Binding of epidermal growth factor (EGF) to its receptor activates the receptor's tyrosine kinase activity which then phosphorylates itself and other intracellular proteins. These proteins are collectively termed EGF receptor tyrosine kinase substrates. A method to isolate these kinase substrates has been developed. This method involved the isolation of phosphotyrosine-containing proteins by anti-phosphotyrosine antibodies from cells treated with EGF. These immunopurified proteins were then used to immunize rabbits for the production of antisera specific for these proteins. These antisera were subsequently used to screen expression cDNA libraries for potential cDNA clones encoding the EGF receptor kinase substrates. Among the isolated cDNA clones, which are designated eps (epidermal receptor putative substrate), the full-length cDNA clone for eps10 encodes a protein of 583 amino acids. Sequence analysis of eps10 has revealed that it is identical to radixin, a previously identified cytoskeletal protein which shares a high degree of similarity with ezrin and mosein, two members of the band 4.1 family of proteins. These three related proteins were further characterized using specific antibodies and it was shown that ezrin is a substrate for both EGF and platelet-derived growth factor (PDGF) receptor. Radixin is specific for the PDGF receptor and moesin is not phosphorylated by these two receptors. It was also demonstrated that a previously described tumor transplantation antigen of the methylcholanthrene-induced sarcoma is ezrin.
