title: Characterization of anti-HIV activity mediated by R88-APOBEC3G mutant fusion proteins in CD4+ T cells, peripheral blood mononuclear cells, and macrophages.
abstract: In this study, we characterized the anti-HIV activities of various R88-APOBEC3G (R88-A3G) mutant fusion proteins in which each A3G mutant was fused with a virus-targeting polypeptide (R14-88, hereafter named R88) derived from HIV-1 Vpr. Our results show that the introduction of the deaminase-defective mutant E259Q into R88-A3G did not affect the virion incorporation of this mutant but blocked the protein's ability to inhibit HIV-1 infection. Our data also reveal that the antiviral effect of A3GY124A, a previously described A3G virus-packaging mutant, was completely rescued when the mutant was fused with R88. In an attempt to identify the most potent R88-A3G fusion proteins against HIV-1 infection, we introduced two Vif-binding mutants (D128K and P129A) into the R88-A3G fusion protein and showed that both R88-A3GD128K and R88-A3GP129A possessed very potent anti-HIV activity. When R88-A3GP129A was transduced into CD4(+) C8166 T cells, HIV-1 infection was completely abolished for at least 24 days. In an attempt to further test the anti-HIV effect of this mutant in primary human HIV susceptible cells, we introduced R88-A3GP129A into human peripheral blood mononuclear cells (PBMCs) and macrophages with a recombinant adeno-associated virus (rAAV2/5) vector. The results demonstrate that a significant inhibition of HIV-1 infection was observed in the transduced PBMCs and macrophages. These results provide evidence for the feasibility of an R88-A3G-based anti-HIV strategy. The further optimization of this system will contribute to the development of new anti-HIV gene therapy approaches.
authors.affiliation: Laboratory of Molecular Human Retrovirology, University of Manitoba, Winnipeg, MB, Canada.
authors.forename: Zhujun
authors.initials: Z
authors.lastname: Ao
authors.forename: Xiaoxia
authors.initials: X
authors.lastname: Wang
authors.forename: Alexander
authors.initials: A
authors.lastname: Bello
authors.forename: Kallesh Danappa
authors.initials: KD
authors.lastname: Jayappa
authors.forename: Zhe
authors.initials: Z
authors.lastname: Yu
authors.forename: Keith
authors.initials: K
authors.lastname: Fowke
authors.forename: Xinying
authors.initials: X
authors.lastname: He
authors.forename: Xi
authors.initials: X
authors.lastname: Chen
authors.forename: Junhua
authors.initials: J
authors.lastname: Li
authors.forename: Gary
authors.initials: G
authors.lastname: Kobinger
authors.forename: Xiaojian
authors.initials: X
authors.lastname: Yao
mesh_terms: D000071480:APOBEC-3G Deaminase; D015153:Blotting, Western; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology* / Q000378:metabolism; D002460:Cell Line; D003564:Cytidine Deaminase / Q000235:genetics / Q000378:metabolism*; D017931:DNA Primers / Q000235:genetics; D000229:Dependovirus; D005455:Fluorescent Antibody Technique; D015316:Genetic Therapy / Q000379:methods*; D015658:HIV Infections / Q000276:immunology / Q000378:metabolism* / Q000517:prevention & control*; D015497:HIV-1 / Q000276:immunology*; D006801:Humans; D047468:Immunoprecipitation; D007963:Leukocytes, Mononuclear / Q000276:immunology / Q000378:metabolism / Q000821:virology; D008264:Macrophages / Q000276:immunology / Q000378:metabolism / Q000821:virology; D020125:Mutation, Missense / Q000235:genetics; D016133:Polymerase Chain Reaction; D060888:Real-Time Polymerase Chain Reaction; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014161:Transduction, Genetic; D054320:vif Gene Products, Human Immunodeficiency Virus; D054325:vpr Gene Products, Human Immunodeficiency Virus / Q000235:genetics / Q000378:metabolism*
chemical_list: D017931:DNA Primers; D011993:Recombinant Fusion Proteins; D054320:vif Gene Products, Human Immunodeficiency Virus; C514847:vif protein, Human immunodeficiency virus 1; D054325:vpr Gene Products, Human Immunodeficiency Virus; D000071480:APOBEC-3G Deaminase; C464924:APOBEC3G protein, human; D003564:Cytidine Deaminase