Cyclic AMP Mediation of Epithelial Cell Function

描述（由申请人提供）：最近的研究已经认识到， 第二信使应答靶被隔离在 牢房这种细胞内世界的观点使人们认识到 广泛的支架蛋白质组，其隔离两种转导酶 以及在电池内的限定位置中的基板。最具特色的， 这些支架蛋白中最广泛的一类是A-激酶锚定蛋白， 结合II型调节亚基二聚体的AKAP蛋白 A-激酶在过去的十年里，我们一直把调查重点放在AKAP上 与胃壁细胞功能有关。这些研究导致了 鉴定Ezrin和AKAP 35 O都是具有重要支架AKAP 在上皮细胞中更普遍重要的功能。本 研究集中在多重剪接的AKAP 350家族的功能上。 除了II型A-激酶，这些蛋白质也是支架蛋白。 磷酸酶1和2a以及蛋白激酶N（Rho激酶）和蛋白激酶N（Rho激酶）。 激酶C-β。最近在极化的HCA-7结肠癌细胞和 在壁细胞中，我们已经证明了一个主要的剪接变体AKAP 35 OA是 特别是与高尔基体有关的。我们假设 位于高尔基体的AKAP 35 OA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，我们最近发现了两个新的AKAP 35 O家族， 蛋白质，氯离子胞内通道（CLIC）和CIP 4 ICIP 5，推定的交叉 肌动蛋白和微管细胞骨架的连接体， AKAP 35 OA位于高尔基体。我们将努力确定和描述 与大分子相关的相互作用蛋白质的共同和特异功能 多蛋白AKAP 350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，我们将描述功能 CIP 4和CIP 5与AKAP 35 OA在高尔基体的结合及其 调节蛋白质运输的作用。第二，我们将调查 AKAP 35 OA与CLIC 5 B锚定在 高尔基体。第三，我们将分离和鉴定 胃壁细胞中的大型非中心体AKAP 5 O支架复合物， HCA-7结肠癌细胞。这些研究将使重点调查 AKAP 35 O支架复合物的光谱，可以调节内高尔基体和 极化上皮细胞的高尔基体后运输。

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.