The 14-3-3 chaperone protein promotes antiviral innate immunity via facilitating MDA5 oligomerization and intracellular redistribution

MDA5 belongs to the RIG-I-like receptor family and plays a non-redundant role in recognizing cytoplasmic viral RNA to induce the production of type I IFNs. Upon RNA ligand stimulation, we observed the redistribution of MDA5 from the cytosol to mitochondrial membrane fractions. However, the molecular mechanisms of MDA5 activation remain less understood. Here we show that 14-3-3 is an essential accessory protein for MDA5-dependent type I IFN induction. We found that several 14-3-3 isoforms may interact with MDA5 through the CARDs (N-MDA5), but 14-3-3 was the only isoform that could enhance MDA5-dependent IFN promoter activities in a dose-dependent manner. Knock-down of 14-3-3 in Huh7 cells impaired and delayed the kinetics of MDA5 oligomerization, which is a critical step for MDA5 activation. Consequently, the MDA5-dependent IFN promoter activities as well as IFN mRNA expression level were also decreased in the 14-3-3 knocked-down cells. We also demonstrated that 14-3-3 is essential in boosting the activation of MDA5-dependent antiviral innate immunity during viral infections. In conclusion, our results uncover a novel function of 14-3-3 to promote the MDA5-dependent IFN induction pathway by reducing the immunostimulatory potential of viral dsRNA within MDA5 activation signaling pathway. Author summary In this study, we utilized biochemistry and molecular biology approaches to defines the molecular mechanisms by which melanoma differentiation-associated protein 5 (MDA5), a cytoplasmic RNA helicase and pattern recognition receptor molecule, is regulated by 14-3-3 to govern its innate immune signaling activity. During viral infection RIG-I-like receptors (RLRs), including MDA5, play essential roles in initiating type I interferon signaling pathway and preventing virus infection or replication in host cells. Besides, the establishment of well functional adaptive immune response to viruses is depending on the timely activation of innate immune antiviral signaling pathway. Our results suggested that the activation of MDA5 is promoted by the chaperone protein 14-3-3. The lack of 14-3-3 in host cells leads to the kinetically-delayed oligomerization of MDA5, which is a key steps of the activation of MDA5-mediated anti-viral signaling pathway. These findings reveal a novel component which participating in the control system of MDA5-dependent signaling pathway. Viral proteins which antagonize 14-3-3 to impair MDA5-dependent antiviral signaling may be suitable targets for antiviral therapy or be modified to generate potential vaccine strains.