Molecular interactions between Hel2 and RNA supporting ribosome-associated quality control [II]
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ABSTRACT:
PROVIDER: PRJNA471331 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR7163430.fastq.gz | Fastqsanger.gz | |||
| SRR7163440.fastq.gz | Fastqsanger.gz | |||
| SRR7163431.fastq.gz | Fastqsanger.gz | |||
| SRR7163441.fastq.gz | Fastqsanger.gz | |||
| SRR7163432.fastq.gz | Fastqsanger.gz |
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Similar Datasets
Project description:Molecular interactions between Hel2 and RNA supporting ribosome-associated quality control
| PRJNA471327 | ENA
Project description:Molecular interactions between Hel2 and RNA supporting ribosome-associated quality control [I]
| PRJNA471315 | ENA
Project description:Ribosome-associated quality control (RQC) pathways monitor and respond to stalling of translating ribosomes. Using a newly developed technique based on in vivo UV crosslinking and mass spectrometry, we identify a C-terminal region in Hel2/Rqt1 as an RNA binding domain, with amino acids L501/K502 directly interacting with RNA. In vivo crosslinking of Hel2 revealed binding to 18S rRNA and translating mRNAs. Consistent with the 18S binding site located between mRNA entrance and exit channels, Hel2 preferentially bound mRNA both upstream and downstream of the termination codon. A C-terminal truncation that deleted L501/K502, abolished crosslinking to 18S rRNA, altered mRNA binding patterns, and reduced Hel2 function comparable to hel2∆. Asc1, also participates in RQC and ASC1 deletion impaired Hel2 18S and mRNA binding. We conclude that Hel2 is recruited or stabilized on translating 40S ribosomal subunits by interactions with 18S rRNA and Asc1. Ribosome-bound Hel2 interacts with mRNA, predominately during translation termination.
2019-01-18 | GSE114420 | GEO
Project description:Ribosome-associated quality control (RQC) pathways monitor and respond to stalling of translating ribosomes. Using a newly developed technique based on in vivo UV crosslinking and mass spectrometry, we identify a C-terminal region in Hel2/Rqt1 as an RNA binding domain, with amino acids L501/K502 directly interacting with RNA. In vivo crosslinking of Hel2 revealed binding to 18S rRNA and translating mRNAs. Consistent with the 18S binding site located between mRNA entrance and exit channels, Hel2 preferentially bound mRNA both upstream and downstream of the termination codon. A C-terminal truncation that deleted L501/K502, abolished crosslinking to 18S rRNA, altered mRNA binding patterns, and reduced Hel2 function comparable to hel2∆. Asc1, also participates in RQC and ASC1 deletion impaired Hel2 18S and mRNA binding. We conclude that Hel2 is recruited or stabilized on translating 40S ribosomal subunits by interactions with 18S rRNA and Asc1. Ribosome-bound Hel2 interacts with mRNA, predominately during translation termination.
2019-01-18 | GSE114415 | GEO
Project description:This SuperSeries is composed of the SubSeries listed below.
2019-01-18 | GSE114429 | GEO
Project description:Ubiquitination of Stalled Ribosome Triggers Ribosome-associated Quality Control
| PRJNA356322 | ENA
Project description:Ribosome-associated quality control (RQC) pathways monitor and respond to ribosome stalling. Using in vivo UV-crosslinking and mass spectrometry, we identified a C-terminal region in Hel2/Rqt1 as an RNA binding domain. Complementary crosslinking and sequencing data for Hel2 revealed binding to 18S rRNA and translated mRNAs. Hel2 preferentially bound mRNAs upstream and downstream of the stop codon. C-terminal truncation of Hel2 abolished the major 18S crosslink and polysome association, and altered mRNA binding. HEL2 deletion caused loss of RQC and, we report here, no-go decay (NGD), with comparable effects for Hel2 truncation including the RNA-binding site. Asc1 acts upstream of Hel2 in RQC and asc1? impaired Hel2 binding to 18S and mRNA. In conclusion: Hel2 is recruited or stabilized on translating 40S ribosomal subunits by interactions with 18S rRNA and Asc1. This 18S interaction is required for Hel2 function in RQC and NGD. Hel2 probably interacts with mRNA during translation termination.
