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Genetic screen for suppressors of lncRNA-mediated transcription interference identifies a gain-of-function mutation in the essential Pol2 termination factor Seb1

ABSTRACT: Genetic screen for suppressors of lncRNA-mediated transcription interference identifies a gain-of-function mutation in the essential Pol2 termination factor Seb1

PROVIDER: PRJNA714532 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR13963582_1.fastq.gz	Fastqsanger.gz
	SRR13963582_2.fastq.gz	Fastqsanger.gz
	SRR13963583_1.fastq.gz	Fastqsanger.gz
	SRR13963583_2.fastq.gz	Fastqsanger.gz
	SRR13963584_1.fastq.gz	Fastqsanger.gz

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Genetic screen for suppressors of lncRNA-mediated transcription interference identifies a gain-of-function mutation in the essential Pol2 termination factor Seb1

Project description:This RNA-Seq analysis compares gene expression of the STF-19 (seb1-G476S gain-of-function mutant) compared to a seb1+ WT in the fission yeast S. pombe. Both strains had WT-rpb1 with a TPI1 terminator sequence and a 3' selection marker.

2021-07-08 | GSE168898 | GEO

Genetic screen for suppression of transcriptional interference identifies a gain-of-function mutation in Pol2 termination factor Seb1.

Project description:The system of long noncoding RNA (lncRNA)-mediated transcriptional interference that represses fission yeast phosphate homoeostasis gene pho1 provides a sensitive readout of genetic influences on cotranscriptional 3'-processing and termination and a tool for discovery of regulators of this phase of the Pol2 transcription cycle. Here, we conducted a genetic screen for relief of transcriptional interference that unveiled a mechanism by which Pol2 termination is enhanced via a gain-of-function mutation, G476S, in the RNA-binding domain of an essential termination factor, Seb1. The genetic and physical evidence for gain-of-function is compelling: 1) seb1-G476S de-represses pho1 and tgp1, both of which are subject to lncRNA-mediated transcriptional interference; 2) seb1-G476S elicits precocious lncRNA transcription termination in response to lncRNA 5'-proximal poly(A) signals; 3) seb1-G476S derepression of pho1 is effaced by loss-of-function mutations in cleavage and polyadenylation factor (CPF) subunits and termination factor Rhn1; 4) synthetic lethality of seb1-G476S with pho1 derepressive mutants rpb1-CTD-S7A and aps1∆ is rescued by CPF/Rhn1 loss-of-function alleles; and 5) seb1-G476S elicits an upstream shift in poly(A) site preference in several messenger RNA genes. A crystal structure of the Seb1-G476S RNA-binding domain indicates potential for gain of contacts from Ser476 to RNA nucleobases. To our knowledge, this is a unique instance of a gain-of-function phenotype in a eukaryal transcription termination protein.

| S-EPMC8379984 | biostudies-literature

Seb1 is recruited co-transcriptionally to non-coding regions to drive transcriptional interference

Project description:Seb1 is recruited co-transcriptionally to non-coding regions to drive transcriptional interference

| PRJNA360639 | ENA

Targeted protein degradation reveals direct roles of SPT6 in POL2 elongation and termination

Project description:SPT6 is both, an important histone chaperone and POL2 elongation factor but its primary role on transcription in mammalian cells remains open, as no acute depletion system is available. We used targeted protein degradation to rapidly deplete SPT6 and analyzed defects in POL2 behavior by a multi-omics approach and estimated POL2 processivity, elongation rates and termination and compared it to gene transcription. Our data indicate that SPT6 is a crucial factor for POL2 elongation. Unexpectedly, SPT6 has also a vital role in POL2 termination, as acute depletion induces POL2 read through at most protein coding genes. In contrast, acute depletion did not induce spurious intragenic initiation, while this behavior can be induced by long-term depletion of SPT6 and can therefore be attributed to its function as a histone chaperone. In conclusion, targeted protein degradation of SPT6 allows the kinetic discrimination of its function as a histone chaperone and POL2 elongation factor.

