FIG 6.

HIV-1 Tat-dependent transcription is inhibited by spironolactone. (A) Effect of SP on the transfected HIV LTR promoter in the presence of Tat. Jurkat T cells were cotransfected with plasmids encoding HIV-LTR-Luc, Renilla luciferase (Ren), and Tat. After 24 h of plasmid expression, the cells were treated with DMSO or SP. P = 0.0121. (B) Effect of SP on transfected CMV promoter in the presence of Tat. Jurkat cells were cotransfected with a plasmid containing CMV-Luc, a second coding for Ren, and 100 ng of a plasmid coding for Tat. After 24 h of plasmid expression, the cells were treated with DMSO or SP. P = 0.1977. ns, no statistically significant difference. (C) Effect of SP on transfected LTR promoter in the presence of Tat. Jurkat cells were cotransfected with a plasmid containing HTLV-LTR-Luc, a second coding for Ren, and 100 ng of a plasmid coding for Tat. After 24 h of plasmid expression, the cells were treated with DMSO or SP. P = 0.1977. (D) Partial rescue of SP-mediated Tat transactivation inhibition by exogenous XPB. Jurkat T cells were transfected with 4 plasmids encoding HIV-LTR-Luc, Ren, XPB, and Tat. After 24 h of transfection, the cells were treated with DMSO or SP. In panels A, B, C, and D, Renilla luciferase was used for normalization. (E) Tat-mediated activation of transcription of a chromosomally integrated HIV-1 LTR promoter. JLTRG-R5 cells were transfected with different concentrations of a plasmid coding for Tat and cultured or not with SP. P = 0.0051. (F) TNF-α activation of transcription of a chromosomal HIV-1 LTR promoter. JLTRG-R5 cells were stimulated with TNF-α and cultured or not with SP before analysis by flow cytometry to determine the percentage of GFP⁺ cells. The results are the means of three experiments performed in triplicate. Unless otherwise indicated, SP was added to the culture medium at a concentration of 10 μM for 24 h. The error bars represent SEM. *, P < 0.05; **, P < 0.01.