Ballotti R, Baron V, Gautier N, Hainaut P, Scimeca J C, Dolais-Kitabgi J, Lammers R, Schlessinger J, Ullrich A, Van Obberghen E
Institut National de la Santé et de la Recherche Médicale INSERM U145, Faculté de Médecine, Nice, France.
Diabete Metab. 1992;18(1 Pt 2):98-103.
For the insulin receptor and the EGF receptor it is believed that ligand occupancy results in interactions within the heterotetrameric alpha 2 beta 2 insulin receptor or between monomeric EGF receptors. These interactions then activate the intracellular receptor tyrosine kinase which induces receptor autophosphorylation and phosphorylation of cellular substrates. In the present study we have approached the nature of this receptor activation and autophosphorylation. We have investigated whether these phenomena occur via an intra--or an intermolecular process. To this end the following receptor model system consisting of two receptors was co-expressed in NIH 3T3 cells: a kinase inactive human insulin receptor (HIR K1018A) and a chimeric (EIR) receptor corresponding to the extracellular and transmembrane domains of the human EGF receptor and the cytosolic domain of the human insulin receptor beta subunit. Using this system we found that stimulation of the cells with EGF induced tyrosine autophosphorylation of the EGF-insulin receptor chimera (150 kd) and tyrosine phosphorylation of the beta-subunit of the kinase-deficient insulin receptor (95 kd). The phosphopeptides of the autophosphorylated cytoplasmic domain of the EGF-insulin receptor chimera were comparable to those of the transphosphorylated beta subunit of the kinase-deficient insulin receptor and the wild type human insulin receptor. When immunoaffinity purified EGF-insulin receptor hybrids and kinase-deficient insulin receptors were used in a cell lysate phosphorylation assay, it was found that addition of EGF produced [32P]-labeling of both receptor species. In conclusion, we have shown that tyrosine transphosphorylation can occur between homologous receptor domains. This transphosphorylation and transactivation could be a possible mechanism for signal amplification.2+ domain could influence interactions between the receptor and cellular structures and, as such, play a key role in signal transduction.
对于胰岛素受体和表皮生长因子（EGF）受体，人们认为配体占据会导致在异源四聚体α2β2胰岛素受体内或单体EGF受体之间发生相互作用。这些相互作用随后激活细胞内受体酪氨酸激酶，从而诱导受体自身磷酸化以及细胞底物的磷酸化。在本研究中，我们探讨了这种受体激活和自身磷酸化的本质。我们研究了这些现象是通过分子内还是分子间过程发生的。为此，由两个受体组成的以下受体模型系统在NIH 3T3细胞中共表达：一个激酶失活的人胰岛素受体（HIR K1018A）和一个嵌合（EIR）受体，其对应于人EGF受体的细胞外和跨膜结构域以及人胰岛素受体β亚基的胞质结构域。使用该系统，我们发现用EGF刺激细胞会诱导EGF - 胰岛素受体嵌合体（150kd）的酪氨酸自身磷酸化以及激酶缺陷型胰岛素受体β亚基（95kd）的酪氨酸磷酸化。EGF - 胰岛素受体嵌合体自身磷酸化的胞质结构域的磷酸肽与激酶缺陷型胰岛素受体和野生型人胰岛素受体的转磷酸化β亚基的磷酸肽相当。当在细胞裂解物磷酸化测定中使用免疫亲和纯化的EGF - 胰岛素受体杂种和激酶缺陷型胰岛素受体时，发现添加EGF会使两种受体都产生[32P]标记。总之，我们已经表明酪氨酸转磷酸化可以在同源受体结构域之间发生。这种转磷酸化和转激活可能是信号放大的一种可能机制。2 + 结构域可能影响受体与细胞结构之间的相互作用，因此在信号转导中起关键作用。
