Incoming RNA Virus Nucleocapsids Containing a 5′-Triphosphorylated Genome Activate RIG-I and Antiviral Signaling

Abstract

Summary Host defense to RNA viruses depends on rapid intracellular recognition of viral RNA by two cytoplasmic RNA helicases, RIG-I and MDA5. RNA transfection experiments indicate that RIG-I responds to naked double-stranded (ds) RNAs with a triphosphorylated 5′ (5′ppp) terminus. However, identity of the RIG-I stimulating viral structures in an authentic infection context remains unresolved. We show that incoming viral nucleocapsids containing a 5′ppp dsRNA “panhandle” structure trigger antiviral signaling that commences with RIG-I, is mediated through the adaptor protein MAVS, and terminates with transcription factor IRF-3. Independent of mammalian cofactors or viral polymerase activity, RIG-I bound to viral nucleocapsids, underwent a conformational switch, and homo-oligomerized. Enzymatic probing and super-resolution microscopy suggest that RIG-I interacts with the panhandle structure of the viral nucleocapsids. These results define cytoplasmic entry of nucleocapsids as the proximal RIG-I-sensitive step during infection, and establish viral nucleocapsids with a 5′ppp dsRNA panhandle as a RIG-I activator.