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. Epub 2020 Oct 19.

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.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Animals
  • Humans
  • Protein Interaction Maps
  • RNA / metabolism*
  • RNA, Long Noncoding / metabolism
  • RNA-Binding Proteins / metabolism*
  • Reproducibility of Results
  • Ribonuclease P / metabolism
  • Ribonucleoproteins / metabolism*

Substances

  • RNA, Long Noncoding
  • RNA-Binding Proteins
  • Ribonucleoproteins
  • XIST non-coding RNA
  • RNA
  • Ribonuclease P