Role of Phospholipase D in Glut-4 translocation

描述(申请人提供)：肌肉和脂肪组织中胰岛素刺激的葡萄糖摄取主要由胰岛素反应葡萄糖转运体亚型GLUT-4介导。在基础状态下，GLUT-4缓慢循环，大部分蛋白质被隔离到细胞内的存储位置。作为对胰岛素的反应，GLUT-4的胞吐速率显著增加，导致GLUT-4在细胞表面大量积聚。最近的发现表明，这种转位过程可能受到磷脂酶D(PLD)的调节，磷脂酶D是一种膜相关酶，也可以被胰岛素激活。磷脂酰胆碱是细胞膜上含量最丰富的磷脂，它能催化磷脂酰胆碱的水解，生成信号转导的磷脂酸(PA)。PA被认为具有多种细胞功能，包括促进膜泡运输。哺乳动物有两个PLD基因，PLD1和PLD2。我们最近发现，在调节胞吐过程中，PLD1促进神经内分泌颗粒与质膜的融合，并且这两个基因都促进含有组胺的肥大细胞颗粒的调节胞吐。重要的是，控制这些分泌颗粒的融合机制与胰岛素刺激的GLUT-4囊泡的质膜融合有许多共同的特征，我们最近发现PLD1和PLD2的激活或抑制改变了胰岛素刺激的GLUT-4到质膜的转位。基于这些新的数据，我们建议通过发展重建分析和建立PLD在生理相关环境中的作用，系统地研究PLD作为胰岛素刺激的GLUT-4运输的关键调节成分的功能。

DESCRIPTION (provided by applicant): Insulin-stimulated glucose uptake in muscle and adipose tissue is primarily mediated by the insulin-responsive glucose transporter isoform, Glut-4. In the basal state, Glut-4 slowly cycles with the majority of the protein sequestered into intracellular storage sites. In response to insulin, the rate of Glut-4 exocytosis becomes markedly increased, resulting in a large accumulation of Glut-4 at the cell surface. Recent findings have suggested that this translocation process may be regulated by Phospholipase D (PLD), a membrane-associated enzyme that is also activated by insulin. PLD catalyzes the hydrolysis of phosphatidylcholine, the most abundant membrane phospholipid, to generate the signaling lipid phosphatidic acid (PA). PA has been proposed to have several cellular functions including the facilitation of membrane vesicle trafficking. There are two mammalian PLD genes, PLD1 and PLD2. We have recently shown that PLD1 facilitates the fusion of neuroendocrine secretory granules into the plasma membrane during regulated exocytosis and that both genes facilitate regulated exocytosis of mast cell histamine-containing granules. Importantly, the fusion mechanisms controlling these secretory granules have many features in common with the insulin-stimulated plasma membrane fusion of Glut-4 vesicles, and we have found recently that PLD1 and PLD2 activation or inhibition alters insulin-stimulated Glut-4 translocation to the plasma membrane. Based upon these novel data, we propose to examine systematically the function of PLD as a key regulatory component in the insulin-stimulated trafficking of Glut-4 by developing reconstitution assays and establishing the role of PLD in physiologically relevant contexts.