title: The autoinhibitory CARD2-Hel2i Interface of RIG-I governs RNA selection.
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.
authors.affiliation: Robert Wood Johnson Medical School, Department of Biochemistry and Molecular Biology, Rutgers University, Piscataway, NJ 08854, USA.
authors.forename: Anand
authors.initials: A
authors.lastname: Ramanathan
authors.affiliation: Robert Wood Johnson Medical School, Department of Biochemistry and Molecular Biology, Rutgers University, Piscataway, NJ 08854, USA.
authors.forename: Swapnil C
authors.initials: SC
authors.lastname: Devarkar
authors.affiliation: Center for Advanced Biotechnology and Medicine, Department of Chemistry and Chemical Biology, Rutgers University, Piscataway, NJ 08854, USA.
authors.forename: Fuguo
authors.initials: F
authors.lastname: Jiang
authors.affiliation: Center for Advanced Biotechnology and Medicine, Department of Chemistry and Chemical Biology, Rutgers University, Piscataway, NJ 08854, USA.
authors.forename: Matthew T
authors.initials: MT
authors.lastname: Miller
authors.affiliation: Center for Advanced Biotechnology and Medicine, Department of Chemistry and Chemical Biology, Rutgers University, Piscataway, NJ 08854, USA.
authors.forename: Abdul G
authors.initials: AG
authors.lastname: Khan
authors.affiliation: Center for Advanced Biotechnology and Medicine, Department of Chemistry and Chemical Biology, Rutgers University, Piscataway, NJ 08854, USA jmarco@cabm-new.rutgers.edu.
authors.forename: Joseph
authors.initials: J
authors.lastname: Marcotrigiano
authors.affiliation: Robert Wood Johnson Medical School, Department of Biochemistry and Molecular Biology, Rutgers University, Piscataway, NJ 08854, USA patelss@rutgers.edu.
authors.forename: Smita S
authors.initials: SS
authors.lastname: Patel
mesh_terms: D000251:Adenosine Triphosphatases / Q000378:metabolism; D001483:Base Sequence; D001665:Binding Sites; D000071457:DEAD Box Protein 58; D053487:DEAD-box RNA Helicases / Q000235:genetics / Q000378:metabolism*; D005454:Fluorescence Polarization; D057809:HEK293 Cells; D006801:Humans; D016899:Interferon-beta / Q000235:genetics / Q000378:metabolism; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary; D012313:RNA / Q000737:chemistry / Q000378:metabolism*; D012330:RNA, Double-Stranded / Q000378:metabolism; D011971:Receptors, Immunologic
chemical_list: D012330:RNA, Double-Stranded; D011971:Receptors, Immunologic; D012313:RNA; D016899:Interferon-beta; D000251:Adenosine Triphosphatases; C452820:RIGI protein, human; D000071457:DEAD Box Protein 58; D053487:DEAD-box RNA Helicases