MODULATION OF CALPASTATIN-CALPAIN INTERACTIONS BY ETHANOL

乙醇对钙蛋白酶抑制素-钙蛋白酶相互作用的调节

• 批准号: 6288658
• 负责人: PAOLO B DE PETRILLO
• 金额: --
• 依托单位: NATIONAL INSTITUTE ON ALCOHOL ABUSE AND ALCOHOLISM
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6288658
• 关键词: PC12 cells; adolescence (12-20); alcoholism /alcohol abuse; bile obstruction; biopsy; blood chemistry; calpain; cholelithiasis; chronic disease /disorder; cytotoxicity; drug withdrawal; endopeptidases; enzyme inhibitors; ethanol; hepatotoxin; human middle age (35-64); human subject; immunochemistry; intermolecular interaction; interview; jaundice; liver disorder diagnosis; male; neurotoxicology; pancreas neoplasms; phosphorylation; posttranslational modifications; protein binding; protein kinase C; protein sequence; young adult human (21-34)

Exposure of PC12 cells to ethanol results in large changes in calcium- stimulated protease activities. Since these proteases are critical modulators of both neurotransmitter release as well as cellular toxicity and death, we are exploring the hypothesis that ethanol- mediated neural toxicity may result from calcium-activated protease dysregulation. Calpastatin, an inhibitor of calpain, is an acidic, hydrophobic protein which interacts with the hydrophobic active site(s) of mu- and m-calpains. A series of post-translational modifications of calpastatin have been described which alter the binding affinity to calpains, among them, PKC-mediated phosphorylations. Using a PC12 model, we are examining the effects of ethanol exposure and withdrawal on protein-protein interactions. Because of the hydrophobic nature of calpastatin-calpain interactions, we examined the possibility that ethanol might modify protease-calpastatin (inhibitor) complex stability. We found that exposure of PC12 cells to ethanol results in an increase in the molecular weight of calpain and calpastatin- containing protein complexes, and that this is associated with a change in protease activity. We are extending these observations by use of immunocytochemical techniques, where the nature of these complexes and their cellular locations will be examined. - alcoholism, neurotoxicity, PC12, post-translational modifications, protein-protein interactions, calpain, calpastatin, proteases

PC12细胞暴露在乙醇中会导致钙刺激的蛋白酶活性发生很大变化。由于这些酶是神经递质释放以及细胞毒性和死亡的关键调节器，我们正在探索一种假设，即乙醇介导的神经毒性可能是由于钙激活的蛋白酶失调所致。钙调蛋白是一种钙蛋白酶的抑制剂，是一种酸性的疏水蛋白，它与u-和m-钙蛋白酶的疏水活性部位(S)相互作用。已经描述了一系列钙调蛋白的翻译后修饰，这些修饰改变了与钙蛋白酶的结合亲和力，其中包括PKC介导的磷酸化。利用PC12模型，我们正在研究酒精暴露和戒断对蛋白质-蛋白质相互作用的影响。由于calastatin-calain相互作用的疏水性，我们研究了乙醇可能改变蛋白酶-calastatin(抑制剂)复合体稳定性的可能性。我们发现，PC12细胞暴露在乙醇中会导致钙蛋白酶和含有钙蛋白酶的蛋白质复合体的分子量增加，这与蛋白酶活性的变化有关。我们正在通过使用免疫细胞化学技术来扩展这些观察，其中将检查这些复合体的性质和它们的细胞位置。-酒精中毒、神经毒性、PC12、翻译后修饰、蛋白质-蛋白质相互作用、钙蛋白酶、钙蛋白酶、蛋白酶