Separate endocytic pathways of kinase-defective and -active EGF receptor mutants expressed in same cells.
Ligand binding to the membrane receptor for EGF induces its clustering and internalization. Both receptor and ligand are then degraded by lysosomal enzymes. A kinase defective point mutant (K721A) of EGF receptor undergoes internalization similarly to the wild-type receptor. However, while internalized EGF molecules bound to either the wild-type or mutant receptors are degraded, the K721A mutant receptor molecules recycle to the cell surface for reutilization. To investigate the mechanism of receptor trafficking, we have established transfected NIH-3T3 cells coexpressing the kinase-negative mutant (K721A) together with a mutant EGF receptor (CD63) with active kinase. CD63 was chosen because it behaves like wild-type EGF receptor with respect to biological responsiveness and cellular routing but afforded immunological distinction between kinase active and inactive mutants. Although expressed in the same cells, the two receptor mutants followed their separate endocytic itineraries. Like wild-type receptor, the CD63 mutant was downregulated and degraded in response to EFG while the kinase-negative mutant K721A returned to the cell surface for reutilization. Intracellular trafficking of EGF receptor must be determined by a sorting mechanism that specifically recognizes EGF receptor molecules according to their intrinsic kinase activity.
Honegger
A M
AM
Rorer Biotechnology, Inc., King of Prussia, Pennsylvania 19406.
Schmidt
A
A
Ullrich
A
A
Schlessinger
J
J
D000818:Animals; D002999:Clone Cells; D003432:Cross-Linking Reagents; D015536:Down-Regulation; D004705:Endocytosis / Q000502:physiology*; D004815:Epidermal Growth Factor / Q000378:metabolism; D066246:ErbB Receptors / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D005022:Ethyldimethylaminopropyl Carbodiimide; D006207:Half-Life; D007457:Iodine Radioisotopes; D007700:Kinetics; D008024:Ligands; D009154:Mutation; D011505:Protein-Tyrosine Kinases / Q000235:genetics / Q000502:physiology*; D014162:Transfection
D003432:Cross-Linking Reagents; D007457:Iodine Radioisotopes; D008024:Ligands; D004815:Epidermal Growth Factor; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases; D005022:Ethyldimethylaminopropyl Carbodiimide