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Regulation of neuronal cell functions by GTP-binding protein, ARF.

GTP 结合蛋白 (ARF) 调节神经细胞功能。

基本信息

批准号：
05808073
负责人：
MURAYAMA Toshihiko
金额：
$ 1.09万
依托单位：
Hokkaido University
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (C)
财政年份：
1993
资助国家：
日本
起止时间：
1993 至 1994
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-05808073/
关键词：
GTP-binding proteins ARF Neuronal cells Cholera toxin ADP-ribosylation

项目摘要

Addition of KCl or ATP stimulated Ca influx and noradrenaline (NA) release in PC12 cells. The activation of protein kinase C by phorbol myristate acetate and increases of cAMP content by stimulants enhanced KCl- and ATP-stimulated NA releases. KCl- and ATP-stimulated NA releases from pertussis toxin-pretreated PC12 cells were same compared with those from control cells. Pretreatment with cholera toxin (CTX) showed stimulatory and inhibitory effects in PC12 cells, depending on pretreatment-time. After 1-2 h, KCl- and ATP-stimulated NA releases were enhanced by CTX.After 24 h, the NA releases were inhibited significantly by CTX,although the level of cAMP in PC12 cells was high.Next we investigated the ADP-rebosylation of membrane proteins by CTX from KCl- and ATP-stimulated PC12 cells. CTX catalyzed [32-P] ADP-ribosylation of 45 kDa protein, alpha subunit of Gs, in membranes. The amounts of ADP-ribosylation of 45 kDa in membranes from KCl- and ATP-stimulated cells increased two-fold, compared with that in the control (non-stimulated) cells. The protein contents of alpha subunit in KCl- and ATP-stimulated cells were same as that in control cells, by immunoblotting analysis. The amount of atuto-ADP-ribosylation by CTX also increased in membranes from KCl- and ATP-stimulated cells. These data suggest that ADP-rebosylation factor (ARF), which was an activator protein of CTX-catalyzed ADP-ribosylation, increased in KCl- and ATP-stimulated cells.

加入KCl或ATP刺激PC 12细胞内钙内流和去甲肾上腺素（NA）释放。蛋白激酶C的激活佛波醇肉豆蔻酸酯乙酸酯和cAMP含量的增加，刺激增强KCl和ATP刺激的NA释放。KCl和ATP刺激的NA释放百日咳毒素预处理的PC 12细胞与对照细胞相比，相同。用霍乱毒素（CTX）预处理的PC 12细胞表现出刺激和抑制作用，依赖于预处理时间。在12 h内，CTX可促进KCl和ATP刺激的PC_（12）细胞NA释放，24 h后，尽管细胞内cAMP水平较高，但CTX可显著抑制NA释放。CTX催化膜中45 kDa蛋白（Gs的α亚基）的[32-P] ADP-核糖基化。与对照（非刺激）细胞相比，KCl和ATP刺激细胞膜中45 kDa ADP-核糖基化的量增加了两倍。免疫印迹分析显示，KCl和ATP刺激后的细胞α亚基蛋白含量与对照组相同。在KCl和ATP刺激的细胞膜中，CTX引起的atuto-ADP-ribosylation量也增加。这些数据表明，ADP-rebosylation因子（ARF），这是一个激活蛋白的CTX催化ADP-核糖基化，增加KCl和ATP刺激的细胞。