Pervasive isoform‐specific translational regulation via alternative transcription start sites in mammals

Wang, Xi; Hou, Jingyi; Quedenau, Claudia; Chen, Wei

doi:10.15252/msb.20166941

Cited by 96 publications

(110 citation statements)

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“…2b and Supplementary Table 2). These results are consistent with previous evidence that alternative 5′-UTRs differ in ribosome association in vivo [24][25][26] . Importantly, DART establishes the direct contribution of 5′-UTR sequences to differences in translation initiation and eliminates confounding effects of transcript localization and co-occurrence of alternative 3′-UTR sequences.…”

Section: Functional Characterization Of Alternative 5'utrssupporting

confidence: 93%

“…Ribosome profiling measures ribosome occupancy per ORF 7 , but the same ORF frequently occurs in multiple transcripts with different regulatory regions [16][17][18][19][20][21][22] . Moreover, alternative 5′-UTRs can produce very large differences in translation activity as shown by reporter assays 23 and by 5′-UTR variant specific polysome profiling [24][25][26] . Thus, the mRNA-specific translational efficiency inferred from ribosome profiling data may not accurately reflect the actual translation activity of any mRNA isoform ( Fig.…”

Section: Functional Characterization Of Alternative 5'utrsmentioning

confidence: 99%

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Direct analysis of ribosome targeting illuminates thousand-fold regulation of translation initiation

Zinshteyn

et al. 2020

Preprint

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Translational control shapes the proteome in normal and pathophysiological conditions. Current high-throughput approaches reveal large differences in mRNA-specific translation activity but cannot identify the causative mRNA features. We developed direct analysis of ribosome targeting (DART) and used it to dissect regulatory elements within 5′ untranslated regions that confer thousand-fold differences in ribosome recruitment in biochemically accessible cell lysates. Using DART, we identified novel translational enhancers and silencers, determined a functional role for most alternative 5′ UTR isoforms expressed in yeast, and revealed a general mode of increased translation via direct binding to a core translation factor. DART enables systematic assessment of the translational regulatory potential of 5′ UTR variants, whether native or disease-associated, and will facilitate engineering of mRNAs for optimized protein production in various systems. Highlights• DART illuminates thousand-fold differences in 5′ UTR-specific translation activity • SNPs and alternative 5′ UTR isoforms affect ribosome recruitment significantly • Inhibitory effects of RNA structures are highly dependent on 5′ UTR context • 5′ UTR motifs bind initiation factors directly, broadly stimulating translation Vogel C, Marcotte EM. Insights into the regulation of protein abundance from proteomic and transcriptomic analyses. Nat Rev Genet.

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Section: Functional Characterization Of Alternative 5'utrssupporting

confidence: 93%

Section: Functional Characterization Of Alternative 5'utrsmentioning

confidence: 99%

Direct analysis of ribosome targeting illuminates thousand-fold regulation of translation initiation

Zinshteyn

et al. 2020

Preprint

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“…This analysis revealed that when considering overall transcript copy number, those transcripts with short 5=UTR mRNAs (up to 40 nt) are abundant in the transcriptome (free plus polysomeassociated transcripts) and the translatome (polysome-associated transcripts). We also analyzed a recently published CapSeq data set of polysome profile with data from each ribosomal fraction (19) and similarly found that short 5=UTR mRNAs constitute a significant fraction of the translatome (see Fig. S1 in the supplemental material).…”

Section: Resultsmentioning

confidence: 97%

Efficient and Accurate Translation Initiation Directed by TISU Involves RPS3 and RPS10e Binding and Differential Eukaryotic Initiation Factor 1A Regulation

