Sense codon-misassociated eRF1 elicits widespread ribosome stalling subjected to quality control
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA1150324 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR30321751_1.fastq.gz | Fastqsanger.gz | |||
| SRR30321751_2.fastq.gz | Fastqsanger.gz | |||
| SRR30321752_1.fastq.gz | Fastqsanger.gz | |||
| SRR30321752_2.fastq.gz | Fastqsanger.gz | |||
| SRR30321753_1.fastq.gz | Fastqsanger.gz |
Items per page: 5 1 - 5 of 54 |
Similar Datasets
Project description:The ribosome-associated quality control (RQC) is a surveillance system for aberrant translation, sensing ribosome collisions. Although the molecular mechanism has been extensively studied, the endogenous targets of RQC in human cells were poorly understood. Here, starting from the study of codon specificity of eukaryotic termination factor eRF1, we unexpectedly find that misrecognition of UUA sense codon by eRF1 leads to ribosome collisions and provides the source of RQC substrates in humans. eRF1-selective Monosome-Seq and Disome-Seq reveal that eRF1 recruitment to ribosome was not restricted to the stop codons but also the sub-cognate sense codons, including the UUA codon. The UUA misrecognition by eRF1 causes ribosome collision without termination reaction. Remarkably, Disome-Seq with the depletion of ASCC3 and 4EHP, key factors in RQC, showed that ribosome stalled at UUA codons are the predominant sub-populations rescued by RQC. Failure to resolve ribosome collisions by RQC triggers p38 phosphorylation and upregulation of stress response transcription factor ATF3. This study presents the impact of sense codon misrecognition by the termination factor on translation homeostasis in human cells.
2024-09-18 | GSE275336 | GEO
Project description:Translation of mRNA into a polypeptide is terminated when the release factor eRF1 recognizes a UAA, UAG, or UGA stop codon in the ribosomal A site and stimulates nascent peptide release. However, stop codon readthrough can occur when a near-cognate tRNA outcompetes eRF1 in decoding the stop codon, resulting in the continuation of the elongation phase of protein synthesis. At the end of a conventional mRNA coding region readthrough allows translation into the mRNA 3’-UTR. Previous studies with reporter systems have shown that the efficiency of termination or readthrough is modulated by cis-acting elements other than stop codon identity, including two nucleotides 5’ of the stop codon, six nucleotides 3’ of the stop codon in the ribosomal mRNA channel, and stem-loop structures in the mRNA 3’-UTR. It is unknown whether these elements are important at a genome-wide level and whether other mRNA features proximal to the stop codon significantly affect termination and readthrough efficiencies in vivo. Accordingly, we carried out ribosome profiling analyses of yeast cells expressing wild-type or temperature-sensitive eRF1 and developed bioinformatics strategies to calculate readthrough efficiency, and to identify mRNA and peptide features which influence that efficiency. We found that the stop codon (nt +1 to +3), the nucleotide after it (nt +4), the codon in the P site (nt -3 to -1), and 3’-UTR length are the most influential features in the control of readthrough efficiency, while nts +5 to +9 and mRNA secondary structure in the 3’-UTR had milder effects. Additionally, we found low readthrough genes to have shorter 3’-UTRs compared to high readthrough genes in cells with thermally inactivated eRF1, while this trend was reversed in wild-type cells. Together, our results demonstrated the general roles of known regulatory elements in genome-wide regulation and identified several new mRNA or peptide features important for translation termination and readthrough.
2020-12-08 | GSE162780 | GEO
Project description:Stalling of the ribosomes on the mRNA has been linked to myriad of molecular and biological functions. While a multitude of algorithms predict codon-usage in ribosome profiling data, no tool exists to predict the context-specific stalling of ribosomes on transcripts. Combining these two features can reveal unprecedented insights in the ribosomal function and their usage across cells and conditions. To overcome this lack of tools to comprehensively analyze these features, we created Bumpfinder, a web-server, that provides comprehensive analytical details on differences in total ribosome occupancy on transcripts, ribosomal occupancy across codons, ribosome-stalling and the underlying causal amino-acids/codons, as well as collisions. Employing Bumpfinder on generated and publicly available ribosome-profiling datasets, we studied the response of various cell types to amino-acid starvation conditions. We observed interesting patterns in response to the biological effects, suggesting the impact of quality control mechanisms of protein synthesis. Complimentary analysis of collided ribosomes demonstrated phased ribosome stalling on the identified sites. Thus, by deciphering context-specific ribosome distribution, Bumpfinder provides unprecedented molecular insights in the ribosome function and mRNA translation during stress response conditions.
