THE GP185 ERBB-2 AND EPIDERMAL GROWTH FACTOR RECEPTOR SIGNALLING PATHWAYS

GP185 ERBB-2 和表皮生长因子受体信号传导途径

• 批准号: 3853535
• 负责人: P P DI FIORE
• 金额: --
• 依托单位: DIVISION OF CANCER EPIDEMIOLOGY AND GENETICS
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/3853535
• 关键词: antibody; biological signal transduction; chimeric proteins; clone cells; enzyme substrate; epidermal growth factor; fibroblasts; growth factor receptors; guanosinetriphosphatases; human tissue; mitogens; oncogenes; phospholipase C; phosphoproteins; phosphorylation; protein purification; protein tyrosine kinase; receptor coupling

The epidermal growth factor receptor (EGFR) and gp185erbB-2 receptor are closely related tyrosine kinases. Despite extensive sequence and structural homology, they display quantitative and qualitative differences in their ability to couple with mitogenic signalling pathways. Utilizing a chimeric EGFR/erbB-2 molecule, a direct comparison of the biological and biochemical events triggered by activation of the erbB-2 and EGFR kinases in NIH/3T3 cells was possible. Under conditions of comparable expression levels in NIH/3T3 cells, the EGFR/erbB-2 chimera conferred increased responsiveness to EGF compared with EGFR, perhaps due to the different intrinsic ability of the two kinases to phosphorylate intracellular substrates. Biochemical analysis of known mitogenic transduction pathways, however, failed to reveal any major difference in the ability of EGFR of gp185erbB-2 to induce tyrosine phosphorylation of PLC-gamma and GAP. Other second messengers, such as PI-3 kinase and c-raf, implicated as substrates for receptor tyrosine kinases, do not efficiently couple with either erbB-2 or EGFR kinase. Results suggest that other, unknown, signal transduction pathways responsible for the different biological effects of EGFR and gp185erbB-2 might exist. To identify and characterize new intracellular substrates, we purified EGF-induced phosphotyrosine proteins from NIH/3T3 fibroblasts overexpressing EGFR. The purified proteins were then used to immunize animals for the production of polyclonal sera. With this antisera we were able to identify six molecular species which phosphorylated all tyrosine residues upon EGF stimulation. V8 protease digestions showed that none were degradation products of the EGFR. Furthermore, the newly identified species do not correspond to any of the known substrates for tyrosine kinase receptors. The polyclonal antisera were also used for screening bacterial expression libraries with which we characterized a number of cDNAs.

表皮生长因子受体（EGFR）和gp 185 erbB-2 受体是密切相关的酪氨酸激酶。 尽管进行了广泛 序列和结构同源性，它们显示出定量和 它们与促有丝分裂素偶联能力的质的差异 信号通路 利用嵌合EGFR/erbB-2分子， 生物和生化事件的直接比较 由NIH/3 T3中erbB-2和EGFR激酶激活触发 细胞是可能的。 在可比表达的条件下 在NIH/3 T3细胞中，EGFR/erbB-2嵌合体赋予了 与EGFR相比，对EGF的反应性增强，这可能是由于 两种激酶磷酸化的不同内在能力 细胞内底物 已知促有丝分裂素的生化分析 然而，转导途径未能揭示任何主要的 gp 185 erbB-2的EGFR诱导酪氨酸的能力的差异 PLC-γ和GAP的磷酸化。 其他第二信使， 例如PI-3激酶和c-raf，涉及作为 受体酪氨酸激酶，不能有效地与 erbB-2或EGFR激酶。 结果表明，其他未知的信号 转导途径负责不同的生物 EGFR和gp 185 erbB-2可能存在作用。 识别和 表征新的细胞内底物，我们纯化了EGF诱导的 来自NIH/3 T3成纤维细胞的过表达的磷酸酪氨酸蛋白 EGFR对 然后将纯化的蛋白质用于免疫动物， 多克隆血清的产生。 有了这种抗血清， 鉴定了六种磷酸化所有酪氨酸的分子种类， EGF刺激的作用V8蛋白酶消化显示， 没有一个是EGFR的降解产物。 此外，新 所鉴定的物质不对应于任何已知的底物 酪氨酸激酶受体。 还制备了多克隆抗血清， 用于筛选细菌表达文库， 鉴定了一些cDNA。