Argetsinger L S, Hsu G W, Myers M G, Billestrup N, White M F, Carter-Su C
Department of Physiology, University of Michigan Medical School, Ann Arbor 48109-0622, USA.
J Biol Chem. 1995 Jun 16;270(24):14685-92. doi: 10.1074/jbc.270.24.14685.
The identification of JAK2 as a growth hormone (GH) receptor-associated, GH-activated tyrosine kinase has established tyrosyl phosphorylation as a signaling mechanism for GH. In the present study, GH is shown to stimulate tyrosyl phosphorylation of insulin receptor substrate 1 (IRS-1), the principle substrate of the insulin receptor. Tyrosyl phosphorylation of IRS-1 is a critical step in insulin signaling and provides binding sites for proteins with the appropriate Src homology 2 domains, including the 85-kDa regulatory subunit of phosphatidylinositol (PI) 3'-kinase. In 3T3-F442A fibroblasts, GH-dependent tyrosyl phosphorylation of IRS-1 was detected by 1 min and at GH concentrations as low as 5 ng/ml (0.23 nM). Tyrosyl phosphorylation of IRS-1 was transient, with maximal stimulation detected at 30 min and diminished signal detected at 60 min. The ability of GH receptor (GHR) to transduce the signal for IRS-1 tyrosyl phosphorylation is mediated by the intracellular region of GHR between amino acids 295 and 380 by a mechanism not involving the two tyrosines in this region. This region of GHR is required for GH-dependent JAK2 association and activation (VanderKuur, J. A., Wang, X., Zhang, L., Campbell, G. S., Allevato, G., Billestrup, N., Norstedt, G., and Carter-Su, C. (1994) J. Biol. Chem. 269, 21709-21717). When other cytokines that activate JAK2 were tested for the ability to stimulate the tyrosyl phosphorylation of IRS-1, stimulation was detected with interferon-gamma and leukemia inhibitory factor. The correlation between JAK2 tyrosyl phosphorylation and IRS-1 tyrosyl phosphorylation in response to GH, interferon-gamma, and leukemia inhibitory factor and in cells expressing different GHR mutants, provides evidence that IRS-1 may interact with JAK2 or an auxiliary molecule that binds to JAK2. GH is also shown to stimulate binding of IRS-1 to the 85-kDa regulatory subunit of PI 3'-kinase. The ability of GH to stimulate tyrosyl phosphorylation of IRS-1 and its association with PI 3'-kinase provides a biochemical basis for responses shared by insulin and GH including the well characterized insulin-like metabolic effects of GH observed in a variety of cell types.
JAK2作为一种与生长激素（GH）受体相关的、受GH激活的酪氨酸激酶的鉴定，确立了酪氨酸磷酸化作为GH的一种信号传导机制。在本研究中，GH被证明可刺激胰岛素受体底物1（IRS-1）的酪氨酸磷酸化，IRS-1是胰岛素受体的主要底物。IRS-1的酪氨酸磷酸化是胰岛素信号传导中的关键步骤，并为具有适当Src同源2结构域的蛋白质提供结合位点，包括磷脂酰肌醇（PI）3'-激酶的85-kDa调节亚基。在3T3-F442A成纤维细胞中，在1分钟时就能检测到GH依赖性的IRS-1酪氨酸磷酸化，且在低至5 ng/ml（0.23 nM）的GH浓度下即可出现。IRS-1的酪氨酸磷酸化是短暂的，在30分钟时检测到最大刺激，60分钟时信号减弱。GH受体（GHR）转导IRS-1酪氨酸磷酸化信号的能力由GHR氨基酸295至380之间的细胞内区域介导，其机制不涉及该区域的两个酪氨酸。GHR的这一区域是GH依赖性JAK2结合和激活所必需的（范德库尔，J.A.，王，X.，张，L.，坎贝尔，G.S.，阿莱瓦托，G.，比勒斯特鲁普，N.，诺尔斯特德，G.，和卡特-苏，C.（1994年）《生物化学杂志》269，21709 - 21717）。当测试其他激活JAK2的细胞因子刺激IRS-1酪氨酸磷酸化的能力时，检测到干扰素-γ和白血病抑制因子可产生刺激作用。在对GH、干扰素-γ和白血病抑制因子的反应中以及在表达不同GHR突变体的细胞中，JAK2酪氨酸磷酸化与IRS-1酪氨酸磷酸化之间的相关性，提供了证据表明IRS-1可能与JAK2或与JAK2结合的辅助分子相互作用。GH还被证明可刺激IRS-1与PI 3'-激酶的85-kDa调节亚基结合。GH刺激IRS-1酪氨酸磷酸化及其与PI 3'-激酶结合的能力，为胰岛素和GH共有的反应提供了生化基础，包括在多种细胞类型中观察到的GH具有特征性的胰岛素样代谢作用。
