Dong Juying, Tsai-Morris Chon-Hwa, Dufau Maria L
Section on Molecular Endocrinology, Endocrinology and Reproduction Research Branch, NICHD, National Institutes of Health, Bethesda, Maryland 20892, USA.
J Biol Chem. 2006 Jul 7;281(27):18825-36. doi: 10.1074/jbc.M512826200. Epub 2006 May 1.
Prolactin exerts diverse functions in target tissues through its membrane receptors, and is a potent mitogen in normal and neoplastic breast cells. Estradiol (E(2)) induces human prolactin receptor (hPRLR) gene expression through stimulation of its generic promoter (PIII). This study identifies a novel E(2)-regulated non-estrogen responsive element-dependent transcriptional mechanism that mediates E(2)-induced hPRLR expression. E(2) stimulated transcriptional activity in MCF7A(2) cells transfected with PIII lacking an estrogen responsive element, and increased hPRLR mRNA and protein. The abolition of the E(2) effect by mutation of Sp1 or C/EBP elements that bind Sp1/Sp3 and C/EBPbeta within PIII indicated the cooperation of these transfactors in E(2)-induced transcription of the hPRLR. DNA affinity protein assay showed that E(2) induced estrogen receptor alpha (ERalpha) binding to Sp1/Sp3 and C/EBPbeta DNA-protein complexes. The ligand-binding domain of ERalpha was essential for its physical interaction with C/EBPbeta, and E(2) promoted this association, and its DNA binding domain was required for transactivation of PIII. Co-immunoprecipitation studies revealed tethering of C/EBPbeta to Sp1 by E(2)-activated ERalpha. Chromatin immunoprecipitation analysis showed that E(2) induced recruitment of C/EBPbeta, ERalpha, SRC1, p300, pCAF, TFIIB, and Pol II, with no change in Sp1/Sp3. E(2) also induced promoter-associated acetylation of H3 and H4. These findings demonstrate that an E(2)/ERalpha, Sp1, and C/EBPbeta complex with recruitment of coactivators and TFIIB and Pol II are required for E(2)-activated transcriptional expression of the hPRLR through PIII. Estradiol produced in breast stroma and adipose tissue, which are major sources of estrogen in post-menopausal women, could up-regulate hPRLR gene expression and stimulate breast tumor growth.
催乳素通过其膜受体在靶组织中发挥多种功能，并且是正常和肿瘤性乳腺细胞中的一种强效促有丝分裂原。雌二醇（E₂）通过刺激其通用启动子（PIII）诱导人催乳素受体（hPRLR）基因表达。本研究确定了一种新的E₂调节的非雌激素反应元件依赖性转录机制，该机制介导E₂诱导的hPRLR表达。E₂刺激了用缺乏雌激素反应元件的PIII转染的MCF7A₂细胞中的转录活性，并增加了hPRLR mRNA和蛋白质。通过突变PIII内结合Sp1/Sp3和C/EBPβ的Sp1或C/EBP元件消除E₂效应，表明这些转录因子在E₂诱导的hPRLR转录中存在协同作用。DNA亲和蛋白分析表明，E₂诱导雌激素受体α（ERα）与Sp1/Sp3和C/EBPβ DNA-蛋白质复合物结合。ERα的配体结合结构域对于其与C/EBPβ的物理相互作用至关重要，E₂促进了这种结合，并且其DNA结合结构域是PIII反式激活所必需的。免疫共沉淀研究揭示了E₂激活的ERα将C/EBPβ与Sp1拴系在一起。染色质免疫沉淀分析表明，E₂诱导C/EBPβ、ERα、SRC1、p300、pCAF、TFIIB和Pol II的募集，而Sp1/Sp3没有变化。E₂还诱导了H3和H4的启动子相关乙酰化。这些发现表明，E₂/ERα、Sp1和C/EBPβ复合物以及共激活因子、TFIIB和Pol II的募集是E₂通过PIII激活hPRLR转录表达所必需的。绝经后女性雌激素的主要来源——乳腺基质和脂肪组织中产生的雌二醇，可能上调hPRLR基因表达并刺激乳腺肿瘤生长。
