Adaptive Regulation in 3T3-L1 Adipocytes

3T3-L1 脂肪细胞的适应性调节

• 批准号: 7252001
• 负责人: SUSAN Cooke FROST
• 金额: $ 20.69万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-05-01 至 2010-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7252001
• 关键词: Ablation; Address; Adipocytes; Adipose tissue; Affect; Animals; Biochemical; Biological Models; Carbon; Cell membrane; Cells; Controlled Environment; Data; Detergents; Disease; Environment; Exhibits; Fluorescence Resonance Energy Transfer; GLUT-1 protein; Glucose; Glucose Transporter; Immunofluorescence Microscopy; Intracellular translocation; Knockout Mice; Lipids; Measures; Membrane; Membrane Microdomains; Metabolism; Movement; Mus; Numbers; Nutrient; Phospholipids; Physiological; Play; Production; Protein Biosynthesis; Protein Overexpression; Proteins; Proteomics; Rate; Recruitment Activity; Regulation; Relative (related person); Research Personnel; Resistance; Role; SLC2A1 gene; Techniques; Time; deprivation; glucose metabolism; glucose transport; glucose uptake; human SLC2A1 protein; novel; programs; protein expression; reconstitution; response

DESCRIPTION (provided by applicant): Most cells depend on glucose as their primary substrate for energy production and carbon storage. In addition, glucose serves a role in regulating the expression of proteins involved in its own metabolism. These include the proteins involved in glucose transport, which is often the rate-limiting step in its metabolism. Animal studies suggest that basal glucose uptake in adipose tissue is directly related to the expression and activity of GLUT1, the "constitutive" glucose transporter. We have focused specifically on the regulation of GLUT1 by glucose, itself. To accomplish this, we have used 3T3-L1 adipocytes which afford characterization over extended time in a controlled environment. In cells deprived of glucose, we have shown that transport activity increases by 20-fold, not an insignificant change. The relevance to this observation is that a 4-fold change is observed over the physiological extremes of circulating glucose. Because GLUT1 expression does not change, this led to the hypothesis that GLUT1 is activated in response to glucose deprivation. We now know that a proportion of the GLUT1 pool in plasma membranes resides in lipid rafts. This percentage increases in response to deprivation. New data demonstrate that GLUT1 reconstituted from lipid rafts isolated from glucose-deprived cells exhibits higher "intrinsic activity" than that observed from control cells. Interestingly, we are able to demonstrate a specific interaction between GLUT1 and stomatin, a protein that exists almost exclusively in the lipid raft fraction. Glucose-deprivation increases this interaction, which infers that stomatin and GLUT1 interact within lipid rafts. Together, these data suggest that the environment of lipid rafts plays a role in GLUT1 function. These novel observations have led to new directions in our attempts to understand the nutrient-dependent control of glucose transport. In Specific Aim 1, we focus on the targeting of GLUT1 to lipid rafts. In Specific Aim 2, we explore the role of lipids in rafts on transport function. In Specific Aim 3, we identify proteins unique to lipid rafts using a novel proteomics approach which may regulate GLUT1. Finally in Specific Aim 4, we use the stomatin knockout mouse to determine the role of stomatin on GLUT1 targeting and function.

描述（由申请人提供）：大多数细胞依赖葡萄糖作为能量产生和碳储存的主要底物。此外，葡萄糖在调节参与其自身代谢的蛋白质的表达中发挥作用。其中包括参与葡萄糖转运的蛋白质，葡萄糖转运通常是其代谢的限速步骤。动物研究表明，脂肪组织中的基础葡萄糖摄取与“组成型”葡萄糖转运蛋白 GLUT1 的表达和活性直接相关。我们特别关注葡萄糖本身对 GLUT1 的调节。为了实现这一目标，我们使用了 3T3-L1 脂肪细胞，它可以在受控环境中长时间提供表征。我们发现，在缺乏葡萄糖的细胞中，转运活性增加了 20 倍，这并不是一个微不足道的变化。与该观察结果相关的是，在循环葡萄糖的生理极值范围内观察到 4 倍的变化。由于 GLUT1 表达不会改变，这导致了 GLUT1 因葡萄糖剥夺而被激活的假设。我们现在知道质膜中 GLUT1 库的一部分存在于脂筏中。这一比例因贫困而增加。新数据表明，从葡萄糖剥夺细胞中分离出的脂筏重建的 GLUT1 表现出比对照细胞观察到的更高的“内在活性”。有趣的是，我们能够证明 GLUT1 和造口素（一种几乎只存在于脂筏部分中的蛋白质）之间的特定相互作用。葡萄糖剥夺会增加这种相互作用，这表明stomatin和GLUT1在脂筏内相互作用。总之，这些数据表明脂筏环境在 GLUT1 功能中发挥作用。这些新颖的观察结果为我们尝试了解葡萄糖转运的营养依赖性控制提供了新的方向。在具体目标 1 中，我们重点关注 GLUT1 靶向脂筏。在具体目标 2 中，我们探讨了筏中脂质对运输功能的作用。在具体目标 3 中，我们使用一种可能调节 GLUT1 的新型蛋白质组学方法鉴定了脂筏特有的蛋白质。最后在具体目标 4 中，我们使用造口素基因敲除小鼠来确定造口素对 GLUT1 靶向和功能的作用。

项目成果

Expression and activity of carbonic anhydrase IX is associated with metabolic dysfunction in MDA-MB-231 breast cancer cells.

• DOI: 10.1080/07357900802653464
• 发表时间: 2009-07
• 期刊: Cancer investigation
• 影响因子: 2.4
• 作者: Li Y;Wang H;Oosterwijk E;Tu C;Shiverick KT;Silverman DN;Frost SC
• 通讯作者: Frost SC

Role of zinc in catalytic activity of carbonic anhydrase IX.

锌在碳酸酐酶 IX 催化活性中的作用。

• DOI: 10.1016/j.abb.2012.03.017
• 发表时间: 2012
• 期刊: Archives of biochemistry and biophysics
• 影响因子: 3.9
• 作者: Tu,Chingkuang;Foster,Lauren;Alvarado,Andrea;McKenna,Robert;Silverman,DavidN;Frost,SusanC
• 通讯作者: Frost,SusanC