CYCLIC AMP-DEPENDENT MEDIATION OF PARIETAL CELL FUNCTION

壁细胞功能的循环AMP依赖性调节

• 批准号: 3464390
• 负责人: JAMES Richard GOLDENRING
• 金额: $ 9.04万
• 依托单位: YALE 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 1996-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3464390
• 关键词: binding proteins; biological signal transduction; cell growth regulation; chemical stimulation; cholinergic receptors; cyclic AMP; cytoplasm; cytoskeletal proteins; enzyme activity; enzyme inhibitors; enzyme mechanism; histamine receptor; immunocytochemistry; laboratory mouse; laboratory rabbit; membrane proteins; membrane reconstitution /synthesis; okadaic acid; pancreatic islets; phosphatase inhibitor; phosphoprotein phosphatase; phosphorylation; protein biosynthesis; protein kinase A; secretion

The manifestations of peptic ulcer disease constitute some of the commonest medical complaints. However, while potent pharmacological therapies targeted at the H2-histamine receptor have evolved over the past decade, the intracellular mechanisms responsible for acid secretion remain elusive. The role of cAMP as a mediator of histamine stimulated acid secretion from parietal cells is well recognized. Nevertheless, the mechanisms actually responsible for the translation of the cAMP signal into the physiological response of secretion remain poorly understood. We have recently found that the dephosphorylation of a subclass of Type II regulatory subunit of cAMP-dependent protein kinase may be a critical step in the processing of the cAMP signal in the parietal cell. It is our working hypothesis that cAMP may act through a coordinated activation of both kinase and phosphatase activities. Inherent in this hypothesis is the belief that cAMP-dependent protein kinase activation II regulatory subunit (RII) of cAMP-dependent protein kinase as a cAMP sensor protein. This proposal will pursue five major lines of investigation: First, the role of protein phosphatase and kinase activities in mediating the histamine stimulation on the phosphorylation of RII in situ will be evaluated in the presence of both a protein phosphatase inhibitor (okadaic acid) and a cAMP-dependent protein kinase inhibitor (H-8). Second, distinct isotypes of RII in membrane-bound and cytosolic compartments of parietal cells will be characterized and the protein kinase systems affecting these proteins will be delineated. Third, the presence of isotypes of RII in distinct subcellular compartments will be established using subcellular fractionation and immunocytochemistry. Fourth, RII protein phosphatase activities in membrane fractions will be isolated and characterized as focal points for histamine action. Fifth, since the effects of cAMP on parietal cell function appear to be in part mediated through activities other than cAMP-dependent protein kinase, the association of RII with other cellular proteins and, in particular, with components of the cytoskeleton will be investigated. Sixth, since secretion by parietal cells is stimulated by both cAMP-dependent and cAMP-independent mechanisms, the common features of histaminergic and cholinergic stimulation will be investigated to determine whether protein phosphatase activities might represent a common mediator. These investigations should elucidate some of the basic mechanisms by which the secretion of acid from the parietal cell is controlled. Expansion of our understanding of the intracellular processes responsible for acid secretion may lead to new methods for the treatment of peptic ulcer disease and other secretory disorders.

消化性溃疡的症状包括： 医疗投诉 然而，虽然有效的药物治疗 靶向H2-组胺受体的药物在过去十年中已经发展， 负责酸分泌的细胞内机制仍然难以捉摸。 cAMP作为组胺刺激的酸分泌介质的作用 壁细胞是公认的。 然而，这些机制实际上 负责将cAMP信号翻译成生理信号。 分泌的反应仍然知之甚少。 我们最近发现 II型调节亚基的一个亚类的去磷酸化， cAMP依赖性蛋白激酶可能是加工中的关键步骤， 壁细胞中的cAMP信号 我们的工作假设是 cAMP可以通过激酶和磷酸化酶的协调激活起作用。 磷酸酶活性 这一假设的内在信念是， cAMP依赖性蛋白激酶激活II调节亚基（RII） cAMP依赖性蛋白激酶作为cAMP传感器蛋白。 这项建议会 主要研究方向有五个：第一，蛋白质的作用 磷酸酶和激酶活性介导组胺刺激 RII的原位磷酸化将在存在以下物质的情况下进行评估： 蛋白磷酸酶抑制剂（冈田酸）和cAMP依赖性 蛋白激酶抑制剂（H-8）。 其次，RII的不同同种型在 壁细胞的膜结合和胞质隔室将被 表征，并且影响这些蛋白质的蛋白激酶系统将 被描绘出来。 第三，RII的同种型在不同的细胞中的存在， 亚细胞区室将使用亚细胞 分级分离和免疫细胞化学。 四、RII蛋白磷酸酶 膜组分中的活性将被分离并表征为 组胺作用的焦点。 第五，由于cAMP对 壁细胞的功能似乎部分通过活动介导， 除了cAMP依赖性蛋白激酶外，RII与其他 细胞蛋白质，特别是细胞骨架的成分 将进行调查。 第六，由于壁细胞的分泌是 在cAMP依赖性和cAMP非依赖性机制的刺激下， 组胺能和胆碱能刺激的共同特征是 研究以确定蛋白磷酸酶活性是否可能 代表一个共同的调解人。 这些调查应阐明一些 壁细胞分泌酸的基本机制 是被控制的 扩展了我们对细胞内 负责酸分泌的过程可能会导致新的方法， 治疗消化性溃疡和其他分泌失调。