BOTH PIT-1 AND THE ESTROGEN-RECEPTOR ARE REQUIRED FOR ESTROGEN RESPONSIVENESS OF THE RAT PROLACTIN GENE
BOTH PIT-1 AND THE ESTROGEN-RECEPTOR ARE REQUIRED FOR ESTROGEN RESPONSIVENESS OF THE RAT PROLACTIN GENE
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DOI：
10.1210/mend-4-12-1964
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发表时间：
1990-12-01
影响因子：
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通讯作者：
MAURER, RA
中科院分区：
文献类型：
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作者：
DAY, RN;KOIKE, S;MAURER, RA
为了研究大鼠PRL基因远端增强子区域内不同顺式活性元件之间的功能关系，我们在GH 3垂体瘤细胞的瞬时转染研究中对该区域进行了缺失和突变分析。 从这些研究的结果表明，该地区的PRL远端增强子包含的Pit-1结合位点是至关重要的，不仅增强子的活性和cAMP的反应，而且对雌二醇的反应。 雌激素受体与赋予基础增强活性的因子的相互作用由突变的远端增强子区域的研究提出，在该区域中PRL雌激素反应元件转化为回文雌激素反应元件。 为了直接检查潜在的相互作用，共转染研究使用PRL远端增强子报告基因构建体和表达载体的Pit-1和大鼠雌激素受体进行了两个异源细胞系。在与胸苷激酶启动子或PRL近端启动子连接的PRL远端增强子的控制下，报告基因的活性通过与COS-1或RAT-1细胞中的Pit-1表达载体共转染没有显著改变。 这些报告结构和雌激素受体的表达载体的共表达导致只有轻微的反应，雌二醇。 然而，当Pit-1和雌激素受体与远端增强子报告基因共转染时，观察到响应雌二醇的显著诱导，并且这种活性依赖于Pit-1表达载体的浓度。 当使用编码含有POU特异性结构域缺失的突变体Pit-1序列的表达载体时，不存在这种应答。 在RAT-1细胞中，Pit-1对PRL近端启动子区域有显著的激活作用，但在雌激素受体不存在的情况下，未观察到远端增强子序列的激活。 这些结果表明，Pit-1和雌激素受体之间的协同相互作用是增强子活性和PRL远端增强子区域赋予的雌激素反应所必需的。 （分子内分泌学4：1964-1972，1990）
To examine the functional relationship between distinct cis-active elements within the distal enhancer region of the rat PRL gene, we have used deletional and mutational analysis of that region in transient transfection studies in GH3 pituitary tumor cells. Results from these studies demonstrate that the region of the PRL distal enhancer containing the Pit-1-binding sites is critical not only for enhancer activity and the response to cAMP, but also for the response to estradiol. An interaction of the estrogen receptor with factors conferring basal enhanced activity is suggested by studies with a mutant distal enhancer region in which the PRL estrogen response element was converted to a palindromic estrogen response element. To directly examine potential interactions, cotransfection studies using PRL distal enhancer reporter gene constructs and expression vectors for Pit-1 and rat estrogen receptor were performed in two heterologous cell lines. The activity of the reporter gene under the control of the PRL distal enhancer linked to either the thymidine kinase promoter or the PRL proximal promoter was not significantly altered by cotransfection with the Pit-1 expression vector in COS-1 or RAT-1 cells. Coexpression of these reporter constructs and an expression vector for estrogen receptor resulted in only a slight response to estradiol. However, when both Pit-1 and estrogen receptor were cotransfected with the distal enhancer reporter gene, a marked induction was observed in response to estradiol, and this activity was dependent upon the concentration of the Pit-1 expression vector. This response was not present when an expression vector encoding a mutant Pit-1 sequence containing a deletion of the POU-specific domain was used. There was substantial activation of the PRL proximal promoter region by Pit-1 in RAT-1 cells, but no activation of the distal enhancer sequence in the absence of estrogen receptor was observed. These results demonstrate that a cooperative interaction between Pit-1 and the estrogen receptor is required for enhancer activity and the estrogen response conferred by the PRL distal enhancer region. (Molecular Endocrinology 4: 1964-1972, 1990)