title: Varicella-zoster virus immediate-early protein ORF61 abrogates the IRF3-mediated innate immune response through degradation of activated IRF3.
abstract: Varicella-zoster virus (VZV) infection of differentiated cells within the host and establishment of latency likely requires evasion of innate immunity and limits secretion of antiviral cytokines. Here we report that its immediate-early protein ORF61 antagonizes the beta interferon (IFN-β) pathway. VZV infection down-modulated the Sendai virus (SeV)-activated IFN-β pathway, including mRNA of IFN-β and its downstream interferon-stimulated genes (ISGs), ISG54 and ISG56. Through a primary screening of VZV genes, we found that ORF61 inhibited SeV-mediated activation of IFN-β and ISRE (IFN-stimulated response element) promoter activities but only slightly affected NF-κB promoter activity, implying that the IFN-β pathway may be blocked in the IRF3 branch. An indirect immunofluorescence assay demonstrated that ectopic expression of ORF61 abrogated the detection of IRF3 in SeV-infected cells; however, it did not affect endogenous dormant IRF3 in noninfected cells. Additionally, ORF61 was shown to be partially colocalized with activated IRF3 in the nucleus upon treatment with MG132, an inhibitor of proteasomes, and the direct interaction between ORF61 and activated IRF3 was confirmed by a coimmunoprecipitation assay. Furthermore, Western blot analysis demonstrated that activated IRF3 was ubiquitinated in the presence of ORF61, suggesting that ORF61 degraded phosphorylated IRF3 via a ubiquitin-proteasome pathway. Semiquantitative reverse transcription-PCR (RT-PCR) analysis demonstrated that the level of ISG54 and ISG56 mRNAs was also downregulated by ORF61. Taken together, our results convincingly demonstrate that ORF61 down-modulates the IRF3-mediated IFN-β pathway by degradation of activated IRF3 via direct interaction, which may contribute to the pathogenesis of VZV infection.
authors.affiliation: Molecular Virology and Viral Immunology Research Group, State Key Laboratory of Virology, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan 430071, China.
authors.forename: Huifang
authors.initials: H
authors.lastname: Zhu
authors.forename: Chunfu
authors.initials: C
authors.lastname: Zheng
authors.forename: Junji
authors.initials: J
authors.lastname: Xing
authors.forename: Shuai
authors.initials: S
authors.lastname: Wang
authors.forename: Shuping
authors.initials: S
authors.lastname: Li
authors.forename: Rongtuan
authors.initials: R
authors.lastname: Lin
authors.forename: Karen L
authors.initials: KL
authors.lastname: Mossman
mesh_terms: D015153:Blotting, Western; D002460:Cell Line; D014645:Herpesvirus 3, Human / Q000276:immunology* / Q000472:pathogenicity*; D006801:Humans; D057131:Immune Evasion; D007113:Immunity, Innate; D047468:Immunoprecipitation; D050838:Interferon Regulatory Factor-3 / Q000037:antagonists & inhibitors* / Q000378:metabolism; D016899:Interferon-beta / Q000037:antagonists & inhibitors; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D011485:Protein Binding; D025941:Protein Interaction Mapping; D020133:Reverse Transcriptase Polymerase Chain Reaction; D029082:Sendai virus / Q000276:immunology; D025801:Ubiquitin / Q000378:metabolism; D014764:Viral Proteins / Q000378:metabolism*
chemical_list: C494232:IRF3 protein, human; D050838:Interferon Regulatory Factor-3; D025801:Ubiquitin; D014764:Viral Proteins; C073411:protein 61, Varicella-zoster virus; D016899:Interferon-beta; D046988:Proteasome Endopeptidase Complex