MEMBRANE EVENTS--FOLLOW HORMONE BINDING TO L H RECEPTOR

在人类中，黄体生成素(LH)和人绒毛膜 促性腺激素(HCG)调节孕酮的分泌 月经周期和月经早期的黄体 怀孕了。这项建议将加强对这些问题的理解 促性腺激素通过增加我们的 关于荷尔蒙周围的生理事件的知识 与细胞表面受体和其他膜的相互作用 组件。人和绵羊黄体细胞的相似性 使母羊成为学习物理的优秀模型系统 伴随人绒毛膜促性腺激素或黄体生成素与黄体生成素受体结合的事件。这个 结合亲和力、内化率和细胞对 短暂和长期的荷尔蒙暴露是有特征的 人绒毛膜促性腺激素和黄体生成素在绵羊黄体细胞上的表达。这项提议的目标是 检查黄体生成素受体的聚集、分布和分子 发情黄体期绵羊黄体细胞的运动 周期，并探索促黄体生成素受体的物理行为 受其与olh或hCG相互作用的影响。第一，范围 将在活细胞上测量促黄体生成素受体聚集率 用荧光能量测定激素结合前后的变化 黄体生成素受体旋转运动的传递和测量。 将使用计算机增强技术来检查受体的分布 微光显微镜和电子显微镜。二、角色定位 HCG和olh的α和β亚基以及 修饰过程中与这些分子相关的碳水化合物 激素占据的黄体生成素受体的侧向和旋转运动 将会被确定。异源二聚体形成的效应 人绒毛膜促性腺激素和脱糖hCG亚基的分布 并观察黄体生成素受体的运动情况。第三，我们的 初步研究表明人绒毛膜促性腺激素-黄体生成素受体 运动受细胞骨架组件的限制。我们会 表征细胞骨架与黄体生成素受体的相互作用 激素结合前后。最后，我们将尝试 确定其他黄体细胞膜蛋白是否 与荷尔蒙占据的黄体生成素受体相互作用。是这样的 相互作用可能会显著限制 黄体生成素受体。科罗拉多州立大学有一项不同寻常的 生殖生理学方面的专业知识和 用于膜蛋白动力学研究的仪器设备。 这种情况为我们提供了一个独特的机会，以获得新的和 关于发生的膜事件的详细信息 在激素与黄体生成素受体结合之后。

In humans, luteinizing hormone (LH) and human chorionic gonadotropin (hCG) regulate the secretion of progesterone from the corpus luteum during the menstrual cycle and early pregnancy. This proposal will enhance understanding of how these gonadotrophs regulate luteal cell function by adding to our knowledge of the physical events surrounding hormone interactions with cell surface receptors and other membrane components. Similarities between human and sheep luteal cells make the ewe an excellent model system for studying physical events accompanying hCG or LH binding to the LH receptor. The binding affinity, rate of internalization, and cellular response to brief and prolonged hormone exposure have been characterized for hCG and LH on ovine luteal cells. The goal of this proposal is to examine LH receptor aggregation, distribution, and molecular motions on ovine luteal cells during the luteal phase of the estrous cycle and to explore how the physical behavior of the LH receptor is affected by its interactions with oLH or hCG. First, the extent of LH receptor aggregation will be measured on viable cells before and after hormone binding using fluorescence energy transfer and measurements of LH receptor rotational motions. Receptor distribution will be examined using computer-enhanced low light microscopy and electron microscopy. Second, the role of the alpha and beta subunits of hCG and oLH and the carbohydrates associated with these molecules in modifying lateral and rotational motions of hormone-occupied LH receptors will be determined. The effects of heterodimers formed from hCG and oLH subunits and deglycosylated hCG on the distribution and motion of LH receptor will be examined. Third, our preliminary studies indicate that hCG-LH receptor lateral motions are constrained by cytoskeletal components. We will characterize cytoskeletal interactions with the LH receptor before and after hormone binding. Finally, we will attempt to determine whether other luteal cell membrane proteins are interacting with the hormone-occupied LH receptor. Such interactions may significantly restrict the lateral movement of the LH receptor. Colorado State University has an unusual confluence of expertise in reproductive physiology and of instrumental facilities for studies of membrane protein dynamics. This situation affords us a unique opportunity to obtain new and detailed information regarding membrane events which occur following hormone binding to the LH receptor.