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

大脑中脂质第二信使代谢酶二酰甘油激酶在分子、定位和功能方面的异质性。

• 批准号: 11480225
• 负责人: GOTO Kaoru
• 金额: $ 8.96万
• 依托单位: Yamagata University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 1999
• 资助国家: 日本
• 起止时间: 1999 至 2001
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-11480225/
• 关键词: phospholipid; diacylglycerol; diacylglycerol kinase; neuron; nucleus; signal transduction; second messenger; 脳; 初代培養神経細胞; 遺伝子導入; 核移行シグナル; アンキリンリピート

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.

二酰甘油激酶（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的激活。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，这是由其一级结构中存在核定位信号所表明的。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。

项目成果

後藤 薫、近藤尚武: "ジアシルグリセロールキナーゼ:その分子多様性と中枢神経系のおける発現局在の多様性"蛋白質・核酸・酵素 脂質の分子生物学と病態生化学 井上圭三・西島正弘 編集、共立出版社. 44・8. 976-982 (1999)

Kaoru Goto、Naotake Kondo：“二酰甘油激酶：其分子多样性和中枢神经系统表达定位的多样性”，蛋白质、核酸、酶和脂质的分子生物学和病理生物化学，井上敬三和西岛正宏编辑，共立出版社 44。・8。976-982（1999）

Shirai,Y.,Segawa,S.,Kuriyama,M.,Goto,K.,Sakai,N.& Saito,N.: "Subtype-specific translocation of diacylglycerol kinase alpha and gamma and its correlation with protein kinase C."J.Biol.Chem.. 275. 24760-24766 (2000)

白井，Y.，濑川，S.，栗山，M.，后藤，K.，酒井，N.

Takeda M., Kagaya Y., Takahashi J., Sugie T., Ohta J., Watanabe J., Shirato K., Kondo H., Goto K.: "Gene expression and in situ localization of diacylglycerol kinase isozymes in normal and infarcted rat hearts: effects of captopril treatment"Circulation R

Takeda M.、Kagaya Y.、Takahashi J.、Sugie T.、Ohta J.、Watanabe J.、Shirato K.、Kondo H.、Goto K.：“正常和中二酰甘油激酶同工酶的基因表达和原位定位

Goto,K.& Kondo,H.: "Diacylglycerol kinase in the central nervous system-heterogeneity and gene expression."Chem. Phys. Lipids. 98. 109-117 (1999)

后藤，K.

Takeda M, Kagaya Y, Takahashi J, Sugie T, Ohta J, Watanabe J, Shirato K, Kondo H, Goto K: "Gene expression and in situ localization of diacylglycerol kinase isozymes in normal and infarcted rat hearts : effects of captopril treatment"Circulation Research.

Takeda M、Kagaya Y、Takahashi J、Sugie T、Ohta J、Watanabe J、Shirato K、Kondo H、Goto K：“正常和梗塞大鼠心脏中二酰甘油激酶同工酶的基因表达和原位定位：卡托普利治疗的效果”