Rab27a silencing does not disrupt Env trafficking. Control and Rab27a-silenced cells were infected with HIV (IIIb strain) and analyzed at day 10 p.i. (A) Representative FACS plot showing Total (top) and cell surface (bottom) Env levels in uninfected, control, and Rab27a-silenced cells. The percentage of Env-positive cells in the gate containing Env-positive cells is indicated. (B) Quantification of FACS data from three independent experiments. (C) LSCM of control and Rab27a-silenced Jurkat cells stained with anti-p24 (green) and anti-Env antibodies. Differential interference contrast (DIC) images are shown on the right. Bars, 2 µm. (D) gp41 immunoblot on cells spinoculated with a high MOI of vesicular stomatitis virus–pseudotyped HIV at day 3 p.i. (E) gp41/gp160 ratio in control and Rab27a-silenced cells (n = 2). (F) Infectivity of the virions produced by control and Rab27a-silenced cells was evaluated using the same amount of p24 (10 ng, as determined by ELISA) to infect GHOST cells (n = 3). Results are normalized to the value obtained by infecting GHOST cells with the virus contained in the supernatant of control cells. Error bars show SDs. KD, knockdown; MFI, mean fluorescence intensity. **, P < 0.01. Bars, 2µm.