Ligand Pharmacology of Estrogen Receptors

描述（由申请人提供）：该项目的长期目标是阐明非基因组雌激素信号传导的分子机制，并创造新的配体，其可以选择性地激活或阻断非基因组信号传导而不是基因组雌激素信号传导。雌激素基因组信号传导涉及核雌激素受体（ERoc和ERP）的激活和随后的雌激素靶基因的转录调节。另一方面，雌激素的非基因组信号传导导致离子通道、酶和第二信使信号级联的快速激活。雌激素的许多非基因组效应发生在中脑，可能与绝经后妇女面临的体温调节和情绪不稳定问题有关。这些反应的快速动力学和不敏感性的转录/翻译抑制剂排除了参与核ER的经典基因组机制的作用，然而，新的细胞机制的非基因组信号转导尚不清楚。本研究计划是围绕一种新的雌激素膜相关G蛋白偶联受体（GPCR）介导至少部分非基因组激素信号传导的假设构建的。支持这一假设的证据表明，雌激素反应性GPCR迅速介导下丘脑神经元中特定钾通道的抑制。此外，这种生理相关的雌激素反应也是由一种新的选择性雌激素受体调节剂（SERM）引起的，该调节剂对ER α或ER β都没有结合亲和力。在具体目标1中，将孤儿GPCR鉴定为雌激素的候选受体，并开发表达该GPCR的稳定细胞系。在具体目标2和3中，开发并进行配体结合和配体活化测定，以建立和表征GPCR的雌激素响应性。有了这些工具，将开发新的SERM，激活（具体目标4）或阻断雌激素响应性GPCR的激活（具体目标5）。这项研究旨在确定快速非基因组雌激素信号传导的分子机制和乌干达激活参数，这可能导致治疗更年期症状的急需的安全疗法。

DESCRIPTION (provided by applicant): The long-term objectives of this project are to elucidate the molecular mechanism of non-genomic estrogen signaling and create novel ligands that can either activate or block non-genomic signaling selectively over genomic estrogen signaling. Estrogen genomic signaling involves activation of the nuclear estrogen receptors (ERoc and ERP) and subsequent transcriptional regulation of estrogen target genes. On the other hand, nongenomic signaling by estrogen results in rapid activation of ion channels, enzymes, and second messenger signaling cascades. Many of these non-genomic effects of estrogen occur in the mid-brain and may be linked to the problems of thermal regulation and mood instability that post-menopausal women face. The rapid kinetics and insensitivity of these responses to transcription/translation inhibitors rules out the involvement of the nuclear ERs acting by the classical genomic mechanism; however, the novel cellular mechanism of nongenomic signaling is unclear. This research plan is constructed around the hypothesis that a novel membrane associated G protein-coupled receptor (GPCR) for estrogen mediates at least some of the non-genomic hormone signaling. Support for this hypothesis comes from evidence that an estrogen-responsive GPCR rapidly mediates the inhibition of a specific potassium channel in hypothalamic neurons. Moreover, this physiologically relevant estrogen response is also elicited by a novel selective estrogen receptor modulator (SERM) that has no binding affinity for either ERa or ERp. In specific aim 1, an orphan GPCR is identified as a candidate receptor for estrogen and a stable cell line expressing this GPCR is developed. In specific aims 2 and 3, ligand binding and ligand activation assays are developed and carried out to establish and characterize the estrogen responsiveness of the GPCR. With these tools in place, novel SERMs will be developed that either activate (specific aim 4) or block activation (specific aim 5) of the estrogen-responsive GPCR. This research aims to define the molecular mechanisms and Uganda-activation parameters of rapid, non-genomic estrogen signaling which could lead to much needed safe therapeutics for treating the symptoms of menopause.