Activation of bone morphogenetic protein-6 gene transcription in MCF-7 cells by estrogen

作者：Zhang M; Yan JD; Zhang L; Wang Q; Lu SJ; Zhang J; Zhu TH^*

来源：Chinese Medical Journal, 2005, 118(19): 1629-1636.

• bone morphogenetic protein-6
• estrogen
• chromatin immunoprecipitation
• site-directed mutagenesis

Background Bone morphogenetic protein-6 (BMP-6) is closely correlated with tumor differentiation and skeletal metastasis. Estrogen is considered as a stimulant for the initiation and promotion of breast cancer. Previous studies demonstrated that 17 beta-estadiol (E2) can selectively increase the expression of BMP-6. This experiment is designed to detect the molecular mechanism of estrogen activating BMP-6 gene transcription in human estrogen receptor positive (ER+) breast cancer cell line MCF-7.
Methods After the treatment of MCF-7 cells with E2 at different concentrations (10(-11) mol/L, 10(-9) mol/L, 10(-7) mol/L), the BMP-6 expression level was examined through real-time polymerase chain reaction. Through restriction enzyme digestion, human BMP-6 1.2 kb long promoter, BMP-6 0.7 kb long promoter was cloned into pGL-3 basic vector; after the treatment with 10-7 mol/L E2, luciferase activities of the two promoters were detected. Site-directed mutagenesis was performed to obtain the mutant forms of estrogen response element half-site (1/2 ERE) element and Spl sites in the BMP-6 promoter, the activities of these mutant form promoters were detected following the methods mentioned above. Chromatin immunoprecipitation ( ChIP) assay was also used to confirm the binding of estrogen receptor alpha (ER alpha) on BMP-6 promoter in the presence of E2.
Results E2 dose dependently increased BMP-6 mRNA expression in human ER+ breast cancer cell line MCF-7. At a dose of 10(-7) mol/L E2, human BMP-6 1.2 kb promoter activity was increased by 90% compared with the control group treated with ethanol (P < 0.05). Both the 1/2 ERE response element mutant form and the Spl site mutant form of the BMP-6 promoter abolished the activation of the BMP-6 promoter's response to E2. Through ChIP assay, the binding of ER alpha on 1/2 ERE response element in BMP-6 promoter was further validated.
Conclusion Estrogen induces BMP-6 expression in human ER+ breast cancer cell line MCF-7 through its receptor ER(x binding on 1/2 ERE element in the BMP-6 promoter.

• 出版日期2005-10-5
• 单位南开大学