title: Conventional protein kinase C-α (PKC-α) and PKC-β negatively regulate RIG-I antiviral signal transduction.
abstract: Retinoic acid-inducible gene I (RIG-I) is a key sensor for viral RNA in the cytosol, and it initiates a signaling cascade that leads to the establishment of an interferon (IFN)-mediated antiviral state. Because of its integral role in immune signaling, RIG-I activity must be precisely controlled. Recent studies have shown that RIG-I CARD-dependent signaling function is regulated by the dynamic balance between phosphorylation and TRIM25-induced K₆₃-linked ubiquitination. While ubiquitination of RIG-I is critical for RIG-I's ability to induce an antiviral IFN response, phosphorylation of RIG-I at S₈ or T₁₇₀ suppresses RIG-I signal-transducing activity under normal conditions. Here, we not only further define the roles of S₈ and T₁₇₀ phosphorylation for controlling RIG-I activity but also identify conventional protein kinase C-α (PKC-α) and PKC-β as important negative regulators of the RIG-I signaling pathway. Mutational analysis indicated that while the phosphorylation of S₈ or T₁₇₀ potently inhibits RIG-I downstream signaling, the dephosphorylation of RIG-I at both residues is necessary for optimal TRIM25 binding and ubiquitination-mediated RIG-I activation. Furthermore, exogenous expression, gene silencing, and specific inhibitor treatment demonstrated that PKC-α/β are the primary kinases responsible for RIG-I S₈ and T₁₇₀ phosphorylation. Coimmunoprecipitation showed that PKC-α/β interact with RIG-I under normal conditions, leading to its phosphorylation, which suppresses TRIM25 binding, RIG-I CARD ubiquitination, and thereby RIG-I-mediated IFN induction. PKC-α/β double-knockdown cells exhibited markedly decreased S₈/T₁₇₀ phosphorylation levels of RIG-I and resistance to infection by vesicular stomatitis virus. Thus, these findings demonstrate that PKC-α/β-induced RIG-I phosphorylation is a critical regulatory mechanism for controlling RIG-I antiviral signal transduction under normal conditions.
authors: Department of Microbiology and Immunobiology, New England Primate Research Center, Harvard Medical School, Southborough, Massachusetts, USA.
authors: Natalya P
authors: NP
authors: Maharaj
authors: Effi
authors: E
authors: Wies
authors: Andrej
authors: A
authors: Stoll
authors: Michaela U
authors: MU
authors: Gack
mesh_terms: D000818:Animals; D002460:Cell Line; D000071457:DEAD Box Protein 58; D053487:DEAD-box RNA Helicases / Q000378:metabolism*; D055785:Gene Knockdown Techniques; D006801:Humans; D007370:Interferon Type I / Q000378:metabolism; D010766:Phosphorylation; D011493:Protein Kinase C / Q000235:genetics / Q000502:physiology*; D064546:Protein Kinase C beta; D051571:Protein Kinase C-alpha / Q000235:genetics / Q000502:physiology*; D011971:Receptors, Immunologic; D015398:Signal Transduction / Q000502:physiology*; D054875:Ubiquitination
chemical_list: D007370:Interferon Type I; D011971:Receptors, Immunologic; D011493:Protein Kinase C; D064546:Protein Kinase C beta; D051571:Protein Kinase C-alpha; C452820:RIGI protein, human; D000071457:DEAD Box Protein 58; D053487:DEAD-box RNA Helicases