Analysis of mRNA decay intermediates in Bacillus subtilis 3' exoribonuclease and RNA helicase mutant strains
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ABSTRACT:
PROVIDER: PRJNA792588 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR17335818_1.fastq.gz | Fastqsanger.gz | |||
| SRR17335818_2.fastq.gz | Fastqsanger.gz | |||
| SRR17335819_1.fastq.gz | Fastqsanger.gz | |||
| SRR17335819_2.fastq.gz | Fastqsanger.gz | |||
| SRR17335820_1.fastq.gz | Fastqsanger.gz |
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Project description:The Bacillus subtilis genome encodes four 3’ exoribonucleases: polynucleotide phosphorylase (PNPase), RNase R, RNase PH, and YhaM. Previous work showed that PNPase, encoded by the pnpA gene, is the major 3’ exonuclease involved in mRNA turnover; in a pnpA deletion strain, numerous mRNA decay intermediates accumulate. Whether B. subtilis mRNA decay occurs in the context of a degradosome complex is controversial. In this study, global mapping of mRNA decay intermediate 3’ ends within coding sequences was performed in strains that were either deleted for, or had an inactivating point mutation in, the pnpA gene. The pattern of 3’ end accumulation in these strains was highly similar, suggesting that mRNA decay was not occurring in the context of a degradosome, whose structure would be affected by the absence of PNPase. A comparison with mapped 3’ ends in a strain lacking CshA, the major RNA helicase, indicated that different mRNAs may require both PNPase and CshA for efficient decay. RNA-seq analysis of strains lacking RNase R suggested that this enzyme did not play a major role in mRNA turnover in the wild-type strain. Strains were constructed that contained only one of the four known 3’ exoribonucleases. When RNase R was the only 3’ exonuclease present, it was able to degrade a model mRNA efficiently, showing processive decay even through a strong stem-loop structure that inhibits PNPase processivity. Strains containing only RNase PH or only YhaM were also insensitive to this RNA secondary structure, suggesting the existence of another, as-yet unidentified, 3’ exoribonuclease.
2022-01-03 | GSE192670 | GEO
Project description:Messenger RNA decay in Bacillus subtilis is accomplished by a combination of exoribonucleases and endoribonucleases. Intermediates in the decay process have not been readily detectable, and previous studies on mRNA decay have used a handful of highly expressed transcripts as models. Here, we use RNA‐Seq analysis to probe mRNA turnover globally. A significant fraction of messages showed differential accumulation of RNA fragments that mapped near the 5′ or 3′ end of the coding sequence, consistent with initiation of decay from either the 5′ end or from an internal cleavage site. Patterns of mRNA decay in the wild type were compared with patterns in a mutant strain lacking polynucleotide phosphorylase (PNPase), which is considered the major 3′ exonuclease activity in mRNA decay and which is one of four known 3′ exonucleases in B. subtilis. The results showed a striking dependence on PNPase for mRNA turnover in many cases, suggesting specificity in the ability of 3′ exonucleases to degrade from 3′‐hydroxyl termini. RNA‐Seq data demonstrated a sharp decrease in expression of Sigma D in the PNPase‐deletion strain. Reduction in sigD regulon expression explained the chain growth phenotype of the PNPase mutant and also predicted a defect in swarming motility.
2018-09-18 | GSE120050 | GEO
Project description:Global analysis of mRNA decay intermediates in Bacillus subtilis wild-type and polynucleotide phosphorylase-deletion strains
| PRJNA491499 | ENA
Project description:The helicase activity of UPF1 remodels miRNPs so as to render miRNAs susceptible to Tudor-SN (TSN)-mediated miRNA decay
2017-08-22 | GSE100068 | GEO
Project description:The general pathways of eukaryotic mRNA decay occur via deadenylation followed by 3’ to 5’ degradation or decapping, although some endonuclease sites have been identified in metazoan mRNAs. To determine the role of endonucleases in mRNA degradation in Saccharomyces cerevisiae, we mapped 5’ monophosphate ends on mRNAs in wild-type and dcp2∆ xrn1∆ yeast cells, wherein mRNA endonuclease cleavage products are stabilized. This led to three important observations. First, only few mRNAs that undergo low level endonucleotyic cleavage were observed suggesting that endonucleases are not a major contributor to yeast mRNA decay. Second, independent of known decapping enzymes, we observed low levels of 5’ monophosphates on some mRNAs suggesting that an unknown mechanism can generate 5' exposed ends, although for all substrates tested Dcp2 was the primary decapping enzyme. Finally, we identified debranched lariat intermediates from intron-containing genes, demonstrating a significant discard pathway for mRNAs during the second step of pre-mRNA splicing, which is a potential new step to regulate gene expression. 5' monophosphorylated ends of poly(A) RNA from wild-type and dcp2D xrn1D strains were identified in duplicates and triplicates, respectively.
