The autoinhibitory CARD2-Hel2i Interface of RIG-I governs RNA selection

Nucleic Acids Res. 2016 Jan 29;44(2):896-909. doi: 10.1093/nar/gkv1299. Epub 2015 Nov 26.

Abstract

RIG-I (Retinoic Acid Inducible Gene-I) is a cytosolic innate immune receptor that detects atypical features in viral RNAs as foreign to initiate a Type I interferon signaling response. RIG-I is present in an autoinhibited state in the cytoplasm and activated by blunt-ended double-stranded (ds)RNAs carrying a 5' triphosphate (ppp) moiety. These features found in many pathogenic RNAs are absent in cellular RNAs due to post-transcriptional modifications of RNA ends. Although RIG-I is structurally well characterized, the mechanistic basis for RIG-I's remarkable ability to discriminate between cellular and pathogenic RNAs is not completely understood. We show that RIG-I's selectivity for blunt-ended 5'-ppp dsRNAs is ≈3000 times higher than non-blunt ended dsRNAs commonly found in cellular RNAs. Discrimination occurs at multiple stages and signaling RNAs have high affinity and ATPase turnover rate and thus a high katpase/Kd. We show that RIG-I uses its autoinhibitory CARD2-Hel2i (second CARD-helicase insertion domain) interface as a barrier to select against non-blunt ended dsRNAs. Accordingly, deletion of CARDs or point mutations in the CARD2-Hel2i interface decreases the selectivity from ≈3000 to 150 and 750, respectively. We propose that the CARD2-Hel2i interface is a 'gate' that prevents cellular RNAs from generating productive complexes that can signal.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Adenosine Triphosphatases / metabolism
  • Base Sequence
  • Binding Sites
  • DEAD Box Protein 58
  • DEAD-box RNA Helicases / genetics
  • DEAD-box RNA Helicases / metabolism*
  • Fluorescence Polarization
  • HEK293 Cells
  • Humans
  • Interferon-beta / genetics
  • Interferon-beta / metabolism
  • Molecular Sequence Data
  • Protein Structure, Tertiary
  • RNA / chemistry
  • RNA / metabolism*
  • RNA, Double-Stranded / metabolism

Substances

  • RNA, Double-Stranded
  • RNA
  • Interferon-beta
  • Adenosine Triphosphatases
  • DDX58 protein, human
  • DEAD Box Protein 58
  • DEAD-box RNA Helicases