Cellular protein maturation and degradation

细胞蛋白质的成熟和降解

• 批准号: 8486450
• 负责人: Daniel N. Hebert
• 金额: $ 27.48万
• 依托单位: UNIVERSITY OF MASSACHUSETTS AMHERST
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-04-01 至 2016-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8486450
• 关键词: Adaptor Signaling Protein; Address; Albinism; Binding; Biological Assay; Calnexin; Carbohydrates; Cells; Cellular biology; Client; Complement; Complex; Consensus; Cystic Fibrosis; Defect; Degradation Pathway; Development; Diabetes Mellitus; Disease; Dislocations; Endoplasmic Reticulum; Enzymes; Etiology; Family; Glucosyltransferase; Glycoproteins; Goals; Heart Diseases; Homeostasis; Human; In Vitro; Investigation; Knowledge; Lectin; Life; Light; Link; Liquid substance; Liver Cirrhosis; Mammalian Cell; Mannose; Mannosidase; Membrane Proteins; Methodology; Modification; Molecular Chaperones; N-Glycosylation Site; Names; Obesity; Pathology; Pathway interactions; Play; Polysaccharides; Process; Property; Proteins; Pulmonary Emphysema; Quality Control; Recruitment Activity; Research; Research Proposals; Role; Series; Side; Signal Transduction; Sorting - Cell Movement; Structure; System; Testing; Therapeutic Agents; UGT1A1 gene; Uridine Diphosphate Glucose; base; biological adaptation to stress; calreticulin; design; insight; novel; overexpression; protein folding; protein function; protein misfolding; protein protein interaction; receptor binding; reconstitution

DESCRIPTION (provided by applicant): The overarching goals of this project are to define mechanisms of protein quality control in the mammalian endoplasmic reticulum (ER). The proposed research will utilize well-defined mammalian cell systems to determine how aberrant secretory cargo is selected and targeted for destruction and how organization of the quality control machinery within the ER coordinates protein transit and maturation. The quality control process is comprised of a series of interlocking steps that involve making critical choices on whether to transport a substrate or divert it for ER retention or degradation. The vast majority of proteins that traverse the mammalian secretory pathway receive N-linked glycans that are added shortly after a consensus N- glycosylation site emerges into the ER lumen. The composition of N-glycans is dynamic because they act as signals to recruit factors that assist the protein maturation and quality control processes. N-glycans support recruitment of the carbohydrate binding chaperones calnexin and calreticulin, which bind to monoglucosylated side chains. Persistent lectin chaperone binding is directed by UDP-glucose: glycoprotein glucosyltransferase (UGT1). Although UGT1 has been studied in reconstituted systems, the cellular role of this critical ER factor is poorly defined. Mannose trimming is proposed to be involved in sorting aberrant proteins to the ER-associated degradation (ERAD) pathway as inhibition of mannosidase activity stabilizes ERAD substrates. However, many questions still remain about the role of mannose trimming in quality control. While the over expression of the ER mannosidase-like protein EDEM1 accelerates the turnover of glycosylated ERAD substrates, its role in the quality control and sorting processes is unclear. Our results indicate that EDEM1 serves as a central link to deliver defective cargo to an ERAD dislocation complex by possessing bipartite binding properties. Finally, the exquisite spatial organization of the ER contributes to the efficiencies by which the ER temporally coordinates processes including protein maturation, quality control and ERAD. Tetratricopeptide repeat (TPR) domains are universal adaptor motifs that support protein-protein interactions involved in post-translational translocation and the formation of chaperone complexes within the cell. We have recently discovered a family of ER resident TPR-rich proteins that we propose to play a role in ER organization. This research proposal has three aims: (1) to define how UGT1 participates in ER quality control in a cellular context; (2) to elucidate the role and mechanism of action for EDEM1 in the ERAD process; and (3) to determine how novel TPR-containing proteins function in ER organization and homeostasis. A deeper knowledge of the processes in the early secretory pathway has widespread implications for understanding basic cell biology and the etiology of a range of human maladies as a growing number of diseases states are caused by defects in protein maturation and ER homeostasis.

描述（由申请人提供）：本项目的首要目标是确定哺乳动物内质网（ER）中蛋白质质量控制的机制。拟议的研究将利用明确的哺乳动物细胞系统来确定如何选择异常分泌货物并将其作为破坏目标，以及ER内的质量控制机制如何协调蛋白质运输和成熟。质量控制过程由一系列连锁步骤组成，涉及对是否运输底物或将其转移用于ER保留或降解进行关键选择。绝大多数 穿过哺乳动物分泌途径的蛋白质接收在共有N-糖基化位点出现到ER腔中后不久添加的N-连接聚糖。N-聚糖的组成是动态的，因为它们作为信号来募集有助于蛋白质成熟和质量控制过程的因子。N-聚糖支持碳水化合物结合分子伴侣钙连接蛋白和钙网蛋白的募集，其与单葡萄糖基化侧链结合。持续的凝集素伴侣结合由UDP-葡萄糖：糖蛋白葡糖基转移酶（UGT 1）指导。虽然UGT 1已在重建系统中进行了研究，但这种关键ER因子的细胞作用尚不清楚。甘露糖修剪被认为参与了ER相关降解（ERAD）途径的异常蛋白的分选，因为甘露糖苷酶活性的抑制稳定了ERAD底物。然而，关于甘露糖修剪在质量控制中的作用仍然存在许多问题。虽然ER甘露聚糖酶样蛋白EDEM 1的过表达加速了糖基化ERAD底物的周转，但其在质量控制和分选过程中的作用尚不清楚。我们的研究结果表明，EDEM 1作为一个中心环节，提供有缺陷的货物ERAD位错复合体具有二分结合性能。最后，ER的精致空间组织有助于ER在时间上协调包括蛋白质成熟、质量控制和ERAD在内的过程的效率。四肽重复序列（TPR）结构域是支持蛋白质-蛋白质相互作用的通用衔接基序，所述蛋白质-蛋白质相互作用参与翻译后易位和细胞内伴侣复合物的形成。我们最近发现了一个家庭的ER居民TPR丰富的蛋白质，我们建议在ER组织中发挥作用。这项研究计划有三个目的：（1）确定UGT 1如何参与细胞环境中的ER质量控制;（2）阐明EDEM 1在ERAD过程中的作用和作用机制;（3）确定新的含TPR蛋白如何在ER组织和稳态中发挥作用。对早期分泌途径中的过程的更深入的了解对于理解基础细胞生物学和一系列人类疾病的病因学具有广泛的意义，因为越来越多的疾病状态是由蛋白质成熟和ER稳态的缺陷引起的。