2021-06-23 | GSE162264 | 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

Seb1 proximity dependent biotinylation (PDB) in Schizosaccharomyces pombe

Project description:Transcription by RNA polymerase I (RNAPI) represents most of the transcriptional activity in eukaryotic cells and is associated with the production of mature ribosomal RNA (rRNA). As several rRNA maturation steps are coupled to RNAPI transcription, the rate of RNAPI elongation directly influences processing of nascent pre-rRNA, and changes in RNAPI transcription rate can result in alternative rRNA processing pathways in response to growth conditions and stress. However, factors and mechanisms that control RNAPI progression by influencing transcription elongation rate remain poorly understood. Our project is to show that the conserved RNA-binding Seb1 is a pausing-promoting factor for RNA polymerases I to control cotranscriptional RNA processing. Here, we did a proximity dependent biotinylation followed by mass spectrometry (PDB-MS) of the Seb1 protein in order to assess for physical interactions with the RNAPI transcription machinery. A mutant E. coli BirA enzyme is fused to the Seb1 protein. This mutant version of BirA uses biotin to catalyze the formation of biotinoyl-5′-AMP (bioAMP), thereby generating a ‘cloud’ of activated biotin molecules that can react with free primary amines (most often lysine residues) of neighboring proteins. This experiment will support the conclusion that Seb1 is located at the rDNA locus.

2023-05-02 | PXD037768 | Pride

The Nrd1-like protein Seb1 coordinates co-transcriptional 3â?? end processing and polyadenylation site decision

Project description:Seb1 promotes co-transcriptional 3â?? end processing events that influence polyadenylation site decision knockdown experiments of Seb1 in Pombe

2016-05-31 | E-GEOD-75753 | biostudies-arrayexpress

The Nrd1-like protein Seb1 coordinates co-transcriptional 3 end processing and polyadenylation site selection

Project description:Three studies comprising of a strand specific RNA-seq analysis of the fission yeast transcriptome upon depletion of the termiantion factor Seb1 (2 repeats). RNAPII ChIP-seq in fission yeast upon depletion of the termiantion factor Seb1 (1 IP and 1 input). Seb1 ChIP-seq in fission yeast (HTP, 2 IP and 2 input).

2016-07-31 | E-MTAB-4827 | biostudies-arrayexpress

The Nrd1-like protein Seb1 coordinates co-transcriptional 3â end processing and polyadenylation site selection

Project description:Termination of RNAPII transcription is associated with RNA 3â end formation. For coding genes, termination is initiated by the cleavage/polyadenylation machinery. In contrast, a majority of noncoding transcription events in S. cerevisiae do not rely on RNA cleavage for termination, but instead terminate via a pathway that requires the Nrd1-Nab3-Sen1 (NNS) complex. Here we show that the S. pombe ortholog of Nrd1, Seb1, does not function in NNS-like termination, but promotes polyadenylation site selection of coding and noncoding genes. We found that Seb1 associates with 3â end processing factors, is enriched at the 3â end of genes, and binds RNA motifs downstream of cleavage sites. Importantly, a deficiency in Seb1 resulted in widespread changes in 3â UTR length as a consequence of increased alternative polyadenylation. Given that Seb1 levels affected the recruitment of conserved 3â end processing factors, our findings indicate that the conserved RNA-binding protein Seb1 co-transcriptionally controls alternative polyadenylation. Two biological replicates of Seb1 and Control (parental strain) CRAC experiments

2016-07-22 | E-GEOD-82326 | biostudies-arrayexpress

Suppressors of PHO regulon hyper-repression by Pol2 CTD Thr4 mutation implicate inositol 1-pyrophosphates as agonists of precocious lncRNA transcription termination

Project description:This RNA-Seq analysis compares gene expression of the suppressors of RNA polyemrase II CTD-T4A (STF6 and STF9 mutants) contrasting to the parental RNA polymerase II CTD-T4A mutant in the fission yeast S. pombe

2020-09-03 | GSE155609 | GEO

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