Cyclic AMP Mediation of Epithelial Cell Function

描述(由申请人提供)：最近的调查发现 第二信使反应靶标被隔离在离散的 牢房。这种对细胞内世界的看法导致了对 广泛的支架蛋白，隔离两种转导酶 以及在细胞内的限定位置处的衬底。最具特色的和 这些支架蛋白中最广泛的基团是A-激酶锚定 与Il型调节亚基二聚体结合的蛋白质(AKAP) A-激酶。在过去的十年里，我们的调查重点是AKAP 与胃壁细胞功能有关。这些研究导致了 Ezrin和AKAP35O均为具有重要支架的AKAP 在上皮细胞中更具有普遍重要性的功能。现在 研究的重点是多重剪接AKAP35O家族的功能。 除了II型A-激酶外，这些蛋白也是支架蛋白 磷酸酶1和2a以及蛋白激酶N(Rho激酶)和蛋白质 激酶C-epsilon。最近在极化的HCa-7结肠癌细胞和 我们已经证明了一个主要的剪接变异体AKAP35OA是 特别是与高尔基装置有关。我们假设过 高尔基器械的AKAP35OA用作多功能脚手架系统 用于锚定极化上皮功能的关键调节器。 事实上，我们最近发现了两个新的AKAP35O家族相互作用 蛋白质，氯离子细胞内通道(CLICS)和CIP4ICIP5，假定交叉 肌动蛋白和微管细胞骨架的连接物，它们与 AKAP35OA在高尔基仪器。我们将努力识别和描述 与大分子相关的相互作用蛋白的共同和特殊功能 多蛋白AKAP35O支架复合体。为了实现这些目标，我们将 追求三个具体目标：第一，我们将功能 CIP4和CIP5与高尔基体AKAP35OA的相关性及其意义 在调节蛋白质交易方面的作用。第二，我们将调查 AKAP35OA与CLIC5B结合的结构和功能 高尔基仪器。第三，我们将分离和识别 胃壁细胞和胃壁细胞中大型非中心体AKAP5O支架复合体 Hca-7结肠癌细胞株。这些研究将使我们能够集中调查 AKAP35O支架复合体的光谱可能调节高尔基体内和 高尔基体后极化上皮细胞的运输。

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.