CHC22 CLATHRIN FUNCTION IN HUMAN GLUCOSE METABOLISM

CHC22 网格蛋白在人体葡萄糖代谢中的功能

• 批准号: 7922790
• 负责人: Frances M. Brodsky
• 金额: $ 38.63万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-20 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7922790
• 关键词: Accounting; Acute; Address; Adipocytes; Affect; Amino Acid Sequence; Animal Model; Autopsy; Behavior; Biochemical; Biochemistry; Biological Assay; Blood Circulation; Cell membrane; Cell surface; Cells; Cellular biology; Chromosomes, Human, Pair 22; Chromosomes, Human, Pair 5; Clathrin; Clathrin Heavy Chains; Data; Development; Diabetes Mellitus; Down-Regulation; Elements; Failure; Fractionation; Frequencies; Functional disorder; GLUT4 gene; Genetic; Genetic Polymorphism; Genetic Screening; Genetic Variation; Glucose; Glucose Transporter; Goals; Habits; Human; Human Characteristics; Human Chromosomes; Incidence; Insecta; Insulin; Insulin Receptor; Insulin Resistance; Laboratories; Lead; Macromolecular Complexes; Mediating; Membrane; Membrane Protein Traffic; Metabolic; Metabolic Diseases; Molecular; Morphology; Mus; Muscle; Muscle Fibers; Names; Non-Insulin-Dependent Diabetes Mellitus; Nutritional; Pathway interactions; Peptide Sequence Determination; Phylogenetic Analysis; Play; Population; Property; Protein Isoforms; Proteins; Recombinants; Regulation; Research; Research Proposals; Resolution; Role; Sampling; Sarcolemma; Signal Transduction; Skeletal Muscle; Source; Structure; Surface; Symptoms; System; Therapeutic; Tissues; Transgenic Mice; Variant; Vertebrates; Vesicle; Work; blood glucose regulation; design; diabetic; diabetic patient; glucose metabolism; glucose uptake; insight; interest; mouse model; novel; nutrition; prevent; research study; response; trafficking; trans-Golgi Network

This proposal focuses on the CHC22 isoform of clathrin, which is preferentially expressed in human skeletal muscle. Recent work from the laboratory indicates that CHC22 is involved in intracellular sequestration of the GLUT4 glucose transporter. Upon insulin stimulation of skeletal muscle and adipocytes, GLUT4 is released from the GLUT4 storage compartment (GSC) in vesicles that fuse with the plasma membrane, where it can import glucose in response to acute metabolic needs. Thus GLUT4 membrane traffic is critical for glucose metabolism and its disruption is associated with type 2 diabetes. The implication of CHC22 clathrin in GLUT4 membrane traffic in humans is interesting because CHC22 is missing from mice and CHC22 expression may therefore account for some of the known differences between glucose metabolism by humans and mice. This proposal aims to characterize the role that CHC22 plays in GLUT4 transport, to determine the molecular mechanisms that control CHC22 function and to explore polymorphisms in CHC22 that would affect its function and thereby contribute to incidence of type 2 diabetes. For Aim 1, basic questions about CHC22-mediated membrane traffic of GLUT4 will be addressed, including molecular details of CHC22 function following insulin stimulation and its distribution in the muscle of diabetic patients. Experiments will compare GLUT4 membrane traffic in humans with its traffic in CHC22-transgenic mice. The presence and role of CHC22 in human adipocytes will also be investigated. For Aim 2, the factors needed for CHC22 to function properly will be established using two assays for its function. Genetic screens will be designed to re-create a functional pathway for CHC22 when it is introduced in mouse cells and to dissect CHC22 function in GSC formation. Aim 3 of this project is to characterize the biochemical properties of CHC22 that mediate its function. CHC22 protein will be purified from both tissue and recombinant sources to determine its morphology and whether it can assemble into macromolecular complexes. CHC22-associated proteins will be identified. Recombinant fragments of CHC22 protein will be expressed and their structure determined at crystallographic resolution. Aim 4 is to determine the distribution of CHC22 polymorphisms within human populations for correlation with incidence of type 2 diabetes. The functional features of common CHC22 variants will be characterized. In addition, phylogenetic analysis of CHC22 will assess whether its function in different species is related to nutrition habits. With these aims the overall goal of this proposal is to further characterize the role that CHC22 plays in GLUT4 transport in order to understand how human glucose metabolism is regulated. While studies of mice have provided major insights into pathways relevant to type 2 diabetes, CHC22 represents a missing species-specific element. The research proposed here will establish novel characteristics of human glucose homeostasis and will enable development of better animal models for studying its dysfunction in diabetes.

该提案重点关注网格蛋白的 CHC22 亚型，该亚型优先在人类骨骼中表达 肌肉。实验室最近的工作表明 CHC22 参与细胞内隔离 GLUT4 葡萄糖转运蛋白。当骨骼肌和脂肪细胞受到胰岛素刺激时，GLUT4 从与质膜融合的囊泡中的 GLUT4 储存室 (GSC) 释放， 它可以输入葡萄糖以满足急性代谢需求。因此 GLUT4 膜流量至关重要 葡萄糖代谢及其破坏与 2 型糖尿病有关。 CHC22的意义 人类 GLUT4 膜运输中的网格蛋白很有趣，因为小鼠中缺少 CHC22，并且 因此，CHC22 表达可能解释了葡萄糖代谢之间的一些已知差异。 由人类和老鼠。该提案旨在描述 CHC22 在 GLUT4 运输中所扮演的角色，以 确定控制 CHC22 功能的分子机制并探索 CHC22 的多态性 这会影响其功能，从而导致 2 型糖尿病的发生。对于目标 1，基本 将解决有关 CHC22 介导的 GLUT4 膜运输的问题，包括分子细节 胰岛素刺激后 CHC22 的功能及其在糖尿病患者肌肉中的分布。 实验将比较人类中的 GLUT4 膜流量与 CHC22 转基因小鼠中的流量。 还将研究 CHC22 在人类脂肪细胞中的存在和作用。对于目标 2，因素 CHC22 正常发挥作用所需的时间将通过两种功能测定来确定。遗传 当 CHC22 被引入小鼠细胞时，屏幕将被设计为重新创建 CHC22 的功能途径 并剖析 CHC22 在 GSC 形成中的功能。该项目的目标 3 是表征生化 CHC22 的特性介导其功能。 CHC22 蛋白将从组织和 重组来源以确定其形态以及是否可以组装成大分子 复合物。 CHC22 相关蛋白将被鉴定。 CHC22蛋白的重组片段将是 在晶体学分辨率下表达并确定其结构。目标 4 是确定 CHC22 多态性在人群中的分布与 2 型发病率的相关性 糖尿病。将描述常见 CHC22 变体的功能特征。此外， CHC22的系统发育分析将评估其在不同物种中的功能是否与营养相关 习惯。出于这些目标，本提案的总体目标是进一步描述 CHC22 所发挥的作用 GLUT4 运输，以了解人类葡萄糖代谢是如何调节的。在研究的同时 小鼠提供了对 2 型糖尿病相关途径的重要见解，CHC22 代表了一个缺失的 物种特异性元素。这里提出的研究将建立人类葡萄糖的新特性 稳态，并将有助于开发更好的动物模型来研究其在糖尿病中的功能障碍。