2C Proteins of Enteroviruses Suppress IKKβ Phosphorylation by Recruiting Protein Phosphatase 1

The NF-kappa B signaling network, which is an ancient signaling pathway, plays a pivotal role in innate immunity and constitutes a first line of defense against invading pathogens, including viruses. However, numerous viruses possess evolved strategies to antagonize the activation of the NF-kappa B signaling pathway. Our previous study demonstrated that the nonstructural protein 2C of enterovirus 71 (EV71), which is the major pathogen of hand, foot, and mouth disease, inhibits tumor necrosis factor alpha (TNF-alpha)-mediated activation of NF-kappa B by suppressing I kappa B kinase beta (IKK beta) phosphorylation. Nevertheless, the mechanism underlying the inhibition of IKK beta phosphorylation by EV71 2C remains largely elusive. We demonstrate that EV71 2C interacts with all isoforms of the protein phosphatase 1 (PP1) catalytic subunit (the PP1 alpha, PP1 beta, and PP1 gamma isoforms) through PP1-docking motifs. EV71 2C has no influence on the subcellular localization of PP1. In addition, the PP1-binding-deficient EV71 2C mutant 3E3L nearly completely lost the ability to suppress IKK beta phosphorylation and NF-kappa B activation was markedly restored in the mutant, thereby indicating that PP1 binding is efficient for EV71 2C-mediated inhibition of IKK beta phosphorylation and NF-kappa B activation. We further demonstrate that 2C forms a complex with PP1 and IKK beta to dephosphorylate IKK beta. Notably, we reveal that other human enteroviruses, including poliovirus (PV), coxsackie A virus 16 (CVA16), and coxsackie B virus 3 (CVB3), use 2C proteins to recruit PP1, leading to the inhibition of IKK beta phosphorylation. Our findings indicate that enteroviruses exploit a novel mechanism to inhibit IKK beta phosphorylation by recruiting PP1 and IKK beta to form a complex through 2C proteins, which ultimately results in the inhibition of the NF-kappa B signaling pathway. IMPORTANCE The innate antiviral immunity system performs an essential function in recognizing and eliminating invading viruses. Enteroviruses include a number of important human pathogens, including poliovirus (PV), EV71, and coxsackieviruses (CVs). As 2C is the most conserved and complex nonstructural protein of enteroviruses, its biological function is largely unclear, whereas the 2A and 3C proteinases of enteroviruses are well characterized. We reveal that EV71 2C forms a complex with PP1 and IKK beta to maintain IKK beta in an unphosphorylated and inactive state, resulting in the inactivation of the TNF-alpha-mediated NF-kappa B signaling pathway. We provide evidence that the 2C proteins of the enteroviruses PV, CVA16, and CVB3 suppress IKK beta phosphorylation through the same mechanism involving PP1. We demonstrate that enteroviruses exploit a novel mechanism involving PP1 to regulate innate antiviral immunity, and our findings may be particularly important for understanding the pathogenicity of enteroviruses.