Fig. 4.

M/C starvation reactivates dormant HIV-1 provirus in HDAC4⁺ ACH-2 cells but not in HDAC4⁻ U1 cells. (A) PCR amplification of HDAC4 and ACTB transcripts in HeLa, ACH-2, and U1 cells; HDAC4 is detected in ACH-2 cells but not in U1 cells. Blank, amplification in the absence of template. (B) Real-time PCR quantification of US HIV-1 RNA in ACH-2 and U1 cells cultivated in the presence (control) or absence of M/C for the indicated times. Results are expressed as the fold change vs. the amount of HIV-1 RNA in control conditions at each time point (mean ± SEM of 2–4 independent experiments; *P < 0.05, paired two-tailed Student t test vs. control cells). HIV-1 RNA expression was significantly up-regulated in starved ACH-2 cells but not in U1 cells. (C) RT activity of daily collected supernatants from ACH-2 and U1 cells cultivated up to 4 d in control medium in the absence of M/C (starved) or presence of TNF-α (mean ± SEM of 5 independent experiments for ACH-2 or 2 independent experiments for U1; *P < 0.05 and **P < 0.01, paired two-tailed Student t test vs. control cells). RT activity accumulation after M/C starvation was observed in ACH-2 cells only. (D) 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay performed on daily collected supernatants from ACH-2 or U1 cells cultured as described in C (mean ± SEM of 3 technical replicates of 1 experiment representative of 3 for ACH-2 or 2 for U1). M/C starvation affects viability and proliferation of ACH-2 and U1 cells to a comparable extent.