Regulation of DHX16-mediated Innate Immunity by Unanchored Polyubiquitin
The innate immune system recognizes pathogens via unique signals present in the invading organism upon infection known as pathogen-associated molecular patterns (PAMPs). Recognition of PAMPs during viral infection requires host molecules capable of interpreting these foreign signals called pattern recognition receptors (PRRs). These PRRs trigger the production of type-I interferon (IFN-I), which is essential to establish antiviral innate immunity, allowing for the clearance of the offending organism. This process is tightly controlled by post-translational modifications (PTMs), like unanchored (or free) polyubiquitin (poly-Ub), which properly regulate IFN-I to elicit responses as needed. Despite its known roles regulating critical IFN-I pathways, the extent to which unanchored poly-Ub is involved in innate immunity is unknown. To identify whether unanchored poly-Ub regulates immune responses through unique systems, host factors interacting with unanchored poly-Ub were isolated from lung tissue utilizing a novel affinity purification method. The DEAH-box RNA helicase DHX16, a pre-mRNA splicing factor, was identified as a new unanchored poly-Ub-interacting factor. Silencing of DHX16 in cells and in vivo diminished IFN-I responses against viruses from multiple families including influenza A (IAV), Zika, and SARS-CoV-2. Recognition of viruses by DHX16 is achieved through the sensing of signals present in viral RNA (vRNA) like the splicing motifs present in IAV vRNA segments. DHX16 requires its RNA helicase motif for direct, high-affinity interactions with specific vRNAs which trigger an association with unanchored K48-poly-Ub synthesized by the E3-Ub ligase TRIM6. These unanchored Ub chains promote the recruitment of RIG-I, which ultimately initiates the downstream production of IFN-I. This study establishes DHX16 as a PRR that partners with RIG-I for optimal activation of antiviral responses and reveals unanchored poly-Ub as having a broad role in the regulation of innate immune pathways through the recruitment of novel host factors.