FIG 7.
Effect of Env changes on gp41 HR1 exposure. 293T cells were transfected with an empty pcDNA3.1 plasmid or a plasmid expressing the indicated cytoplasmic-tail-deleted HIV-1JR-FL Env variants. (A) Two days posttransfection, Env expression was evaluated with the 2G12 antibody, which recognizes the gp120 outer domain, by flow cytometry as described in Materials and Methods. (B) HR1 exposure was assessed with an Alexa Fluor 647 (AF647)-conjugated C34-Ig recombinant protein in the absence (gray) or presence (green) of 10 μg/ml of sCD4 by flow cytometry, as described in Materials and Methods. (C) Means and standard deviations of AF647-C34-Ig recognition of Env variants in the presence (green) or absence (gray) of sCD4, presented as the mean fluorescence intensity (MFI) detected with AF647-C34-Ig divided by the MFI detected with the 2G12 antibody. Data are the averages from at least two independent experiments. Statistical significance was evaluated using a two-way analysis of variance (ANOVA), with multiple comparisons. ****, P < 0.0001; ns, not significant. (D) Means of CD4-Ig recognition of Env variants are presented as the MFI detected with CD4-Ig divided by the MFI detected with the 2G12 antibody. Data are the averages from four independent experiments. Statistical significance was evaluated using Student's t test. *, P < 0.05; **, P < 0.01; ns, not significant.