Cyclic AMP Mediation of Epithelial Cell Function

环 AMP 介导上皮细胞功能

• 批准号: 7031523
• 负责人: JAMES Richard GOLDENRING
• 金额: $ 29.86万
• 依托单位: VANDERBILT UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-04-01 至 2008-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7031523
• 关键词: A kinase anchoring protein; Golgi apparatus; biological signal transduction; cell line; cellular polarity; chloride channels; cyclic AMP; cytoskeleton; enzyme mechanism; enzyme substrate; gastrointestinal epithelium; goats; immunologic assay /test; intracellular transport; protein binding; protein kinase A; protein protein interaction; protein structure function; protein transport; proteomics

DESCRIPTION (provided by applicant): Recent investigations have recognized that second messenger responsive targets are sequestered within discrete domains of the cell. This view of the intracellular world has led to the recognition of broad groups of scaffolding proteins, which sequester both transducing enzymes and substrates in defined locations within cells. The best-characterized and most extensive groups of these scaffolding proteins are the A-kinase anchoring proteins (AKAPs), which bind a dimer of regulatory subunits of Type Il A-kinase. We have focused our investigations over the past decade on AKAPs associated with gastric parietal cell function. These studies have led to the identification of both ezrin and AKAP35O as AKAPs with important scaffolding functions of more general importance in epithelial cells. The present investigations center on the function of the multiply spliced AKAP35O family. In addition to Type II A-kinase, these proteins also scaffold protein phosphatases 1 and 2a as well as protein kinase N (Rho kinase) and protein kinase C-epsilon. Recently in polarized HCA-7 colon carcinoma cells and parietal cells, we have demonstrated that a major splice variant AKAP35OA is specifically associated with the Golgi apparatus. We have hypothesized that AKAP35OA at the Golgi apparatus serves as a multi-functional scaffolding system for the anchoring of critical regulators of polarized epithelial function. Indeed, we have recently identified two novel families of AKAP35O interacting proteins, Chloride Intracellular Channels (CLICs) and CIP4ICIP5, putative cross linkers of the actin and microtubule cytoskeletons, which associate with AKAP35OA at the Golgi apparatus. We will seek to identify and characterize common and specific functions of interacting proteins associated with the large multiprotein AKAP35O scaffolded complexes. To accomplish these goals we will pursue three specific aims: First, we will characterize the functional association of CIP4 and CIP5 with AKAP35OA at the Golgi apparatus and their roles in regulating protein trafficking. Second, we will investigate the structural and functional association of AKAP35OA with CLIC5B anchoring at the Golgi apparatus. Third, we will isolate and identify the components of the large non-centrosomal AKAP5O scaffolded complexes in gastric parietal cells and HCA-7 colon carcinoma cells. These studies will allow focused investigation of the spectrum of AKAP35O scaffolding complexes that may regulate intra-Golgi and post-Golgi trafficking in polarized epithelial cells.

描述（由申请人提供）：最近的调查发现 第二信使响应目标被隔离在离散的域内 细胞。这种细胞内世界的观点导致人们认识到 广泛的支架蛋白组，隔离两种转导酶 和细胞内指定位置的底物。最具特色和 这些支架蛋白中最广泛的组是 A 激酶锚定蛋白 蛋白质（AKAP），其结合 II 型调节亚基的二聚体 A-激酶。过去十年我们的调查重点是 AKAP 与胃壁细胞功能有关。这些研究导致 将 ezrin 和 AKAP35O 鉴定为具有重要支架的 AKAP 在上皮细胞中具有更普遍重要的功能。现在的 研究重点是多重剪接的 AKAP35O 家族的功能。 除了 II 型 A 激酶外，这些蛋白质还充当支架蛋白 磷酸酶 1 和 2a 以及蛋白激酶 N（Rho 激酶）和蛋白质 激酶C-ε。最近在极化的 HCA-7 结肠癌细胞和 壁细胞，我们已经证明主要剪接变体 AKAP35OA 是 与高尔基体特别相关。我们假设 高尔基体上的 AKAP35OA 作为多功能支架系统 用于锚定极化上皮功能的关键调节因子。 事实上，我们最近发现了 AKAP35O 相互作用的两个新家族 蛋白质、氯离子细胞内通道 (CLIC) 和 CIP4ICIP5，假定的交叉 肌动蛋白和微管细胞骨架的连接体，与 高尔基体上的 AKAP35OA。我们将寻求识别和表征 与大分子相关的相互作用蛋白的共同和特定功能 多蛋白 AKAP35O 支架复合物。为了实现这些目标，我们将 追求三个具体目标：首先，我们将表征功能 CIP4 和 CIP5 与高尔基体 AKAP35OA 的关联及其 调节蛋白质运输的作用。其次，我们将调查 AKAP35OA 与 CLIC5B 锚定的结构和功能关联 高尔基体。第三，我们将分离并识别其组成部分。 胃壁细胞中的大型非中心体 AKAP5O 支架复合物和 HCA-7结肠癌细胞。这些研究将有助于集中调查 AKAP35O 支架复合物的范围可能调节高尔基体内部和 极化上皮细胞的后高尔基体运输。