REGULATION OF PROTEIN TRAFFICKING IN ADIPOCYTES

描述(申请人提供)：GLUT4葡萄糖转运体催化餐后全身葡萄糖处置的限速步骤。胰岛素刺激的GLUT4重新分布到质膜的破坏是与2型糖尿病相关的外周胰岛素抵抗的近端原因。因此，阐明胰岛素刺激GLUT4急性重分布到质膜的确切分子途径对于理解胰岛素抵抗状态的发病机制具有重要意义。GLUT4的亚细胞转运调控部分由胰岛素反应的AS160/Rab10 GTP酶周期决定，但其他未知因素是必要的，以完全解释其基本的细胞内隔离和胰岛素刺激的向质膜的移动。此外，指导其受管制的亚细胞贩运的GLUT4的具体结构特征仍然知之甚少。我们在GLUT4胞质尾部的羧基末端12个残基上发现了一个新的亚细胞靶向基序(LXXLXP)，该基序与胰岛素反应性氨基肽酶共享。与其他GLUT4靶向基序不同，LXXLXP基序(这里称为IRM，胰岛素反应基序)的改变对转运蛋白的稳态分布有深远的影响，似乎完全取消了转运蛋白的基础循环和胰岛素刺激的细胞表面转位。这项建议的目标是通过描述IRM和假定的Akt调节的GTPase激活蛋白AS250在这一过程中的作用，获得对GLUT4调控的基本见解。为了实现这一目标，将采取以下具体目标：1)准确地定义IRM以及它如何与其他已知的GLUT4靶向基序相互作用；2)鉴定和表征GLUT4通过的新的亚细胞膜室；以及3)阐明AS250复合体的功能，胰岛素如何影响其功能，并确定AS250/KIAA1219复合体的新成分。

DESCRIPTION (provided by applicant): The Glut4 glucose transporter catalyzes the rate-limiting step in postprandial whole body glucose disposal. A disruption in the insulin-stimulated redistribution of Glut4 to the plasma membrane is the proximal cause of the peripheral insulin resistance associated with type 2 diabetes mellitus. Elucidation of the precise molecular pathways involved in the mechanism by which insulin stimulates the acute redistribution of Glut4 to the plasma membrane may thus be of considerable importance in understanding the pathogenesis of insulin resistant states. The regulated subcellular trafficking of Glut4 is partially dictated by the insulin- responsive AS160/Rab10 GTPase cycle, but additional unidentified factors are necessary to fully account for its basal intracellular sequestration and insulin-stimulated movement to the plasma membrane. Additionally, the specific structural features of Glut4 that direct its regulated subcellular trafficking remain poorly understood. We have identified a novel subcellular targeting motif (LXXLXP) within the carboxy-terminal 12 residues of the cytoplasmic tail of Glut4 that is shared with the insulin-responsive aminopeptidase. Unlike other Glut4 targeting motifs, alteration of the LXXLXP motif (herein referred to as IRM, insulin-responsive motif) has a profound effect on the steady-state distribution of the transporter and appears to completely abolish its basal recycling and insulin-stimulated translocation to the cell surface. The goal of this proposal is to gain fundamental insights into Glut4 regulation by delineating the role of the IRM and of a putative Akt-regulated GTPase activating protein, AS250, in this process. In order to accomplish this goal, the following specific aims will be undertaken: 1) to precisely define the IRM and how it interacts with other known Glut4 targeting motifs; 2) to identify and characterize novel subcellular membrane compartments through which Glut4 moves; and 3) to elucidate the function of the AS250 complex, how insulin affects its function, and identify novel components of the AS250/KIAA1219 complex.