Saccharomyces cerevisiae
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ABSTRACT:
PROVIDER: PRJNA117539 | ENA |
REPOSITORIES: ENA
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Project description:RNAse III is an evolutionarily conserved family of endoribonuclease that cleaves dsRNA structures. Here we studied RNAse III homolog Pac1 In fission yeast to shed light on underappreciated roles of Drosha. We found that Pac1 co-transcriptional cleavage of nascent hairpin RNA structures trigger transcriptional termination by creating an entry point for “torpedo” exonucleases – that is, exonuclease that degrade the nascent RNA until they bump into the transcribing polymerases. As such termination pathway decouple termination from pre-mRNA polyadenylation, the nascent transcript are destabilized upon Pac1 cleavage, therefore expending the paradigm of post-transcriptional regulation of gene expression.
2021-02-19 | GSE167041 | GEO
Project description:RNAse III is an evolutionarily conserved family of endoribonuclease that cleaves dsRNA structures. Here we studied RNAse III homolog Pac1 In fission yeast to shed light on underappreciated roles of Drosha. We found that Pac1 co-transcriptional cleavage of nascent hairpin RNA structures trigger transcriptional termination by creating an entry point for “torpedo” exonucleases – that is, exonuclease that degrade the nascent RNA until they bump into the transcribing polymerases. As such termination pathway decouple termination from pre-mRNA polyadenylation, the nascent transcript are destabilized upon Pac1 cleavage, therefore expending the paradigm of post-transcriptional regulation of gene expression.
2021-02-19 | GSE167040 | GEO
Project description:Genome-wide prediction and analysis of yeast RNase III-dependent snoRNA processing signals
| PRJNA273006 | ENA
Project description:This project aims to identify novel RNA binding proteins in the baker's yeast, Saccharomyces cerevisiae. Since interactions between RNAs and proteins may be transient, yeast cells were crosslinked with UV light at 254 nm which promotes the covalent link between proteins and RNAs. After this, polyadenylated mRNAs were purified via oligo(dT) coupled to magentic beads under stringet conditions. Finally, samples were subjected to mass spectrometry analysis. To rule out the possibility of RNA-independent binding we also analysed other samples: i) samples digested with RNase one; ii) samples where we performed competition assays with polyadenylic acid.
2015-10-26 | PXD002293 | Pride
Project description:Transcription termination of messenger RNA (mRNA) is normally achieved by polyadenylation followed by Rat1p dependent 5’-3’ exoribonuleolytic degradation of the downstream transcript. Here we show that the yeast orthologue of the dsRNA-specific ribonuclease III (Rnt1p) may trigger Rat1p dependent termination of RNA transcripts that fail to terminate near polyadenylation signals. Rnt1p cleavage sites were found downstream of several genes and the deletion of RNT1 resulted in transcription read-through. Inactivation of Rat1p impaired Rnt1p dependent termination and resulted in the accumulation of 3’ end cleavage products. These results support a new model for transcription termination in which co-transcriptional cleavage by Rnt1p provides access for exoribonucleases in the absence of polyadenylation signals.
2009-10-09 | GSE16784 | GEO
Project description:MicroRNA (miRNA) play a major role in the post-transcriptional regulation of gene expression. In mammals most miRNA derive from the introns of protein coding genes where they exist as hairpin structures in the primary gene transcript, synthesized by RNA polymerase II (Pol II). These are cleaved co-transcriptionally by the Microprocessor complex, comprising DGCR8 and the RNase III endonuclease Drosha, to release the precursor (pre-)miRNA hairpin, so generating both miRNA and spliced messenger RNA1-4. However, a substantial minority of miRNA originate from Pol II-synthesized long non coding (lnc) RNA where transcript processing is largely uncharacterized5. Here, we show that most lnc-pri-miRNA do not use the canonical cleavage and polyadenylation (CPA) transcription termination pathway6, but instead use Microprocessor cleavage both to release pre-miRNA and terminate transcription. We present a detailed characterization of one such lnc-pri-miRNA that generates the highly expressed liver-specific miR-1227. Genome-wide analysis then reveals that Microprocessor-mediated transcription termination is commonly used by lnc-pri-miRNA but not by protein coding miRNA genes. This identifies a fundamental difference between lncRNA and pre-mRNA processing. Remarkably, inactivation of the Microprocessor can lead to extensive transcriptional readthrough of lnc-pri-miRNA, resulting in inhibition of downstream genes by transcriptional interference. Consequently we define a novel RNase III-mediated, polyadenylation-independent mechanism of Pol II transcription termination in mammalian cells.
