Heterogeneity of lipid second messenger-metabolizing enzyme, diacylglycerol kinase, in the brain in terms of molecule, localization, and function.

二酰基甘油激酶（DGK）催化脂质第二信使DG磷酸化为磷脂酸（PA），被认为是调节DG水平的关键酶，因此，负责减弱PKC的活化。我们已从大鼠脑cDNA文库中克隆了几种同工酶，并利用原位杂交组织化学方法对它们在脑中的基因表达进行了详细的研究。本研究利用绿色荧光蛋白融合表达载体对cDNA转染的COS细胞中的同工酶进行了精确的亚细胞定位。DGK-alpha弥散分布于细胞质和细胞核中。DGK-beta在细胞质中呈丝状分布，与肌动蛋白丝的位置非常相似。DGK-gamma在细胞核内呈网状结构，与高尔基体标记一致。DGK-zeta主要位于细胞核中，这表明其一级结构中存在核定位信号。DGK-HRP分布于核周区，类似于内质网的定位模式。这些数据清楚地显示了DGK在亚细胞定位以及分子结构上的异质性，表明每种同工酶可能负责DG的代谢，DG在特定的亚细胞位点受到刺激时产生。因此，很可能每种DGK同工酶在信号转导中起着独特的作用。

Diacylglycerol kinase (DGK), which catalyzes phosphorylation of lipid second messenger, DG to phosphatidic acid (PA), is thought to be a key enzyme in the regulation of DG levels and, as a result, to be responsible for attenuating the activation of PKC. We have so far cloned several isozymes from rat brain cDNA library, and performed detailed examination of their gene expression in the brain by in situ hybridization histochemistry. In this study we analyzed precise subcellular localization of the isozymes in cDNA-transfected COS cells using green fluorescent protein-fusion expression vector. DGK-alpha was localized diffusely in both cytoplasm and nucleus in the cells. DGK-beta was distributed throughout the cytoplasm in the shape of filamentous structure, which is quite similar to the location of actin filament. DGK-gamma was seen as a reticular structure in the juxtanuclear position, which is coincided with Golgi apparatus marker. DGK-zeta was located mostly in the nucleus, as suggested by the presence of nuclear localizing signal in its primary structure. DGK-epsilon was widespread to the perinuclear area, which resembles the location pattern of endoplasmic reticulum. These data clearly show the heterogeneity of DGK in subcellular localization as well as molecular structure, suggesting that each isozyme may be responsible for the metabolism of DG, which is produced at specific subcellular site upon stimulation. Therefore it is highly conceivable that each DGK isozyme plays a unique role in the signal transduction.