Molecular Mechanism Study Of Feline Infectious Peritonitis Virus Inhibiting Type ? IFN Signaling

Feline infectious peritonitis(FIP),caused by feline infectious peritonitis virus(FIPV),is one of the leading infectious cause of death in wild and domestic cats.It kills approximately 5% cats in multi-cat households per year.FIPV causes systemic infection,effective and sustainable viral replication in monocytes,and activation of infected monocytes.The innate immune system utilizes pattern-recognition receptors(PRRs)to detect the invasion of pathogens and initiate host antimicrobial responses such as the production of type ? interferons and proinflammatory cytokines.Accumulating evidence shows that coronaviruses have abilities to evade host IFN response.However,whether FIPV antagonizes the type ? IFN signaling and the mechanism of evading natural immune response remains unclear.This study aims to provide a further explanation that how FIPV nsp5 efficiently inhibits host IFN response and contribute to our understanding of the mechanism of FIPV to evade innate immunity strategies,providing scientific support for accelerating the development of more effective control strategies.To explore whether the FIPV-DF2 infection can activate the host innate immune responses,the effect of FIPV DF2 infection on IFN-? promoter activation and IFN-? and ISGs m RNA levels were evaluated by qRT-PCR and dual luciferase reporter assay.We found that FIPV-DF2 infection not only failed to induce IFN-? and ISGs(Viperin,ISG15,IFITMI)production,but also inhibited SeV or poly(I:C)mediated IFN-? production.Then,we found that the IRF3 promoter activation induced by SeV or poly(I:C)were significantly inhibited by FIPV-DF2 infection in a dose-dependent manner.Furthermore,we examined the effect of FIPV DF2 infection on the phosphorylation level and translocation of IRF3 induced by SeV in CRFK cells.The results showed that FIPV DF2 infection blocked IRF3 phosphorylation and translocation.In conclusion,FIPV-DF2 interrupts the SEV-mediated activation of IFN-? by blocking the IRF3 pathway.Of the several known viral evasion strategies,the cleavage of innate immune adaptor is a particularly powerful means of inactivating antiviral response.Here,we report that FIPV nsp5 was a negative regular of type ? IFN production.The overexpression of FIPV nsp5 inhibited SeV and poly(I:C)mediated type ? IFN response.We found that FIPV nsp5 inhibited IFN signaling by inhibiting upstream of TBK1.Then we found that FIPV nsp5,interrupted type ? IFN signaling by cleaving NEMO at three sites-glutamine132(Q132),Q205 and Q231.FIPV nsp5 cleaved NEMO in a protease-dependent manner and apoptotic caspases were not required for NEMO cleavage.We constructed a series of truncated mutants based on the identified nsp5 cleavage sites.Further investigation revealed that the cleavage products of NEMO lost the ability to activate IFN-? promoter.Mechanistically,the nsp5-mediated NEMO cleavage disrupted the recruitment of TANK to NEMO,which reduced the phosphorylation of interferon regulatory factor 3(IRF3),leading to the inhibition of type ? IFN production.