Figure 7.
PR acetylation mimics display altered transcriptional activation. A, Acetylation mimics exhibit delayed phosphorylation. T47D cells stably expressing wt, K-Q, or K-T PR-B were serum starved and treated with R5020 (10−8 m) for 0, 1, or 4 h. Cells were subjected to Western blotting for PR phosphorylation on Ser345 and Ser400 as well as total PR. B, Acetylation mimics display a lag in nuclear retention. Serum starved T47D cells stably expressing wt, K-Q, or K-T PR-B were treated with R5020 (10−8 m) for 0, 45, or 240 min. Coverslips incubated with PR antibody and stained with DAPI were subjected to confocal microscopy. Fluorescence intensity of whole cell and nuclear PR-B was calculated using ImageJ software. P values represent statistical significance between wt PR-B and K-Q or K-T PR-B (±sd; *, P < 0.01; **, P < 0.002). C, K-Q and K-T PR-B exhibit delayed c-myc induction. T47D cells stably expressing wt, K-Q, or K-T PR-B were starved in 5% DCC and treated with vehicle or R5020 (10−8 m) for 1 or 6 h. Triplicate cultures were subjected to qPCR, c-myc mRNA levels were normalized to β-actin, and statistical significance was evaluated by comparing vehicle and R5020-treated conditions within cell lines (±sem; *, P < 0.03). D, PR acetylation mimics exhibited suppressed activation of SGK expression. T47D cells stably expressing wt, K-Q, or K-T PR-B were plated in triplicate cultures, serum starved, and treated for 18 h with vehicle or R5020 (10−8 m). SGK mRNA levels were normalized to β-actin and measured using qPCR. Statistical significance between groups was evaluated (±sd; *, P < 0.002). All experiments were completed three times. EtOH, Ethanol.