Reconstitution of human epidermal growth factor receptors and its 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: it exhibits typical high affinity (10%; Kd = 3 X 10(-10) M) and low affinity (90%; Kd = 3 X 10(-9) M) binding sites for 125I-EGF; 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; 125I-EGF is rapidly internalized into the CHO cells by receptor mediated endocytosis and; 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.
D000818:Animals; D002478:Cells, Cultured; D002872:Chromosome Deletion; D006224:Cricetinae; D004247:DNA / Q000378:metabolism; D004261:DNA Replication / Q000187:drug effects; D004815:Epidermal Growth Factor / Q000378:metabolism; D066246:ErbB Receptors; D006801:Humans; D009154:Mutation; D010957:Plasmids; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000235:genetics* / Q000378:metabolism; D013936:Thymidine / Q000378:metabolism
D011956:Receptors, Cell Surface; D004815:Epidermal Growth Factor; D004247:DNA; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases; D013936:Thymidine