Regulation of lymphocyte phospholipase C activity by a low molecular weight GTP-binding protein and its physiological role

低分子量GTP结合蛋白对淋巴细胞磷脂酶C活性的调节及其生理作用

• 批准号: 02670990
• 负责人: TOYOSHIMA Satoshi
• 金额: $ 1.47万
• 依托单位: University of Tokyo
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1990
• 资助国家: 日本
• 起止时间: 1990 至 1991
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-02670990/
• 关键词: Phospholipase C; Butulinum ADP-ribosyltransferase; Lymphocyte activation; T cell antigen receptor; MRL; lpr mouse; low molecular weight GTP-binding protein; ボツリヌスADPーリボシルトランスフェラ-ゼ; lprマウス; イノシト-ルリン脂質

As reported previously, there is a low molecular weight GTP-binding protein(G21K)in calf thymocytes which is associated with not only physically but also functionally with soluble phosphoinositide-specific phospholipase C(PLC). In the present study, its physiological role was investigated and following results were obtained.1. G21K was found to be ADP-ribosylated by ADP-ribosyltransferase from Clostridum botulinum type C strain and ^<32>P -ADP-ribosylated G21K was easily identified on two dimensional gel electrophoresis.2. G21K was also detected in mouse thymocytes.3. Botulinum ADP-ribosyltransferase induced an increase of inositol phosphates(PLC)formation in mouse thymocyte membranes. Incubation of electropermeabilized mouse thymocytes with the enzyme also caused an inprease of IPs formation in the cells. These results suggest that the stimulation of PLC was related to ADP-ribosylation of G21K by the enzyme.4. G21K was found to be associated with T cell antigen receptor(TCR)/CD3 complex in mouse thymocytes. This result suggests that G21K plays some important role in the T cell activation pathway via TCR/CD3 complex.5. It was found that very little amounts of G21K were associated with Con A receptor in T cells of autoimmune model mouse, MRL/Ipr. As reported previously, MRL/Ipr T cells hardly respond to Con A and an increase of phosphoinositide turnover is not observed in MRL/Ipr T cells stimulated with Con A. However, Con A binding to the receptor is normal and stimulation of G-protein by GTPYS induces PLC activation. These results suggest that the coupling of Con A-receptor and the G-protein is abnormal.

如前所述，小牛胸腺细胞中存在一种低分子量的GTP结合蛋白（G21 K），它不仅在物理上而且在功能上与可溶性磷酸肌醇特异性磷脂酶C（PLC）相关。本研究主要探讨了其生理作用，并获得以下结果. G21 K被C型肉毒梭菌的ADP-核糖基转移酶标记为ADP-核糖基化，<32>并在二维凝胶电泳上易于鉴定.在小鼠胸腺细胞中也检测到G21 K.肉毒素ADP-核糖基转移酶诱导小鼠胸腺细胞膜磷酸肌醇（PLC）的形成增加。用酶孵育电透化的小鼠胸腺细胞也引起细胞内IP形成的增加。这些结果表明，PLC的刺激作用与G21 K的ADP-核糖基化有关.在小鼠胸腺细胞中发现G21 K与T细胞抗原受体（TCR）/CD 3复合物相关。这一结果表明，G21 K通过TCR/CD 3复合物在T细胞活化途径中起重要作用.结果发现，在自身免疫模型小鼠MRL/Ipr的T细胞中，极少量的G21 K与Con A受体相关。如前所述，MRL/Ipr T细胞几乎不响应Con A，并且在Con A刺激的MRL/Ipr T细胞中未观察到磷酸肌醇周转的增加。然而，Con A与受体的结合是正常的，GTPYS对G蛋白的刺激诱导PLC活化。这些结果表明，ConA受体与G蛋白的偶联是异常的。

项目成果

豊島 聰: "各種細胞における細胞内情法伝達系の異常ーリンパ球" Clinical Neuroscience. 9. 870-872 (1991)

Satoshi Toyoshima：“各种细胞 - 淋巴细胞的细胞内信号传输系统异常”临床神经科学 9. 870-872 (1991)。

豊島 聡: "サイトカインネットワ-クと免疫調節" ファルマシア. 27. 323-328 (1991)

Satoshi Toyoshima：“细胞因子网络和免疫调节”Pharmacia 27. 323-328 (1991)。

P. Wang: "Roles of phospholipase C and GTP-binding proteins in lymphocyte activation." J. Pharmacobio-Dyn.13. s-105 (1990)

P. Wang：“磷脂酶 C 和 GTP 结合蛋白在淋巴细胞激活中的作用。”

M.Higuchi: "TNFーmediated cytotoxicity.Importance of intracellular cGMP level for determining TNFーsensitivity." Molecular Immumology. 28. 1039-1044 (1991)

M. Higuchi：“TNF 介导的细胞毒性。细胞内 cGMP 水平对于确定 TNF 敏感性的重要性。”28. 1039-1044 (1991)

P. Wang: "Low-molecular weight GTP-binding proteins syrving as ADP-ribosylation substrate for ADP-ribosyltransferase from Clostridium botulinum and their relation to phosphoinositides metabolism in thymocytes." J. Biochem.108. 87-885 (1990)

P. Wang：“低分子量 GTP 结合蛋白可作为肉毒杆菌 ADP-核糖基转移酶的 ADP-核糖基化底物，及其与胸腺细胞中磷酸肌醇代谢的关系。”