POWRE: Estrogen Receptor-Beta Activation of Prolactin Gene Expression

POWRE：催乳素基因表达的雌激素受体β激活

• 批准号: 9806217
• 负责人: Rosemary Steinmetz
• 金额: $ 7.48万
• 依托单位: Indiana University
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-09-01 至 1999-08-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=9806217&HistoricalAwards=false
• 关键词: POWRE; Estrogen; Receptor; Beta; Activation

Rosemary Steinmetz Project Title: Estrogen Receptor-Beta Activation of Prolactin Gene Expression Estrogens modulate the growth and differentiation of many body tissues by influencing the activation or deactivation of estrogen-responsive genes. The first, and most critical, step that is involved in this activation is the binding of estrogen to its endogenous intracellular receptor (ER). ER- alpha (ERa), the first identified and best characterized ER, is composed of several regions or functional domains. Each of these domains carry out a specific function which is absolutely essential for the ER to induce gene activation. Recently, a new ER variant ER-Beta (ER-B) has been discovered. ER-B differs from ERa both in size and the sequence of the amino acids which comprise it. These differences may cause this new receptor to interact with new genes or new ligands or allow it to interact with already identified estrogen-sensitive genes in a manner different from that of ERa. Little is known concerning how this new receptor interacts physically with the genes it governs. The overall objective of this study is to determine the functional domains of ER-B that are required for the activation of the estrogen-responsive prolactin gene. Based on information concerning the structure of ER-B and its homology to ERa, the first specific aim is to construct a series of expression vectors which will express either truncated or mutated ER-B receptors. Using theses mutants, for in vitro transfection assays, we will be able to identify which domain(s) of the ER-B molecule bind various estrogenic compounds and which are required for gene activation. ER-B has been detected in a variety of tissues including the pituitary. Prolactin is a pituitary hormone that is exquisitely estrogen-sensitive. Prolactin is involved in many physiologic functions including reproduction, lactation, sexual development, cellular immunity and behavior. Little is known concerning the regulation of the prolactin gene by ER-B. Estrogens bind to the ER and in synergy with the pituitary-specific factor Pit-1 stimulate prolactin synthesis and release. While the nature of the interaction between Pit-1 and the ERa variant has been well characterized, the interaction between Pit-1 and ER-B has not been determined. The second specific aim of this study is to characterize the molecular interactions between Pit-1 and ER-B that are involved in prolactin gene activation. This will be accomplished by using the prepared mutant receptors in gene transfer studies. Each of the mutant receptors will be introduced into heterologous cell cultures in combination with Pit-1 and a prolactin reporter gene. These cells will then be treated with selected estrogenic compounds and the transcriptional activation of the prolactin reporter gene will be quantified. These studies will determine which regions of the ER-B work synergistically with Pit-1 and which are required for induction of prolactin by estrogenic compounds. Results of these studies will begin to determine the physical interactions between this new receptor isoform and the genes that it controls, as well as, expanding our existing knowledge concerning the mechanisms which govern prolactin homeostasis. These results will advance our knowledge concerning the actions of this newly discovered receptor.

Rosemary Steinmetz 项目名称：催乳素基因表达的雌激素受体β激活雌激素通过影响雌激素反应基因的激活或失活来调节许多身体组织的生长和分化。 该激活过程中的第一个也是最关键的步骤是雌激素与其内源性细胞内受体 (ER) 的结合。 ER-α (ERa) 是第一个被识别和最充分表征的 ER，由多个区域或功能域组成。这些结构域中的每一个都执行特定的功能，这对于 ER 诱导基因激活来说是绝对必要的。 最近，发现了一种新的 ER 变体 ER-Beta (ER-B)。 ER-B 与 ERa 的不同之处在于其大小和组成氨基酸的序列。 这些差异可能导致这种新受体与新基因或新配体相互作用，或者使其以不同于 ERa 的方式与已识别的雌激素敏感基因相互作用。 关于这种新受体如何与其控制的基因发生物理相互作用，人们知之甚少。 本研究的总体目标是确定激活雌激素反应催乳素基因所需的 ER-B 功能域。 基于有关ER-B结构及其与ERa同源性的信息，第一个具体目标是构建一系列表达截短或突变的ER-B受体的表达载体。 使用这些突变体进行体外转染测定，我们将能够识别 ER-B 分子的哪些结构域结合各种雌激素化合物以及基因激活所需的结构域。 ER-B 已在包括垂体在内的多种组织中检测到。催乳素是一种垂体激素，对雌激素非常敏感。催乳素参与许多生理功能，包括生殖、哺乳、性发育、细胞免疫和行为。 关于 ER-B 对催乳素基因的调节知之甚少。 雌激素与 ER 结合，并与垂体特异性因子 Pit-1 协同作用，刺激催乳素的合成和释放。 虽然 Pit-1 和 ERa 变体之间相互作用的性质已得到很好的表征，但 Pit-1 和 ER-B 之间的相互作用尚未确定。 本研究的第二个具体目的是表征参与催乳素基因激活的 Pit-1 和 ER-B 之间的分子相互作用。 这将通过在基因转移研究中使用制备的突变受体来实现。 每个突变受体将与 Pit-1 和催乳素报告基因结合引入异源细胞培养物中。 然后用选定的雌激素化合物处理这些细胞，并对催乳素报告基因的转录激活进行定量。 这些研究将确定 ER-B 的哪些区域与 Pit-1 协同作用以及雌激素化合物诱导催乳素所需的区域。 这些研究的结果将开始确定这种新受体亚型与其控制的基因之间的物理相互作用，并扩展我们关于控制催乳素稳态机制的现有知识。这些结果将增进我们对这种新发现的受体的作用的了解。