Impact of Modulated Exocyst Activity on Glut4 Trafficking in Metabolic Tissues

调节外囊活性对代谢组织中 Glut4 运输的影响

基本信息

批准号：
10252773
负责人：
Noemi Polgar
金额：
$ 20.44万
依托单位：
UNIVERSITY OF HAWAII AT MANOA
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-08-01 至 2024-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10252773
关键词：
Adipocytes Adipose tissue Affect Alleles American Area Blood Glucose Cell Line Cell membrane Cells Centers of Research Excellence Complex Defect Diabetes Mellitus Diabetic mouse Disease Exocytosis Future GLUT 4 protein Generations Genetic study Glucose Transporter Glycogen Goals Hyperglycemia Injections Instruction Insulin Insulin Resistance Insulin deficiency Kinetics Knockout Mice Label Lead LoxP-flanked allele Mediating Metabolic Metabolic Diseases Modeling Molecular Molecular Genetics Mouse Strains Mus Muscle Muscle Fibers Myoblasts Non-Insulin-Dependent Diabetes Mellitus Outcome Pathogenesis Phenotype Phosphorylation Physiological Plasmids Play Regulation Reporting Research Role SLC2A1 gene Secretory Vesicles Skeletal Muscle Specificity Strategic Planning Surface System Tamoxifen Testing Tetracyclines Tissues Transgenes Transgenic Mice United States National Institutes of Health Validation Vesicle base blood glucose regulation cell type design diabetic gene delivery system glucose metabolism glucose tolerance glucose transport glucose uptake improved in vivo insulin signaling insulin tolerance kidney cell knock-down knockout animal mouse model novel overexpression protein complex protein degradation protein transport rab GTP-Binding Proteins recombinase-mediated cassette exchange response therapeutic target trafficking

项目摘要

Abstract Type-2 diabetes mellitus (T2DM) is a metabolic disorder characterized by hyperglycemia, relative insulin deficiency and insulin resistance. Insulin resistant cells show defects in insulin-induced exocytosis of the glucose transporter GLUT4, affecting glucose uptake. The exocyst, a highly conserved eight protein trafficking complex was identified in cultured adipocytes as a key regulator of GLUT4 trafficking in response to insulin. However, it is not known if the molecular mechanisms through which the exocyst orchestrates exocytosis of this glucose transporter in adipocytes are conserved in other, insulin-responsive tissues, such as the skeletal muscle. Skeletal muscle cells are responsible for the vast majority of insulin-induced glucose uptake, therefore investigating glucose transporter trafficking in these cells will be key to better understanding the mechanism of glucose homeostasis. Sec10 is a central subunit of the exocyst complex, and its overexpression can lead to overall increased exocyst activity, while its knockdown leads to protein degradation of several of the other subunits. We have recently generated a new transgenic mouse to analyze exocyst-regulated cellular trafficking in vivo: the first mouse strain with a conditional Sec10 allele. This unique model enables us to investigate the consequences of tissue-specific Sec10 inactivation by facilitating generation of adipose and muscle-specific conditional Sec10 knockout animals. We hypothesize that in vivo modulation of the exocyst complex can independently regulate GLUT4 exocytosis and glucose uptake in insulin-responsive cells and tissues, affecting the diabetic phenotype. To test this hypothesis, we propose the following Specific Aims: (1) Determine if the exocyst mediated regulatory mechanism of insulin-induced GLUT4 exocytosis is conserved in skeletal muscle cells. (2) Test for defects in glucose homeostasis in vivo using adipocyte and skeletal muscle-specific Sec10 knockout mouse strains. (3) Determine T2DM-associated metabolic defects in mice can be ameliorated by increasing GLUT4 exocytosis with a novel tissue-specific in vivo gene delivery system. The anticipated outcome of this project is the first in vivo evidence of the exocyst's role in regulating insulin-induced glucose uptake, and verification whether this regulatory mechanism is conserved in muscle tissues as well. These studies will also show if the exocyst, and other molecules that directly control GLUT4 exocytosis, are potential therapeutic targets for insulin resistance and diabetes.

抽象的 2 型糖尿病 (T2DM) 是一种代谢性疾病，其特征是高血糖、相对胰岛素缺乏和胰岛素抵抗。胰岛素抵抗细胞在胰岛素诱导的胞吐作用中表现出缺陷葡萄糖转运蛋白 GLUT4，影响葡萄糖摄取。外囊，高度保守的八种蛋白质运输在培养的脂肪细胞中，复合物被鉴定为响应胰岛素的 GLUT4 运输的关键调节因子。然而，尚不清楚胞吐体协调胞吐作用的分子机制是否脂肪细胞中的这种葡萄糖转运蛋白在其他胰岛素响应组织中是保守的，例如骨骼肌肉。骨骼肌细胞负责绝大多数胰岛素诱导的葡萄糖摄取，因此研究这些细胞中的葡萄糖转运蛋白运输将是更好地理解葡萄糖转运蛋白机制的关键葡萄糖稳态。 Sec10 是外囊复合体的中心亚基，其过度表达可导致总体上增加了外囊活性，而其敲低则导致其他几种蛋白的降解亚单位。我们最近培育了一种新的转基因小鼠来分析外囊调节的细胞运输体内：第一个具有条件 Sec10 等位基因的小鼠品系。这种独特的模型使我们能够研究通过促进脂肪和肌肉特异性的生成，组织特异性 Sec10 失活的后果有条件的Sec10基因敲除动物。我们假设外囊复合物的体内调节可以独立调节胰岛素反应细胞和组织中的 GLUT4 胞吐作用和葡萄糖摄取，影响糖尿病表型。为了检验这一假设，我们提出以下具体目标： (1) 确定是否外囊介导的胰岛素诱导的 GLUT4 胞吐作用的调节机制在骨骼肌中是保守的细胞。 (2) 使用脂肪细胞和骨骼肌特异性 Sec10 测试体内葡萄糖稳态缺陷基因敲除小鼠品系。 (3) 确定小鼠 T2DM 相关代谢缺陷可以通过以下方式改善通过新型组织特异性体内基因传递系统增加 GLUT4 胞吐作用。预计的该项目的成果是外囊在调节胰岛素诱导的葡萄糖中作用的第一个体内证据摄取，并验证这种调节机制在肌肉组织中是否也保守。这些研究还将表明外囊和其他直接控制 GLUT4 胞吐作用的分子是否具有潜在的潜力胰岛素抵抗和糖尿病的治疗目标。