2023-12-31 | GSE156443 | GEO
Project description:The control of mRNA stability plays a central role in regulating gene expression patterns. While much is known about the roles of 5´ and 3´ untranslated regions in the mRNA stability control, the impact of protein-coding sequences on mRNA stability had been obscure. Recently, several groups reported that codon composition in the ORF affects mRNA deadenylation and degradation rates in a translation-dependent manner. Hence, codons define not only the amino acid sequences to be synthesized but also the stability of mRNAs. However, how 61 codons differently affect mRNA stability remains unclear. Besides, aberrant stalling of the ribosome induces ribosome quality control (RQC) and No-go decay. The relationship between the two co-translational mRNA decay pathways is not systematically analyzed. To precisely characterize the effects of 61 codons on mRNA stability, we developed a simplified reporter system that allows detection of the effect of every single codon on mRNA stability in zebrafish embryos. Using this system, we show that the effect of codons on mRNA stability is partially but significantly correlated with the translation elongation rate and tRNA abundance. Interestingly, the codon effect is still maintained in zebrafish embryos lacking Znf598, an essential mediator of RQC and NGD. Znf598-dependent NGD targets a particular type of ribosome stalling but has limited impact on endogenous mRNA stability. Our study thus defines two related co-translational mRNA decay pathways during animal development.
2020-05-05 | GSE133392 | GEO
Project description:DNA replication forks in eukaryotic cells stall at a variety of replication barriers. Stalling forks require strict cellular regulations to prevent fork collapse. However, the mechanism underlying these cellular regulations is poorly understood. In this study, a cellular mechanism was uncovered that regulates chromatin structures to stabilize stalling forks. When replication forks stall, H2BK33, a newly identified acetylation site, is deacetylated and H3K9 trimethylated in the nucleosomes surrounding stalling forks, which results in chromatin compaction around forks. Acetylation-mimic H2BK33Q and its deacetylase clr6-1 mutations compromise this fork stalling-induced chromatin compaction, cause physical separation of replicative helicase and DNA polymerases, and significantly increase the frequency of stalling fork collapse. Furthermore, this fork stalling-induced H2BK33 deacetylation is independent of checkpoint. In summary, these results suggest that eukaryotic cells have developed a cellular mechanism that stabilizes stalling forks by targeting nucleosomes and inducing chromatin compaction around stalling forks. This mechanism is named the "Chromsfork" control: Chromatin Compaction Stabilizes Stalling Replication Forks.
| S-EPMC6642376 | biostudies-literature
Project description:In bacteria, translation-transcription coupling inhibits RNA polymerase (RNAP) stalling. We present evidence suggesting that, upon amino acid starvation, inactive ribosomes promote rather than inhibit RNAP stalling. We developed an algorithm to evaluate genome-wide polymerase progression independently of local noise, and used it to reveal that the transcription factor DksA inhibits promoter-proximal pausing and increases RNAP elongation when uncoupled from translation by depletion of charged tRNAs. DksA has minimal effect on RNAP elongation in vitro and on untranslated RNAs in vivo. In these cases, transcripts can form RNA structures that prevent backtracking. Thus, the effect of DksA on transcript elongation may occur primarily upon ribosome slowing/stalling or at promoter-proximal locations that limit the potential for RNA structure. We propose that inactive ribosomes prevent formation of backtrackblocking mRNA structures and that, in this circumstance, DksA acts as a transcription elongation factor in vivo. Chromatin immunoprecipitation (ChIP) experiments were performed by using antibodies against RNA polymerase b subunit in wild-type and DdksA cells treated with 0.5mg/ml serine hydroxamate (SHX) or untreated. DksA and s70 enrichments were compared to RNAP enrichment by ChIP experiments using antibodies against s70 and DksA in wild-type cells (also in DdksA cells as a negative control for DksA ChIP-chip). Differentially labeled ChIP DNA and genomic DNA were competitively hybridized to an E. coli K-12 MG1655 tiling array with overlapping probes at ~12bp spacing across the entire genome. The series contains 19 datasets.
