FIGURE 5.

The disruption of MDM2 binding restores the protein levels of the Vif variant that does not bind to CBFβ, but it does not fully restore the function to counteract APOBEC3G. Single-cycle infection experiments were performed by using VSV-G pseudotyped luciferase reporter viruses produced in 293T cells by co-transfection of pNL43/ΔEnv-Luc with the indicated vif phenotype and pVSV-G in the presence or absence of co-transfection of pcDNA3/HA-A3G. Virus-containing supernatant was added to fresh 293T cells, and the luciferase activity of the cell lysates was measured by adding substrate and using a luminometer. The value of the virus without vif mutation in the absence of APOBEC3G was normalized to 100%, and mean ± S.E. of three independent experiments is shown (top panel). The levels of APOBEC3G in cells and virions and Vif in cells were also analyzed by immunoblotting (bottom panel). Capsid (CA) and tubulin were used as control.