Transmembrane signaling by epidermal growth factor receptors lacking autophosphorylation sites.
Mutant epidermal growth factor (EGF) receptors in which the five known tyrosine autophosphorylation sites (tyrosines 992, 1068, 1086, 1148, and 1173) were replaced with phenylalanine residues were expressed in NIH-3T3 cells (5F-EGFR) and transmembrane signaling parameters compared with cells expressing wild-type EGF receptor (WT-EGFR). Mutant and wild-type clones were chosen expressing similar numbers of receptors and Scatchard analysis of 125I-EGF binding showed high and low affinity binding of equal affinities for both receptor types. EGF stimulated tyrosine phosphorylation of proteins to a much lesser degree in cells expressing 5F-EGFR relative to cells expressing WT-EGFR. Tyrosine phosphorylation of the 5F-EGFR was 2-4% of WT-EGFR. Surprisingly, cells expressing WT-EGFR or 5F-EGFR showed little difference in dose response of EGF-stimulated [3H]thymidine incorporation or EGF stimulation of mitogen-activated protein kinase activity. However, EGF did not induce anchorage-independent growth of cells expressing 5F-EGFR to the same extent as it did for cells expressing WT-EGFR. EGF treatment of 5F-EGFR cells failed to elicit an increase in phosphatidylinositol 3-kinase activity or to stimulate hydrolysis of phosphoinositides or tyrosine phosphorylation of phospholipase C-gamma 1. These data suggest that a significant proportion of EGF receptor signaling can occur through receptors with altered capacity to interact with src homology 2 domain-containing proteins.
Decker
S J
SJ
Parke-Davis Pharmaceuticals, Ann Arbor, Michigan 48106.
D016475:3T3 Cells; D000818:Animals; D002448:Cell Adhesion; D002455:Cell Division / Q000187:drug effects; D004261:DNA Replication / Q000187:drug effects*; D004305:Dose-Response Relationship, Drug; D004815:Epidermal Growth Factor / Q000378:metabolism* / Q000494:pharmacology*; D066246:ErbB Receptors / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D051379:Mice; D016297:Mutagenesis, Site-Directed; D019869:Phosphatidylinositol 3-Kinases; D010716:Phosphatidylinositols / Q000378:metabolism; D010766:Phosphorylation; D010770:Phosphotransferases / Q000378:metabolism; D011994:Recombinant Proteins / Q000187:drug effects / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013936:Thymidine / Q000378:metabolism; D014162:Transfection; D014443:Tyrosine
D010716:Phosphatidylinositols; D011994:Recombinant Proteins; D014443:Tyrosine; D004815:Epidermal Growth Factor; D010770:Phosphotransferases; D019869:Phosphatidylinositol 3-Kinases; D066246:ErbB Receptors; D013936:Thymidine