2014-04-21 | E-GEOD-54669 | biostudies-arrayexpress
Project description:Control of messenger RNA (mRNA) decay rate is intimately connected to translation elongation, but the spatial coordination of these events is poorly understood. The Ccr4-Not complex initiates mRNA decay through deadenylation and activation of decapping. We used a combination of cryo–electron microscopy, ribosome profiling, and mRNA stability assays to examine the recruitment of Ccr4-Not to the ribosome via specific interaction of the Not5 subunit with the ribosomal E-site in Saccharomyces cerevisiae. This interaction occurred when the ribosome lacked accommodated A-site transfer RNA, indicative of low codon optimality. Loss of the interaction resulted in the inability of the mRNA degradation machinery to sense codon optimality. Our findings elucidate a physical link between the Ccr4-Not complex and the ribosome and provide mechanistic insight into the coupling of decoding efficiency with mRNA stability.
2020-04-16 | GSE144250 | GEO
Project description:Control of messenger RNA (mRNA) decay rate is intimately connected to translation elongation, but the spatial coordination of these events is poorly understood. The Ccr4-Not complex initiates mRNA decay through deadenylation and activation of decapping. We used a combination of cryo–electron microscopy, ribosome profiling, and mRNA stability assays to examine the recruitment of Ccr4-Not to the ribosome via specific interaction of the Not5 subunit with the ribosomal E-site in Saccharomyces cerevisiae. This interaction occurred when the ribosome lacked accommodated A-site transfer RNA, indicative of low codon optimality. Loss of the interaction resulted in the inability of the mRNA degradation machinery to sense codon optimality. Our findings elucidate a physical link between the Ccr4-Not complex and the ribosome and provide mechanistic insight into the coupling of decoding efficiency with mRNA stability.
2020-04-16 | GSE131408 | GEO
Project description:Somatic hypermutation (SHM) drives the genetic diversity of immunoglobulin genes in activated B cells and supports the generation of antibodies with increased affinity for antigen. SHM is targeted to Ig genes by their enhancers (DIVACs; diversification activators) but how the enhancers mediate this activity is unknown. We show that DIVACs that strongly stimulate SHM increase the phosphorylation of RNA polymerase 2 (Pol2) and Pol2 occupancy in the mutating gene with little or no accompanying increase in elongation-competent Pol2 or production of full length transcripts, indicating the accumulation of stalled Pol2. DIVAC-induced stalling is weakly associated with an increase in the detection of single-stranded DNA bubbles in the mutating target gene. We did not find evidence for anti-sense transcription or an association of H3K27ac with DIVAC activity in the mutating gene. These findings argue for a connection between Pol2 stalling and cis-acting targeting elements in the context of SHM and thus define a mechanistic basis for locus-specific targeting of SHM in the genome. Our results suggest that DIVACs make the target gene a suitable platform for AID-mediated mutation without the need for increasing transcriptional output.
2021-10-22 | GSE180176 | GEO
Project description:Somatic hypermutation (SHM) drives the genetic diversity of immunoglobulin genes in activated B cells and supports the generation of antibodies with increased affinity for antigen. SHM is targeted to Ig genes by their enhancers (DIVACs; diversification activators) but how the enhancers mediate this activity is unknown. We show that DIVACs that strongly stimulate SHM increase the phosphorylation of RNA polymerase 2 (Pol2) and Pol2 occupancy in the mutating gene with little or no accompanying increase in elongation-competent Pol2 or production of full length transcripts, indicating the accumulation of stalled Pol2. DIVAC-induced stalling is weakly associated with an increase in the detection of single-stranded DNA bubbles in the mutating target gene. We did not find evidence for anti-sense transcription or an association of H3K27ac with DIVAC activity in the mutating gene. These findings argue for a connection between Pol2 stalling and cis-acting targeting elements in the context of SHM and thus define a mechanistic basis for locus-specific targeting of SHM in the genome. Our results suggest that DIVACs make the target gene a suitable platform for AID-mediated mutation without the need for increasing transcriptional output.
2021-10-22 | GSE180177 | GEO