EPIDERMAL GROWTH-FACTOR STIMULATES A PROTEIN-TYROSINE KINASE WHICH ISSEPARABLE FROM THE EPIDERMAL GROWTH-FACTOR RECEPTOR

The intrinsic protein tyrosine kinase activity of the epidermal growth factor (EGF) receptor is necessary for ligand-induced signaling. To d etermine whether cellular protein tyrosine kinases are substrates for EGF-activated receptors, phosphotyrosine-containing proteins were isol ated from EGF-treated cells and assayed for tyrosine kinase activity u sing peptide substrates. A tyrosine kinase activity that is distinct f rom the EGF receptor was adsorbed to monoclonal anti-phosphotyrosine a ntibody columns and eluted with phenyl phosphate. Near-maximal tyrosin e phosphorylation of this kinase occurred within 1 min of cell stimula tion with an ED50 for EGF of 2.5 nM. The kinase was deactivated by inc ubation with purified CD45 tyrosine phosphatase in vitro, but activity could be restored by incubation with purified EGF receptor and Mn2+ A TP. These results suggest a cascade of tyrosine kinase signaling analo gous to well characterized serine/threonine kinase cascades.