencing study of (1) nucleosome core particles and under-digested chromatin from MNase-treated nuclei; (2) ChIP samples for HA-tagged histone H4 and H2B; (3) ChIP for the Rpb3 subunit of Pol II. Nucleosomal DNA and immunopurified sonicated DNA fragments were subjected to paired-end sequencing.

2014-10-28 | E-GEOD-54524 | biostudies-arrayexpress

A study of Mediator subunit enhancer-promoter binding

Project description:Mediator is a transcriptional co-activator recruited to enhancers by DNA-binding activators, and it also interacts with RNA polymerase (Pol) II as part of the preinitiation complex (PIC). We demonstrate that a single Mediator complex associates with the enhancer and core promoter in vivo, indicating that it can physically bridge these transcriptional elements. However, the Mediator kinase module associates strongly with the enhancer, but not with the core promoter, and it dissociates from the enhancer upon depletion of the TFIIH kinase. Severing the kinase module from Mediator by removing the connecting subunit Med13 does not affect Mediator association at the core promoter, but increases occupancy at enhancers. Thus, Mediator undergoes a compositional change in which the kinase module, recruited via Mediator to the enhancer, dissociates from Mediator to permit association with Pol II and the PIC. As such, Mediator acts as a dynamic bridge between the enhancer and core promoter.

2016-10-31 | GSE82082 | GEO

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