Insulin control of GLUT4 traffic to the plasma membrane of adipocytes

葡萄糖向脂肪和肌肉的转运受到严格调控，由质膜（PM）中GLUT4葡萄糖转运蛋白的数量决定。在胰岛素水平较低的禁食状态下，GLUT4通过快速内吞和缓慢循环的方式被积极地隔离在细胞内。餐后状态的胰岛素升高诱导GLUT4重新分配到PM，主要是通过加速GLUT4的循环。在胰岛素升高的餐后期间，通过快速的GLUT4循环来积极维持PM GLUT4水平的升高。当循环胰岛素水平降低时，GLUT4 PM水平恢复到刺激前的水平。因此，脂肪和肌肉细胞对葡萄糖摄取的控制依赖于细胞内部和PM之间GLUT4运输的调节。受损的GLUT4再分配到PM有助于与胰岛素抵抗和2型糖尿病相关的高血糖。阐明GLUT4转运及其胰岛素调控的分子机制将显著影响我们对胰岛素抵抗的理解，从而为未来治疗干预的发展奠定基础。虽然GLUT4转运的现象已经被很好地描述了，但关于这一过程的分子机制还有很多需要了解的。GLUT4主要分布于2个细胞膜腔室：专门负责GLUT4运输的胰岛素反应囊泡（IRVs）和GLUT4 TGN核周腔室。irv已被深入研究，因为它们在未刺激（基础）和胰岛素刺激的细胞中将GLUT4运送到PM。核周隔室在基底脂肪细胞内GLUT4的隔离和GLUT4的动员中起关键作用，以支持胰岛素刺激细胞增加的需求。控制GLUT4进出核周围部位的分子机制尚未被详细描述。在此，我建议解决这一知识差距。该工作计划以我的实验室过去的成就为基础，解决了GLUT4贩运领域的几个关键问题。该项目的主要目标是：定义核周隔室蛋白质组，从而识别在调节GLUT4交通中起作用的蛋白质，以及识别储存在其上的GLUT4以外的货物蛋白质；发现和表征负责调节GLUT4通过核周腔室通量的蛋白质机制。为了实现这些目标，我们将使用最先进的质谱分析方法，以及一系列功能完整的细胞分析来表征调节进出核周腔室的分子机制。

Glucose transport into fat and muscle, which is tightly regulated, is determined by the amount of the GLUT4 glucose transporter in the plasma membrane (PM). In the fasted state, when insulin levels are low, GLUT4 is actively sequestered intracellularly by rapid endocytosis and slow recycling. Elevated insulin in the postprandial state induces a redistribution of GLUT4 to the PM, predominantly by accelerating GLUT4 recycling. Increased levels of PM GLUT4 are actively maintained by rapid GLUT4 recycling during the postprandial period of elevated insulin. GLUT4 PM levels return to pre-stimulation amounts when circulating insulin levels decrease. Thus, the control of glucose uptake by fat and muscle cells is dependent upon regulation of GLUT4 trafficking between the interior and PM of cells. Compromised GLUT4 redistribution to the PM contributes to hyperglycemia associated with insulin-resistance and type 2 diabetes. Elucidating the molecular mechanism underlying GLUT4 traffic and its regulation by insulin will significantly impact our understanding of insulin resistance and thereby provide a foundation for the future development of therapeutic interventions. Although the phenomenon of GLUT4 trafficking is well described, there is much to be learned about the molecular mechanisms of the process. GLUT4 is largely distributed among 2 intracellular membrane compartments: insulin-responsive vesicles (IRVs) that are specialized for the traffic of GLUT4 and the GLUT4 TGN perinuclear compartment. IRVs have been intensively studied because they ferry GLUT4 to the PM in both unstimulated (basal) and insulin-stimulated cells. The perinuclear compartment has a pivotal role in the intracellular sequestration of GLUT4 in basal adipocytes and in the mobilization of GLUT4 to support the increased demand in insulin-stimulated cells. The molecular mechanisms controlling GLUT4 trafficking to and from the perinuclear site have not been described in detail. Here I propose to address that gap in knowledge. The workplan, building on my labs past accomplishments, addresses several key questions in the field of GLUT4 trafficking. The major objectives of the project are to: define the perinuclear compartment proteome, thereby identifying proteins that function in the regulation of GLUT4 traffic as well as identifying cargo proteins other than GLUT4 that are stored their; discover and characterize the protein machinery responsible for regulating flux of GLUT4 through this perinuclear compartment. To accomplish these objectives, we will use state-of-the-art mass spectrometry profiling methods, and a comprehensive battery of functional intact cell assays to characterize the molecular mechanism regulating trafficking to and from the perinuclear compartment.