Sauvage M, Mazière P, Fathallah H, Giraud F
Laboratoire des Biomembranes et Messagers Cellulaires, Université Paris XI, Orsay, France.
Eur J Biochem. 2000 Feb;267(4):955-62. doi: 10.1046/j.1432-1327.2000.01084.x.
The signaling cascade linking insulin receptor stimulation to the activation of Na/H exchanger (NHE) was investigated in human erythrocytes, a simple cell model expressing the NHE1 isoform and protein kinase C (PKC) alpha and zeta isoforms only. Our results demonstrate the presence of phosphatidylinositol (PtdIns) 3-kinase in these cells and its activation by insulin. With a similar time-course, insulin also promoted both the translocation and activation of PKC zeta, but had no effect on PKC alpha. Inhibition of PtdIns 3-kinase with wortmannin prevented the activation of PKC zeta by insulin. Stimulation of NHE1 was observed after 10 min of insulin treatment and persisted for at least 60 min. This effect was totally abolished by wortmannin or GF 109203X, an inhibitor of all PKC isoforms, but not by Gö 6976, a specific inhibitor of conventional and novel PKCs (e.g. PKC alpha). These data indicate that PKC zeta activation is mediated by a PtdIns 3-kinase-dependent mechanism and that NHE1 stimulation involves the sequential activation of PtdIns 3-kinase and PKC zeta. In addition, insulin stimulation of NHE1 occurred without altering the phosphorylation state of the exchanger, suggesting that the phosphorylation of an ancillary protein by PKC zeta would be responsible for activation of the transporter.
在人类红细胞（一种仅表达NHE1亚型以及蛋白激酶C（PKC）α和ζ亚型的简单细胞模型）中，研究了将胰岛素受体刺激与钠/氢交换体（NHE）激活联系起来的信号级联反应。我们的结果表明这些细胞中存在磷脂酰肌醇（PtdIns）3激酶及其被胰岛素激活的情况。胰岛素以相似的时间进程促进了PKCζ的转位和激活，但对PKCα没有影响。用渥曼青霉素抑制PtdIns 3激酶可阻止胰岛素对PKCζ的激活。胰岛素处理10分钟后观察到NHE1的刺激，且这种效应持续至少60分钟。渥曼青霉素或GF 109203X（一种所有PKC亚型的抑制剂）可完全消除这种效应，但Gö 6976（一种传统和新型PKC（如PKCα）的特异性抑制剂）则不能。这些数据表明PKCζ的激活是由一种PtdIns 3激酶依赖性机制介导的，并且NHE1的刺激涉及PtdIns 3激酶和PKCζ的顺序激活。此外，胰岛素对NHE1的刺激在不改变交换体磷酸化状态的情况下发生，这表明PKCζ对辅助蛋白的磷酸化可能负责转运体的激活。
