Fig. 2

Effects of Gag cleavage mutation on Gag-Pol trans incorporation into virus particles. a Schematic representations of HIV-1 Gag and Gag-Pol expression constructs. Gag domains, pol-encoded products, and Gag cleavage site mutations are identical to those described in the Fig. 1 caption. GPfs and CSMfs both contained the same gag/pol frameshifting mutation to force Gag-Pol expression without Gag. b 293T cells were transfected with indicated amounts of GPfs or CSMfs plasmid DNA, either alone or with 10 µg Gag plasmid DNA at a ratio of 1:10 (lanes 2 and 5) or 1:1 (lanes 3 and 6). c 293T cells were transfected with 10 µg GPfs or CSMfs plasmid DNA, either alone or with 10 µg Gag (lanes 5–6) or CSMGag (lanes 8–9) plasmid DNA. Ten µg of pBluescript (SK) plasmid DNA was added to give a total of 20 µg DNA for each transfection. d 293T cells were co-transfected with either Gag (10 µg) or CSMGag (10 µg) plus either GPfs (1 µg) or CSMfs (1 µg) expression vectors. At 18 h post-transfection, transfectants were equally split, placed on two plates, and either treated or mock-treated with 1 µM efavirenz (EFV). After an additional 4 h (22 h post-transfection), culture supernatants were removed and replaced with medium containing EFV. Cells and supernatants were harvested for Western immunoblot analysis 48 h post-medium replacement. b-d Show is a representative immunoblot from three similar independent experiments