EGF RECEPTOR FUNCTION AND CONTROL BY PHOSPHORYLATION

有关细胞生长控制的分子机制的信息是 重要的是了解周围的正常发展的事件， 这些潜在的致癌作用。 表皮生长因子（EGF）是一种 配体刺激的蛋白酪氨酸激酶，经历快速EGF诱导 在其末端羧基-（C-）末端自我磷酸化。 的EGF 受体与逆转录病毒转化蛋白v-erbB同源，并且 它在某些人类肿瘤中过表达。 感兴趣的是两个一般 erbB突变增加其致癌能力：氨基末端 EGF结合位点的缺失，以及各种C-末端 截断。 在这方面，我们的初步研究显示， C-末端自身磷酸化结构域的去除降低了高 亲和结合，增加体内酪氨酸激酶活性并破坏 受体与细胞细胞骨架的结合。 此外，这些和其他 研究强烈表明，受体C-末端结构域和 受体与细胞骨架的相互作用既可以控制 EGF受体结合和激酶活性 因为EGF受体激酶 活动是必要的适当的生物功能，以下 具体目标是：1）检查EGF的机制， 受体C末端调节蛋白酪氨酸激酶活性。 这些 研究将涉及底物特异性、动力学 机制和敏感性的蛋白激活剂正常和C- 表皮生长因子受体 2)EGF的作用 受体C-末端在促进细胞骨架附着，并确定 细胞骨架结合影响受体高亲和力的过程 结合和蛋白酪氨酸激酶活性。 这项工作将侧重于 C端结构域和蛋白激酶C激活对受体影响 细胞骨架分布，也将评估细胞骨架的影响， 对受体高亲和力结合、酪氨酸激酶活性和 底物特异性 3)识别和分析相关的特定蛋白质 在使用直接结合研究的EGF受体-细胞骨架缔合中， 以及免疫共沉淀和交联分析。 这些 预计经验将更严格地建立机制， 哪些特定的结构域，以及它们与细胞 细胞骨架，可以发挥关键作用，在调节EGF受体 在正常和异常细胞生长中起作用。

Information on the molecular mechanisms involved in cell growth control is important for understanding the events surrounding normal development and those underlying carcinogenesis. The epidermal growth factor (EGF) is a ligand-stimulated protein-tyrosine kinase that undergoes rapid EGF-induced self-phosphorylation in its extreme carboxy- (C-) terminus. The EGF receptor is homologous to the retroviral transforming protein v-erbB, and it is overexpressed in certain human tumors. Of interest are two general mutations in erbB that increase its oncogenic capacity: an amino-terminal deletion that removes the EGF binding site, and various C-terminal truncations. In this regard, our initial studies have suggested that removal of the C-terminal self-phosphorylation domain decreases high affinity binding, increases tyrosine kinase activity in vivo and disrupts receptor association with the cell cytoskeleton. Moreover, these and other investigations strongly suggest that the receptor C-terminal domain and receptor interaction with the cell cytoskeleton can both exert control over EGF receptor binding and kinase activities. Because EGF receptor kinase activity is essential for proper biological function, the following specific aims are proposed: 1) Examine the mechanisms by which the EGF receptor C-terminus regulates protein-tyrosine kinase activity. These studies will involve an analysis of the substrate specificity, kinetic mechanism and sensitivity to protein activators of both normal and C- terminally mutated EGF receptors. 2) Characterize the role of the EGF receptor C-terminus in promoting cytoskeletal attachment, and ascertain the processes by which cytoskeletal association affects receptor high affinity binding and protein-tyrosine kinase activity. This work will focus on the influence of C-terminal domains and protein kinase C activation on receptor cytoskeletal distribution, and will also asses the effects of cytoskeletal association on receptor high affinity binding, tyrosine kinase activity and substrate specificity. 3) Identify and analyze specific proteins involved in the EGF receptor-cytoskeletal association using direct binding studies, as well as co-immunoprecipitation and crosslinking analyses. These experiences are expected to more rigorously establish the mechanisms by which specific domains, and their interaction with the cellular cytoskeleton, can play a key role in the regulation of EGF receptor function in both normal and abnormal cell growth.