TUG-mediated GLUT4 glucose transporter trafficking and regulation by post-transla

描述（由申请人提供）：2型糖尿病是美国最常见的疾病之一，对公众健康有着巨大的影响，在世界范围内已达到流行病的程度。2型糖尿病的特点是对胰岛素的反应有缺陷，这阻碍了餐后葡萄糖进入肌肉和脂肪组织的正常摄取，并导致高血糖和长期并发症。葡萄糖转运蛋白4 （GLUT4）是胰岛素应答的主要葡萄糖转运蛋白。GLUT4从细胞膜转位到细胞表面是胰岛素增强葡萄糖摄取所必需的，这一过程中的缺陷有助于2型糖尿病的发病。许多研究表明胰岛素信号通路在这一缺陷中的重要性，但越来越多的证据指向GLUT4运输本身的作用。然而，控制GLUT4运输的确切机制在很大程度上尚不清楚。我的导师发现了含有GLUT4 UBX结构域（TUG）的Tether，并证明它是GLUT4运输的重要调节剂，对于不受胰岛素刺激的细胞内GLUT4的细胞内保留至关重要。虽然TUG的c端对于这种保持是必需的，但是我们不知道TUG是如何完成这个功能的。我的博士论文和本次提案的目标是研究TUG c -末端如何介导GLUT4在细胞内的保留，以及TUG c -末端的乙酰化如何调节其功能。作为其中的一部分，我们希望测试我们最喜欢的模型，该模型基于初步和已发表的数据，假设TUG在GLUT4的细胞内锚定机制中起作用，涉及反式高尔基网络（TGN）蛋白GCC185，这是一种已知的囊泡运输介质。此外，我们认为TUG的乙酰化对这种锚定功能很重要。我们将用两个具体目标来检验这些假设。在第一个目标中，我将采用基因敲低GCC185来测试这是否会破坏未受刺激的3T3-L1脂肪细胞中GLUT4的细胞内隔离。在第二个Aim中，我将研究TUG乙酰化如何调节其功能，并将验证乙酰化控制胰岛素反应性GLUT4库的大小，并且是TUG与GCC185结合所必需的假设。我们将根据需要研究我们实验室已经确定的其他候选锚和TUG乙酰化的其他功能。这项研究的结果将阐明GLUT4如何在细胞内运输，并将加强对胰岛素敏感性如何被乙酰化控制的理解。这些都是2型糖尿病的重要问题，也可能对细胞生物学和生理学有更根本的意义。

DESCRIPTION (provided by applicant): Type 2 diabetes is one of the most common diseases in America and carries an enormous impact on public health, having reached epidemic proportions worldwide. Type 2 diabetes is characterized by a defective response to insulin, which prevents normal uptake of post-prandial glucose into muscle and adipose tissue and contributes to hyperglycemia and long-term complications. Glucose transporter 4 (GLUT4) is the major insulin responsive glucose transporter. Translocation of GLUT4 from intracellular membranes to the cell surface is required for insulin to enhance glucose uptake, and defects in this process in this process contribute to type 2 diabetes pathogenesis. Much research has indicated the importance of insulin signaling pathways in this defect but increasing evidence points to a role in GLUT4 trafficking itself. However, the precise mechanisms controlling GLUT4 trafficking are largely unclear. Tether containing a UBX domain for GLUT4 (TUG) was discovered by my mentor and shown to be an important regulator of GLUT4 trafficking, essential for intracellular retention of GLUT4 within cells not stimulated by insulin. Although the TUG C-terminus is necessary for this retention, it is not known how TUG performs this function. The goal of my PhD thesis and this proposal is to study how the TUG C-terminus mediates intracellular retention of GLUT4 and how acetylation of the TUG C-terminus modulates its function. As part of this, we look to test our favored model that, based on both preliminary and published data, hypothesizes TUG to function in an intracellular anchoring mechanism for GLUT4 involving the trans-Golgi network (TGN) protein GCC185, a known mediator of vesicle trafficking. Further, we propose that acetylation of TUG is important for this anchoring function. We will test these hypotheses with two Specific Aims. In the first Aim, I will employ genetic knockdown of GCC185 to test if this disrupts intracellular sequestration of GLUT4 in unstimulated 3T3-L1 adipocytes. In the second Aim, I will study how TUG acetylation modulates its function, and will test the hypotheses that acetylation controls the size of the insulin- responsive GLUT4 pool and is required for TUG to bind to GCC185. We will investigate other candidate anchors already identified by our lab and other functions of TUG acetylation as needed. The results of this research will elucidate how GLUT4 traffics within cells, and will enhance the understanding of how insulin sensitivity may be controlled by acetylation. These are important questions for type 2 diabetes, and may also have more fundamental significance for cell biology and physiology.