Figure 4
CSs suppress HIV-1 gene expression by altering viral RNA processing. HeLa rtTA-HIV-ΔMls cells were treated with ~IC80s of CSs, RNAs extracted, quantitated by qRT-PCR or RT-PCR, and levels displayed relative to DMSO (+). (a,b) CSs induce oversplicing of HIV-1 RNAs (n ≥ 3, mean, s.e.m.). (a) Diagram of the primer positions (arrow heads) used for qRT-PCR. (b) Graph of the relative amount of US (black), SS (white), and MS (gray) HIV-1 RNAs in cells treated with various CSs. (c) Nuclear export of HIV-1 US RNAs in cells is altered by CSs (representative n ≥ 4). HIV-1 US RNAs were localized by FISH after treating HeLa rtTA-HIV(Gag-GFP) cells as described above but with ~IC90s of CSs. Nuclei were stained by DAPI and images captured at 630 × magnification. (d) CSs induce differential post-translational modification of SR proteins (representative n ≥ 3). SRp20, Tra2β, and tubulin were analyzed by immunoblot of HeLa rtTA-HIV-ΔMls cells treated as described above except cells of this representative blot were treated 3 d with ~IC50s of CSs. (g) Lanes were cropped and assembled from the same gel/blot per box (Supplementary Fig. S4d).
