Figure 2

PIs activate latent HIV-1 gene expression in tissue culture model systems. A. Schematic of the RLUC/RFP construct used to establish latently infected HeLa#14 cells. The vpu gene start codon is mutated for robust reporter gene expression. The black box represents a deletion and the patterned boxes represent RLUC and RFP reporter gene insertions. Together, the deletion in pol and the insertion in env render the vector replication-incompetent. B. Schematic of the SEAP/GFP construct used to establish latently infected 24ST1NLESG cells. The vpu gene start codon is mutated for robust reporter gene expression. The black box represents a deletion and the patterned boxes represent SEAP and GFP reporter gene insertions. Together, the deletion in pol and the insertion in env render the vector replication-incompetent. C. HeLa#14 cells were treated for 48 hours with PIs as indicated and RLUC activity was measured. The values shown indicate RLUs normalized to protein concentrations. D. 24ST1NLESG cells were treated with CLBL for 48 hours and MG-132 and Velcade for 72 hours at the indicated concentrations, and SEAP activity was analyzed. The values shown indicate RLUs normalized to the number of live cells. E. OM-10.1 cells were treated for 72 hours with PIs as indicated and p24 levels in the supernatant were analyzed via p24 ELISA. p24 levels (ng/mL) were calculated using standard curve values. Error bars indicate SEM. Asterisks indicate significant differences (** p<0.01; *** p<0.001) between drug-treated cells and DMSO-treated (negative control) cells. P-values were calculated using one-tailed Student’s t test. Cells treated with SAHA served as positive controls. The figure represents average values from three independent experiments.