The apolipoprotein B messenger RNA editing enzyme, catalytic polypeptide-like (APOBEC) family of single-stranded DNA (ssDNA) cytosine deaminases provides innate immunity against virus and transposon replication1–4. A well-studied mechanism is APOBEC3G restriction of human immunodeficiency virus type 1, which is counteracted by a virus-encoded degradation mechanism1–4. Accordingly, most work has focused on retroviruses with obligate ssDNA replication intermediates and it is unclear whether large double-stranded DNA (dsDNA) viruses may be similarly susceptible to restriction. Here, we show that the large dsDNA herpesvirus Epstein–Barr virus (EBV), which is the causative agent of infectious mononucleosis and multiple cancers5, utilizes a two-pronged approach to counteract restriction by APOBEC3B. Proteomics studies and immunoprecipitation experiments showed that the ribonucleotide reductase large subunit of EBV, BORF26,7, binds APOBEC3B. Mutagenesis mapped the interaction to the APOBEC3B catalytic domain, and biochemical studies demonstrated that BORF2 stoichiometrically inhibits APOBEC3B DNA cytosine deaminase activity. BORF2 also caused a dramatic relocalization of nuclear APOBEC3B to perinuclear bodies. On lytic reactivation, BORF2-null viruses were susceptible to APOBEC3B-mediated deamination as evidenced by lower viral titres, lower infectivity and hypermutation. The Kaposi’s sarcoma-associated herpesvirus homologue, ORF61, also bound APOBEC3B and mediated relocalization. These data support a model where the genomic integrity of human γ-herpesviruses is maintained by active neutralization of the antiviral enzyme APOBEC3B.
Bibliographical noteFunding Information:
We thank B. Anderson for technical advice and RT–qPCR data; Y.-F. Chiu for sharing BAC EBV B95.8; H.-J. Delecluse for M81-transformed B cells; K. Hogquist and S. Dunmire for providing the Akata cells; T. Ikeda and C. Richards for cell culture assistance; M. Sanders and staff at the University of Minnesota Imaging Center for assistance with fluorescence microscopy and live cell imaging; A. Serebrenik for the gRNA construct targeting UNG2; G. Starrett for technical programming advice; and R. Khanna, S. Rice, S. Simon, and P. Southern for thoughtful comments. This work was supported by NCI R21-CA206309 (R.S.H.), the University of Minnesota (College of Biological Sciences, Academic Health Center, and Masonic Cancer Center to R.S.H.), and Canadian Institutes of Health Research grant 153014 (to L.F.). National Institutes of Health training grants provided salary support for A.Z.C. (F30 CA200432 and T32 GM008244) and M.C.J. (T32 CA009138). Salary support for J.L.M. was provided by a National Science Foundation Graduate Research Fellowship. J.Y.M. was funded by Secretaría Nacional de Educación Superior, Ciencia, Tecnología e Innovación. L.F. is a tier 1 Canada Research Chair in Molecular Virology. R.S.H. is the Margaret Harvey Schering Land Grant Chair for Cancer Research, a Distinguished University McKnight Professor, and an Investigator of the Howard Hughes Medical Institute.
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