Regulatory mechanisms of multi-functional sheddase

多功能脱落酶的调控机制

• 批准号: 15570121
• 负责人: SAKANE Fumio
• 金额: $ 2.05万
• 依托单位: Sapporo Medical University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2003
• 资助国家: 日本
• 起止时间: 2003 至 2004
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-15570121/
• 关键词: Diacylglycerol kinas; Phosphatidic acid; Osmotic shock; Phorbol ester; Phosphorylation; Oligomer; SAM; Pleckstrin homology; TACE; ホルボールエスチル; ジアシルグリセロール; 腫瘍壊死因子α; トランスロケーション; プロテインキナーゼC

We had ahrady obtained several lit s of evidence suggesting chat diacylglycerol kinase (DGK) 6 participates m the regulation of a multi-functional sheddase, tumor necrosis α-converning enzyme (TACE). As a next step, we by modification of DGK6 and its closely related isofoms (DGKs η and κ) in stimulated cells. DGKδ1 or its pleckstrin homology (PH) domain alone has been shown to be translocated to the plasma membranes from the cytoplasm in phorbol myristate acetate (PMA)-treated cells. In the present work we identified Ser-22 and Ser-26 within the PH domain as the PMA-and epidermal growth factor-dependent phosphorylation sites of DGKδ1. Experiments in vitro and with intact cells suggested that the conventional protein kinase C (cPKC) directly phosphorylated these Ser residues. Interestingly, the Asp-mutation, which mimics phosphoserine, at Ser-22 or Ser-26, markedly inhibited the translocation of full-length DGKδ1 and the PH domain, suggesting that the phosphorylation regulates negatively the enzyme translocation.We identified another DGKη isoform (η2, 135 kDa) that shared the same sequence with DGKη1 except for a sterile a motif (SAM) domain added at the C-terminus. DGKη2 was shown to form through its SAM domain homo-oligomers as well as hetero-oligomers with other SAM-containing DGKs (81 and 62). Interestingly, DGKη1 and DGKη2 were rapidly translocated from the cytoplasm to endosomes in response to stress stimuli. In this case, DGKη1 l was rapidly relocated back to the cytoplasm upon removal of stress stimuli, whereas DGKη2 exhibited sustained endosomal association.Recently, we identified a tenth member of the DGK family designated DGKκ. The new DGK isozyme has additionally 33 tandem repeats of Glu-Pro-Ala-Pro at the N-terminus. Interestingly, DGKκ, but not other type II DGKs, was specifically tyrosine-phosphorylated by Src-family kinase in H_2O_2-treated cells.

我们已经获得了一些证据，表明二酰甘油激酶 (DGK) 6 参与多功能脱落酶、肿瘤坏死 α-转换酶 (TACE) 的调节作为下一步，我们对 DGK6 及其进行了修饰。受刺激细胞中密切相关的异构体（DGK η 和 κ）已被证明可以单独使用 DGKδ1 或其 pleckstrin 同源 (PH) 结构域。在佛波醇肉豆蔻酸酯乙酸酯 (PMA) 处理的细胞中，Ser-22 和 Ser-26 被从细胞质转移到质膜。 DGKδ1。体外实验和完整细胞实验表明，传统的蛋白激酶 C (cPKC) 直接磷酸化这些 Ser 残基，即 Asp 突变。在 Ser-22 或 Ser-26 处模拟磷酸丝氨酸，显着抑制全长 DGKδ1 和 PH 结构域的易位，表明磷酸化对酶易位产生负调节。与 DGKη1 相同的序列，除了在 C 末端添加的不育 a 基序 (SAM) 结构域已显示形成。通过其 SAM 结构域同源寡聚体以及与其他含有 SAM 的 DGK（信息丰富）的异源寡聚体，DGKη1 和 DGKη2 响应应激刺激而快速从细胞质转移到内体。在去除应激刺激后，DGKη2 迅速重新定位回细胞质，而 DGKη2 显示出持续的内体关联。最近，我们发现了一个DGK 家族的第十个成员命名为 DGKκ。新的 DGK 同工酶在 N 末端另外有 33 个 Glu-Pro-Ala-Pro 串联重复，这表明 DGKκ（而非其他 II 型 DGK）被 Src 特异性酪氨酸磷酸化。 -H_2O_2 处理的细胞中的家族激酶。

项目成果

生物薬科学実験講座 情報伝達物質[II](石橋貞彦, 市川厚, 堅田利明編)

生物制药科学实验课程信息传递物质[II]（石桥定彦、市川厚、片田俊明编）

• 作者: 坂根郁夫;山田恵子;加納英雄(分担執筆)
• 通讯作者: 加納英雄(分担執筆)

The plasma membrane translocation of diacylglycerol kinase δ1 is negatively regulated by conventional protein kinase C-dependent phosphorylation at Ser-22 and Ser-26 within the pleckstrin homology domain

• DOI: 10.1042/bj20040681
• 发表时间: 2004-09-15
• 期刊: BIOCHEMICAL JOURNAL
• 影响因子: 4.1
• 作者: Imai, S;Kai, M;Sakane, F
• 通讯作者: Sakane, F

Jia, Y-J., Kai, M., Wada, I., Sakane, F., Kanoh, H.: "Differential localization of lipid phosphate phosphatases 1 and 3 to cell surface subdomains in polarized MDCK cells"FEBS Lett.. 552(2-3). 240-246 (2003)

Jia, Y-J.、Kai, M.、Wada, I.、Sakane, F.、Kanoh, H.：“脂质磷酸磷酸酶 1 和 3 在极化 MDCK 细胞中细胞表面子结构域的差异定位”FEBS Lett.. 552（

Gene expression, cellular localization and enzymatic activity of diacylglycerol kinase isozymes in rat ovary and placenta

大鼠卵巢和胎盘中二酰甘油激酶同工酶的基因表达、细胞定位和酶活性

• 期刊: Cell Tissue Res. (in press)
• 作者: Toya;M.;Hozumi;Y.;Ito;T.;Takeda;M.;Sakane;F.;Kanoh;H.;Saito;H.;Hiroi;M.;Kurachi;H.;Kondo;H.;Goto;K.
• 通讯作者: K.

Expression and localization of diacylglycerol kinase isozymes and enzymatic features in rat lung.

二酰甘油激酶同工酶的表达和定位以及大鼠肺中的酶学特征。

• 期刊: Am.J.Physiol.Lung Cell Mol.Physiol. (in press)
• 作者: Katagiri;Y.;Ito;T.;Saino-Saito;S.;Hozumi;Y.;Suwabe;A.;Otake;K.;Sata;M.;Kondo;H.;Sakane;F.;Kanoh;H.;Kubota;I.;Goto;K.
• 通讯作者: K.