NHERF-Mediated PTH Signaling in Mineral Ion Homeostasis

在其对骨和肾以及整体矿物质离子稳态的生物学作用中，PTH刺激PTH/PTHrP受体并激活靶细胞内的独特信号传导途径（包括腺苷酸环化酶、磷脂酶C、磷脂酶-C非依赖性蛋白激酶C和钙通道）。越来越多的证据表明，这些独特的途径中的一些在骨和肾细胞功能中具有明显的差异，这导致了对信号选择性及其细胞作用的研究增加。Na+/H+交换调节因子（NHERFs）（信号调节细胞内支架蛋白的组分）最近已被我们的小组显示充当“分子开关”，其增加PTH对PLC途径的激活并抑制腺苷酸环化酶途径。由合作者进行的NHERF-1基因敲除具有骨质减少骨骼表型。现已在骨细胞系中鉴定出NHERF。缺乏NHERF的负鼠肾细胞系（OK）缺乏通常的PTH刺激的磷酸盐摄取阻断。通过NHERF转染逆转该缺陷并恢复野生型OK细胞系功能。该细胞系作为PTH介导的抑制肾小管再狭窄的模型。 磷的吸收。因此，我们的具体目标是：在负鼠肾细胞模型中分析NHERF在PTH信号传导和细胞形态方面的功能作用，比较NHERF完整、缺陷和补充的细胞系。AIM II.研究NHERF/PTH 1 R复合物在体外骨细胞系中PTH信号传导中的作用。AIM III.研究NHERF-1在体外和体内介导PTH在骨中的作用，使用我们的同事提供的NHERF- 1 -/-小鼠评估负责骨骼表型的细胞机制。

In its biological actions on bone and kidney and overall mineral ion homeostasis, PTH stimulates the PTH/PTHrP receptor and activates distinctive signalling pathways within target cells (including adenylyl cyclase, phospholipase C, phospholipase-C independent protein kinase C and calcium channels). Growing evidence that some of these distinctive pathways have discernable differences in bone and renal cell function has led to increased study of signal selectivity and its cellular role. Na+/H+exchange regulatory factors (NHERFs) (components of signal modulating intracellular scaffold proteins) have recently been shown by our group to act as "molecular switches" that increase activation by PTH of the PLC pathway and suppress the adenylyl cyclase pathway. A gene knockout of NHERF-1, performed by collaborators, has an osteopenic skeletal phenotype. NHERFs have now been identified in bone cell lines. An opossum kidney cell line (OK) which is deficient in NHERF lacks the usual PTH stimulated blockade of phosphate uptake. The deficiency is reversed and wild type OK cell line function restored by NHERF transfection. This cell line serves as a model of PTH-mediated inhibition of tubular re- absorption of phosphate. Our specific aims are therefore: AIM I. Analyze the functional role of NHERFs with respect to PTH signaling and cell morphology in an opossum kidney cell model, comparing cell lines that are NHERF intact, defective and repleted. AIM II. Study the role of the NHERF/PTH1R complex in PTH signaling in bone cell lines in vitro. AIM III. Study the role of NHERF-1 in mediating the actions of PTH in bone, both in vitro and in vivo, working with the NHERF- 1 -/- mice provided by our colleagues for assessment of cellular mechanisms responsible for the skeletal phenotype.