Grass carp reovirus activates fish Ppp1r3g to suppress innate immunity and exacerbate hemorrhage via the 14-3-3ζ/Itgb3 signaling axis

Abstract

Protein phosphatase 1 regulatory subunit 3G (PPP1R3G) is one of the regulatory subunits of protein phosphatase 1 (PP1). Current research on PPP1R3G is primarily focused on glycogen synthesis and lipid metabolism. However, the role of ppp1r3g in the innate immune response of fish has not been completely elucidated. This study demonstrated that ppp1r3g regulates RLR signaling pathway through interactions with Rig-I, Mda5, and Traf6. In parallel, Ppp1r3g inhibits platelet activation by regulating 14-3-3 zeta (14-3-3ζ) and consequently modulating the formation of the 14-3-3ζ-integrin-β3 (Itgb3) complex. In vivo analysis revealed that compared with the wild-type controls, ppp1r3g-null rare minnow (Gobiocypris rarus) and ppp1r3g-chimera grass carp (Ctenopharyngodon idella) exhibited significantly enhanced resistance to grass carp reovirus (GCRV), with markedly alleviated hemorrhagic symptoms post-infection. Furthermore, knockdown of ppp1r3g suppressed GCRV replication and upregulated type I interferon (IFN-1) expression in grass carp kidney (CIK) cells. Additionally, Ppp1r3g binds to the -ESKVFYLKMKGDYYRYL- fragment (EL17) of 14-3-3ζ via the Ser36/37 phosphorylation site located on the PP1c domain. This study also demonstrated that 14-3-3ζ interacts with Itgb3 through this fragment, indicating that Ppp1r3g competes with Itgb3 to bind 14-3-3ζ. Furthermore, Ppp1r3g dose-dependently promoted Itgb3 degradation via a proteolytic pathway. These findings indicate that Ppp1r3g negatively regulates innate antiviral immunity after GCRV stimulation and suppresses platelet and coagulation cascade activation by modulating the interaction between 14-3-3ζ and Itgb3 in fish.