HDX-MS reveals dysregulated checkpoints that compromise discrimination against self RNA during RIG-I mediated autoimmunity

Nat Commun. 2018 Dec 18;9(1):5366. doi: 10.1038/s41467-018-07780-z.

Abstract

Retinoic acid inducible gene-I (RIG-I) ensures immune surveillance of viral RNAs bearing a 5'-triphosphate (5'ppp) moiety. Mutations in RIG-I (C268F and E373A) lead to impaired ATPase activity, thereby driving hyperactive signaling associated with autoimmune diseases. Here we report, using hydrogen/deuterium exchange, mechanistic models for dysregulated RIG-I proofreading that ultimately result in the improper recognition of cellular RNAs bearing 7-methylguanosine and N1-2'-O-methylation (Cap1) on the 5' end. Cap1-RNA compromises its ability to stabilize RIG-I helicase and blunts caspase activation and recruitment domains (CARD) partial opening by threefold. RIG-I H830A mutation restores Cap1-helicase engagement as well as CARDs partial opening event to a level comparable to that of 5'ppp. However, E373A RIG-I locks the receptor in an ATP-bound state, resulting in enhanced Cap1-helicase engagement and a sequential CARDs stimulation. C268F mutation renders a more tethered ring architecture and results in constitutive CARDs signaling in an ATP-independent manner.

Publication types

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

MeSH terms

  • Adenosine Triphosphatases / metabolism
  • Autoimmunity / genetics*
  • Caspase Activation and Recruitment Domain / immunology
  • DEAD Box Protein 58 / chemistry
  • DEAD Box Protein 58 / genetics*
  • DEAD Box Protein 58 / immunology
  • DEAD Box Protein 58 / metabolism
  • Deuterium Exchange Measurement / methods
  • Gain of Function Mutation
  • Guanosine / analogs & derivatives
  • Guanosine / chemistry
  • Guanosine / immunology
  • Guanosine / metabolism
  • Immunity, Innate / genetics*
  • Interferon-Induced Helicase, IFIH1 / immunology
  • Interferon-Induced Helicase, IFIH1 / metabolism
  • Mass Spectrometry / methods
  • Methylation
  • Models, Molecular
  • Mutagenesis, Site-Directed
  • Protein Binding / genetics
  • Protein Binding / immunology
  • RNA Caps / chemistry
  • RNA Caps / immunology*
  • RNA Caps / metabolism
  • RNA, Double-Stranded / chemistry
  • RNA, Double-Stranded / immunology*
  • RNA, Double-Stranded / metabolism
  • RNA, Viral / immunology
  • Recombinant Proteins / chemistry
  • Recombinant Proteins / genetics
  • Recombinant Proteins / immunology
  • Recombinant Proteins / metabolism
  • Signal Transduction / genetics
  • Signal Transduction / immunology

Substances

  • RNA Caps
  • RNA, Double-Stranded
  • RNA, Viral
  • Recombinant Proteins
  • Guanosine
  • 7-methylguanosine
  • Adenosine Triphosphatases
  • DDX58 protein, human
  • IFIH1 protein, human
  • DEAD Box Protein 58
  • Interferon-Induced Helicase, IFIH1