Reconstitution of human epidermal growth factor receptors and the deletion mutants in cultured hamster cells

DNA sequences encoding the human epidermal growth factor (EGF) receptor and various EGF-receptor deletion mutants were transfected into Chinese hamster ovary (CHO) cells devoid of endogenous EGF receptors. A functional human EGF-receptor is expressed on the surface of heterologous CHO cells with the following properties: 1) it exhibits typical high affinity (10%; K(d) = 3 x 10^-10 M) and low affinity (90%; K(d) = 3 x 10^-9 M) binding sites for ¹²⁵I-EGF; 2) it is expressed as a polypeptide of 170,000 molecular weight with intrinsic protein tyrosine kinase activity. EGF stimulates the kinase activity leading to self-phosphorylation and to phosphorylation of exogenous substrate; 3) ¹²⁵I-EGF is rapidly internalized into the CHO cells by receptor mediated endocytosis and 4) EGF stimulates DNA synthesis in the cells expressing the human EGF-receptor. Deletion of 63 amino acids from the C-terminal end of EGF-receptor, which removes two autophosphorylation sites, abolishes the high affinity state of the receptor. Nevertheless, this receptor mutant is able to undergo endocytosis and to respond mitogenically to EGF to a similar extent as the 'wild type' receptor. Further deletions from the cytoplasmic domain give rise to low affinity endocytosis-defective receptor mutants. Finally, deletion of the transmembrane domain of the human receptor yields an EGF-receptor ligand binding domain which is secreted from the cells.