2012-06-17 | E-GEOD-33712 | biostudies-arrayexpress
Project description:The general pathways of eukaryotic mRNA decay occur via deadenylation followed by 3’ to 5’ degradation or decapping, although some endonuclease sites have been identified in metazoan mRNAs. To determine the role of endonucleases in mRNA degradation in Saccharomyces cerevisiae, we mapped 5’ monophosphate ends on mRNAs in wild-type and dcp2∆ xrn1∆ yeast cells, wherein mRNA endonuclease cleavage products are stabilized. This led to three important observations. First, only few mRNAs that undergo low level endonucleotyic cleavage were observed suggesting that endonucleases are not a major contributor to yeast mRNA decay. Second, independent of known decapping enzymes, we observed low levels of 5’ monophosphates on some mRNAs suggesting that an unknown mechanism can generate 5' exposed ends, although for all substrates tested Dcp2 was the primary decapping enzyme. Finally, we identified debranched lariat intermediates from intron-containing genes, demonstrating a significant discard pathway for mRNAs during the second step of pre-mRNA splicing, which is a potential new step to regulate gene expression.
2012-06-17 | GSE33712 | GEO
Project description:Transcriptome-wide methodology is developed to capture and sequence decay intermediates deriving from direct NMD targets. Results demonstrate that mammalian-cell NMD generates decay intermediates in a process that involves the covalent addition of nontemplated U, C, G and A nucleotides to 3'-ends. Variations of this methodology demonstrate that these decay intermediates arise co-translationally.
2018-07-05 | GSE111818 | GEO
Project description:Messenger RNA decay in Bacillus subtilis is accomplished by a combination of exoribonucleases and endoribonucleases. Intermediates in the decay process have not been readily detectable, and previous studies on mRNA decay have used a handful of highly expressed transcripts as models. Here, we use RNA-Seq analysis to probe mRNA turnover globally. A significant fraction of messages showed differential accumulation of RNA fragments that mapped near the 5' or 3' end of the coding sequence, consistent with initiation of decay from either the 5' end or from an internal cleavage site. Patterns of mRNA decay in the wild type were compared with patterns in a mutant strain lacking polynucleotide phosphorylase (PNPase), which is considered the major 3' exonuclease activity in mRNA decay and which is one of four known 3' exonucleases in B. subtilis. The results showed a striking dependence on PNPase for mRNA turnover in many cases, suggesting specificity in the ability of 3' exonucleases to degrade from 3'-hydroxyl termini. RNA-Seq data demonstrated a sharp decrease in expression of Sigma D in the PNPase-deletion strain. Reduction in sigD regulon expression explained the chain growth phenotype of the PNPase mutant and also predicted a defect in swarming motility.
| S-EPMC4177450 | biostudies-literature
Project description:mRNA decay in E. Coli degradosome mutants and their parental strains following transcriptional arrest with rifampicin. Abstract: RNase E, an essential endoribonuclease of Escherichia coli, interacts through its C-terminal region with multiple other proteins to form a complex termed the RNA degradosome. To investigate the degradosome's proposed role as an RNA decay machine, we used DNA microarrays to globally assess alterations in the steady-state abundance and decay of 4,289 E. coli mRNAs at single-gene resolution in bacteria carrying mutations in the degradosome constituents RNase E, polynucleotide phosphorylase, RhlB helicase, and enolase. Our results show that the functions of all four of these proteins are necessary for normal mRNA turnover. We identified specific transcripts and functionally distinguishable transcript classes whose half-life and abundance were affected congruently by multiple degradosome proteins, affected differentially by mutations in degradosome constituents, or not detectably altered by degradosome mutations. Our results, which argue that decay of some E. coli mRNAs in vivo depends on the action of assembled degradosomes, whereas others are acted on by degradosome proteins functioning independently of the complex, imply the existence of structural features or biochemical factors that target specific classes of mRNAs for decay by degradosomes. An RNA stablity experiment design type examines stability and/or decay of RNA transcripts. User Defined
2006-01-05 | E-GEOD-3978 | biostudies-arrayexpress
Project description:mRNA decay in E. Coli degradosome mutants and their parental strains following transcriptional arrest with rifampicin. Abstract: RNase E, an essential endoribonuclease of Escherichia coli, interacts through its C-terminal region with multiple other proteins to form a complex termed the RNA degradosome. To investigate the degradosome's proposed role as an RNA decay machine, we used DNA microarrays to globally assess alterations in the steady-state abundance and decay of 4,289 E. coli mRNAs at single-gene resolution in bacteria carrying mutations in the degradosome constituents RNase E, polynucleotide phosphorylase, RhlB helicase, and enolase. Our results show that the functions of all four of these proteins are necessary for normal mRNA turnover. We identified specific transcripts and functionally distinguishable transcript classes whose half-life and abundance were affected congruently by multiple degradosome proteins, affected differentially by mutations in degradosome constituents, or not detectably altered by degradosome mutations. Our results, which argue that decay of some E. coli mRNAs in vivo depends on the action of assembled degradosomes, whereas others are acted on by degradosome proteins functioning independently of the complex, imply the existence of structural features or biochemical factors that target specific classes of mRNAs for decay by degradosomes. An RNA stablity experiment design type examines stability and/or decay of RNA transcripts. Keywords: RNA_stability_design
2006-01-06 | GSE3978 | GEO