Mechanism Of Porcine Pseudorabies Virus UL24 Antagonizing CGAS-STING Signaling Pathway

Innate immunity is the body's first line of defense against viral infections.After the virus invades cells,the pattern recognition receptors(PRRs)expressed by the cells recognize and present viral pathogen-associated molecular patterns(PAMPs).Cells sense pathogen invasion and activate downstream natural immune signaling pathways.Cyclic GMP-AMP synthase(cGAS)is a newly discovered important DNA recognition receptor that can recognize foreign DNA.By combining with viral DNA,it can catalyze ATP and GTP to produce cGAMP.cGAMP can recruit TANK1-binding kinasel and TGF-activated kinasel after interacting with the stimulator of interferon genes,further activate the downstream pathway IRF3 and NF-?B pathway,and activate anti-viral genes such as transcription of type I interferon and inflammatory factors.These cytokines are important for the body generating antiviral immunity and regulating adaptive immunity.Pseudorabies virus(PRV)is a double-stranded DNA virus.The viral protein UL24 encoded by different PRV strains is highly conserved.However,the function of UL24 protein still unclear so far.The escape of cGAS-STING signaling pathway after PRV infection is the key to its successful infection of host cells.Systematic screening of PRV antagonizing c GAS-STING signaling pathway viral proteins is expected to comprehensively analyze the molecular mechanisms by which PRV escapes the host's natural immunity and pathogenicity.This provides a scientific basis for more effective PRV vaccine design and new drug targets.The research is of great significance to the prevention and control of Porcine Pseudorabies.In this study,we first constructed various expression plasmids of PRV and systematically screened them by dual luciferase reporter gene detection,and found that UL24 protein is one of the viral coding proteins that PRV antagonizes the cGAS-STING signaling pathway,and further explore the molecular mechanism of the pseudorabies virus UL24 involved in the natural immune response of the virus to escape the host.In this project,the dual luciferase reporter gene detection system was used to verify that the PRV-encoded protein UL24 has the ability to antagonize the cGAS-STING signaling pathway,and found that UL24 can significantly inhibit the activation of this signaling pathway.At the same time,the results of the dual luciferase reporter gene detection system showed that overexpression of UL24 could significantly inhibit the activation of the NF-?B signaling pathway mediated by TNF-?,indicating that the protein is involved in the antiviral effect of the viral escape host.By constructing the UL24 gene knockout virus,it was proved that the NF-?B activation of cells infected with the UL24-deficient PRV strain(HeN1-UL24-KO)was significantly higher than that of the wild-type virus(HeN1).The overexpression experiment transfected the linker molecule of c GAS-STING signaling pathway,which showed that the expression of NF-?B pathway related protein in cGAS-STING signaling pathway could be suppressed by UL24 protein,which further confirmed that the expression of P65 protein was affected by UL24 protein.In addition,co-immunoprecipitation analysis showed that UL24 protein interacts with P65 protein.Indirect immunofluorescence experiments showed that UL24 can reduce the expression ofP65 protein,and we further proved that UL24 can significantly degrade P65 protein through the ubiquitin-proteasome pathway.In summary,this study has increased our understanding of PRV,and for the first time proved that UL24 plays an important role in inhibiting cGAS-STING-mediated natural immune signaling pathways after PRV infection,escaping the host's antiviral immune response.