Feline calicivirus strain 2280 p30 antagonizes type I interferon-mediated antiviral innate immunity through directly degrading IFNAR1 mRNA

Feline calicivirus (FCV) belongs to theCaliciviridae, which comprises small RNA viruses of both medical and veterinary importance. Once infection has occurred, FCV can persist in the cat population, but the molecular mechanism of how it escapes the innate immune response is still unknown. In this study, we found FCV strain 2280 to be relatively resistant to treatment with IFN-beta. FCV 2280 infection inhibited IFN-induced activation of the ISRE (Interferon-stimulated response element) promoter and transcription of ISGs (Interferon-stimulated genes). The mechanistic analysis showed that the expression of IFNAR1, but not IFNAR2, was markedly reduced in FCV 2280-infected cells by inducing the degradation of IFNAR1 mRNA, which inhibited the phosphorylation of downstream adaptors. Further, overexpression of the FCV 2280 nonstructural protein p30, but not p30 of the attenuated strain F9, downregulated the expression of IFNAR1 mRNA. His-p30 fusion proteins were produced inEscherichia coliand purified, and anin vitrodigestion assay was performed. The results showed that 2280 His-p30 could directly degrade IFNAR1 RNA but not IFNAR2 RNA. Moreover, the 5'UTR of IFNAR1 mRNA renders it directly susceptible to cleavage by 2280 p30. Next, we constructed two chimeric viruses: rFCV 2280-F9 p30 and rFCV F9-2280 p30. Compared to infection with the parental virus, rFCV 2280-F9 p30 infection displayed attenuated activities in reducing the level of IFNAR1 and inhibiting the phosphorylation of STAT1 and STAT2, whereas rFCV F9-2280 p30 displayed enhanced activities. Animal experiments showed that the virulence of rFCV 2280-F9 p30 infection was attenuated but that the virulence of rFCV F9-2280 p30 was increased compared to that of the parental viruses. Collectively, these data show that FCV 2280 p30 could directly and selectively degrade IFNAR1 mRNA, thus blocking the type I interferon-induced activation of the JAK-STAT signalling pathway, which may contribute to the pathogenesis of FCV infection. Author summary Vaccination against FCV has been available for many years and has effectively reduced the incidence of clinical disease. However, vaccines cannot prevent infection, and vaccinated cats can still become persistently infected by FCV, suggesting that FCV has evolved several strategies for counteracting various components of the innate and adaptive immune systems. Here, we show that FCV strain 2280 is resistant to the antiviral effect of IFN. The molecular mechanism by which this occurs is that FCV 2280 infection blocks the JAK-STAT pathway through promoting the degradation of IFNAR1 mRNA by FCV p30 protein. Anin vitrodegradation assay demonstrated that 2280 p30, but not p30 of the vaccine strain F9, could directly and selectively decay IFNAR1 RNA. The exchange of p30 between 2280 and F9 strains using a reverse genetic system also showed that 2280 p30 is a key factor that contributes to the resistance to IFN and enhances virulence. Our findings reveal a new mechanism evolved by FCV to circumvent the host antiviral response.