TmRNADB
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REPOSITORIES: ENA
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Project description:We used a combination of genetic and proteomic approaches to characterize tmRNA (ssrA) activity in the genome-reduced bacterium Mycoplasma pneumoniae. For this, we generated tmRNA mutants encoding a tag resistant to proteolysis. Endogenous protein tagging by the mutant tmRNA gene (ssrAmk) was then examined by immunoprecipitation (IP) enrichment followed by LC-MS/MS analysis. Additionally, RNA-seq differential expression analysis of the mutants compared to the wild-type strain was assessed.
2021-08-09 | E-MTAB-10111 | biostudies-arrayexpress
Project description:We used a combination of genetic and proteomic approaches to characterize tmRNA (ssrA) activity in the genome-reduced bacterium Mycoplasma pneumoniae. For this, we generated tmRNA mutants encoding a tag resistant to proteolysis. Endogenous protein tagging by the mutant tmRNA gene (ssrAmk) was then examined by immunoprecipitation (IP) enrichment followed by LC-MS/MS analysis.
2021-08-23 | PXD023973 | Pride
Project description:Organisms possessing genetic codes with unassigned codons raise the question of how cellular machinery resolves such codons and how this could impact horizontal gene transfer. Here, we use a genomically recoded Escherichia coli to examine how organisms address translation at unassigned UAG codons, which obstruct propagation of UAG-containing viruses and plasmids. Using mass spectrometry, we show that recoded organisms resolve translation at unassigned UAG codons via near-cognate suppression, dramatic frameshifting from at least -3 to +19 nucleotides, and rescue by ssrA-encoded tmRNA, ArfA, and ArfB. We then demonstrate that deleting tmRNA restores expression of UAG-ending proteins and propagation of UAG-containing viruses and plasmids in the recoded strain, indicating that tmRNA rescue and nascent peptide degradation is the cause of impaired virus and plasmid propagation. The ubiquity of tmRNA homologs suggests that genomic recoding is a promising path to impair horizontal gene transfer and confer genetic isolation in diverse organisms.
2018-09-18 | PXD009643 | Pride
Project description:We assess the mRNA-enrichment performance of a custom-made non-mRNA depletion protocol in comparison to a commercially available mRNA-enrichment kit (Ribo-off, Vazyme). Whereas most available kits focus only on removal of rRNA, our method also targets the transfer-messenger RNA (tmRNA). tmRNA was shown to consume up to 25% of the reads in RNA-sequencing data of Pseudomonas aeruginosa. Our established depletion technique is based on the targeting of overly abundant RNA species (16S and23S rRNA, tmRNA) in total RNA preparations of Pseudomonas aeruginosa PA14 by hybridization with organism-specific DNA probes and subsequent degradation by RNase H treatment. While introducing no systematic bias into the gene expression profile we were able to increase the mRNA read share of the total reads in the samples treated with our mRNA-enrichment protocol to 93% - 99%. Therefore, our custom-made depletion technique outcompeted the commercially available reference kit (72% mRNA share) and represents a cost-efficient mRNA-enrichment method for high-throughput next-generation sequencing.
2020-03-31 | GSE147748 | GEO
Project description:In our study, we found that lack of tmRNA affects the biofilm formation in Bacillus subtilis, which has important research significance. Then, we obtained a revertant strain MB from tmRNA mutant strain. And MB strain could restore biofilm production capacity. Therefore, the transcriptomes of the three strains were sequenced. Based on the statistical analysis of gene expression (P < 0.05), there were 756 genes up regulated while 992 genes down regulated by at least two folds when comparing TM to WT. Secondly, there were 974 genes up regulated while 563 genes down regulated by at least two folds when comparing MB to TM. And there were 179 genes up regulated while 307 genes down regulated by at least two folds when comparing MB to WT.
2022-08-03 | GSE199151 | GEO
Project description:To elucidate whole-transcriptome changes in stationary-phase of growth of L. monocytogenes induced by inactivation of lmo0946 gene, we performed transcriptome comparison of wild-type and its derivative mutant in lmo0946 gene. The analysis revealed very high abundance of two non-coding RNAs, namely Rli47 (sRNA) and SsrA (tmRNA) in both studied strains.
