IFN induction in MDDCs by HIV viruses with Vpr but not Vif deleted from their genome. (A) Day 2 MDDCs were treated with VSVG-pseudotyped HIV-1_NLAD8ΔVpr, HIV-1_NLAD8ΔVif, HIV-1_{NLAD8ΔVpr,Vif}, or HIV-1_NLAD8 at an MOI of 1 for 48 and 96 hours. (A) IFNβ mRNA expression was determined by qPCR. The mean data from 3 independent donors are shown with standard error bars. (B-D) Day 6 MDDCs were treated with HIV-1_BaL or mock-treated for 24 and 48 hours. (B) IRF-3 intracellular expression levels were determined by flow cytometry; the isotype control is shown as filled curve, mock treated cells are shown with a solid line, and HIV-1–treated cells are shown with a broken line. (C) The percentage of HIV-1–infected cells was determined by flow cytometry with the use of a PE-conjugated p24 antibody, and IRF-3 expression was determined in p24⁻ cells and p24⁺ cells separately (mean of 5 experiments shown with standard error bars). (D-E) IRF-3 expression in response to HIV-1_BaL was also determined by Western blot in both MDDCs (D) and SupT1 cells (E). (F) IRF3 cellular localization was determined by confocal microscopy in mock- and HIV-1–treated cells compared with Sendai virus–treated TZM-bl cells.