Sakaue H, Hara K, Noguchi T, Matozaki T, Kotani K, Ogawa W, Yonezawa K, Waterfield M D, Kasuga M
Second Department of Internal Medicine, Kobe University School of Medicine, Japan.
J Biol Chem. 1995 May 12;270(19):11304-9. doi: 10.1074/jbc.270.19.11304.
Activation of glycogen synthase is one of the major metabolic events triggered by exposure of cells to insulin. The molecular mechanism by which insulin activates glycogen synthase was investigated. The possible role of Ras and mitogen-activated protein kinase cascade was investigated with a stable cell line, CHO-IR-C/S 46, that overexpresses insulin receptors and a catalytically inactive SH-PTP 2 protein phosphatase and in which insulin does not induce the formation of the Ras-GTP complex or the subsequently activation of the mitogen-activated protein kinase cascade. Insulin activated glycogen synthase in this cell line to a similar extent as in parental CHO-IR cells. The importance of heteromeric phosphoinositide (PI) 3-kinase in insulin activation of glycogen synthase was examined in a stable cell line, CHO-IR/delta p85, that overexpresses insulin receptors and a dominant negative mutant (delta p85) of the 85-kDa subunit of PI 3-kinase that lacks the binding site for the catalytic 110-kDa subunit. Insulin-dependent activation of PI-3 kinase and glucose transport, but not the formation of the Ras-GTP complex, are markedly attenuated in this cell line. In CHO-IR/delta p85 cells, insulin activated glycogen synthase to a similar extent as in parental CHO-IR cells. The failure of overproduction of the mutant (delta p85) protein to inhibit insulin activation of glycogen synthase was also confirmed by transient expression in Rat 1 cells with the use of a recombinant vaccinia virus. However, wortmannin abolished insulin activation of glycogen synthase in all cell lines. These data suggest that existence of a Ras-independent and wortmannin-sensitive pathway for activation of glycogen synthase by insulin.
糖原合酶的激活是细胞暴露于胰岛素后触发的主要代谢事件之一。研究了胰岛素激活糖原合酶的分子机制。利用稳定细胞系CHO-IR-C/S 46研究了Ras和丝裂原活化蛋白激酶级联反应的可能作用，该细胞系过表达胰岛素受体和催化失活的SH-PTP 2蛋白磷酸酶，且胰岛素不会诱导Ras-GTP复合物的形成或随后丝裂原活化蛋白激酶级联反应的激活。胰岛素在该细胞系中激活糖原合酶的程度与在亲本CHO-IR细胞中相似。在稳定细胞系CHO-IR/delta p85中研究了异源磷酸肌醇（PI）3激酶在胰岛素激活糖原合酶中的重要性，该细胞系过表达胰岛素受体和PI 3激酶85 kDa亚基的显性负突变体（delta p85），该突变体缺乏与催化性110 kDa亚基的结合位点。在该细胞系中，胰岛素依赖性PI-3激酶激活和葡萄糖转运，但不是Ras-GTP复合物的形成，均明显减弱。在CHO-IR/delta p85细胞中，胰岛素激活糖原合酶的程度与在亲本CHO-IR细胞中相似。使用重组痘苗病毒在大鼠1细胞中瞬时表达也证实了突变蛋白（delta p85）过量表达未能抑制胰岛素对糖原合酶的激活。然而，渥曼青霉素消除了所有细胞系中胰岛素对糖原合酶的激活。这些数据表明存在一条不依赖Ras且对渥曼青霉素敏感的胰岛素激活糖原合酶的途径。
