Regulation of T Cell and Mast Cell Function by DGKzeta

描述（由申请人提供）：近年来，关于免疫细胞上许多细胞表面受体介导的信号转导最关键的生化事件，已经了解了很多。一个关键的第二信使是二酰基甘油（DAG），它是在接合的ITAM受体激活磷脂酰肌醇级联反应时形成的。DAG反过来刺激至少两个关键的信号级联; Ras途径和几个蛋白激酶C家族成员。因此，理解DAG的形成及其代谢是理解免疫细胞活化的关键。终止DAG功能的一种方法是通过DAG激酶（DGK）将DAG转化为磷脂酸。该提案集中于DGKzeta，这是造血细胞中高水平表达的两种DGKs之一。我们的初步数据表明DGKzeta是体内T细胞和肥大细胞功能的重要负调节因子。本提案中描述的实验将利用细胞系和基因改变的小鼠来研究DGKzeta如何发挥这一关键作用。将追求三个具体目标。首先，我们提出了DGKzeta的细胞生物学和生化分析，以确定其亚细胞定位和影响信号通路的最关键的功能。第二个目标将调查DGKzeta缺陷对体内T细胞功能的影响，第三个目标将研究肥大细胞区室中的类似问题。目标2和3将扩展结构/功能分析，以确定这种酶对免疫挑战反应最重要的特征。我们预计这项工作将进一步阐明T细胞和肥大细胞功能的复杂调控，并有望提出调节这些细胞类型活性的新方法。

DESCRIPTION (provided by applicant): Much has been learned in recent years regarding the biochemical events most critical for signal transduction mediated by numerous cell surface receptors on immune cells. A key second messenger is diacylglycerol (DAG), which is formed when engaged ITAM-bearing receptors activate the phosphatidylinositol cascade. DAG, in turn, stimulates at least two critical signaling cascades; the Ras pathway and several protein kinase C family members. Thus, understanding the formation of DAG and its metabolism is key to understanding immune cell activation. One means by which DAG function is terminated is the conversion of DAG to phosphatidic acid by DAG kinases (DGKs). This proposal focuses on DGKzeta, one of two DGKs expressed at high levels in hematopoietic cells. Our preliminary data indicate that DGKzeta is an important in vivo negative regulator of T cell and mast cell function. The experiments described in this proposal will make use of cell lines and genetically altered mice to investigate how DGKzeta serves this key role. Three specific aims will be pursued. First, we propose a cell biologic and biochemical analysis of DGKzeta to determine its subcellular localization and features most critical for affecting signaling pathways. The second aim will investigate the impact of DGKzeta deficiency on in vivo T cell function and the third aim will examine similar questions in the mast cell compartment. Both aims 2 and 3 will extend the structure/function analysis to determine features of this enzyme most essential for responses to immunologic challenges. We anticipate that this work will shed additional light on the complex regulation of T cell and mast cell function and will hopefully suggest novel approaches to modulate activity of these cell types.