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Photoactivatable ribonucleosides mark base-specific RNA-binding sites

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Title
Photoactivatable ribonucleosides mark base-specific RNA-binding sites
Author(s)
Jong Woo Bae; Kim, Sangtae; V. Narry Kim; Jong-Seo Kim
Publication Date
2021-12-01
Journal
NATURE COMMUNICATIONS, v.12, no.1
Publisher
NATURE PORTFOLIO
Abstract
RNA-protein interactions play critical roles in post-transcriptional gene regulation. Here the authors demonstrate pRBS-ID, an updated MS/MS-based method that combines the benefits of photoactivatable ribonucleosides and the chemical cleavage of RNA. RNA-protein interaction can be captured by crosslinking and enrichment followed by tandem mass spectrometry, but it remains challenging to pinpoint RNA-binding sites (RBSs) or provide direct evidence for RNA-binding. To overcome these limitations, we here developed pRBS-ID, by incorporating the benefits of UVA-based photoactivatable ribonucleoside (PAR; 4-thiouridine and 6-thioguanosine) crosslinking and chemical RNA cleavage. pRBS-ID robustly detects peptides crosslinked to PAR adducts, offering direct RNA-binding evidence and identifying RBSs at single amino acid-resolution with base-specificity (U or G). Using pRBS-ID, we could profile uridine-contacting RBSs globally and discover guanosine-contacting RBSs, which allowed us to characterize the base-specific interactions. We also applied the search pipeline to analyze the datasets from UVC-based RBS-ID experiments, altogether offering a comprehensive list of human RBSs with high coverage (3,077 RBSs in 532 proteins in total). pRBS-ID is a widely applicable platform to investigate the molecular basis of posttranscriptional regulation.
URI
https://pr.ibs.re.kr/handle/8788114/10548
DOI
10.1038/s41467-021-26317-5
ISSN
2041-1723
Appears in Collections:
Center for RNA Research(RNA 연구단) > 1. Journal Papers (저널논문)
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