title: Identification of Kaposi Sarcoma Herpesvirus (KSHV) vIRF1 Protein as a Novel Interaction Partner of Human Deubiquitinase USP7.
abstract: Viral interferon regulatory factor 1 (vIRF1), a Kaposi sarcoma herpesvirus protein, destabilizes p53 by inhibiting p53 acetylation and Hdm2 phosphorylation. This leads to increased ubiquitination and degradation of p53 by Hdm2, which cripples the cellular p53-mediated antiviral response. Ubiquitin-specific protease 7 (USP7) deubiquitinates p53 and Hdm2 and regulates their stability. We identified an EGPS consensus sequence in vIRF1, which is identical to that found in Epstein-Barr virus nuclear antigen 1 (EBNA1) that interacts with the N-terminal domain of USP7 (USP7-NTD). GST pulldown assays demonstrated that vIRF1 interacts with USP7-NTD via its EGPS motif. NMR heteronuclear single quantum correlation (HSQC) analysis revealed chemical perturbations after titration of USP7-NTD with vIRF1 (44)SPGEGPSGTG(53) peptide. In contrast, these perturbations were reduced with a mutant vIRF1 peptide, (44)SPGEGPAGTG(53). Fluorescence polarization analysis indicated that the vIRF1 peptide interacted with USP7-NTD with a Kd of 2.0 μm. The crystal structure of the USP7-NTD·vIRF1 peptide complex revealed an identical mode of binding as that of the EBNA1 peptide to USP7-NTD. We also showed that USP7 interacts with vIRF1 in U2OS cells. Decreased levels of p53, but not Hdm2 or ataxia telangiectasia-mutated (ATM), were seen after expression of vIRF1, but not with a vIRF1 mutant protein. Our results support a new role for vIRF1 through deregulation of the deubiquitinating enzyme USP7 to inhibit p53-mediated antiviral responses.
authors.affiliation: From the Department of Biology, York University, Toronto, Ontario M3J 1P3, Canada.
authors.forename: Sara
authors.initials: S
authors.lastname: Chavoshi
authors.affiliation: From the Department of Biology, York University, Toronto, Ontario M3J 1P3, Canada.
authors.forename: Olga
authors.initials: O
authors.lastname: Egorova
authors.affiliation: From the Department of Biology, York University, Toronto, Ontario M3J 1P3, Canada.
authors.forename: Ira Kay
authors.initials: IK
authors.lastname: Lacdao
authors.affiliation: From the Department of Biology, York University, Toronto, Ontario M3J 1P3, Canada.
authors.forename: Sahar
authors.initials: S
authors.lastname: Farhadi
authors.affiliation: From the Department of Biology, York University, Toronto, Ontario M3J 1P3, Canada.
authors.forename: Yi
authors.initials: Y
authors.lastname: Sheng
authors.affiliation: From the Department of Biology, York University, Toronto, Ontario M3J 1P3, Canada vsaridak@yorku.ca.
authors.forename: Vivian
authors.initials: V
authors.lastname: Saridakis
mesh_terms: D020816:Amino Acid Motifs; D020134:Catalytic Domain; D045744:Cell Line, Tumor; D018360:Crystallography, X-Ray; D019288:Herpesvirus 8, Human; D006801:Humans; D050835:Interferon Regulatory Factors / Q000737:chemistry* / Q000502:physiology; D008958:Models, Molecular; D011485:Protein Binding; D054730:Protein Interaction Domains and Motifs; D055550:Protein Stability; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D043222:Ubiquitin Thiolesterase / Q000737:chemistry* / Q000502:physiology; D000076084:Ubiquitin-Specific Peptidase 7; D014764:Viral Proteins / Q000737:chemistry* / Q000502:physiology
chemical_list: D050835:Interferon Regulatory Factors; C495901:TP53 protein, human; D016159:Tumor Suppressor Protein p53; D014764:Viral Proteins; C413486:viral interferon regulatory factors; C464569:USP7 protein, human; D043222:Ubiquitin Thiolesterase; D000076084:Ubiquitin-Specific Peptidase 7
keywords: DNA viruses; KSHV; USP7; X-ray crystallography; deubiquitylation (deubiquitination); vIRF1; viral protein; virology