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.

在PC12细胞中添加KCL或ATP刺激CA流入和去甲肾上腺素（Na）释放。佛比尔肉豆蔻酸酯乙酸肉豆蔻酸酯激活蛋白激酶C的激活，并通过刺激剂增强了cAMC含量的增加，从而增强了KCL和ATP刺激的Na释放。与对照细胞的KCL和ATP刺激的NA释放相比，与对照细胞的PC12细胞相比。霍乱毒素（CTX）预处理在PC12细胞中显示出刺激性和抑制作用，具体取决于预处理。 1-2 h后，CTX通过CTX增强了KCL和ATP刺激的Na释放。尽管PC12细胞中PC12细胞中的CAMP水平很高，但NA释放被显着抑制。Next我们研究了CTX从KCL-PC-PC12细胞中CTX对膜蛋白的ADP rebosylation。 CTX在膜中催化45 kDa蛋白的α蛋白，α亚基的催化[32-P] ADP-核糖基化。与对照（未刺激）细胞相比，来自KCL和ATP刺激的细胞的45 kDa的ADP-核糖基化量增加了两倍。通过免疫印迹分析，KCL和ATP刺激细胞中α亚基的蛋白质含量与对照细胞中的蛋白质含量相同。 CTX和ATP刺激的细胞中CTX的ATUTO-ADP-核糖基化量也增加了。这些数据表明，在KCL和ATP刺激的细胞中，ADP-烯酰化因子（ARF）是CTX催化的ADP-核糖基化的激活蛋白。