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Proteome-wide quantitative RNA-interactome capture identifies phosphorylation sites with regulatory potential in RBM20

Item Type:Article
Title:Proteome-wide quantitative RNA-interactome capture identifies phosphorylation sites with regulatory potential in RBM20
Creators Name:Vieira e Vieira, C.H. and Dauksaite, V. and Sporbert, A. and Gotthardt, M. and Selbach, M.
Abstract:Cellular mRNA-binding proteins (mRBPs) are major posttranscriptional regulators of gene expression. Although many posttranslational modification sites in mRBPs have been identified, little is known about how these modifications regulate mRBP function. Here, we developed quantitative RNA-interactome capture (qRIC) to quantify the fraction of mRBPs pulled down with polyadenylated mRNAs. Combining qRIC with phosphoproteomics allowed us to systematically compare pull-down efficiencies of phosphorylated and nonphosphorylated forms of mRBPs. Almost 200 phosphorylation events affected pull-down efficiency compared with the unmodified mRBPs and thus have regulatory potential. Our data capture known regulatory phosphorylation sites in ELAVL1, SF3B1, and UPF1 and identify potential regulatory sites. Follow-up experiments on the splicing regulator RBM20 revealed multiple phosphorylation sites in the C-terminal disordered region affecting nucleocytoplasmic localization, association with cytoplasmic ribonucleoprotein granules, and alternative splicing. Together, we show that qRIC in conjunction with phosphoproteomics is a scalable method to identify functional posttranslational modification sites in mRBPs.
Keywords:RNA-Binding Proteins, RNA-Interactome Capture, Cell Signaling, Phosphorylation, Proteomics, RBM20, Alternative Splicing, Cardiomyopathy
Source:Molecular Cell
Publisher:Cell Press
Page Range:2069-2083.e8
Date:2 June 2022
Official Publication:https://doi.org/10.1016/j.molcel.2022.03.024
PubMed:View item in PubMed
Related to:
https://edoc.mdc-berlin.de/20475/Preprint version

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