Jeng M H, Kao C, Sivaraman L, Krnacik S, Chung L W, Medina D, Conneely O M, O'Malley B W
Department of Internal Medicine, University of Virginia, Health Sciences Center, Charlottesville 22908, USA.
Endocrinology. 1998 Jun;139(6):2916-25. doi: 10.1210/endo.139.6.6073.
Estrogen regulates proliferation and morphogenesis of mammary ductal epithelium by interacting with a specific intracellular estrogen receptor (ER) that acts as a hormone-dependent transcriptional regulator of gene expression. The mechanisms by which ER regulates transcription in response to estrogen have been analyzed extensively in tissue culture and in cell-free systems. These studies have demonstrated that the transcriptional activity of ER is strongly influenced by cellular context and highlight the need to address ER transcriptional activity in an appropriate cellular background. Thus, to gain insight into the mechanistic role of ER in mammary epithelial morphogenesis, we have used an adenoviral gene delivery strategy to introduce an estrogen-responsive reporter gene into the mammary epithelium and to monitor the activity of endogenous ERs in their natural environment where cellular context including stromal-epithelial interactions can be taken into account. Using this approach, we first demonstrated highly efficient adenoviral delivery throughout the mammary epithelium using a beta-galactosidase (betagal) reporter gene under the control of the constitutively active cytomegalovirus (CMV) promoter. Next, we constructed an adenoviral vector by substituting the CMV promoter with an estrogen-dependent promoter fragment-linked betagal (Ad-ERE-tk-betagal). This adenoviral reporter system provides evidence that ER positive mammary epithelial cells display a differential sensitivity in a region-specific manner toward estrogen induction. Our data suggest that the availability of factor(s) other than ER is necessary for ER-mediated gene activation and may be important in modulating the differential responses of mammary epithelial cells to estrogen.
雌激素通过与特定的细胞内雌激素受体（ER）相互作用来调节乳腺导管上皮的增殖和形态发生，该受体作为基因表达的激素依赖性转录调节因子。在组织培养和无细胞系统中，人们对ER响应雌激素调节转录的机制进行了广泛分析。这些研究表明，ER的转录活性受到细胞环境的强烈影响，并突出了在适当的细胞背景下研究ER转录活性的必要性。因此，为了深入了解ER在乳腺上皮形态发生中的机制作用，我们采用了腺病毒基因递送策略，将雌激素反应性报告基因导入乳腺上皮，并在能够考虑包括基质-上皮相互作用在内的细胞环境的自然环境中监测内源性ER的活性。使用这种方法，我们首先使用组成型活性巨细胞病毒（CMV）启动子控制下的β-半乳糖苷酶（betagal）报告基因，证明了腺病毒在整个乳腺上皮中的高效递送。接下来，我们构建了一个腺病毒载体，用雌激素依赖性启动子片段连接的betagal（Ad-ERE-tk-betagal）取代CMV启动子。这个腺病毒报告系统提供了证据，表明ER阳性乳腺上皮细胞对雌激素诱导以区域特异性方式表现出不同的敏感性。我们的数据表明，除ER外的其他因子的可用性对于ER介导的基因激活是必要的，并且可能在调节乳腺上皮细胞对雌激素的不同反应中起重要作用。
