ROLES OF OST3/6 THIOL OXIDOREDUCTASES IN N-LINKED PROTEIN GLYCOS/REDOX HOMEOSTAS

OST3/6 硫醇氧化还原酶在 N-连接蛋白糖/氧化还原稳态中的作用

• 批准号: 8168314
• 负责人: Dmitri Fomenko
• 金额: $ 14.17万
• 依托单位: UNIVERSITY OF NEBRASKA LINCOLN
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-08-01 至 2011-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8168314
• 关键词: Address; Biological Assay; Biological Process; Brain Diseases; Colorectal; Complex; Computer Retrieval of Information on Scientific Projects Database; Enzymes; Eukaryota; Family; Funding; Gene Expression; Goals; Grant; Homeostasis; Human; Institution; Knock-out; Knockout Mice; Link; Malignant Neoplasms; Malignant neoplasm of pancreas; Mammalian Cell; Mediating; Membrane; Mental Retardation; Metastatic Prostate Cancer; Methods; Modeling; Modification; N33 gene; Oxidation-Reduction; Oxidative Stress; Oxidoreductase; Process; Protein Deficiency; Protein Glycosylation; Proteins; Regulation; Research; Research Personnel; Resources; Role; Saccharomyces cerevisiae; Series; Signal Transduction; Source; Stress; Sulfhydryl Compounds; Thioredoxin; Tumor Suppressor Proteins; United States National Institutes of Health; Yeasts; disulfide bond; dolichyl-diphosphooligosaccharide - protein glycotransferase; glycosylation; link protein; mouse model; protein folding; research study; tool

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The overall goal of this project is to provide understanding of the roles of OST3/6 thiol oxidoreductases in Nlinked protein glycosylation and ER redox homeostasis. Thiol-dependent redox processes are involved in oxidative stress defense, signal transduction, and protein folding, modification and regulation, and are catalyzed by structurally distinct families of enzymes known as thiol oxidoreductases. Numerous ER thiol oxidoreductases involved in protein folding have been characterized; however, the overall machinery of folding and glycosylation remains poorly characterized. The N-linked protein glycosylation in the ER is an essential process and a key step in the control of protein folding in eukaryotes. OST3/6 proteins are abundant ER membrane-linked thioredoxin-fold thiol oxidoreductases involved in the redox control of N-linked protein glycosylation in the oligosaccharyltransferase complex. OST3/6 deficiency is associated with severe protein underglycosylation and ER stress. Homozygous deletion of human OST3/6 like protein, N33, correlates with metastatic prostate cancer and its allelic deletion is associated with human colorectal and pancreatic cancers. This observation suggests a possible tumor suppressor function of N33. In addition, there are two known cases of a natural knockout of N33 in humans which are associated with nonsyndromic mental retardation. In the proposed study, we will systematically characterize the biological function of OST3/6 proteins. The effect of OST3/6 deficiency will be examined with regard to efficiency of protein glycosylation and ER stress. Possible targets of OST3/6 proteins will be identified using thiol-mediated substrate-trapping method and global gene expression analysis. The OST3/6 roles in disulfide bond formation will be addressed in series of thiol oxidoreductase assays. These experiments will be carried in yeast Saccharomyces cerevisiae and mammalian cells. We also would like to develop OST3/6 knockout mouse models. These models will provide tools in a better understanding of the consequences of OST3/6 protein deficiency on cancer and brain disorders and will be useful in the analysis of OST3/6 biological function.

该子项目是利用该技术的众多研究子项目之一 资源由 NIH/NCRR 资助的中心拨款提供。子项目和 研究者 (PI) 可能已从 NIH 的另一个来源获得主要资金， 因此可以在其他 CRISP 条目中表示。列出的机构是 对于中心来说，它不一定是研究者的机构。 该项目的总体目标是了解 OST3/6 硫醇氧化还原酶在 Nlinked 中的作用 蛋白质糖基化和 ER 氧化还原稳态。硫醇依赖性氧化还原过程参与 氧化应激防御、信号转导、蛋白质折叠、修饰和调节等 由结构不同的酶家族（称为硫醇氧化还原酶）催化。大量的ER硫醇 参与蛋白质折叠的氧化还原酶已得到表征；然而，折叠的整体机械 糖基化的特征仍然很少。内质网中的 N 连接蛋白糖基化是必不可少的 过程和控制真核生物蛋白质折叠的关键步骤。 OST3/6 蛋白在 ER 中含量丰富 膜连接硫氧还蛋白折叠硫醇氧化还原酶参与 N 连接蛋白的氧化还原控制 寡糖转移酶复合物中的糖基化。 OST3/6 缺陷与严重的蛋白质相关 糖基化和内质网应激下。人类 OST3/6 样蛋白 N33 的纯合缺失与 转移性前列腺癌及其等位基因缺失与人类结直肠癌和胰腺癌相关。 这一观察结果表明 N33 可能具有肿瘤抑制功能。另外，还有两个已知的案例 人类中 N33 的自然敲除与非综合征性智力低下有关。在 在拟议的研究中，我们将系统地表征OST3/6蛋白的生物学功能。的效果 将检查 OST3/6 缺陷的蛋白质糖基化效率和 ER 应激。可能的 将使用硫醇介导的底物捕获方法和全局基因来鉴定 OST3/6 蛋白的靶标 表达分析。 OST3/6 在二硫键形成中的作用将在一系列硫醇中得到解决 氧化还原酶测定。这些实验将在酿酒酵母和哺乳动物中进行 细胞。我们还想开发 OST3/6 敲除小鼠模型。这些模型将提供工具 更好地了解 OST3/6 蛋白缺乏对癌症和脑部疾病的影响以及意愿 可用于OST3/6生物学功能的分析。