Fig. 8.

Validation and functional tests of hormone depletion-insensitive AR recruitment sites in the chromatin. A, Hormone-depleted HeLa cells were transfected with either a minimal promoter-Luc reporter (pG5luc) in which the indicated genomic DNA fragments were inserted upstream of the promoter. The chromosomal locations of the insert sequences are indicated on the x-axis. The cells were cotransfected with either an expression plasmid for wtAR, mutAR, or the vector control; 48 h after transfection, the cells were harvested to measure luciferase activity. The promoter activities are plotted as the ratio of luciferase activities to that of the vector control. Triplicate samples were included in each experimental set. *, P < 0.001. B, Hormone-depleted LP50 cells were transfected with either AR shRNA or control shRNA in combination with mutAR or the vector control; 48 h later, the cells were subjected to ChIP analysis using antibody to AR. TaqMan probes were used to quantify immunoprecipitation of the DNA fragments encompassing the indicated chromatin sites. The TaqMan probe for GAPDH was used for the nontarget control. Chr, Chromosome; *, P < 0.01.