(A) SKIP enhances the activity of transduced HIV-1 Tat101 in vivo. HeLa cells were transfected with HIV-1 LTR-luciferase together with pHA-SKIP or an empty vector, as indicated. At 24 h post-transfection, HeLa cells were transduced with either 500 ng GST or GST-Tat101 in the presence of 100 μM of chloroquine, and luciferase activity was analyzed 48 h after transfection. Results are presented as fold activation relative to transduction with GST. (B) SKIP-specific siRNA blocks activation of the integrated HIV-1 LTR by transduced GST-Tat101. siRNA-treated HeLa P4 cells were transduced with the indicated amount of either GST or GST-Tat101 in the presence of 100 μM of chloroquine. Fold activation was calculated relative to transduction in the presence of chloroquine alone. (C) The SNW domain of SKIP enhances Tat transactivation in vivo. The HIV-1 LTR-Luc vector was transfected together with vectors expressing different HA-tagged SKIP mutants (as indicated schematically at the bottom of the panel), and HeLa cells were transduced with 500 ng GST-Tat101 at 24 h post-transfection. Luciferase activity was analyzed 48 h post-transfection, and results are presented as fold activation relative to transduction with GST-Tat101. Levels of expression of HA-tagged SKIP proteins were determined by immunoblot using an anti-HA antibody, and GADPH levels were analyzed as a loading control.