Homo sapiens
Ontology highlight
ABSTRACT:
PROVIDER: PRJDB11308 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| DRR277015.fastq.gz | Fastqsanger.gz | |||
| DRR277016.fastq.gz | Fastqsanger.gz | |||
| DRR277017.fastq.gz | Fastqsanger.gz | |||
| DRR277018.fastq.gz | Fastqsanger.gz | |||
| DRR277019.fastq.gz | Fastqsanger.gz |
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Project description:Combinatorial analyis of translation dynamics reveals eIF2 dependence of translation initiation at near-cognate codons
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Project description:Although ribosome-profiling and translation initiation sequencing (TI-seq) analyses have identified many noncanonical initiation codons, the precise detection of translation initiation sites (TISs) remains a challenge, mainly because of experimental artifacts of such analyses. Here we describe a new method, TISCA (TIS detection by translation Complex Analysis), for the accurate identification of TISs. TISCA proved to be more reliable for TIS detection compared with existing tools, and it identified a substantial number of near-cognate codons in Kozak-like sequence contexts. Analysis of proteomics data revealed the presence of methionine at the NH2-terminus of most proteins derived from near-cognate initiation codons. Although eukaryotic initiation factor 2 (eIF2), eIF2A, and eIF2D have previously been shown to contribute to translation initiation at near-cognate codons, we found that most noncanonical initiation events are dependent on eIF2, consistent with the initial amino acid being methionine. Comprehensive identification of TISs by TISCA should facilitate characterization of the mechanism of noncanonical initiation.
2021-07-01 | GSE174329 | GEO
Project description:So far, the annotation of translation initiation sites (TISs) has been based mostly upon bioinformatics rather than experimental evidence. We adapted ribosomal footprinting to puromycin-treated cells to generate a transcriptome-wide map of TISs in a human monocytic cell line. A neural network was trained on the ribosomal footprints at previously annotated AUG translation initiation codons (TICs), and used for the ab initio prediction of TISs in 5062 transcripts with sufficient sequence coverage. Functional interpretation suggested 2994 novel upstream open reading frames (uORFs) in the 5´ UTR (924 AUG, 2070 near-cognate codons), 1406 uORFs overlapping with the coding sequence (116 AUG, 1290 near-cognate) and 546 N-terminal protein extensions (6 AUG, 540 near-cognate). The TIS detection method was validated on the basis of previously published alternative TISs and uORFs. On average, TICs in newly annotated TISs were significantly more conserved among primates than control codons, both for AUGs (p<10-10) and near-cognate codons (p=3.8×10-3). The derived transcriptome-wide map of novel candidate TISs will help to explain how human proteome diversity is influenced by alternative translation initiation and regulation. Examination of translational initiation in human cell lines using ribosomal footprinting
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Project description:The conserved and essential DEAD-box RNA helicase Ded1p from yeast and its mammalian ortholog DDX3 are critical for translation initiation. Mutations in DDX3 are linked to tumorigenesis and intellectual disability, and the enzyme is targeted by diverse viruses. How Ded1p and its orthologs engage RNAs to impact translation initiation has been a longstanding, unresolved question. Here we show that Ded1p associates with the pre-initiation complex at the mRNA entry channel of the small ribosomal subunit and that the helicase unwinds mRNA structure ahead of the scanning pre-initiation complex. Defective Ded1p causes pervasive translation in 5’UTRs, starting from near-cognate initiation codons located 5' of mRNA structures and concomitant decrease of protein synthesis from of the main ORFs. The data indicate that Ded1p functions to suppress translation initiation on near-cognate codons proximal to mRNA structure and show how the helicase is targeted to specific RNA sites without common sequence signatures. Our results reveal a straightforward mechanism for the activation of upstream open reading frames and suggest that mRNA structure and proximal near-cognate initiation codons encode a widespread regulatory program for translation initiation that is sensitive to RNA helicase function.
