Analysis of RNA-protein networks with RNP-MaP defines functional hubs on RNA

  • Nat Biotechnol. 2021 Mar;39(3):347-356. doi: 10.1038/s41587-020-0709-7.
Chase A Weidmann  1  2 ,  Anthony M Mustoe  1 ,  Parth B Jariwala  1 ,  J Mauro Calabrese  2  3 ,  Kevin M Weeks  4
Affiliations
  • 1. Department of Chemistry, University of North Carolina, Chapel Hill, NC, USA.
  • 2. Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, NC, USA.
  • 3. Department of Pharmacology, University of North Carolina, Chapel Hill, NC, USA.
  • 4. Department of Chemistry, University of North Carolina, Chapel Hill, NC, USA. [email protected].
PMID: 33077962 DOI: 10.1038/s41587-020-0709-7
Abstract

RNA-protein interaction networks govern many biological processes but are difficult to examine comprehensively. We devised ribonucleoprotein networks analyzed by mutational profiling (RNP-MaP), a live-cell chemical probing strategy that maps cooperative interactions among multiple proteins bound to single RNA molecules at nucleotide resolution. RNP-MaP uses a hetero-bifunctional crosslinker to freeze interacting proteins in place on RNA and then maps multiple bound proteins on single RNA strands by read-through reverse transcription and DNA Sequencing. RNP-MaP revealed that RNase P and RMRP, two sequence-divergent but structurally related non-coding RNAs, share RNP networks and that network hubs define functional sites in these RNAs. RNP-MaP also identified protein interaction networks conserved between mouse and human XIST long non-coding RNAs and defined protein communities whose binding sites colocalize and form networks in functional regions of XIST. RNP-MaP enables discovery and efficient validation of functional protein interaction networks on long RNAs in living cells.

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