Cysteine String Protein and Acute Pancreatitis

半胱氨酸串蛋白与急性胰腺炎

• 批准号: 6558604
• 负责人: GUY E GROBLEWSKI
• 金额: $ 14.33万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-03-01 至 2005-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6558604
• 关键词: acinar cell; adenosinetriphosphatase; antisense nucleic acid; cysteine; cytotoxicity; enzyme activity; gene induction /repression; heat shock proteins; laboratory rat; lysozyme; membrane activity; molecular chaperones; pancreatitis; protein engineering; protein structure function; proteolysis; secretion; tissue /cell culture; zymogens

DESCRIPTION (provided by applicant): Acute pancreatitis is an inflammatory disease that is triggered by the premature activation of proteolytic zymogens in acinar cells. As a protective mechanism, zymogens are synthesized as inactive pro-enzymes and packaged in specialized secretory granules destined for exocytosis into the pancreatic duct. During pancreatitis, proteolytic zymogens are prematurely activated in acinar cells by aberrantly mixing with hydrolases present in lysosomes. The mixing of lysosomal hydrolases with zymogens occurs in large cytoplasmic vacuoles as a result of pathogenic alterations in acinar cell membrane trafficking. It is well established that the stress-induced expression of heat shock protein-70 (HSP70) in acini provides a natural protective effect against the pathogenic mixing of lysosomal and secretory granule membrane compartments. This proposal is designed to elucidate the mechanism by which HSP70 inhibits the intra-acinar cell activation of zymogens in preventing the on-set of acute pancreatitis. HSP70 is a molecular chaperone in the cytosol that requires a co-chaperone protein to stimulate ATPase activation and substrate binding. We have 1) identified an HSP70 co-chaperone protein called cysteine string protein (CSP) in acinar cells, 2) localized this membrane associated molecule throughout the secretory pathway and 3) developed the TAT-fusion protein system to manipulate CSP expression in acinar cells and thereby study its function. We hypothesize that CSP mediates the protective effects of HSP70 against pancreatitis by targeting HSPT0 chaperone activity from the cytosol to the secretory pathway. In Specific Aim 1, mutant forms of CSP that no longer anchor to secretory granule membranes will be overexpressed acinar cells to determine the importance of CSP in targeting HSP70 to these organelles during pancreatitis. In Specific Aim II, short interference RNAs will be used to inhibit CSP expression in pancreatic Iobules and the protective effects of HSP70 on secretagogue-induced acinar cell damage will be evaluated. These studies will allow us to directly evaluate the role of CSP in the secretory pathway of acini under physiological and pathophysiological conditions.

描述（由申请人提供）：急性胰腺炎是一种炎症性疾病，是由腺泡细胞中蛋白水解Zymogen的过早激活引起的。作为一种保护性机制，将酶基因合成为无活性的促酶，并包装在原始的分泌颗粒中，这些分泌颗粒原定为胞吞作用到胰管中。在胰腺炎期间，通过与溶酶体中存在的水解酶异常混合，在腺泡细胞中过早地激活蛋白水解酶。溶酶体水解酶与酶原的混合在大细胞质液泡中发生，这是由于腺泡细胞膜运输的致病性改变。众所周知，ACINI中应力诱导的热休克蛋白-70（HSP70）的表达具有自然保护作用，以抵抗溶酶体和分泌颗粒膜室的致病性混合。该建议旨在阐明HSP70抑制酶内细胞激活Zymogen的机制，以防止急性胰腺炎的现场。 HSP70是细胞质中的分子伴侣，需要辅助蛋白刺激ATPase激活和底物结合。我们有1）在腺泡细胞中鉴定出一种称为半胱氨酸弦蛋白（CSP）的HSP70共伴酮蛋白，2）在整个分泌途径中将这种膜相关的分子定位，并且3）开发了TAT融合蛋白系统，以在腺泡细胞中操纵CSP表达，从而研究其功能。我们假设CSP通过将HSPT0伴侣活性从细胞质到分泌途径介导HSP70对胰腺炎的保护作用。在特定的目标1中，不再锚定向分泌颗粒膜的CSP的突变形式将被过表达腺泡细胞，以确定CSP在胰腺炎期间将CSP靶向这些细胞器的重要性。在特定的目标II中，将使用短干扰RNA来抑制胰腺iObules中的CSP表达，并评估Hsp70对促分泌物诱导的腺泡细胞损伤的保护作用。这些研究将使我们能够直接评估CSP在生理和病理生理条件下ACINI的分泌途径中的作用。