Retinoic-acid-inducible gene-I (RIG-I; also called DDX58) is a cytosolic viral RNA receptor that interacts with MAVS (also called VISA, IPS-1 or Cardif) to induce type I interferon-mediated host protective innate immunity against viral infection. Furthermore, members of the tripartite motif (TRIM) protein family, which contain a cluster of a RING-finger domain, a B box/coiled-coil domain and a SPRY domain, are involved in various cellular processes, including cell proliferation and antiviral activity. Here we report that the amino-terminal caspase recruitment domains (CARDs) of RIG-I undergo robust ubiquitination induced by TRIM25 in mammalian cells. The carboxy-terminal SPRY domain of TRIM25 interacts with the N-terminal CARDs of RIG-I; this interaction effectively delivers the Lys 63-linked ubiquitin moiety to the N-terminal CARDs of RIG-I, resulting in a marked increase in RIG-I downstream signalling activity. The Lys 172 residue of RIG-I is critical for efficient TRIM25-mediated ubiquitination and for MAVS binding, as well as the ability of RIG-I to induce antiviral signal transduction. Furthermore, gene targeting demonstrates that TRIM25 is essential not only for RIG-I ubiquitination but also for RIG-I-mediated interferon- production and antiviral activity in response to RNA virus infection. Thus, we demonstrate that TRIM25 E3 ubiquitin ligase induces the Lys 63-linked ubiquitination of RIG-I, which is crucial for the cytosolic RIG-I signalling pathway to elicit host antiviral innate immunity.

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SUMMARY TRIM25 mediates Lys 63-linked ubiquitination of the N-terminal CARDs of the viral RNA sensor RIG-I, leading to type I interferon (IFN) production. Here, we report that the influenza A virus non-structural protein 1 (NS1) specifically inhibits TRIM25-mediated RIG-I CARD ubiquitination, thereby suppressing RIG-I signal transduction. A novel domain in NS1 comprising E96/E97 residues mediates its interaction with the coiled-coil domain of TRIM25, thus blocking TRIM25 multimerization and RIG-I CARD ubiquitination. Furthermore, a recombinant influenza A virus expressing an E96A/E97A NS1 mutant is defective in blocking TRIM25-mediated anti-viral IFN response and loses virulence in mice. Our findings reveal a novel mechanism of influenza virus to inhibit host IFN response and also emphasize the vital role of TRIM25 in modulating viral infections.

| Retinoic acid-inducible gene I (RIG-I)-like receptors (RLRs) are key sensors of virus infection, mediating the transcriptional induction of type I interferons and other genes that collectively establish an antiviral host response. Recent studies have revealed that both viral and host-derived RNAs can trigger RLR activation; this can lead to an effective antiviral response but also immunopathology if RLR activities are uncontrolled. In this Review , we discuss recent advances in our understanding of the types of RNA sensed by RLRs in the contexts of viral infection, malignancies and autoimmune diseases. We further describe how the activity of RLRs is controlled by host regulatory mechanisms, including RLR-interacting proteins, post-translational modifications and non-coding RNAs. Finally , we discuss key outstanding questions in the RLR field, including how our knowledge of RLR biology could be translated into new therapeutics.

