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起关键作用。因此，它们与细胞膜的组织和连接对红细胞生理学具有根本的重要性。我们最近的证据表明，dematin执行其功能，连接血影蛋白肌动蛋白连接到细胞膜。在这里，我们提出，dematin链接血影蛋白肌动蛋白连接到细胞膜通过葡萄糖转运蛋白，GLUT 1，在人类红细胞和通过相关的转运蛋白在小鼠红细胞。我们将通过以下三个目标来检验这一假设。A1：人红细胞中GLUT 1与血影蛋白-肌动蛋白连接的相互作用的表征。缺失dematin的头部结构域可导致小鼠代偿性贫血伴小红细胞增多和球形红细胞增多。我们确定GLUT 1作为人红细胞中dematin和内收蛋白的主要膜受体。我们将确定其相互作用的结构域和关键的序列元素，并通过破坏的GLUT 1-细胞骨架桥的生化手段，评估其对膜的稳定性，从而对人类红细胞的形状功能的影响。A2.小鼠红细胞和脂肪细胞中去毛蛋白和内收蛋白结合受体的鉴定。GLUT 1受体不将血影蛋白-肌动蛋白连接点连接到小鼠红细胞膜。我们预测，dematin和内收蛋白结合到一个新的膜受体（S）在小鼠红细胞。我们将使用一系列生物化学和蛋白质组学方法鉴定这种膜受体。我们将确定dematin和adducin是否结合GLUT 4，胰岛素敏感的葡萄糖转运蛋白，通常发现在脂肪和肌肉细胞，并研究其在脂肪细胞中的定位和回收GLUT 4的作用。这些实验将阐明连接血影蛋白-肌动蛋白连接点的替代受体在小鼠红细胞和脂肪细胞中的作用。A3.测定完全性地马丁缺乏症的影响。由于dematin的核心结构域仍然在头部缺失小鼠中表达，因此在携带全长dematin的纯合基因破坏的小鼠中仍然存在更深刻的血液学和代谢表型的可能性。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GLUT 4运输和糖尿病的潜在发展。总之，这些研究将阐明dematin的头部和核心结构域在红细胞生成和中间代谢中的特定作用。公共卫生相关性：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。本计画将探讨dematin、adducin及葡萄糖转运蛋白在溶血性贫血、糖尿病及肥胖症中形成一种新的膜-细胞骨架桥的作用。