Nucleotide sequences and modifications that determine RIG-I/RNA binding and signaling activities

• Main Authors: Urzi, Dina (Contributor), Gehrke, Lee (Contributor)
• Other Authors: Harvard University- (Contributor)
• Format: Article
• Language: English
• Published: American Society for Microbiology, 2012-03-28T19:20:47Z.
• Subjects: Article
• Online Access: Get fulltext

 Cytoplasmic viral RNAs with 5' triphosphates (5'ppp) are detected by the RNA helicase RIG-I, initiating downstream signaling and alpha/beta interferon (IFN-α/β) expression that establish an antiviral state. We demonstrate here that the hepatitis C virus (HCV) 3' untranslated region (UTR) RNA has greater activity as an immune stimulator than several flavivirus UTR RNAs. We confirmed that the HCV 3'-UTR poly(U/UC) region is the determinant for robust activation of RIG-I-mediated innate immune signaling and that its antisense sequence, poly(AG/A), is an equivalent RIG-I activator. The poly(U/UC) region of the fulminant HCV JFH-1 strain was a relatively weak activator, while the antisense JFH-1 strain poly(AG/A) RNA was very potent. Poly(U/UC) activity does not require primary nucleotide sequence adjacency to the 5'ppp, suggesting that RIG-I recognizes two independent RNA domains. Whereas poly(U) 50-nt or poly(A) 50-nt sequences were minimally active, inserting a single C or G nucleotide, respectively, into these RNAs increased IFN-β expression. Poly(U/UC) RNAs transcribed in vitro using modified uridine 2' fluoro or pseudouridine ribonucleotides lacked signaling activity while functioning as competitive inhibitors of RIG-I binding and IFN-β expression. Nucleotide base and ribose modifications that convert activator RNAs into competitive inhibitors of RIG-I signaling may be useful as modulators of RIG-I-mediated innate immune responses and as tools to dissect the RNA binding and conformational events associated with signaling.