2015-03-02 | GSE58838 | GEO
Project description:MicroRNA (miRNA) play a major role in the post-transcriptional regulation of gene expression. In mammals most miRNA derive from the introns of protein coding genes where they exist as hairpin structures in the primary gene transcript, synthesized by RNA polymerase II (Pol II). These are cleaved co-transcriptionally by the Microprocessor complex, comprising DGCR8 and the RNase III endonuclease Drosha, to release the precursor (pre-)miRNA hairpin, so generating both miRNA and spliced messenger RNA1-4. However, a substantial minority of miRNA originate from Pol II-synthesized long non coding (lnc) RNA where transcript processing is largely uncharacterized5. Here, we show that most lnc-pri-miRNA do not use the canonical cleavage and polyadenylation (CPA) transcription termination pathway6, but instead use Microprocessor cleavage both to release pre-miRNA and terminate transcription. We present a detailed characterization of one such lnc-pri-miRNA that generates the highly expressed liver-specific miR-1227. Genome-wide analysis then reveals that Microprocessor-mediated transcription termination is commonly used by lnc-pri-miRNA but not by protein coding miRNA genes. This identifies a fundamental difference between lncRNA and pre-mRNA processing. Remarkably, inactivation of the Microprocessor can lead to extensive transcriptional readthrough of lnc-pri-miRNA, resulting in inhibition of downstream genes by transcriptional interference. Consequently we define a novel RNase III-mediated, polyadenylation-independent mechanism of Pol II transcription termination in mammalian cells. Chromatin associated RNA-seq from sicntrl,siDrosha,siDGCR8 treated Hela cells. Same for sicntrl and siDGCR8 from Huh7 cells. Nuclear polyA + and polyA- RNA-seq from sicntrl and siDGCR8 in HeLa cells. Chromatin associated RNA-seq from siDicer treated Hela cells.
2015-03-02 | E-GEOD-58838 | biostudies-arrayexpress
Project description:THO/TREX is a conserved nuclear complex that functions in mRNP biogenesis and prevents transcription-associated recombination. Whether or not it has a ubiquitous role in the genome is an open question. ChIP-chip studies reveal that the Hpr1 component of THO and the Sub2 RNA-dependent ATPase have genome wide-distributions at active ORFs in yeast. In contrast to RNAPII, evenly distributed from promoter to termination regions, THO and Sub2 are absent at promoters and distributed in a sharp 5’→3’ gradient. Importantly, ChIP-chips reveal an over-recruitment of Rrm3 in active genes in THO mutants that is reduced by overexpression of RNase H1. Our work establishes a genome-wide function for THO-Sub2 in transcription elongation and mRNP biogenesis that function to prevent the accumulation of transcription-mediated replication obstacles, including R-loops.
2011-06-29 | GSE24960 | GEO
Project description:In the yeast Saccharomyces cerevisiae, cleavage factor I (CFI) and cleavage and polyadenylation factor (CPF) build the core of the transcription termination machinery. CFI comprises the Rna14, Rna15, Pcf11, and Clp1 proteins, as well as the associated Hrp5 RNA-binding protein. We found that CFI participates in the DNA damage response and that rna14-1 shows synthetic growth defects with mutants of different repair pathways, including homologous recombination, non-homologous end joining, post replicative repair, mismatch repair, and nucleotide excision repair, implicating that impaired RNAPII termination and 3â-end processing decreases the cellular tolerance for DNA damage. Beyond replication progression defects, we found that bypass of the G1/S checkpoint in rna14-1 cells leads to synthetic sickness, accumulation of phosphorylated H2A, as well as increase in Rad52-foci and in recombination. Our data provide evidence that CFI dysfunction impairs RNAPII turnover, leading to replication hindrance and lower tolerance to exogenous DNA damage. These findings underscore the importance of coordination between transcription termination, DNA repair and replication in the maintenance of genomic stability. S. cerevisiae strains were grown in YPAD liquid culture at 30°C, total RNA was isolated and hybridized on Affymetrix microarrays.
2014-03-31 | E-GEOD-50947 | biostudies-arrayexpress
Project description:In the yeast Saccharomyces cerevisiae, cleavage factor I (CFI) and cleavage and polyadenylation factor (CPF) build the core of the transcription termination machinery. CFI comprises the Rna14, Rna15, Pcf11, and Clp1 proteins, as well as the associated Hrp5 RNA-binding protein. We found that CFI participates in the DNA damage response and that rna14-1 shows synthetic growth defects with mutants of different repair pathways, including homologous recombination, non-homologous end joining, post replicative repair, mismatch repair, and nucleotide excision repair, implicating that impaired RNAPII termination and 3’-end processing decreases the cellular tolerance for DNA damage. Beyond replication progression defects, we found that bypass of the G1/S checkpoint in rna14-1 cells leads to synthetic sickness, accumulation of phosphorylated H2A, as well as increase in Rad52-foci and in recombination. Our data provide evidence that CFI dysfunction impairs RNAPII turnover, leading to replication hindrance and lower tolerance to exogenous DNA damage. These findings underscore the importance of coordination between transcription termination, DNA repair and replication in the maintenance of genomic stability.
2014-03-31 | GSE50947 | GEO