Sakane F, Imai S, Kai M, Wada I, Kanoh H
Department of Biochemistry, Sapporo Medical University School of Medicine, South-1, West-17, Sapporo 060, Japan.
J Biol Chem. 1996 Apr 5;271(14):8394-401. doi: 10.1074/jbc.271.14.8394.
A fourth member of the diacylglycerol kinase (DGK) gene family termed DGK delta was cloned from the human testis cDNA library. The cDNA sequence contains an open reading frame of 3,507 nucleotides encoding a putative DGK protein of 130,006 Da. Interestingly, the new DGK isozyme contains a pleckstrin homology domain found in a number of proteins involved in signal transduction. Furthermore, the C-terminal tail of this isozyme is very similar to those of the EPH family of receptor tyrosine kinases. The primary structure of the delta-isozyme also has two cysteine-rich zinc finger-like structures (C3 region) and the C-terminal C4 region, both of which have been commonly found in the three isozymes previously cloned (DGKs alpha, beta and gamma). However, DGK delta lacks the EF-hand motifs (C2) and contains a long Glu- and Ser-rich insertion (317 residues), which divides the C4 region into two portions. Taken together, these structural features of DGK delta indicate that this isozyme belongs to a DGK subfamily distinct from that consisting of DGKs alpha, beta, and gamma. Increased DGK activity without marked preference to arachidonoyl type of diacylglycerol was detected in the particulate fraction of COS-7 cells expressing the transfected DGKdelta cDNA. The enzyme activity was independent of phosphatidylserine, which is a common activator for the previously sequenced DGKs. Northern blot analysis showed that the DGK delta mRNA (approximately 6.3 kilobases) is most abundant in human skeletal muscle but undetectable in the brain, thymus, and retina. This expression pattern is different from those of the previously cloned DGKs. Our results show that the DGK gene family consists of at least two subfamilies consisting of enzymes with distinct structural characteristics and that each cell type probably expresses its own characteristic repertoire of DGKs whose functions may be regulated through different signal transduction pathways.
从人睾丸cDNA文库中克隆出二酰基甘油激酶（DGK）基因家族的第四个成员，即DGKδ。该cDNA序列包含一个3507个核苷酸的开放阅读框，编码一个推定的130006 Da的DGK蛋白。有趣的是，新的DGK同工酶含有一个在许多参与信号转导的蛋白质中发现的普列克底物蛋白同源结构域。此外，该同工酶的C末端尾巴与受体酪氨酸激酶EPH家族的非常相似。δ同工酶的一级结构也有两个富含半胱氨酸的锌指样结构（C3区）和C末端C4区，这两个结构在先前克隆的三种同工酶（DGKα、β和γ）中都很常见。然而，DGKδ缺乏EF手基序（C2），并含有一个长的富含谷氨酸和丝氨酸的插入序列（317个残基），该序列将C4区分为两部分。综上所述，DGKδ的这些结构特征表明该同工酶属于一个与由DGKα、β和γ组成的亚家族不同的DGK亚家族。在表达转染的DGKδ cDNA的COS-7细胞的微粒体部分检测到DGK活性增加，且对花生四烯酰基型二酰基甘油没有明显偏好。该酶活性不依赖于磷脂酰丝氨酸，而磷脂酰丝氨酸是先前测序的DGK的常见激活剂。Northern印迹分析表明，DGKδ mRNA（约6.3千碱基）在人骨骼肌中最丰富，但在脑、胸腺和视网膜中检测不到。这种表达模式与先前克隆的DGK不同。我们的结果表明，DGK基因家族至少由两个亚家族组成，这些亚家族的酶具有不同的结构特征，并且每种细胞类型可能表达其自身特征性的DGK库，其功能可能通过不同的信号转导途径进行调节。
