Demonstration of epidermal growth factor-induced receptor dimerization in living cells using a chemical covalent cross-linking agent.
We have used the soluble covalent cross-linking agent 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide (EDAC) to examine the capacity of epidermal growth factor (EGF) to stimulate the dimerization of purified EGF receptor, of EGF receptor in membrane preparations and in intact A431 cells. The addition of EGF either to membranes from A431 cells or to EGF receptor which was purified from A431 cells by immunoaffinity chromatography caused the appearance of a cross-linked product of Mr 340,000 which was identified using EGF receptor-specific antibodies as an EGF receptor dimer. Three independent approaches including biosynthetic labeling, surface iodination, and immunoblotting experiments were utilized to follow EGF receptor dimerization in living A431 cells. These approaches provided consistent results indicating that EGF induced rapid dimerization of EGF receptor in living cells, suggesting that this process may play a role in transmembrane signalling mediated by EGF.
Cochet
C
C
Department of Chemical Immunology, Weizmann Institute of Science, Rehovot, Israel.
Kashles
O
O
Chambaz
E M
EM
Borrello
I
I
King
C R
CR
Schlessinger
J
J
D002234:Carbodiimides; D002294:Carcinoma, Squamous Cell; D002846:Chromatography, Affinity; D003432:Cross-Linking Reagents; D004591:Electrophoresis, Polyacrylamide Gel; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D066246:ErbB Receptors / Q000378:metabolism; D005022:Ethyldimethylaminopropyl Carbodiimide; D006801:Humans; D007118:Immunoassay; D007700:Kinetics; D046911:Macromolecular Substances; D008970:Molecular Weight; D010766:Phosphorylation; D014407:Tumor Cells, Cultured
D002234:Carbodiimides; D003432:Cross-Linking Reagents; D046911:Macromolecular Substances; D004815:Epidermal Growth Factor; D066246:ErbB Receptors; D005022:Ethyldimethylaminopropyl Carbodiimide