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

甘油二酯激酶(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，因此可减弱PKC的激活。到目前为止，我们已经从大鼠脑cDNA文库中克隆了几种同工酶，并用原位杂交组织化学方法详细检测了它们在脑内的基因表达。在本研究中，我们利用绿色荧光蛋白融合表达载体分析了转基因COS细胞中同工酶的精确亚细胞定位。DGK-α弥漫分布于胞浆和胞核。DGK-β以丝状结构分布于细胞质内，与肌动蛋白细丝的定位十分相似。DGK-γ在核旁呈网状结构，与高尔基体标记相吻合。DGK-Zeta的一级结构中存在核定位信号，表明DGK-Zeta主要位于细胞核内。DGK-epsilon广泛分布于核周，类似于内质网的定位模式。这些数据清楚地显示了DGK在亚细胞定位和分子结构上的异质性，提示每种同工酶可能负责DG的代谢，DG是在刺激时在特定的亚细胞位置产生的。因此，很可能每种DGK同工酶在信号转导中都发挥着独特的作用。

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.