Figure 3

A packaging-defective APOBEC3G mutant. A. Infectivity of Vif-defective HIV-1 particles (expressed in HeLa P4.2 transducing units normalized for RT activity) produced in the presence of wild-type (A3G) or W127L APOBEC3G by transfection of 293T cells. Numbers below columns represent amount of transfected DNA in μg. Representative of five independent experiments. B. Western blot analysis of these particles and of cytoplasmic extracts of the transfected cells, using indicated antibodies, PCNA and capsid serving as loading controls. Although well expressed, APOBEC3GW127L is not packaged into virions. C. APOBEC3GW127L is sensitive to HIV-1 Vif-induced degradation. HIV-1 Vif was co-expressed in 293T cells with either wild type or W127L HA-tagged APOBEC3G, and extracts were analyzed by HA-specific Western blotting. D. HA-specific indirect immunofluorescence analysis of cells expressing HA-tagged versions of wild-type or W127L APOBEC3G. Both proteins localize to the cytoplasm, although the W127L mutant may exhibit a slightly more diffuse distribution.