Insulin inhibits the ubiquitin-dependent degrading activity of the 26S proteasome
Insulin inhibits the ubiquitin-dependent degrading activity of the 26S proteasome
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DOI：
10.1210/en.141.7.2508
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发表时间：
2000-07-01
期刊：
影响因子：
4.8
通讯作者：
Duckworth, WC
中科院分区：
文献类型：
--
作者：
Bennett, RG;Hamel, FG;Duckworth, WC
胰岛素的主要代谢作用是抑制细胞蛋白水解，但涉及的蛋白水解系统尚不清楚。组织有多种蛋白水解系统，包括ATP和泛素依赖的蛋白酶体途径。胰岛素在体外和培养细胞中对这一途径的影响进行了研究。胰岛素对网状细胞提取物的ATP和泛素依赖性溶菌酶降解的抑制作用超过90%，呈剂量依赖性（TC50类似于50 nM）。胰岛素不降低泛素与溶菌酶的结合，本身也不与泛素结合。在HepG2细胞中，胰岛素使泛素偶联物积累增加80%。26S蛋白酶体与细胞内蛋白酶胰岛素降解酶（IDE）之间的关系，通过一种旨在富集26S蛋白酶体的纯化方案进行了检测。通过几个色谱步骤检测IDE活性和与蛋白酶体的免疫反应性。甘油梯度分析显示IDE与20S蛋白酶体共沉淀，也可能与26S蛋白酶体共沉淀。当用胰岛素处理样品时，蛋白酶体相关的IDE被置换。这些结果表明胰岛素至少部分通过降低泛素介导的蛋白酶体活性来调节蛋白质分解代谢，并为胰岛素的作用提供了新的靶点。从蛋白酶体中置换IDE为胰岛素的作用提供了一种机制。
A major metabolic effect of insulin is inhibition of cellular proteolysis, but the proteolytic systems involved are unclear. Tissues have multiple proteolytic systems, including the ATP- and ubiquitin-dependent proteasome pathway. The effect of insulin on this pathway was examined in vitro and in cultured cells. Insulin inhibited ATP- and ubiquitin-dependent lysozyme degradation more than 90% by reticulocyte extract, in a dose-dependent manner (TC50 similar to 50 nM). Insulin did not reduce the conjugation of ubiquitin to lysozyme and was not itself ubiquitin-conjugated. In HepG2 cells, insulin increased ubiquitin-conjugate accumulation 80%. The association between the 26S proteasome and an intracellular protease, the insulin-degrading enzyme (IDE), was examined by a purification scheme designed to enrich for the 26S proteasome. Copurification of IDE activity and immunoreactivity with the proteasome were detected through several chromatographic steps. Glycerol gradient analysis revealed cosedimentation of IDE with the 20S proteasome and possibly with the 26S proteasome. The proteasome-associated IDE was displaced when the samples were treated with insulin. These results suggest that insulin regulates protein catabolism, at least in part, by decreasing ubiquitin-mediated proteasomal activity, and provides a new target for insulin action. The displacement of IDE from the proteasome provides a mechanism for this insulin action.