Role of Caveolae in Signaling in Fat Cells

描述（由申请人提供）：脂肪酸是哺乳动物最富含卡路里的燃料来源。脂肪酸被吸收并储存在脂肪细胞中，当其他组织需要时，可以从脂肪细胞中动员出来。过量的循环脂肪酸是胰岛素抵抗和2型糖尿病的近端原因，因此调节其细胞摄取和释放的因素与这些病理状况有潜在的直接联系。适当的燃料平衡需要脂肪酸在细胞表面，即质膜上快速而大量地移动。脂肪细胞质膜中含有大量的称为小洞的结构，这是一种小的膜内陷（“小洞”），富含胆固醇和鞘磷脂，由小洞蛋白-1的表达形成。小泡是一种脂质筏，其特点是不受温和洗涤剂（如脂肪酸）的破坏。缺乏小窝蛋白-1的小鼠具有胰岛素抵抗和脂肪酸代谢异常。我们假设脂肪细胞中的小窝蛋白/小窝在脂肪酸跨质膜运动的调节中发挥作用，并可能减轻其潜在的病理作用。一系列不同程度表达小窝蛋白-1的模型细胞系已经被创造出来，导致脂质筏（如脂肪细胞中的小泡）的膜胆固醇水平升高。初步数据表明，这些细胞是脂肪细胞样的，它们具有增强的急性脂肪酸跨膜摄取和增强的脂肪酸储存。我们的两个具体目标是：1。目的：确定新型囊泡蛋白cavin在囊泡功能中的生理生化作用。我们发现这种蛋白经历了化学计量泛素化，我们希望确定这种独特的修饰在小泡功能和动力学中的作用。2. 研究小泡膜脂质和蛋白质组成对脂肪酸通量和储存速率和程度的影响。为此，我们将使用多种检测方法，实时监测现有细胞系、脂肪细胞和其他模型细胞系中脂肪酸的运输速率，这些模型细胞系将通过转染与小泡、跨膜脂肪酸通量和脂肪酸储存相关的蛋白质来创建。该应用程序的长期目标是了解脂肪细胞的细胞区室化，因为它适用于脂肪细胞作为能量储存的主要部位，以及作为生物体（包括智人）营养状态的传感器。

DESCRIPTION (provided by applicant): Fatty acids comprise the most calorie-rich source of fuel for mammals. Fatty acids are taken up and stored in adipocytes and mobilized from this depot when needed by other tissues. Excess circulating fatty acids are a proximal cause of insulin resistance and type 2 diabetes, and therefore factors that modulate their cellular uptake and release have potentially direct connections with these pathological conditions. Proper fuel homeostasis requires that fatty acids move across the cell surface, the plasma membrane, rapidly and in large amounts. The adipocyte plasma membrane is particularly abundant in structures called caveolae, which are small membrane invaginations ("little caves") enriched in cholesterol and sphingomyelin that are formed by the expression of caveolin-1 protein. Caveolae are a type of lipid raft characterized by resistance to dispruption by mild detergents such as fatty acids. Mice lacking caveolin-1 are insulin resistant and have abnormal fatty acid metabolism. We hypothesize that caveolin/caveolae in adipocytes play a role in the regulation of fatty acid movement across the plasma membrane and may mitigate their potentially pathological actions. A series of model cell lines have been created that express caveolin-1 to varying extents, resulting in the increased membrane cholesterol levels characteristic of lipid rafts such as caveolae in adipocyte. Preliminary data show that these cells are adipocyte-like in that they have enhanced acute fatty acid transmembrane uptake as well as enhanced fatty acid storage. Our two specific aims are 1. To determine the physiological and biochemical role of the novel caveolar protein, cavin, in caveolae function. We found that this protein undergoes stoichiometric ubiquitination, and we wish to determine the role of this unique modification in caveolae function and dynamics. 2. To study the effects of caveolar membrane lipid and protein composition on the rate and extent of fatty acid flux and storage. To this end, we will use a number of assays that allow real time monitoring of fatty acid transport rates in our extant cell lines, in adipocytes and in other model cell lines that we will create by transfection with proteins relevant to caveolae, transmembrane fatty acid flux and fatty acid storage. The long term goals of the application are understand the cellular compartmentalization of adipocytes as it applies to their role as the principle site of energy storage, and as sensors for the nutritional state of the organism, including homo sapiens.