Negative Regulation of Estrogen Receptors

雌激素受体的负调节

• 批准号: 6724340
• 负责人: CAROLYN Louise SMITH
• 金额: $ 25.59万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-02-01 至 2008-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6724340
• 关键词: SDS polyacrylamide gel electrophoresis; cell line; estrogen receptors; estrogens; genetic transcription; ligands; proteasome; protein degradation; protein structure function; receptor expression; tissue /cell culture; transcription factor; ubiquitin

DESCRIPTION (provided by applicant): Although great effort has been spent examining the mechanisms through which ligands activate estrogen receptor-alpha (ERalpha) transcriptional activity, relatively little is known about the molecular events through which Estrogen receptor action is negatively regulated. It is clear that estrogen treatment of many cell types leads to the down regulation of ERalpha expression through polyubiquitination and degradation of the receptor by the 26S proteasome. However, it is unknown how ligand binding to receptor targets it for degradation. Paradoxically, proteasome inhibitors block ERalpha-dependent gene expression, even though there is more receptor present within the cell, and this indicates an important link between the proteasome and gene expression. The overall goal of the experiments outlined in this application is to provide a more detailed understanding of the mechanisms utilized by ligands to induce ERalpha degradation by the proteasome, how cell-specific regulation of this is achieved, and how this contributes to the cell specificity of ERalpha transcriptional activity. Our planned studies are based on five key observations. First, the ligand binding domain of ERalpha is sufficient to mediate ligand-dependent down regulation. Second, induction of ERalpha degradation by estradiol and ICI 182,780 is blocked by proteasome inhibitors. Third, proteasome-mediated down regulation of ERalpha is cell-type specific. Fourth, estradiol and ICI 182,780 induce interactions between the ERalpha ligand binding domain and CBP/p300 in a cell-specific manner that correlates with ERalpha degradation. Lastly, inhibition of CBP/p300 function blocks ligand-dependent ERalpha down regulation. These findings support the hypothesis that the ability of CBP and/or p300 to be recruited to ERalpha by agonist or pure antagonist ligands is an important molecular event necessary for ERa degradation by the 26S proteasome, and that cell specific ERa interactions with CBP and/or p300 therefore contribute to cell-type dependent ERa down regulation as well as transcriptional activity. This will be tested in the following specific aims: 1) Examine the relationship of the 26S proteasome with ligand-dependent down regulation of ERalpha expression and ERalpha-dependent transcriptional activity; 2) Determine the role of AF2 interacting factors in ligand-induced down regulation of ERalpha and 3) Examine the relationship between cell-type specific receptor down regulation and transcriptional activity, particularly with respect to the relative transcriptional strength of ERalpha's AF-1 and AF-2 domains.

描述（由申请人提供）：尽管人们花费了大量的精力来研究配体激活雌激素受体-α（ERα）转录活性的机制，但对于雌激素受体作用被负调节的分子事件知之甚少。很明显，许多细胞类型的雌激素处理通过多泛素化和 26S 蛋白酶体降解受体导致 ERα 表达下调。然而，目前尚不清楚配体与受体的结合如何靶向其降解。矛盾的是，即使细胞内存在更多受体，蛋白酶体抑制剂也会阻断 ERα 依赖性基因表达，这表明蛋白酶体和基因表达之间存在重要联系。本申请中概述的实验的总体目标是更详细地了解配体诱导蛋白酶体降解 ERα 的机制、如何实现细胞特异性调节以及这如何促进 ERα 转录活性的细胞特异性。我们计划的研究基于五个关键观察结果。首先，ERα 的配体结合结构域足以介导配体依赖性下调。其次，雌二醇和 ICI 182,780 诱导的 ERα 降解被蛋白酶体抑制剂阻断。第三，蛋白酶体介导的 ERα 下调具有细胞类型特异性。第四，雌二醇和 ICI 182,780 以细胞特异性方式诱导 ERα 配体结合结构域和 CBP/p300 之间的相互作用，这与 ERα 降解相关。最后，抑制 CBP/p300 功能可阻断配体依赖性 ERα 下调。这些发现支持这样的假设：CBP和/或p300被激动剂或纯拮抗剂配体招募到ERα的能力是26S蛋白酶体降解ERa所必需的重要分子事件，并且细胞特异性ERa与CBP和/或p300的相互作用因此有助于细胞类型依赖性ERa下调以及转录活性。这将在以下具体目标中进行测试：1）检查26S蛋白酶体与配体依赖性ERα表达下调和ERα依赖性转录活性的关系； 2) 确定 AF2 相互作用因子在配体诱导的 ERalpha 下调中的作用，3) 检查细胞类型特异性受体下调和转录活性之间的关系，特别是 ERalpha 的 AF-1 和 AF-2 结构域的相对转录强度。