MOLECULAR BIOLOGY OF HORMONES THAT REGULATE REPRODUCTION

本项目的目的是研究 编码大脑和性腺激素的基因， 哺乳动物的生殖 四种多肽激素调节 已经描述了垂体促性腺激素的合成和分泌。 这些是下丘脑肽促性腺激素释放激素（GnRH）， 与之相关的性腺激素类物质，如：促性腺激素和激活素， 描述了卵巢激素卵泡抑素。 抑制素、激活素和卵泡抑素 调节促卵泡激素（FSH）和GnRH的分泌 调节卵泡刺激素和黄体生成素（LH）的分泌， 脑垂体 因为FSH和LH在调节 生殖腺的配子发生和类固醇生成功能，这些调节 激素可能对生殖系统的稳态很重要， 系统 我们已经从大鼠中分离了所有这些肽的cDNA克隆 激素：促性腺激素释放激素（GnRH）、卵泡抑素以及 白蛋白/激活素。 我们已经使用这些cDNA来检查表达， 原位杂交检测生殖组织中相应的mRNA 组织化学，并已开始探索这些基因的调控 在大鼠发情周期和妊娠期间。 我们现建议 把这些研究向两个方向延伸。 首先，我们计划探讨 调控这些基因的分子机制。 我们将完成 体内实验，以检查GnRH、β-受体结合蛋白和激活素mRNA的变化 在发情周期和特定激素的反应水平。 我们 将开发体外试验，包括卵巢器官培养和原代培养。 大鼠颗粒细胞培养物，以直接检查类固醇和 多肽类激素对拟南芥素基因表达的影响。 颗粒细胞将 也可用于转染研究， 将表征调节元件。 其次，我们计划研究 这些基因表达在几种新的 组织中 已在多种细胞中发现了β-受体结合素/激活素β链mRNA。 组织包括脑、垂体、造血系统和 胎盘 这是特别有趣的，因为它与 转化生长因子-β家族的生长和分化 监管部门 我们将研究何时在发展中的reflobin/激活素 β-链基因在这些组织中表达，并决定是否 基因的调节方式与在性腺中观察到的类似。 我们 还将检测大鼠卵泡抑素基因的性腺外表达， 根据我们最近的观察，这种基因在 妊娠早期的蜕膜组织。 为了帮助这些研究，我们将 产生对激活素和卵泡抑素特异的免疫试剂 proteins. 我们希望这些实验能增强我们对 这些激素在调节正常生殖中发挥作用，并提供 一个框架，用于审查其活动的改变如何可能导致 生殖障碍或功能障碍。

The goal of this project is to examine the expression and regulation of genes encoding brain and gonadal hormones that are involved in controlling mammalian reproduction. Four polypeptide hormones that modulate the synthesis and secretion of the pituitary gonadotropins have been described. These are the hypothalamic peptide gonadotropin-releasing hormone (GnRH), the related gonadal hormones inhibin and activin, and the recently described ovarian hormone follistatin. Inhibin, activin, and follistatin regulate the secretion of follicle-stimulating hormone (FSH) and GnRH regulates the secretion of FSH and lutienizing hormone (LH) from the pituitary. Because FSH and LH play a crucial role in modulating the gametogenic and steroidogenic functions of the gonads, these regulatory hormones are likely to be important for homeostasis in the reproductive system. We have isolated cDNA clones from the rat for all of these peptide hormones: GnRH, follistatin, and the alpha- and beta-chains of inhibin/activin. We have used these cDNAs to examine expression of the corresponding mRNAs in reproductive tissues using in situ hybridization histochemistry, and have begun to explore the regulation of these genes during the rat estrous cycle and during pregnancy. We now propose to extend these studies in two directions. Firstly, we plan to explore the molecular mechanisms by which these genes are regulated. We will complete in vivo experiments to examine changes in GnRH, inhibin, and activin mRNA levels during the estrous cycle and in response to specific hormones. We will develop in vitro assays, including ovarian organ cultures and primary rat granulosa cell cultures, to directly examine the effects of steroid and polypeptide hormones on inhibin gene expression. The granulosa cells will also be utilized for transfection studies in which inhibin and activin gene regulatory elements will be characterized. Secondly, we plan to examine the biological significance of expression of these genes in several novel tissues. The inhibin/activin beta-chain mRNA has been found in diverse tissues including the brain, pituitary, hematopoietic system, and the placenta. This is particularly intriguing in light of its similarity to the transforming growth factor-beta family of growth and differentiation regulators. We will investigate when in development the inhibin/activin beta-chain gene is expressed in these tissues, and determine whether the gene is regulated in a manner similar to that observed in the gonads. We will also examine extra-gonadal expression of the rat follistatin gene, pursuing our recent observation that this gene is highly expressed in decidual tissue early in pregnancy. To aid in these studies, we will generate immunological reagents specific for the activin and follistatin proteins. We expect these experiments to enhance our understanding of the roles these hormones play in regulating normal reproduction, and to provide a framework for examining how alterations in their activities might lead to reproductive disorders or dysfunction.