Fig. 4.

Mutating the DBD of PR eliminates progestin inhibition of Stat5a binding to the β-casein promoter and enhancer. HC-11 cells were transduced with 50 pfu/cell of adenovirus encoding hPR-B containing a point mutation in the zinc finger (C587A) region of the DBD and then treated for 1 h with either: ethyl alcohol (EtOH) (−control), ovine PRL (3 μg/ml), R5020 (100 nm), or ovine PRL (3 μg/ml) + R5020 (100 nm). Cells were processed for ChIP assay using antibodies to PR (1294) or to Stat5 (L-20). A, hPR-B (C587A) binding at the proximal promoter and distal enhancer. B, Stat5a binding at the proximal promoter and distal enhancer. Graphical representation of ChIP data was performed as in Figs. 2 and 3. The values are averages ± sem from three experiments. Insets are representative agarose gels of PCR products from one of three replicate experiments.