| S-EPMC6362110 | biostudies-literature
Project description:Translation elongation stalling has the potential to produce toxic truncated protein fragments. Translation of either non-stop mRNA or transcripts coding for poly-basic sequences induces ribosome stalling, and the arrest product is degraded by the ribosome-mediated quality control (RQC) system. During this process, the stalled ribosome is dissociated into subunits, and the polypeptide is ubiquitinated by the E3 ubiquitin ligase Listerin on the 60S large ribosomal subunit, leading to subsequent proteasomal degradation. However, it is largely unknown how the specific stalled ribosomes are recognized as aberrant to engage the RQC system. Here, we report that ubiquitination of the ribosomal protein uS10 of the 40S small ribosomal subunit, by the E3 ubiquitin ligase Hel2 (or RQC-trigger (Rqt) 1) initiates RQC. We identified a novel RQC-trigger (RQT) complex composed of the RNA helicase-family protein Slh1/Rqt2, the ubiquitin binding protein Cue3/Rqt3, and yKR023W/Rqt4 that is required for RQC. The defects in RQC of the RQT mutants correlated with sensitivity to anisomycin, which stalls ribosome at the rotated form, suggesting that RQT factors rescue ribosomes stalled by this drug. Our un-biased survey by ribosome profiling revealed that ribosomes stalled at the rotated state with specific pairs of codons at P-A sites serve as RQC substrates. Rqt1 specifically ubiquitinates these arrested ribosomes to target them to the RQT complex, allowing subsequent RQC reactions including dissociation of the stalled ribosome into subunits. Our results provide mechanistic insight into the surveillance system for aberrant proteins induced by ribosome stalling and mediated by ribosome ubiquitination.
2017-07-31 | GSE90919 | GEO
Project description:Translation elongation rates are regulated to ensure proper conformation and biological function of proteins. Translation of either non-stop mRNA or transcripts coding for poly-basic sequences induces ribosome stalling, and the arrest product is degraded by the ribosome-mediated quality control system (RQC). During this process, the stalled ribosome is dissociated into subunits, and the polypeptide is ubiquitinated by the E3 ubiquitin ligase Listerin on the 60S large ribosomal subunit (LSU) leading to subsequent proteasomal degradation. However, it is largely unknown how stalled ribosomes are recognized and dissociated into subunits. Here we report that ubiquitination of the ribosomal protein uS10 by the E3 ubiquitin ligase Hel2 is required for the production of the RQC substrate. RQC-trigger (RQT) factors, a RNA helicase-family protein Slh1/Rqt2, ubiquitin binding protein Cue3/Rqt3 and yKR023W/Rqt4, were also required for the primary steps of RQC, and associated with Hel2-ribosome complexes. Rqt2-4 factors were dispensable for the ubiquitination of uS10 by Hel2/Rqt1 and associated with ribosomes independent of the ubiquitination of uS10. However, the ubiquitin-binding activity of Rqt3 were crucial to trigger RQC. Cryo-electron microscopy (cryo-EM) analysis revealed that Hel2 bound ribosomes are in an rotated state containing hybrid state AP- and PE-tRNAs. Furthermore, ribosome profiling revealed that short footprints, hallmarks of hybrid state ribosomes18, were accumulated at tandem CGA rare codons at the beginning of the poly arginine stalling sequence and long footprints at subsequent codons, respectively. Short footprints at CGA codons were decreased in rqt1 mutant but drastically increased in uS10 mutants defective in the ubiquitination or rqt2 mutant. Collectively, our results demonstrate that Hel2 stabilizes ratcheted ribosomes leading to ubiquitination of uS10. Subsequently, Rqt2-4 factors target these hybrid state ribosomes specifically, allowing subsequent RQC reactions.
2017-07-31 | GSE79622 | GEO
Project description:Ribosome stalling occurring on aberrant mRNA activates quality control pathways to maintain proteostasis. Recently, ribosome stalling has also been linked to the activation of Gcn2 and the subsequent integrated-stress response (ISR). How the two processes are coordinated is not completely clear. Here we show that activation of ribosome-quality control by Hel2 suppresses that of Gcn2 in yeast. In the absence of Hel2, we observe a gene-expression signature indicative of ISR activation, suggesting that factor is used to suppress premature activation of Gcn2 in the absence of stress conditions. We further show that Hel2 and Gcn2 are activated by similar set of agents that cause ribosome stalling, with Hel2’s maximal activation occurring at lower frequency of stalling. Interestingly, inactivation of one pathway was found to result in the overactivation of the other, suggesting that both are activated by the same signal. Indeed, we provide evidence that suggests that, similar to Hel2, Gcn2 is activated by ribosome collisions. Collectively, our findings provide interesting details about how the multiple pathways that recognize stalled ribosomes coordinate to mount the appropriate response.
2020-05-19 | GSE150790 | GEO