Autophagic and endocytic pathways are tightly regulated membrane rearrangement processes that are crucial for homeostasis, development and disease. Autophagic cargo is delivered from autophagosomes to lysosomes for degradation through a complex process that topologically resembles endosomal maturation. Here, we report that a Beclin1-binding autophagic tumour suppressor, UVRAG, interacts with the class C Vps complex, a key component of the endosomal fusion machinery. This interaction stimulates Rab7 GTPase activity and autophagosome fusion with late endosomes/lysosomes, thereby enhancing delivery and degradation of autophagic cargo. Furthermore, the UVRAG-class-C-Vps complex accelerates endosome-endosome fusion, resulting in rapid degradation of endocytic cargo. Remarkably, autophagosome/endosome maturation mediated by the UVRAG-class-C-Vps complex is genetically separable from UVRAG-Beclin1-mediated autophagosome formation. This result indicates that UVRAG functions as a multivalent trafficking effector that regulates not only two important steps of autophagy -autophagosome formation and maturation -but also endosomal fusion, which concomitantly promotes transport of autophagic and endocytic cargo to the degradative compartments.© 2008 Macmillan Publishers Limited. All rights reserved. 6 Correspondence should be addressed to J.U.J. (jaeujung@usc.edu). AUTHOR CONTRIBUTIONS C.L. performed all aspects of this study; L.S., K.I., M.G., Q.L. and P.F. assisted with the experimental design and in collecting the data; E.R., I.V. and V.D. assisted with the autophagic protein degradation and in vitro endosome fusion assay; C.A. provided Vps constructs and their antibodies; C.L. and J.J. organized this study and wrote the paper. All authors discussed the results and commented on the manuscript.Note: Supplementary Information is available on the Nature Cell Biology website. COMPETING FINANCIAL INTERESTSThe authors declare no competing financial interests.Reprints and permissions information is available online at http://npg.nature.com/reprintsandpermissions/ NIH Public Access Author ManuscriptNat Cell Biol. Author manuscript; available in PMC 2010 May 31. Published in final edited form as:Nat Cell Biol. 2008 July ; 10(7): 776-787. doi:10.1038/ncb1740. NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author ManuscriptAutophagy is a tightly regulated membrane rearrangement process that ensures lysosomedependent bulk degradation of cytosolic proteins or organelles, and is highly conserved in eukaryotic cells, as seen with the endocytic pathway 1 . In response to environmental stresses, portions of cytoplasmic constituents are engulfed by a unique membrane structure, the phagophore, as it elongates to form a double-or multiple-membrane-bound compartment called the autophagosome. Newly synthesized autophagosomes then undergo extensive remodelling to acquire degradative capabilities. The remodelling process, also known as autophagosomal maturation, involves sequential fusion of autophagosomes with endocytic vesicles...

Infection with SARS-CoV-2 can lead to excessive production of pro-inflammatory cytokines, but the production of type I interferons, which are key antiviral mediators, is reportedly blunted. Here, we discuss how imbalanced interferon responses may contribute to the pathology of COVID-19.

RIG-I is a cytosolic multi-domain protein that detects viral RNA and elicits an antiviral immune response. Two N-terminal caspase activation and recruitment domains (CARDs) transmit the signal and the regulatory domain prevents signaling in the absence of viral RNA. 5′-triphosphate and double stranded (ds) RNA are two molecular patterns that enable RIG-I to discriminate pathogenic from self-RNA. However, the function of the ATPase domain that is also required for activity is less clear. Using single-molecule fluorescence assays we discovered a robust, ATP-powered dsRNA translocation activity of RIG-I. The CARDs dramatically suppress translocation in the absence of 5′f-triphosphate and the activation by 5′-triphosphate triggers RIG-I to translocate preferentially on dsRNA in cis. This functional integration of two RNA molecular patterns may provide a means to specifically sense and counteract replicating viruses.

The co-evolution of viruses with their hosts has led to the emergence of viral pathogens that are adept at evading or actively suppressing host immunity. Pattern recognition receptors (PRRs) are key components of antiviral immunity that detect conserved molecular features of viral pathogens and initiate signalling that results in the expression of antiviral genes. In this Review, we discuss the strategies that viruses use to escape immune surveillance by key intracellular sensors of viral RNA or DNA, with a focus on RIG-I-like receptors (RLRs), cyclic GMP–AMP synthase (cGAS) and interferon-γ (IFNγ)-inducible protein 16 (IFI16). Such viral strategies include the sequestration or modification of viral nucleic acids, interference with specific post-translational modifications of PRRs or their adaptor proteins, the degradation or cleavage of PRRs or their adaptors, and the sequestration or relocalization of PRRs. An understanding of viral immune-evasion mechanisms at the molecular level may guide the development of vaccines and antivirals.