Tyrosine kinase receptors play a central role in normal growth and development as well as in pathophysiologic processes, notably cancer. When activated by ligand these receptors induce a diverse array of responses, and these responses differ between different cell types expressing the same receptor and between different receptor types expressed in the same cell. Understanding the molecular mechanisms of signalling by these receptors is fundamental to understanding the growth and differentiation of cells. The important roles of autophosphorylation and interaction with SH2 domains in signalling by tyrosine kinase receptors are well established. However, it is uncertain whether this accounts for all of the pleiotropic effects of these receptors. In this application we propose to examine two alternative pathways for EGF receptor activation and signalling. I. Phosphorylation and activation of the EGF receptor by pp60src. We have found that the EGF receptor in src transformed cells is constitutively phosphorylated on novel sites and functionally activated. The goals to extend this analysis are: a. Identification of the novel sites of src-induced phosphorylation. B. Analysis of the function of those phosphorylation sites in EGF receptor signalling. C. Determination of the role of these phosphorylation sites in the synergistic interactions between pp60src and the EGF receptor. II. Signalling by kinase-defective EGF receptors. Although receptors coupled to heterotrimeric G-proteins can send regulatory signals without the involvement of tyrosine kinase activity, it has generally been thought that tyrosine kinase receptors depend on their kinase activity for cell signalling. This may not be true in all cases, as it seems clear that a kinase-defective EGF receptor can activate MAP kinase. Since EGF receptors in hepatocytes can signal to G-proteins, and since G-protein agonists can activate MAP kinase, we propose the following parallel investigations. A. Critical investigation of whether tyrosine phosphorylation is activated by """"""""kinase defective"""""""" EGF receptors. B. Direct examination of the interaction of EGF receptors with G- proteins. C. Biochemical analysis of the mechanisms by which G-proteins activate MAP kinase.