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Project description:So far, the annotation of translation initiation sites (TISs) has been based mostly upon bioinformatics rather than experimental evidence. We adapted ribosomal footprinting to puromycin-treated cells to generate a transcriptome-wide map of TISs in a human monocytic cell line. A neural network was trained on the ribosomal footprints at previously annotated AUG translation initiation codons (TICs), and used for the ab initio prediction of TISs in 5062 transcripts with sufficient sequence coverage. Functional interpretation suggested 2994 novel upstream open reading frames (uORFs) in the 5´ UTR (924 AUG, 2070 near-cognate codons), 1406 uORFs overlapping with the coding sequence (116 AUG, 1290 near-cognate) and 546 N-terminal protein extensions (6 AUG, 540 near-cognate). The TIS detection method was validated on the basis of previously published alternative TISs and uORFs. On average, TICs in newly annotated TISs were significantly more conserved among primates than control codons, both for AUGs (p<10-10) and near-cognate codons (p=3.8×10-3). The derived transcriptome-wide map of novel candidate TISs will help to explain how human proteome diversity is influenced by alternative translation initiation and regulation.
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Project description:The RNA helicase Ded1p suppresses translation initiation from near-cognate start codons
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Project description:The Ribo-seq analysis demonstrated that eIF1A is predominantly essential for translation of genes with long 5'UTR genes including cell proliferation and cell cycle progression genes. eIF1A depletion causes broad stimulation of initiation in 5’UTRs at near-cognate AUG codons that diminshes the translation initiation fidelity
2018-07-13 | GSE116982 | GEO
Project description:The Ribo-seq analysis demonstrated that eIF1A is predominantly essential for translation of genes with long 5'UTR genes including cell proliferation and cell cycle progression genes. eIF1A depletion causes broad stimulation of initiation in 5’UTRs at near-cognate AUG codons that diminshes the translation initiation fidelity
2018-07-13 | GSE116981 | GEO
Project description:Translation termination is an essential cellular process that is also of therapeutic interest for diseases that manifest from premature stop codons. In eukaryotes, translation termination requires eRF1, which recognizes stop codons, catalyzes the release of nascent proteins fom ribosomes, and facilitates ribosome recycling. The small molecule SRI-41315 triggers eRF1 degradation and enhances translational readthrough of premature stop codons. SRI-41315 promotes retention of eRF1 on ribosomes and leads to a higher frequency of translation termination at near-cognate stop codons. In this study, the systematic effect of SRI-41315 on translation termination at cognate and near-cognate stop codons is evaluated using a rabbit reticulocyte lysate-based in vitro translation system with endogenous transcript as well as reporter transcripts containing defined near-cognate stop codon.
2023-12-31 | GSE248946 | GEO
Project description:The translation pre-initiation complex (PIC) scans the mRNA for an AUG codon in favorable context. Previous findings suggest that the factor eIF1 discriminates against non-AUG start codons by impeding full accommodation of Met-tRNAi in the P site of the 40S ribosomal subunit, necessitating eIF1 dissociation for start codon selection. Consistent with this, yeast eIF1 substitutions that weaken its binding to the PIC increase initiation at UUG codons on a mutant his4 mRNA and particular synthetic mRNA reporters; and also at the AUG start codon of the mRNA for eIF1 itself owing to its poor Kozak context. It was not known however whether such eIF1 mutants increase initiation at suboptimal start codons genome-wide. By ribosome profiling, we show that the eIF1-L96P variant confers increased translation of numerous upstream open reading frames (uORFs) initiating with either near-cognate codons (NCCs) or AUGs in poor context. The increased uORF translation is frequently associated with reduced translation of the downstream main coding sequences (CDS). Initiation is also elevated at the NCCs initiating N-terminal extensions on GRS1 and ALA1 mRNAs, and at a small set of main CDS AUG codons with especially poor context, including that of eIF1 itself. Thus, eIF1 acts throughout the yeast translatome to discriminate against NCC start codons and AUGs in poor context; and impairing this function enhances the repressive effects of uORFs on CDS translation and alters the ratios of protein isoforms translated from near-cognate versus AUG start codons.
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