Molecular Cell
Volume 64, Issue 2, 20 October 2016, Pages 416-430
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High-Resolution Mapping of RNA-Binding Regions in the Nuclear Proteome of Embryonic Stem Cells

https://doi.org/10.1016/j.molcel.2016.09.034Get rights and content
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Highlights

  • RBR-ID identifies RNA-binding regions by 4SU photocrosslinking and mass spectrometry

  • RBRs were mapped in 803 nuclear RNA-binding proteins (RBPs) in embryonic stem cells

  • Many previously unknown RBPs regulate chromatin structure and transcription

  • RBRs were found in disordered regions and domains associated with chromatin function

Summary

Interactions between noncoding RNAs and chromatin proteins play important roles in gene regulation, but the molecular details of most of these interactions are unknown. Using protein-RNA photocrosslinking and mass spectrometry on embryonic stem cell nuclei, we identified and mapped, at peptide resolution, the RNA-binding regions in ∼800 known and previously unknown RNA-binding proteins, many of which are transcriptional regulators and chromatin modifiers. In addition to known RNA-binding motifs, we detected several protein domains previously unknown to function in RNA recognition, as well as non-annotated and/or disordered regions, suggesting that many functional protein-RNA contacts remain unexplored. We identified RNA-binding regions in several chromatin regulators, including TET2, and validated their ability to bind RNA. Thus, proteomic identification of RNA-binding regions (RBR-ID) is a powerful tool to map protein-RNA interactions and will allow rational design of mutants to dissect their function at a mechanistic level.

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