Skolnik E Y, Lee C H, Batzer A, Vicentini L M, Zhou M, Daly R, Myers M J, Backer J M, Ullrich A, White M F
New York University Medical Center, Department of Pharmacology, NY 10016.
EMBO J. 1993 May;12(5):1929-36. doi: 10.1002/j.1460-2075.1993.tb05842.x.
GRB2, a small protein comprising one SH2 domain and two SH3 domains, represents the human homologue of the Caenorhabditis elegans protein, sem-5. Both GRB2 and sem-5 have been implicated in a highly conserved mechanism that regulates p21ras signalling by receptor tyrosine kinases. In this report we show that in response to insulin, GRB2 forms a stable complex with two tyrosine-phosphorylated proteins. One protein is the major insulin receptor substrate IRS-1 and the second is the SH2 domain-containing oncogenic protein, Shc. The interactions between GRB2 and these two proteins require ligand activation of the insulin receptor and are mediated by the binding of the SH2 domain of GRB2 to phosphotyrosines on both IRS-1 and Shc. Although GRB2 associates with IRS-1 and Shc, it is not tyrosine-phosphorylated after insulin stimulation, implying that GRB2 is not a substrate for the insulin receptor. Furthermore, we have identified a short sequence motif (YV/IN) present in IRS-1, EGFR and Shc, which specifically binds the SH2 domain of GRB2 with high affinity. Interestingly, both GRB2 and phosphatidylinositol-3 (PI-3) kinase can simultaneously bind distinct tyrosine phosphorylated regions on the same IRS-1 molecule, suggesting a mechanism whereby IRS-1 could provide the core for a large signalling complex. We propose a model whereby insulin stimulation leads to formation of multiple protein--protein interactions between GRB2 and the two targets IRS-1 and Shc. These interactions may play a crucial role in activation of p21ras and the control of downstream effector molecules.
GRB2是一种由一个SH2结构域和两个SH3结构域组成的小蛋白，是秀丽隐杆线虫蛋白sem-5的人类同源物。GRB2和sem-5都参与了一种高度保守的机制，该机制通过受体酪氨酸激酶调节p21ras信号传导。在本报告中，我们表明，在胰岛素刺激下，GRB2与两种酪氨酸磷酸化蛋白形成稳定复合物。一种蛋白是主要的胰岛素受体底物IRS-1，另一种是含SH2结构域的致癌蛋白Shc。GRB2与这两种蛋白之间的相互作用需要胰岛素受体的配体激活，并由GRB2的SH2结构域与IRS-1和Shc上的磷酸酪氨酸结合介导。尽管GRB2与IRS-1和Shc结合，但在胰岛素刺激后它不会被酪氨酸磷酸化，这意味着GRB2不是胰岛素受体的底物。此外，我们在IRS-1、表皮生长因子受体（EGFR）和Shc中鉴定出一个短序列基序（YV/IN），它能以高亲和力特异性结合GRB2的SH2结构域。有趣的是，GRB2和磷脂酰肌醇-3（PI-3）激酶都能同时结合同一IRS-1分子上不同的酪氨酸磷酸化区域，这提示了一种机制，即IRS-1可以为一个大型信号复合物提供核心。我们提出了一个模型，即胰岛素刺激导致GRB2与两个靶点IRS-1和Shc之间形成多种蛋白质-蛋白质相互作用。这些相互作用可能在p21ras的激活和下游效应分子的控制中起关键作用。
