The mechanisms of glucose-induced catabolite degradation in the yeast Saccharomyces cerevisiae

酿酒酵母中葡萄糖诱导的分解代谢物降解的机制

• 批准号: 235347673
• 负责人: Professor Dr. Dieter H. Wolf
• 金额: --
• 依托单位: Abteilung für Biochemie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2013
• 资助国家: 德国
• 起止时间: 2012-12-31 至 2018-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/235347673?language=en
• 关键词: mechanisms; glucose; induced; catabolite; degradation

The sugar glucose is essential for energy metabolism and the generation of building blocks in cells. Besides this, glucose is a central signalling molecule. It has a strong impact on the entire cell metabolism up to the cell cycle. The consequences of a disturbed glucose metabolism are impressively demonstrated by the widespread human disease diabetes. Besides the regulation of transcription and translation glucose induces the activation and inactivation of enzymes and especially their degradation. In the project applied, the regulation of the two central antagonistic pathways, glycolysis and gluconeogenesis, are analyzed in the eukaryotic model organism yeast. Central to this project is the elucidation of the glucose induced degradation of three key enzymes of gluconeogenesis, fructose-1,6-bisphosphatase, phosphoenolpyruvate carboxykinase and cytoplasmic malate dehydrogenase, by two different degradation pathways: the ubiquitin proteasome system and the vacuole (lysosome). Glucose addition to cells growing for up to 24 hours on a nonfermentable carbon source induces a signalling pathway which leads to degradation of the enzymes via the ubiquitin proteasome pathway. Specific for this degradation pathway is a novel RING E3 ligase complex, the Gid complex, which catalyzes ubiquitination of the enzymes and thus triggers their degradation by the proteasome. The glucose signal induces the rapid synthesis of one of the subunits of the Gid ligase complex, Gid4, which binds to the core complex and thereby stimulates ubiquitination of the gluconeogenic enzymes and their subsequent proteasomal degradation. The aim of this grant proposal is to elucidate the entire signal cascade, which leads to degradation of the enzymes as well as to examine the regulation of Gid4 by proteolysis. In addition, the mechanistic events occurring at the Gid ubiquitin ligase complex during polyubiquitination of the enzymes will be investigated. Glucose addition to cells which have been grown for about 48 to 72 hours on a non-fermentable carbon source until they reach stationary phase induces degradation of the gluconeogenic enzymes in the vacuole (lysosome). Obviously, during this growth phase a switch occurs which channels degradation of the enzymes from proteasomal proteolysis to vacuolar (lysosomal) proteolysis. Preliminary results indicate that the Gid ubiquitin ligase complex is also required for the vacuolar (lysosomal) proteolysis of the enzymes and that this process is related to selective autophagy. Our aim is to understand in which manner the trigger molecule glucose can induce these two different degradation pathways.The studies are not only meant to better understand the regulation of gluconeogenesis but are additionally supposed to shed light on the glucose induced proteolysis of gluconeogenic enzymes by two completely different proteolytic systems, the proteasome and the vacuole.

葡萄糖是能量代谢和细胞生成的基本物质。除此之外，葡萄糖是一个中心信号分子。它对整个细胞代谢乃至细胞周期都有很强的影响。葡萄糖代谢紊乱的后果在广泛的人类疾病糖尿病中得到了令人印象深刻的证明。除了调控转录和翻译外，葡萄糖还能诱导酶的活化和失活，尤其是酶的降解。本项目拟对真核模式生物酵母的糖酵解和糖异生两种主要拮抗途径的调控进行分析。该项目的核心是阐明葡萄糖诱导糖异生的三个关键酶，果糖-1,6-双磷酸酶，磷酸烯醇丙酮酸羧激酶和细胞质苹果酸脱氢酶，通过两种不同的降解途径：泛素蛋白酶体系统和液泡（溶酶体）。将葡萄糖添加到在不可发酵碳源上生长长达24小时的细胞中，可诱导通过泛素蛋白酶体途径导致酶降解的信号通路。这种降解途径的特异性是一种新的环E3连接酶复合物，Gid复合物，它催化酶的泛素化，从而触发蛋白酶体对其的降解。葡萄糖信号诱导Gid连接酶复合体的一个亚基Gid4的快速合成，Gid4与核心复合体结合，从而刺激糖异生酶的泛素化及其随后的蛋白酶体降解。这项拨款提案的目的是阐明导致酶降解的整个信号级联，并检查蛋白质水解对Gid4的调节。此外，在酶的多泛素化过程中，在Gid泛素连接酶复合物上发生的机制事件将被研究。在不可发酵的碳源上生长约48至72小时直至达到固定阶段的细胞中添加葡萄糖可诱导液泡（溶酶体）中糖异生酶的降解。显然，在这个生长阶段发生了一个开关，该开关引导酶的降解从蛋白酶体蛋白水解到液泡（溶酶体）蛋白水解。初步结果表明，Gid泛素连接酶复合体也是酶的空泡（溶酶体）蛋白水解所必需的，并且这一过程与选择性自噬有关。我们的目的是了解触发分子葡萄糖以何种方式诱导这两种不同的降解途径。这些研究不仅是为了更好地理解糖异生的调控，而且还应该揭示糖异生酶在两种完全不同的蛋白质水解系统，蛋白酶体和液泡中的蛋白质水解。