Haimov

Sinvani

Martin

et al. 2017

Molecular and Cellular Biology

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Canonical translation initiation involves ribosomal scanning, but short 5= untranslated region (5=UTR) mRNAs are translated in a scanning-independent manner. The extent and mechanism of scanning-independent translation are not fully understood. Here we report that short 5=UTR mRNAs constitute a substantial fraction of the translatome. Short 5=UTR mRNAs are enriched with TISU (translation initiator of short 5=UTR), a 12-nucleotide element directing efficient scanning-independent translation. Comprehensive mutagenesis revealed that each AUG codon-flanking nucleotide of TISU contributes to translational strength, but only a few are important for accuracy. Using site-specific UV cross-linking of ribosomal complexes assembled on TISU mRNA, we demonstrate specific binding of TISU to ribosomal proteins at the E and A sites. We identified RPS3 as the major TISU binding protein in the 48S complex A site. Upon 80S complex formation, RPS3 interaction is weakened and switched to RPS10e (formerly called RPS10). We further demonstrate that TISU is particularly dependent on eukaryotic initiation factor 1A (eIF1A) which interacts with both RPS3 and RPS10e. Our findings suggest that the cap-recruited ribosome specifically binds the TISU nucleotides at the A and E sites in cooperation with eIF1A to promote scanning arrest.KEYWORDS translation initiation, TISU, RPS3, RPS10e, eIF1A, short 5=UTR, RPS10 C anonical translation initiation in eukaryotes starts with binding of eukaryotic initiation factor 4F (eIF4F) to the m 7 G-capped 5= end of the mRNA, which is then followed by the recruitment of the 43S preinitiation complex (PIC) consisting of the 40S small ribosomal subunit, initiator Met-tRNA, and a subset of initiation factors (eIF2, eIF1, eIF1A, eIF3, and eIF5). The 43S PIC then scans the mRNA 5= untranslated region (5=UTR) and inspects it for an AUG start codon. During scanning, the 43S PIC is in an "open" state, and following AUG recognition, the 48S initiation complex is switched to a "closed" conformation that arrests scanning and promotes joining of the 60S large subunit to create the 80S elongation-competent complex (reviewed in references 1, 2, 3, 4, and 5). Translation usually initiates at the first 5=-proximal AUG codon, but in certain cases, translation initiates at a downstream AUG (DS-AUG) codon, a phenomenon known as leaky scanning. The extent of leaky scanning depends on the 5=UTR length and on the AUG nucleotide context (6,7,8,9). For mRNA with a 5=UTR that is more than 30 nucleotides (nt) long, the optimal AUG context is the Kozak element, RCCAUGG, in which the most significant nucleotides (shown in boldface type) are the

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“…Predicting various steps of gene expression from sequence alone has long been a subject of study (Beer and Tavazoie 2004;Vogel et al 2010;Zur and Tuller 2013;Wang et al 2016). To this end, two distinct classes of models have been proposed: the biophysical models on the one hand and the machine learning models on the other hand (Zur and Tuller 2016).…”

Section: Discussionmentioning

confidence: 99%

Cis-regulatory elements explain most of the mRNA stability variation across genes in yeast

Cheng

Maier

Avsec

et al. 2017

RNA

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The stability of mRNA is one of the major determinants of gene expression. Although a wealth of sequence elements regulating mRNA stability has been described, their quantitative contributions to half-life are unknown. Here, we built a quantitative model for Saccharomyces cerevisiae based on functional mRNA sequence features that explains 59% of the half-life variation between genes and predicts half-life at a median relative error of 30%. The model revealed a new destabilizing 3 ′ ′ ′ ′ ′ UTR motif, ATATTC, which we functionally validated. Codon usage proves to be the major determinant of mRNA stability. Nonetheless, single-nucleotide variations have the largest effect when occurring on 3 ′ ′ ′ ′ ′ UTR motifs or upstream AUGs. Analyzing mRNA half-life data of 34 knockout strains showed that the effect of codon usage not only requires functional decapping and deadenylation, but also the 5 ′ ′ ′ ′ ′ -to-3 ′ ′ ′ ′ ′ exonuclease Xrn1, the nonsense-mediated decay genes, but not no-go decay. Altogether, this study quantitatively delineates the contributions of mRNA sequence features on stability in yeast, reveals their functional dependencies on degradation pathways, and allows accurate prediction of half-life from mRNA sequence.

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Pervasive isoform‐specific translational regulation via alternative transcription start sites in mammals

Cited by 96 publications

References 70 publications

Direct analysis of ribosome targeting illuminates thousand-fold regulation of translation initiation

Direct analysis of ribosome targeting illuminates thousand-fold regulation of translation initiation

Efficient and Accurate Translation Initiation Directed by TISU Involves RPS3 and RPS10e Binding and Differential Eukaryotic Initiation Factor 1A Regulation

Cis-regulatory elements explain most of the mRNA stability variation across genes in yeast

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