EBP50 REGULATION OF PTH RECEPTOR IN BONE AND KIDNEY

描述（由申请人提供）：本项目的长期目标是阐明甲状旁腺激素受体（PTH1R）作用的细胞机制。PTH1R通过其对肾脏和骨骼的作用来调节细胞外钙和磷酸盐稳态。与其他G蛋白偶联受体一样，PTH1R表现出激活、脱敏、内化和再敏化的循环过程。受体脱敏提供了一种机制，以保护细胞免受过度刺激，而再敏化保护细胞免受长期不活动和激素抵抗。然而，与大多数其他受体不同，PTH1R在其激活中表现出相当大的细胞和组织特异性差异。这些差异不能归因于选择性剪接受体形式，受体丰度，或G蛋白的可用性。最近的证据表明，胞质衔接蛋白ezrin结合蛋白50 kD（EBP 50）可能有助于细胞特异性PTH1R信号转导和内化。所提出的研究的中心目标是检查EBP 50对PTH1R循环的各个方面的相互作用和调节活性。四个具体的目标是测试统一的假设，EBP 50调节配体特异性反应的PTH1R。目的1将表征EBP 50对细胞特异性PTH1R活化的影响。目的2将描述EBP 50对PTH1R脱敏的作用。目的3将确定参与PTH1R内化的EBP 50的结构决定因素。目的4将确定EBP 50对PTH1R再循环的影响。计划中的研究采用了一系列细胞生物学、生物化学和分子生物学技术，这些技术将应用于特定的肾脏和骨细胞，这些细胞是PTH作用的主要靶点。初步数据提供了临时支持，并确定了许多拟议工作的可行性。计划中的研究将产生新的和重要的信息的机制和作用，PTH1R调节细胞外钙稳态。结果将提供更多的了解的启动和终止PTH1R的行动。这些结果可能提示导致PTH抵抗的其他病理生理机制，并导致新的治疗机会。

DESCRIPTION (provided by applicant): The long-term goal of this project is to elucidate the cellular mechanisms of parathyroid hormone receptor (PTH1R) action. The PTH1R regulates extracellular calcium and phosphate homeostasis by its actions on kidney and bone. Like other G protein-coupled receptors, the PTH1R exhibits a cyclical process of activation, desensitization, internalization, and resensitization. Receptor desensitization provides a mechanism to protect cells against excessive stimulation, while resensitization guards cells against prolonged inactivity and hormone resistance. Unlike most other receptors, however, the PTH1R exhibits considerable cell- and tissue-specific differences in its activation. These differences cannot be ascribed to alternatively spliced receptor forms, receptor abundance, or G protein availability. Recent evidence suggests that the cytoplasmic adaptor protein ezrin-binding protein 50 kD (EBP50) may contribute to cell-specific PTH1R signaling and internalization. The central goal of the proposed studies is to examine the interaction and modulatory activity of EBP50 on all aspects of PTH1R cycling. Four specific aims are developed to test the unifying hypothesis that EBP50 regulates ligand-specific responses of the PTH1R. Aim 1 will characterize the effects of EBP50 on cell-specific PTH1R activation. Aim 2 will describe EBP50 effects on PTH1R desensitization. Aim 3 will identify structural determinants of EBP50 that are involved in PTH1R internalization. Aim 4 will determine the effects of EBP50 on PTH1R recycling. The planned studies employ an array of cell biological, biochemical, and molecular biological techniques that will be applied to specific kidney and bone cells that are the primary targets of PTH action. Preliminary data provide provisional support and establish the feasibility for much of the proposed work. The planned studies will yield novel and important information on the mechanism and role by which the PTH1R regulates extracellular calcium homeostasis. The results will provide greater understanding of the initiation and termination of PTH1R action. The outcomes may suggest additional pathophysiological mechanisms causing PTH resistance and lead to new treatment opportunities.