Until recently, breast cancer carcinogenesis has not been fully understood, but the roles of estrogen receptors(ERs) and growth factor receptors(like HER2) were known to be important. Growth factors have been shown to synergize in the E2 signaling pathway, although the actual molecular mechanism remains largely unknown. To invers-tigate the effect of HER2 overexpression on the ERE(estrogen responsive element) mediated tran-scriptional activity of the ERs, this study was designed.

NIH3T3 cells, T6-17 cells (NIH3T3 cells with stably transfected with HER2), and MCF-7 cells were maintained in dextran-coated charcoal stripped 10% Dulbecco's Modified Eagle Medium(DMEM). Transient transfection of constructs (pcDNA 3-ERα, pcDNA3-ERβ, pERE-luc, Pap-1-luciferase, and pcDNA-HER2) into each cells was performed using the Lipofectamine PLUS™ system. Reporter gene assays using ERE-luciferase or AP-1-luciferase were used to measure the ER tran-scriptional activities after treatment with estradiol(EZ) and tamoxifen.

Reporter gene assay using ERE-luciferase in both ERα and ERβ, showed much less responsiveness to estrogen in HER2 overexpressing T6-17cells than in NIH3T3 cells, but there was no remarkable difference after treatment with tamoxifen. The AP-1-mediated transcriptional activity was increased in ERβ after tamoxifen treatment, but it disappeared in HER2-expressing T6-17 cells. The responsiveness to estrogen in HER2-transfected MCF-7 cells was also slightly less than in the control MCF-7 cells, and the ERE-mediated transcriptional activity of estrogen in MCF-7 cells was decreased, in a dose-dependent manner, after HER2 transfection.

Coexpression of HER2 and ER seems to make cells less responsive to estrogen stimulation, and decrease the ERE-mediated transcriptional activity in both ERα and ERβ. These results suggest that the expression of HER2 reduces the estrogen dependency in cell growth and eventually induces estrogen independent-growth.