Vif-negative HIV-1 produced in non-permissive human cells incorporate both APOBEC3F (hA3F) AND APOBEC3G (hA3G), and have a severely reduced ability to produce viral DNA in newly infected cells. While it has been proposed that this reduction is due to deamination of deoxycytidine in viral DNA by either hA3G or hA3F, followed by DNA degradation, recent evidence indicates that the inhibition of viral DNA production can occur independently of DNA editing by either hA3F or hA3G. We have reported that the presence of hA3G in Vif-negative HIV-1 produced from either the non-permissive cell line, H9, or from transfected 293T cells transiently or stably expressing hA3G, results in a >or=50% reduction in the ability of primer tRNA(Lys3) to initiate reverse transcription in these virions, and that this is correlated with a similar reduction in the production of early DNA transcripts in the infected cells. In this work, we show that, like hA3G, hA3F in Vif-negative virions also results in a similar reduction in the initiation of reverse transcription in HIV-1, which is correlated with the inhibition of early viral DNA synthesis in the cell, and which does not require cytidine-deamination of DNA.