Large-scale tethered function assays identify factors that regulate mRNA stability and translation

Luo, En‐Ching; Nathanson, Jason L.; Tan, Frederick E.; Schwartz, Joshua L.; Schmok, Jonathan C.; Shankar, Archana; Markmiller, Sebastian; Yee, Brian A.; Sathe, Shashank; Pratt, Gabriel A.; Scaletta, Duy B.; Ha, Yuanchi; Hill, David E.; Aigner, Stefan; Yeo, G

doi:10.1038/s41594-020-0477-6

Cited by 80 publications

(93 citation statements)

References 85 publications

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“…In line with this, several researches had implied that MTDH acted as an RNA-binding protein. 36 , 37 , 38 Also illustrated was the decreased mRNAs that could bind to MTDH including a large amount of E2 and E3 ubiquitin ligases such as FBXW2 on a dual PI3K/mTOR inhibitor treatment. 36 This indicates that MTDH could accelerate the degradation of these mRNAs in response to drug stimuli.…”

Section: Resultsmentioning

confidence: 99%

TAK1 Is a Novel Target in Hepatocellular Carcinoma and Contributes to Sorafenib Resistance

Xia

Tao

et al. 2021

Cellular and Molecular Gastroenterology and Hepatology

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Section: Resultsmentioning

confidence: 99%

TAK1 Is a Novel Target in Hepatocellular Carcinoma and Contributes to Sorafenib Resistance

Xia

Tao

et al. 2021

Cellular and Molecular Gastroenterology and Hepatology

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“…Mechanistic insight into the quantitative effects of RBP expression on mRNA translation - i.e., when translation rates of the endogenous RBP targets respond directly to changes in RBP abundance - was recently provided by several studies (Chothani et al, 2019; Luo et al, 2020; Sharma et al, 2021). These studies used this quantitative relationship to assign novel functions to known RBPs and to investigate the kinetics through which RBPs regulate their targets.…”

Section: Introductionmentioning

confidence: 99%

Multifunctional RNA-binding proteins influence mRNA abundance and translational efficiency of distinct sets of target genes

Schneider-Lunitz

Ruiz-Orera

Hübner

et al. 2021

Preprint

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RNA-binding proteins (RBPs) are key regulators of RNA metabolism. Many RBPs possess uncharacterized RNA-binding domains and localize to multiple subcellular compartments, suggesting their involvement in multiple biological processes. We searched for such multifunctionality within a set of 143 RBPs by integrating experimentally validated target genes with the transcriptomes and translatomes of 80 human hearts. This revealed that RBP abundance is predictive of the extent of target regulation in vivo, leading us to newly associate 27 RBPs with translational control. Amongst those were several splicing factors, of which the muscle specific RBM20 modulated target translation rates through switches in isoform production. For 21 RBPs, we newly observed dual regulatory effects impacting both mRNA levels and translation rates, albeit for virtually independent sets of target genes. We highlight a subset, including G3BP1, PUM1, UCHL5, and DDX3X, where dual regulation is achieved by differential affinity for targets of distinct length and functionality. Strikingly, in a manner very similar to DDX3X, the known splicing factors EFTUD2 and PRPF8 selectively influence target translation rates depending on 5' UTR structure. Our results indicate unanticipated complexity of protein-RNA interactions at consecutive stages of gene expression and implicate multiple core splicing factors as key regulators of translational output.

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“…Indeed, tethering has frequently been used in the literature to study a wide variety of RNA binding proteins in diverse biological settings. For example, tethering has been used to investigate nonsense-mediated decay (Hickey et al, 2020), to screen for effects caused by RNA binding proteins (Luo et al, 2020), and to study specific protein domains (Fabian et al, 2011; Liu, Valencia-Sanchez, et al, 2005; Piao et al, 2010) or specific amino acid modifications (Golden et al, 2017). With the TnT biosensor, these studies can be expanded to the single-molecule level, where their impact can be more thoroughly investigated with higher spatiotemporal resolution.…”

Section: Discussionmentioning

confidence: 99%

Imaging translational control by Argonaute with single-molecule resolution in live cells

Cialek

Morisaki

Zhao

et al. 2021

Preprint

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A major challenge to our understanding of translational control has been deconvolving the individual impact specific regulatory factors have on the complex dynamics of mRNA translation. MicroRNAs (miRNAs), for example, guide Argonaute and associated proteins to target mRNAs, where they direct gene silencing in multiple ways that are not well understood. To better deconvolve these dynamics, we have developed technology to directly visualize and quantify the impact of human Argonaute2 (Ago2) on the translation and subcellular localization of individual reporter mRNAs in living cells. We show that our combined translation and Ago2 tethering system reflects endogenous miRNA-mediated gene silencing. Using the system, we find that Ago2 association leads to progressive silencing of translation at individual mRNA. The timescale of silencing was similar to that of translation, consistent with a role for Ago2 in blocking translation initiation, leading to ribosome runoff. At early time points, we observed occasional brief bursts of translational activity at Ago2-tethered mRNAs undergoing silencing, suggesting that translational repression may initially be reversible. At later time points, Ago2-tethered mRNAs cluster and coalesce with endogenous P-bodies, where a translationally silent state is maintained. These results provide a framework for exploring miRNA-mediated gene regulation in live cells at the single-molecule level. Furthermore, our tethering-based, single-molecule reporter system will likely have wide-ranging application in studying general RNA-protein interactions.

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Large-scale tethered function assays identify factors that regulate mRNA stability and translation

Cited by 80 publications

References 85 publications

TAK1 Is a Novel Target in Hepatocellular Carcinoma and Contributes to Sorafenib Resistance

TAK1 Is a Novel Target in Hepatocellular Carcinoma and Contributes to Sorafenib Resistance

Multifunctional RNA-binding proteins influence mRNA abundance and translational efficiency of distinct sets of target genes

Imaging translational control by Argonaute with single-molecule resolution in live cells

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