Composite glucocorticoid regulation at a functionally defined negative glucocorticoid response element of the human corticotropin-releasing hormone gene.
Composite glucocorticoid regulation at a functionally defined negative glucocorticoid response element of the human corticotropin-releasing hormone gene.
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DOI：
10.1210/mend.13.10.0351
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发表时间：
1999-10
影响因子：
--
通讯作者：
Stephen P. Malkoski;R. Dorin
中科院分区：
文献类型：
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作者：
Stephen P. Malkoski;R. Dorin
下丘脑-垂体-肾上腺轴的糖皮质激素依赖性负反馈部分是通过直接抑制下丘脑CRH基因转录而实现的。在本研究中，我们试图进一步定位和表征糖皮质激素受体(GR)和AP-1在CRH启动子的功能定义的负糖皮质激素反应元件(NGRE)上的相互作用。在小鼠促肾上腺皮质激素ATT-20细胞中的瞬时转基因研究表明，在1kb完整CRH启动子的背景下，nGRE(-278到-249个核苷酸)的内部缺失导致8-BrcAMP刺激减少和糖皮质激素依赖性抑制CRH启动子活性。NGRE通过cAMP和过表达的c-jun或c-fos AP-1核蛋白进行转录激活，并对异源启动子进行糖皮质激素依赖的特异性抑制。来自小鼠增殖素启动子的复合GRE也观察到了类似的调节曲线。CRH nGRE与一致性cAMP反应元件(CRE)在-224个核苷酸上明显不同，后者增加了异源启动子的基础活性和cAMP反应，但不提供糖皮质激素依赖性抑制。在nGRE内的相邻元件上发现了GR和AP-1核蛋白的高亲和力结合部位。破坏GR或AP-1结合活性的突变与糖皮质激素依赖性抑制的丧失有关。这些结果与糖皮质激素依赖性抑制的复合机制一致，该机制涉及CRH启动子内离散相邻位置的GR和AP-1核蛋白直接与DNA结合。
Glucocorticoid-dependent negative feedback of the hypothalamic-pituitary-adrenal axis is mediated in part through direct inhibition of hypothalamic CRH gene transcription. In the present study, we sought to further localize and characterize glucocorticoid receptor (GR) and AP-1 interactions at a functionally defined negative glucocorticoid response element (nGRE) of the CRH promoter. Transient transfection studies in mouse corticotroph AtT-20 cells demonstrated that internal deletion of the nGRE (-278 to -249 nucleotides) within the context of 1 kb of the intact CRH promoter resulted in decreased 8-BrcAMP stimulation and glucocorticoid-dependent repression of CRH promoter activity. The nGRE conferred transcriptional activation by both cAMP and overexpressed c-jun or c-fos AP-1 nucleoproteins as well as specific glucocorticoid-dependent repression to a heterologous promoter. A similar profile of regulation was observed for the composite GRE derived from mouse proliferin promoter. The CRH nGRE was clearly distinct from the consensus cAMP response element (CRE) at -224 nucleotides, which increased basal activity and cAMP responsiveness of a heterologous promoter but did not confer glucocorticoid-dependent repression. High-affinity binding sites for both GR and AP-1 nucleoproteins were identified at adjacent elements within the nGRE. Mutations that disrupted either GR or AP-1 binding activity were associated with loss of glucocorticoid-dependent repression. These results are consistent with a composite mechanism of glucocorticoid-dependent repression involving direct DNA binding of GR and AP-1 nucleoproteins at discrete adjacent sites within the CRH promoter.