Lim Y P, Low B C, Ong S H, Guy G R
Signal Transduction Laboratory, Institute of Molecular and Cell Biology, National University of Singapore, Singapore 119076, Republic of Singapore.
J Biol Chem. 1997 Nov 21;272(47):29892-8. doi: 10.1074/jbc.272.47.29892.
The growth factor receptor-binding protein (Grb2) has a key role in initiating the mitogen-activated protein kinase signaling cascade in major cell regulatory pathways. The binding of proteins to the SH2 domain of Grb2 has been reported to occur mainly after they are tyrosine-phosphorylated following receptor activation. Using an in vitro binding assay, immunoprecipitation, and Far Western techniques, we report that in quiescent cells a 75-kDa protein binds directly to the SH2 domain of Grb2. All of the tyrosine-phosphorylated p75 protein co-localizes with Grb2.Sos complex in the cytosolic fraction of the cell in vivo and undergoes tyrosine dephosphorylation when cells are treated with mitogenic ligands such as epidermal, platelet-derived, and fibroblast growth factors, endothelin-1, and bombesin but not tumor necrosis factor-alpha, interferon-alpha and -gamma, interleukein-6, and leukemic inhibitory factor, which are either cell growth inhibitory or not significantly mitogenic. The dephosphorylation of p75 and the ensuing dissociation from Grb2 is rapid, occurring within 30 s following mitogenic stimulation by ligands such as epidermal growth factor, suggesting p75 to be an early component in the signal transduction pathways involving Grb2.
生长因子受体结合蛋白（Grb2）在主要细胞调节途径中启动丝裂原活化蛋白激酶信号级联反应方面发挥着关键作用。据报道，蛋白质与Grb2的SH2结构域结合主要发生在受体激活后它们被酪氨酸磷酸化之后。通过体外结合试验、免疫沉淀和Far Western技术，我们报道在静止细胞中一种75 kDa的蛋白质直接与Grb2的SH2结构域结合。所有酪氨酸磷酸化的p75蛋白在体内细胞的胞质部分与Grb2-Sos复合物共定位，并且当细胞用有丝分裂配体如表皮生长因子、血小板衍生生长因子、成纤维细胞生长因子、内皮素-1和蛙皮素处理时会发生酪氨酸去磷酸化，但用细胞生长抑制或无明显有丝分裂作用的肿瘤坏死因子-α、干扰素-α和-γ、白细胞介素-6和白血病抑制因子处理时则不会。p75的去磷酸化以及随后与Grb2的解离很快，在表皮生长因子等配体的有丝分裂刺激后30秒内发生，这表明p75是涉及Grb2的信号转导途径中的早期成分。
