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PreprintCited by 36RNA Research and SplicingRNA and protein synthesis mechanismsBiotin and Related Studies

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TREX reveals proteins that bind to specific RNA regions in living cells

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Dodel, Martin, Guiducci, Giulia, Dermit, Maria, Krishnamurthy, Sneha, Stojic, Lovorka, Mardakheh, Faraz K.·2023·Nature Methods
DOI 10.1101/2023.06.30.547259PMC PMC10927567

Preprint — published as 10.1038/s41592-024-02181-1

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Abstract

Summary: Different regions of RNA molecules can often engage in specific interactions with distinct RNA-binding proteins (RBPs), giving rise to diverse modalities of RNA regulation and function. However, there are currently no methods for unbiased identification of RBPs that interact with specific RNA regions in living cells under endogenous settings. Here, we introduce TREX (Targeted RNase H-mediated extraction of crosslinked RBPs), a highly sensitive approach for identifying proteins that directly bind to specific RNA regions in living cells. We demonstrate that TREX outperforms existing methods in identifying known interactors of U1 snRNA, and reveals endogenous region-specific interactors of NORAD lncRNA. Using TREX, we generated a comprehensive region-by-region interactome for 45S rRNA, uncovering both established and novel interactions that regulate ribosome biogenesis. With its applicability to any RNA in any cell-type, TREX is the first RNA-centric tool for unbiased positional mapping of endogenous RNA-protein interactions in living cells.

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Metadata

Authors
Dodel, M., Guiducci, G., Dermit, M., Krishnamurthy, S., Stojic, L., & Mardakheh, F. K.
Affiliations
Queen Mary University of London (GB)
Corresponding author
  • Stojic, Lovorka
  • Mardakheh, Faraz K.
ORCID
Venue
Nature Methods
Publisher
openRxiv
Published
2023-06-30
Volume
21
Issue
3
Pages
423-434
ISSN
1548-7091 (ISSN-L)
DOI
10.1101/2023.06.30.547259
PMC
PMC10927567
OpenAlex
W4391941875
Cited by
36
References
1
Citations by year
2026: 13 · 2025: 15 · 2024: 7 · 2023: 1
FWCI
4.71
Type
journal article
Language
en
Open access
Licence
cc-by
Locations
  • Nature Methods (publishedVersion, cc-by)
  • bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • Queen Mary Research Online (Queen Mary University of London) (submittedVersion, cc-by)
Funders
  • Barts Charity (MGU0404, MGU0346)
  • Cancer Research UK (RCCFEL\100007)
  • Academy of Medical Sciences (SBF006\1026)
  • Associazione Italiana per la Ricerca sul Cancro
  • Medical Research Council (MR/P009417, MR/W001500/1, MGU0346, MR/P009417/1)
  • Biotechnology and Biological Sciences Research Council (BB/X007820/1)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA Research and Splicing
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA and protein synthesis mechanisms
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Cell Biology › Biotin and Related Studies
Keywords
TREX complex, RNA-binding proteins, RNA-protein interactions, crosslinking, U1 snRNA, 45S rRNA, ribosome biogenesis
MeSH terms
RNA, RNA-Binding Proteins

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