Parathyroid Hormone Receptor Type 1: Dynamics, Traffic and Recycling

甲状旁腺激素受体 1 型：动力学、交通和回收

• 批准号: 8002407
• 负责人: GUILLERMO G ROMERO
• 金额: $ 22.73万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-02-19 至 2011-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8002407
• 关键词: Adaptor Signaling Protein; Address; Arrestins; Arts; Behavior; Binding; Biochemical; Biochemistry; Cell model; Cell surface; Cells; Chronic Kidney Failure; Clathrin; Clathrin Adaptors; Complement; DLG4 gene; Data; Endocytosis; Event; Exhibits; Family; Family member; G-Protein-Coupled Receptors; Goals; Homeostasis; Hormone Receptor; Hormones; Image; Immunoprecipitation; Individual; Kidney; Laboratories; Life; Ligand Binding; Ligands; Measures; Mediating; Membrane; Microscopy; Molecular Biology; Nature; Organism; Outcome; Parathyroid Hormone Receptor; Parathyroid Hormones; Pathway interactions; Phosphorylation; Play; Process; Property; Protein Binding; Proteins; Recycling; Regulation; Relative (related person); Renal Osteodystrophy; Role; Signal Transduction; Spectrum Analysis; Surface; TFAP2A gene; Techniques; Testing; Time; Tissues; Work; base; bone; bone cell; calcium metabolism; calcium phosphate; cellular imaging; coated pit; desensitization; extracellular; genetic regulatory protein; hormone analog; hormone resistance; human PTH protein; innovation; member; mutant; novel strategies; receptor; receptor function; receptor internalization; response; single molecule; sodium-hydrogen exchanger regulatory factor; tool; trafficking

The traffic and fate of internalized G protein coupled receptors (GPCR) play a central role in the regulation of their function. Although the general pathways of GPCR internalization are shared among most members of the family, the precise mechanisms by which these pathways are regulated remain elusive. An extreme example of this is the Type I parathyroid hormone receptor (PTH1R). The PTH1R exhibits remarkable celland tissue-specific differences regarding the regulation of its activity and traffic by different ligands. Previous work from our laboratory and others has shown that the PTH1R is internalized by a combination of arrestindependent and independent processes. However, the relative contributions of these processes and the mechanisms that regulate them are still largely unknown. This application will test the hypothesis that PTH1R internalization is tightly regulated by the interactions of the PTH1R with PDZ (PSD95, discs-large, Zo-1) proteins. Abundant preliminary work presented here shows that the traffic the PTH1R is regulated by the PDZ proteins NHERF1 and Dvl2. This proposal will address the mechanisms by which these regulatory proteins perform their function in bone cell models. These studies will be conducted using state-of-the-art imaging (time-lapsed confocal and total internal reflection microscopy, single molecule tracking, and image correlation spectroscopy) in combination with classical membrane biochemistry and molecular biology strategies (analysis of the behavior of point mutants, ligand binding studies, immunoprecipitation techniques). A major innovation of the approach proposed is that it relies substantially on the analysis of individual endocytic steps to complement the global biochemical analyses. The tools developed and the conclusions obtained from this work will have applications to other members of the GPCR family.

内化G蛋白偶联受体(GPCR)的运输和去向在调节中起着核心作用 它们的功能。尽管大多数成员共享GPCR内化的一般途径 在该家族中，调控这些途径的确切机制仍然难以捉摸。一种极端的 这方面的例子是I型甲状旁腺激素受体(PTH1R)。PTH1R展示了非凡的细胞和 不同配体对其活性和运输的调节存在组织特异性差异。上一首 我们实验室和其他实验室的研究表明，PTH1R是通过与逮捕无关的组合内化的 和独立的进程。然而，这些进程的相对贡献和 监管它们的机制在很大程度上仍是未知的。这个应用程序将测试以下假设 PTH1R内化受到PTH1R与PDZ(PSD95，Diss-Large， ZO-1)蛋白质。大量的前期工作表明，PTH1R的交通是由 PDZ蛋白NHERF1和Dvl2。这项提案将解决这些监管机构通过哪些机制 蛋白质在骨细胞模型中发挥其功能。这些研究将使用最先进的技术进行 成像(延时共焦和全内反射显微镜、单分子跟踪和成像 相关光谱学)与经典膜生物化学和分子生物学相结合 策略(点突变的行为分析、配基结合研究、免疫沉淀 技术)。提出的方法的一个主要创新是，它在很大程度上依赖于对 个别内吞步骤，以补充全球生化分析。开发的工具和 从这项工作中获得的结论将适用于GPCR家族的其他成员。