Cytoskeletal Regulation of Erythrocyte Glucose Transporter-1

红细胞葡萄糖转运蛋白 1 的细胞骨架调节

• 批准号: 7741124
• 负责人: Athar H. Chishti
• 金额: $ 38.96万
• 依托单位: UNIVERSITY OF ILLINOIS AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-08-05 至 2013-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7741124
• 关键词: Actins; Adipocytes; Adipose tissue; Anemia; Binding; Biochemical; Biochemical Genetics; Biological Assay; Biological Process; Brain; Cell Shape; Cell membrane; Chemicals; Data; Development; Diabetes Mellitus; Elements; Erythroblasts; Erythrocyte Membrane; Erythrocytes; Erythroid Cells; Erythropoiesis; GLUT-1 protein; GLUT4 gene; Gene Targeting; Genes; Glucose Transporter; Glycophorin C; Heart; Hemolytic Anemia; Human; Immunoprecipitation; Insulin; Knockout Mice; Length; Lesion; Link; Maintenance; Mass Spectrum Analysis; Membrane; Metabolic; Metabolism; Molecular; Mus; Muscle Cells; Mutant Strains Mice; Mutation; Obesity; Phenotype; Physiology; Play; Property; Proteins; Proteomics; Recycling; Regulation; Reporting; Reticulocytes; Role; SLC2A1 gene; Series; Shapes; Skeletal Muscle; Spectrin; Techniques; Testing; Tissues; Vesicle; adducin; base; crosslink; design; mouse model; novel; protein 4.1; public health relevance; receptor; receptor binding; receptor function; research study; trafficking

DESCRIPTION (provided by applicant): Spectrin-actin junctions play critical roles in both the maintenance of red blood cell shape and membrane properties. Their organization and linkage to the cell membrane is therefore of fundamental importance to erythrocyte physiology. Our recent evidence indicates that dematin performs its function by linking spectrin-actin junctions to the cell membrane. Here, we propose that dematin links spectrin-actin junctions to the cell membrane via the glucose transporter, GLUT1, in human erythrocytes and via a related transporter in mouse erythrocytes. We will test this hypothesis with the following three aims. A1: Characterization of GLUT1 interactions with spectrin-actin junctions in human erythrocytes. Deletion of the headpiece domain of dematin results in a compensated anemia with microcytosis and spherocytosis in mice. We identified GLUT1 as the primary membrane receptor for both dematin and adducin in human erythrocytes. We will identify their interacting domains and the critical sequence elements and, through disruption of the GLUT1-cytoskeletal bridge by biochemical means, assess its functional impact on membrane stability and thus the shape of human erythrocytes. A2. Identification of dematin- and adducin-binding receptor(s) in mouse erythrocytes and adipocytes. The GLUT1 receptor does not link spectrin-actin junctions to the membrane in mouse erythrocytes. We predict that dematin and adducin bind to a novel membrane receptor(s) in mouse erythrocytes. We will identify this membrane receptor using a series of biochemical and proteomic approaches. We will determine whether dematin and adducin bind to GLUT4, the insulin-sensitive glucose transporter generally found in adipose and muscle cells, and investigate their role in localization and recycling of GLUT4 in adipocytes. These experiments will clarify the role of alternate receptors that link the spectrin-actin junctions in mouse erythrocytes and adipocytes. A3. Determination of the effects of complete dematin deficiency. As the core domain of dematin is still expressed in headpiece-null mice, there remains the possibility that more profound hematological and metabolic phenotypes develop in mice carrying a homozygous gene disruption of full-length dematin. We propose to generate the latter mouse model and conduct a comprehensive analysis of the hematological and metabolic lesions, focusing particularly on GLUT4 trafficking and the potential development of diabetes. Together, these studies will elucidate the specific role(s) of the headpiece and core domains of dematin in erythropoiesis and intermediary metabolism. PUBLIC HEALTH RELEVANCE: Erythrocyte membrane has served as a paradigm for discovering the function of its proteins in many non-erythroid cells. This project will investigate the role of dematin, adducin, and glucose transporters in the formation of a novel membrane-cytoskeletal bridge with functional implications in hemolytic anemia, diabetes, and obesity.

描述（由申请人提供）：血影蛋白-肌动蛋白连接在维持红细胞形状和膜特性方面发挥着关键作用。因此，它们的组织和与细胞膜的连接对于红细胞生理学至关重要。我们最近的证据表明，脱蛋白通过将血影蛋白-肌动蛋白连接与细胞膜连接来发挥其功能。在这里，我们提出脱蛋白通过人红细胞中的葡萄糖转运蛋白 GLUT1 以及小鼠红细胞中的相关转运蛋白将血影蛋白-肌动蛋白连接与细胞膜连接起来。我们将通过以下三个目标来检验这一假设。 A1：GLUT1 与人红细胞中血影蛋白-肌动蛋白连接相互作用的表征。删除脱蛋白的头状结构域会导致小鼠代偿性贫血，伴有小红细胞增多症和球形红细胞增多症。我们确定 GLUT1 是人红细胞中脱蛋白和内收蛋白的主要膜受体。我们将鉴定它们的相互作用域和关键序列元件，并通过生化手段破坏 GLUT1-细胞骨架桥，评估其对膜稳定性的功能影响，从而评估人类红细胞的形状。 A2。小鼠红细胞和脂肪细胞中脱蛋白和内收蛋白结合受体的鉴定。 GLUT1 受体不会将血影蛋白-肌动蛋白连接与小鼠红细胞膜连接起来。我们预测脱蛋白和内收蛋白与小鼠红细胞中的新型膜受体结合。我们将使用一系列生化和蛋白质组学方法来识别这种膜受体。我们将确定脱蛋白和内收蛋白是否与 GLUT4（通常存在于脂肪和肌肉细胞中的胰岛素敏感性葡萄糖转运蛋白）结合，并研究它们在脂肪细胞中 GLUT4 的定位和再循环中的作用。这些实验将阐明连接小鼠红细胞和脂肪细胞中血影蛋白-肌动蛋白连接的替代受体的作用。 A3。确定完全脱蛋白缺乏的影响。由于脱蛋白的核心结构域仍然在头戴缺失小鼠中表达，因此在携带全长脱蛋白纯合基因破坏的小鼠中仍然有可能出现更深刻的血液学和代谢表型。我们建议生成后一种小鼠模型，并对血液学和代谢病变进行全面分析，特别关注 GLUT4 运输和糖尿病的潜在发展。这些研究将共同​​阐明脱蛋白的头部和核心域在红细胞生成和中间代谢中的具体作用。公众健康相关性：红细胞膜已成为发现其蛋白质在许多非红细胞中的功能的范例。该项目将研究脱蛋白、内收蛋白和葡萄糖转运蛋白在新型膜-细胞骨架桥形成中的作用，该桥对溶血性贫血、糖尿病和肥胖症具有功能意义。