Kohn A D, Kovacina K S, Roth R A
Department of Molecular Pharmacology, Stanford University School of Medicine, CA 94305, USA.
EMBO J. 1995 Sep 1;14(17):4288-95. doi: 10.1002/j.1460-2075.1995.tb00103.x.
In the present study, insulin is shown to rapidly stimulate by 8- to 12-fold the enzymatic activity of RAC-PK alpha, a pleckstrin homology domain containing ser/thr kinase. In contrast, activation of protein kinase C by phorbol esters had almost no effect on the enzymatic activity of RAC-PK alpha. Insulin activation was accompanied by a shift in molecular weight of the RAC-PK alpha protein, and the activated kinase was deactivated by treatment with a phosphatase, indicating that insulin activated the enzyme by stimulating its phosphorylation. This insulin-induced shift in RAC-PK was also observed in primary rat epididymal adipocytes, as well as in a muscle cell line called C2C12 cells. The insulin-stimulated increase in RAC-PK alpha activity was inhibited by wortmannin (an inhibitor of phosphatidylinositol 3-kinase) in a dose-dependent manner with a half-maximal inhibition of 10 nM, but not by 20 ng/ml of rapamycin. Activation of RAC-PK alpha activity was also observed in a variant RAC lacking the pleckstrin homology domain. These results indicate that RAC-PK alpha activity can be regulated by the insulin receptor. RAC-PK alpha may therefore play a general role in intracellular signaling mediated by receptor tyrosine kinases.
在本研究中，胰岛素可使含pleckstrin同源结构域的丝氨酸/苏氨酸激酶RAC-PKα的酶活性迅速增强8至12倍。相比之下，佛波酯激活蛋白激酶C对RAC-PKα的酶活性几乎没有影响。胰岛素激活伴随着RAC-PKα蛋白分子量的改变，并且用磷酸酶处理可使激活的激酶失活，这表明胰岛素通过刺激其磷酸化来激活该酶。在原代大鼠附睾脂肪细胞以及一种名为C2C12细胞的肌肉细胞系中也观察到了胰岛素诱导的RAC-PK这种变化。渥曼青霉素（一种磷脂酰肌醇3激酶抑制剂）以剂量依赖方式抑制胰岛素刺激的RAC-PKα活性增加，半数最大抑制浓度为10 nM，但20 ng/ml雷帕霉素无此作用。在缺乏pleckstrin同源结构域的RAC变体中也观察到了RAC-PKα活性的激活。这些结果表明，RAC-PKα活性可受胰岛素受体调节。因此，RAC-PKα可能在受体酪氨酸激酶介导的细胞内信号传导中发挥普遍作用。
