Fig. 5

17-β-estradiol treatment induces BRK gene and protein expression. a Schematic of the BRK promoter showing the positions of three EREs and binding sites for SP1 and NFκ-B. b MCF7 cells were treated with increasing concentrations of 17-β-estradiol (E2) for 24 h. Cellular proteins were detected in total cell lysates by immunoblotting analysis with anti-BRK and anti-ERα antibodies and β-actin expression served as loading control. c Luciferase activity assay of the reporter constructs in MCF7 cells transfected with and without ERα and ERβ and with and without E2 treatment shows E2 responsive transcriptional activation of the BRK promoter. ETOH (ethanol) was used as a vehicle control. d Plasmids expressing ERα or ERβ were transiently transfected into ER-negative breast cancer cell lines SKBR3 and BT20 and the cell lines treated with either E2 or with a DMSO. Cell lysates were analyzed by immunoblotting using antibodies against BRK and ER. The expression of β-tubulin was used as a loading control