Fig. 5

SAHA counteracts the PML-dependent inhibition of HIV-1 gene expression. a Effects of SAHA on HIV-1 LTR-driven GFP expression. PML-KO and WT MEFs were infected with increasing doses of HIV-1_NL-GFP. Ten days later, the cells were treated with 5 µM SAHA or with DMSO for 48 h and the MFI was then measured by FACS (P = 0.0001, one-tailed paired Student’s t test for SAHA vs. DMSO treatment in WT cells). b Analysis of HIV-1 p24 expression levels. PML-KO and WT MEFs were infected in triplicate with HIV-1_NL-GFP at a CRFK MOI of 0.1 and then treated with either SAHA or DMSO at 10 dpi. Cellular lysates were prepared 48 h later and analyzed by WB using an anti-p24 antibody. Uninfected extracts were used as a negative control and actin was analyzed as a loading control. c The p24 and actin bands in the WB analysis shown in b were quantified by densitometry. The values represent the means of p24/actin ratios from the three data points for each condition with standard deviations (*P < 0.05; **P < 0.01, two-tailed Student’s t test). d qRT-PCR analysis of HIV-1 transcription. WT or PML-KO MEFs were infected with HIV-1_NL-GFP in triplicate. Ten days after infection, the cells were treated with either DMSO or SAHA for 48 h. Total RNA was purified from the cells and the level of GFP transcript was quantified by qRT-PCR. Total RNA from uninfected cells was used as a negative control. The values represent the means of three independent experiments with standard deviations (*P < 0.05; **P < 0.01, two-tailed Student’s t test). ND not detected