FIGURE 2.

Vpu displaces BST2 from Gag in a Trp-76-dependent manner. A, HeLa P4-R5 cells were transfected with proviral plasmids, either pNL4-3 (“WT”) or the indicated Vpu-mutants. The following day, the cells were fixed, permeabilized, and stained for p24 and BST2. Wide field images were acquired as a Z-series, and the plane along the cover glass that captures the cell surface is shown after deconvolution of the image using a “nearest neighbors” algorithm. The left column shows anti-BST2 staining; the middle column shows anti-Gag/p24; and the right column shows a merged image in which the overlapping Gag and BST2 appears yellow. Wild-type Vpu and the S52N,S56N mutant are each able to displace BST2 from Gag (compared with ΔVpu), and this activity is lost when Trp-76 is mutated to Gly. B, Pearson's correlation coefficients for the colocalization of BST2 and Gag along the plasma membrane in the presence of Vpu or mutants from the experiment shown in A. At least 25 cells were assessed for each condition, using the image plane adjacent to the cover glass. Error bars are the standard deviations. p values for unpaired two-tailed t tests were as noted on the graph and for ΔVpu versus WT, W76G, and S52N,S56N were <0.0001; for WT versus S52N,S56N, p = 0.0094; for WT versus W76G and S52N,S56N + W76G, p < 0.0001; and for W76G versus S52N,S56N and S52N,S56N + W76G, p < 0.0001.