Tsakiridis T, Wang Q, Taha C, Grinstein S, Downey G, Klip A
Division of Cell Biology, Hospital for Sick Children, Toronto, Ontario, Canada.
Soc Gen Physiol Ser. 1997;52:257-71.
The purpose of the studies included in this chapter was to examine the role of the actin network in the propagation of insulin action leading to stimulation of glucose transport and activation of the mitogen-activated protein kinase cascade. The active insulin receptor phosphorylates tyrosine residues of intracellular proteins such as the insulin receptor substrate-1 (IRS-1) which acts as docking sites for molecules containing Src homology 2 (SH2) domains. One such molecule is phosphatidylinositol 3-kinase (PI 3-kinase) which becomes activated by binding to IRS-1. PI 3-kinase activity is required for the insulin-stimulation of glucose transport and glycogen synthesis. Grb2, a small adaptor molecule, can bind IRS-1 and, through the guanine nucleotide exchange factor Sos, leads to the activation of the small GTP binding protein Ras. Through a cascade of protein kinases, activation of Ras results in activation of the Erk 1 and 2 mitogen-activated protein kinases (MAPKs) which appear to control important nuclear and metabolic events. To investigate the role of the actin network in the propagation of insulin action leading to stimulation of glucose transport and the activation of the Erk MAPKs, we used the fungal metabolite cytochalasin D which disassembles the actin network. Actin disassembly abolished almost completely the ability of insulin to increase the rate of glucose transport into L6 muscle cells (myotubes) through prevention of the insulin-induced recruitment of glucose transporters to the plasma membrane which is the event that mediates the increase in the rate of transport. Actin disassembly did not affect either the insulin-mediated phosphorylation of IRS-1, the association of PI 3-kinase with this molecule, or the activation of IRS-1-associated PI 3-kinase. These results were also verified in another insulin responsive cell line, the 3T3-L1 adipocytes. In these cells, actin disassembly inhibited the insulin-induced recruitment of PI 3-kinase to intracellular membranes containing glucose transporters. Moreover, actin disassembly abolished the insulin-mediated phosphorylation of the Erk MAPKs. We conclude that the cellular actin network of insulin responsive cells is not required for the activation of PI 3-kinase but prevents its cellular redistribution. In contrast, intact actin filaments are essential for the propagation of insulin signals leading to the the activation of the MAPKs.
本章所包含的研究目的是考察肌动蛋白网络在胰岛素作用传导过程中的作用，该传导过程可导致葡萄糖转运的刺激以及丝裂原活化蛋白激酶级联反应的激活。活性胰岛素受体使细胞内蛋白质的酪氨酸残基磷酸化，如胰岛素受体底物-1（IRS-1），它作为含有Src同源2（SH2）结构域分子的对接位点。其中一个这样的分子是磷脂酰肌醇3-激酶（PI 3-激酶），它通过与IRS-1结合而被激活。PI 3-激酶活性是胰岛素刺激葡萄糖转运和糖原合成所必需的。Grb2是一种小的衔接分子，它可以结合IRS-1，并通过鸟嘌呤核苷酸交换因子Sos，导致小GTP结合蛋白Ras的激活。通过一系列蛋白激酶，Ras的激活导致Erk 1和2丝裂原活化蛋白激酶（MAPKs）的激活，它们似乎控制着重要的核和代谢事件。为了研究肌动蛋白网络在胰岛素作用传导过程中的作用，该传导过程可导致葡萄糖转运的刺激以及Erk MAPKs的激活，我们使用了可使肌动蛋白网络解体的真菌代谢产物细胞松弛素D。肌动蛋白解体几乎完全消除了胰岛素通过阻止胰岛素诱导的葡萄糖转运蛋白募集到质膜来增加葡萄糖转运到L6肌肉细胞（肌管）速率的能力，而这一事件介导了转运速率的增加。肌动蛋白解体既不影响胰岛素介导的IRS-1磷酸化、PI 3-激酶与该分子的结合，也不影响与IRS-1相关的PI 3-激酶的激活。这些结果在另一种胰岛素反应性细胞系3T3-L1脂肪细胞中也得到了验证。在这些细胞中，肌动蛋白解体抑制了胰岛素诱导的PI 3-激酶募集到含有葡萄糖转运蛋白的细胞内膜上。此外，肌动蛋白解体消除了胰岛素介导的Erk MAPKs的磷酸化。我们得出结论，胰岛素反应性细胞的细胞肌动蛋白网络对于PI 3-激酶的激活不是必需的，但可阻止其细胞内重新分布。相反，完整的肌动蛋白丝对于胰岛素信号传导导致MAPKs激活是必不可少的。