2023-08-05 | E-MTAB-10810 | biostudies-arrayexpress
Project description:Ribosome stalling is a major problem in all domains of life. When a ribosome stalls, trailing ribosomes may catch up to and collide with the stalled ribosome, depleting protein synthesis capacity. Here, we describe a novel pathway used by Gram-positive bacteria to rescue ribosome collisions. We used the ATPase defective ABCF protein YdiF(EQ2) to induce ribosome stalling and collisions in Bacillus subtilis. Ribosome profiling (Ribo-seq) of YdiF(EQ2)-expressing cells revealed that collided ribosomes are enriched for tmRNA, a functional RNA involved in trans-translation. We confirmed that tmRNA tagging activity is globally increased upon expression of any ATPase defective ABCF as well as in cells treated with the collision-inducing antibiotic erythromycin, suggesting this is a generalizable mechanism to rescue stalled and collided ribosomes. The global increase in tmRNA tagging that occurred in response to both erythromycin and YdiF(EQ2) induced collisions was dependent on the Rae1 endonuclease. Loss of trans-translation in cells experiencing widespread ribosome collisions leads to a severe fitness defect, consistent with the importance of this pathway in rescuing ribosomes stalled on truncated mRNAs that result from ribosome collisions. Altogether, our work supports a model in which Rae1 cleaves mRNA on collided ribosomes, thereby generating a truncated mRNA substrate for trans-translation and leading to rescue and recycling of the collided ribosomes. We term this mechanism Collision-Associated Rae1-induced trans-Translation (CART). CART broadens the repertoire of tools that bacteria use to manage ribosome collisions.
2026-09-23 | GSE342019 | GEO
Project description:Ribosome stalling is a major problem in all domains of life. When a ribosome stalls, trailing ribosomes may catch up to and collide with the stalled ribosome, depleting protein synthesis capacity. Here, we describe a novel pathway used by Gram-positive bacteria to rescue ribosome collisions. We used the ATPase defective ABCF protein YdiF(EQ2) to induce ribosome stalling and collisions in Bacillus subtilis. Ribosome profiling (Ribo-seq) of YdiF(EQ2)-expressing cells revealed that collided ribosomes are enriched for tmRNA, a functional RNA involved in trans-translation. We confirmed that tmRNA tagging activity is globally increased upon expression of any ATPase defective ABCF as well as in cells treated with the collision-inducing antibiotic erythromycin, suggesting this is a generalizable mechanism to rescue stalled and collided ribosomes. The global increase in tmRNA tagging that occurred in response to both erythromycin and YdiF(EQ2) induced collisions was dependent on the Rae1 endonuclease. Loss of trans-translation in cells experiencing widespread ribosome collisions leads to a severe fitness defect, consistent with the importance of this pathway in rescuing ribosomes stalled on truncated mRNAs that result from ribosome collisions. Altogether, our work supports a model in which Rae1 cleaves mRNA on collided ribosomes, thereby generating a truncated mRNA substrate for trans-translation and leading to rescue and recycling of the collided ribosomes. We term this mechanism Collision-Associated Rae1-induced trans-Translation (CART). CART broadens the repertoire of tools that bacteria use to manage ribosome collisions.
2026-09-23 | GSE342018 | GEO
Project description:Bacterial persisters are a small proportion of phenotypically heterogeneous variants with the transient capability to survive in high concentrations of antibiotics, causing recurrent infections in both human and aquatic animals. Transfer-messenger RNA (tmRNA), which was encoded by the <i>ssrA</i> gene, was identified as a determinant regulator mediating the persistence to β-lactams in the pathogenic <i>Aeromonas veronii</i> C4. The deletion of tmRNA exhibited the increased ability of persister formation most probably due to the reduction of protein synthesis. Transcriptomic and metabolomic analyses revealed that the absence of tmRNA not only significantly elevated the intercellular levels of metabolite GlcNAc and promoted NaCl osmotic tolerance, but also upregulated the expression of metabolic genes in both the upstream biosynthesis pathway and the downstream metabolic flux of peptidoglycan (PG) biosynthesis. Finally, exogenous GlcNAc stimulated significant bacterial growth, enhanced content of GlcNAc in the cell wall, higher resistance to osmotic response, and higher persistence to cefotaxime in a concentration-dependent manner, implying its potential role in promoting the multiple phenotypes observed in tmRNA deletion strains. Taken together, these results hint at a potential mechanism of persister formation mediated by tmRNA against the β-lactam challenges in <i>A. veronii</i>.
2021-10-18 | MTBLS1182 | MetaboLights