Figure 4

Blocking endogenous ps20 inhibits HIV-1 transfer. Jwt ps20^inter or clone 3 (ps20^inter) was treated with either a non-silencing (siNS) siRNA or a WFDC1/ps20-silencing siRNA for 6 days. ps20, GAPDH and HPRT mRNA was then measured by qRT-PCR relative to β-actin expression in either (A) Jurkat population or (B) Clone 3. Normalized relative expression was calculated in reference to siNS control. (C) 8 × 10⁴ siRNA treated cells were dye-labelled and co-cultured with 40% 2044-infected donor Jurkat cells at a T:D ratio of 1:0.2. The mean percentage of Gag+ target cells in a 4 hour transfer assay is shown. (D) 2 × 10⁵ Jwt cells and C3 clone from were pre-cultured for 3 days with 5 μg/ml of either control mouse IgG1 or the anti-ps20 Ab IG7, then washed, dye-labelled and co-cultured with 40% 2044-infected donor cells at a T:D ratio of 1:0.2 in the presence of a further addition of each Ab. Mean percentage of Gag+ ps20high target cells is shown after 4 hours of co-culture. (E) 2 × 10⁵ Jwt cells and C3 clone cells were cultured in the absence (control, con) or presence of 1 ug/ml of rps20 for 16 hours, washed, dye-labelled and co-cultured with donor cells infected with 40% 2044 at a T:D cell ratio of 1:0.2. The percentage of Gag+ ps20 target cells is shown after 4 hours of co-culture. All data represent the mean of three replicate assays. Asterisk denotes statistically significant data as calculated using a paired t-test. *P ≤0.05; **P ≤0.01.