Molecular Mechanism Of Herpes Simplex Virus 1 UL2 Protein Inhibits RIG-Ⅰ Like Receptor Pathway

Background and ObjectivesHerpes simplex virus type 1(HSV-1)belongs to the subfamily of Alphaherpesvirus and is one of the common viruses to which humans are susceptible,and is widely prevalent worldwide.The genome of HSV-1 is a linear double-stranded DNA(dsDNA)with a size of 152 kbp(kilobase pairs)which is latently infected in the host cells and causes diseases such as cold sores,neonatal herpes and encephalitis.At present,the specific mechanism of HSV-1 pathogenesis is still being explored.Innate immune response is a critical defense against viral invasion of the host.Previous studies have shown that interferon type Ⅰ(IFN-Ⅰ)plays a dominant antiviral role in the early stages of viral infection.Upon viral infection of host cells,triggering an intracellular signaling cascade response,the downstream junction protein IRF3 and IRF7 were activated to induce the expression of IFN-β.Multiple HSV-1 encoded viral proteins were found to negatively regulate the IFN-β signaling pathway,but there are other HSV-1 encoded viral proteins whose regulation of the IFN-β signaling pathway is not yet clear.UL2,an early protein encoded by HSV-1,is uracil DNA glycosylase(UDG),which is an indispensable enzyme for DNA repair during viral DNA replication.UDG is able to cut the uracil residues in the process of replication,ensuring the correct replication of DNA.In addition,UL2 may play a significant role in latent and recurrent viral infections.However,it is unclear whether UL2 regulates IFN-β signaling pathway.Therefore,this study can further expand the biological function of HSV-1 UL2 protein and elucidate the molecular mechanism of HSV-1 escape type I interferon signaling pathway,which will lay the foundation for future treatment and vaccine development for HSV-1 related diseases.Methods(1)UL2 recombinant expression plasmids were constructed by polymerase chain reaction experiments.(2)The UL2 protein inhibits SeV-induced IFN-β promoter activity by dual-luciferase reporter assay.(3)UL2 inhibits the transcription expression level of IFN-β,ISG54 and ISG56 by semi-Quantitative RT-PCR.(4)The interacting proteins of UL2 was demonstrated by Co-IP experiments.(5)The phosphorylation and dimerization of IRF3 by UL2 through western blot.(6)The nuclear translocation of IRF3 by UL2 through IF A experiments.Results(1)Successful construction of UL2-related label plasmids.(2)UL2 inhibits SeV-induced IFN-β-Luc and(pRDⅢ-Ⅰ)4-Luc promoter activity.Moreover,UL2 can inhibit the mRNA expression levels of IFN-β,ISG54 and ISG56.(3)UL2 inhibits the RLR signaling pathway through targeting IRF3 or IRF7.(4)UL2 interacts with IRF3 activated forms,TRAF3,TBK1 and IRF7.(5)UL2 has little effect on the formation of the IKKi-TBK1-IRF3 complex.(6)UL2 can inhibit the phosphorylation of IRF3 Ser396 site and the formation of IRF3 dimerization by SeV infection.(7)UL2 blocks SeV-induced IRF3 nuclear translocation.(8)The deubiquitination of IRF3-activated forms was mediated by UL2.ConclusionsOur study shows that UL2 protein of HSV-1 can inhibit the activity of SeV-induced IFN-β promoter,which provide an important connection between UL2 and IFN-β signaling pathway.Above all,these results can help us to understand the pathogenic mechanism of HSV-1 more comprehensively.