Figure 7. Env is not required for Gag localization to the uropod.

A) P2 cells expressing KFS/Gag-YFP (green) were immunostained after fixation with anti-PSGL-1 (red, upper panel) or anti- α-tubulin (red, lower panel, arrow indicates MTOC) as described for Figure 1. B) KFS/Gag-YFP-expressing P2 cells were immunostained for PSGL-1 or CD43 (red) using the co-patching method as described for Figure 6, and Z-series of images of unpolarized cells were acquired. Maximum projections of each color were generated from the Z stacks and merged to examine colocalization (yellow). Several small copatching puncta are indicated by arrows. C) The Gag polarization index was determined as described in the Materials and Methods section for cells expressing Gag-YFP and KFS/Gag-YFP. Four separate experiments (32, 23, 39, and 37 cells each) for a total of 131 cells for Gag-YFP and 3 separate experiments (34, 48, and 30 cells each) for a total of 112 cells for KFS/Gag-YFP were used for quantification of Gag polarization. P values were determined using Student's t test. NS, not significant. D) Cell-cell contact assays were performed as in Figure 3. This graph compares the results of Figure 3 to the results obtained with cells expressing KFS/Gag-YFP, which had been obtained concurrently. P values were determined using Student's t test. *, P<0.05.