IFI16 directly senses viral RNA and enhances RIG-I transcription and activation to restrict influenza virus infection

The retinoic acid-inducible gene I (RIG-I) receptor senses cytoplasmic viral RNA and activates type I interferons (IFN-I) and downstream antiviral immune responses. How RIG-I binds to viral RNA and how its activation is regulated remains unclear. Here, using IFI16 knockout cells and p204-deficient mice, we demonstrate that the DNA sensor IFI16 enhances IFN-I production to inhibit influenza A virus (IAV) replication. IFI16 positively upregulates RIG-I transcription through direct binding to and recruitment of RNA polymerase II to the RIG-I promoter. IFI16 also binds to influenza viral RNA via its HINa domain and to RIG-I protein with its PYRIN domain, thus promoting IAV-induced K63-linked polyubiquitination and RIG-I activation. Our work demonstrates that IFI16 is a positive regulator of RIG-I signalling during influenza virus infection, highlighting its role in the RIG-I-like-receptor-mediated innate immune response to IAV and other RNA viruses, and suggesting its possible exploitation to modulate the antiviral response. IFI16 enhances the type I IFN response to inhibit influenza virus replication by two mechanisms: it directly binds viral RNA to promote RIG-I activation and upregulates RIG-I expression via recruiting RNA polymerase II and binding to the RIG-I promoter.