Analysis of NRAS RNA G-quadruplex binding proteins reveals DDX3X as a novel interactor of cellular G-quadruplex containing transcripts

Nucleic Acids Res. 2018 Nov 30;46(21):11592-11604. doi: 10.1093/nar/gky861.

Abstract

RNA G-quadruplexes (rG4s) are secondary structures in mRNAs known to influence RNA post-transcriptional mechanisms thereby impacting neurodegenerative disease and cancer. A detailed knowledge of rG4-protein interactions is vital to understand rG4 function. Herein, we describe a systematic affinity proteomics approach that identified 80 high-confidence interactors that assemble on the rG4 located in the 5'-untranslated region (UTR) of the NRAS oncogene. Novel rG4 interactors included DDX3X, DDX5, DDX17, GRSF1 and NSUN5. The majority of identified proteins contained a glycine-arginine (GAR) domain and notably GAR-domain mutation in DDX3X and DDX17 abrogated rG4 binding. Identification of DDX3X targets by transcriptome-wide individual-nucleotide resolution UV-crosslinking and affinity enrichment (iCLAE) revealed a striking association with 5'-UTR rG4-containing transcripts which was reduced upon GAR-domain mutation. Our work highlights hitherto unrecognized features of rG4 structure-protein interactions that highlight new roles of rG4 structures in mRNA post-transcriptional control.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • 5' Untranslated Regions
  • Cytoplasm / genetics
  • Cytoplasm / metabolism
  • DEAD-box RNA Helicases / genetics
  • DEAD-box RNA Helicases / metabolism*
  • G-Quadruplexes*
  • Genes, ras / genetics*
  • HeLa Cells
  • Humans
  • Protein Domains
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • Reproducibility of Results

Substances

  • 5' Untranslated Regions
  • RNA, Messenger
  • DDX3X protein, human
  • DEAD-box RNA Helicases