The Molecular Mechanism Of Feline Calicivirus Infection Inhibiting Type ? Interferon Signaling Pathway

The innate immune system plays an essential role in the host's first line of defense against microbial invasion.Host pattern recognition receptors(PRRs)can readily recognize the pathogen associated molecular patterns(PAMPs),which can activate the innate immune system,and ultimately release a series of cytokines such as interferons and inflammatory factors.Type I interferon can activate the transcription of hundreds and thousands of interferon stimulated genes(ISGs)through the JAK/STAT pathway,which exerting an efficiently antiviral effect.while,in order to survive,the virus has also evolved many strategies combating against the host antiviral response,The study of the relationship between FCV and innate immunity is of great importance for the study of Caliciviridae.The feline calicivirus(FCV)is a member of the Vesivirus genu in Caliciviridae family.A cat can cause upper respiratory tract diseases and acute stomatitis ulcers and occasionally even would be die after infected by FCV.Human norovirus of caliciviridae is one of the most important pathogens which could cause acute gastroenteritis and diarrhea.The Immunocompromised and the organ transplant patients may cause death after infected by norovirus.There are still not effective drugs and vaccines against norovirus currently.The reason is that the norovirus can not be cultured readily In vitro.In addition,the interaction between calicivirus and host immune system is relatively rare.FCV is currently one of the best virus models for Norovirus study because FCV can be readily cultured in vitro.The study of the relationship between FCV and innate immunity facilitate the study of the relationship between Caliciviridae and innate immune system.Firstly,differential proteomics was applied to screen differentially expressed proteins when CRFK cells was infected by FCV.A total of 429 differentially expressed proteins were identified,229 proteins expression were upregulated and 200 proteins expression were downregulated.The proteins associated with the innate immune system were IFNAR1,IRF1,IRF9,IFI35,IFT3,PKR and ISG15,and their expression were all downregulated to some degree.Among them,IFNAR1 is a component of type I interferon receptor,and IRF1 is an important regulatory molecule of type I interferon signaling pathway.This finding lays the foundation for the study of FCV inhibition of type I interferon pathway.Secondly,Antiviral assay of IFN-? against FCV were conducted and the result showed that the IFN-? could not inhibit the replication of FCV.Western blot and flow cytometry results showed that the expression of IFNAR1 on the cell surface was down-regulated after FCV infection.Quantitative Real-time PCR,semi-quantitative PCR and Northern blot results showed that the expression of IFNAR1 mRNA was decreased after FCV infection.Finally,we found that FCV nonstructural proteins PP and p30 can inhibit the expression of co-transfected plasmid and can reduce the endogenous mRNA expression of IFNAR1.In addition,the feline IRF1(Fe-IRF1)was cloned and the transient transfection of Fe-IRF1 can significantly inhibit the replication of FCV,The dual luciferase assay results showed that Fe-IRF1 exerts antiviral activity through the activation of type I interferon pathway.The Western blot result showed that FCV infection could down-regulate the expression of IRF1,Quantitative Real-time PCR showed that FCV infection could down-regulate the mRNA expression of IRF1.In conclusion,the differential proteomic methods successfully identified the downregulated innate immune-related proteins IFNAR1,IRF1,IRF9,IFI35,IFIT3,PKR and ISG15 when CRFK were infected by FCV.FCV infection can downregulate the expression of IFNAR1 mRNA and IRF1 mRNA,which block the activation of type I IFN pathway.Ultimately this is the first study that FCV inhibiting the antiviral effects of type I interferon pathway through rpomotion the degration of mRNA.