The characterization of the receptor for endothelial cell growth factor by covalent ligand attachment.
Cellular receptors for endothelial cell growth factor (ECGF) have been demonstrated on several cell types by binding of 125I-ECGF in a specific and saturable manner (Schreiber, A. B., Kennedy, J., Kowalski, J., Friesel, R., Mehlman, T., and Maciag, T. (1985) Proc. Natl. Acad. Sci. U. S. A. 82, 6138-6142). Here we report the covalent cross-linking of 125I-ECGF to a polypeptide present on the surface of the plasma membrane of murine lung capillary endothelial cells by the homobifunctional reagent, disuccinimidyl suberate. Cross-linking of cell surface associated 125I-ECGF yields a major polypeptide with an apparent molecular weight of 150,000. Experiments demonstrated that the cross-linked polypeptide complex represents 125I-ECGF covalently bound specifically to a cell surface receptor because: covalent modification of the polypeptide was inhibited by excess, unlabeled ECGF; preincubation of cells with unlabeled ECGF at 37 degrees C significantly reduced cross-linking while incubation at 4 degrees C did not; other polypeptide growth factors do not compete with 125I-ECGF for cross-linking to the ECGF receptor; labeling of the polypeptide did not take place in the absence of DSS; and cells previously shown to have a paucity of ECGF receptors did not yield a cross-linked labeled receptor. These data suggest that the mitogenic events mediated by ECGF occur after occupancy of the specific cell surface polypeptide and suggest that these events are relevant to ECGF-induced signal transduction across the endothelial cell plasma membrane.
D000818:Animals; D001921:Brain / Q000378:metabolism; D002196:Capillaries / Q000378:metabolism; D002417:Cattle; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D016228:Endothelial Growth Factors; D004727:Endothelium / Q000378:metabolism; D006133:Growth Substances / Q000378:metabolism*; D007457:Iodine Radioisotopes; D051379:Mice; D008970:Molecular Weight; D011652:Pulmonary Circulation; D011975:Receptors, Mitogen / Q000302:isolation & purification / Q000378:metabolism*; D040262:Receptors, Vascular Endothelial Growth Factor; D013388:Succinimides / Q000378:metabolism
D016228:Endothelial Growth Factors; D006133:Growth Substances; D007457:Iodine Radioisotopes; D011975:Receptors, Mitogen; D013388:Succinimides; D040262:Receptors, Vascular Endothelial Growth Factor; C019358:disuccinimidyl suberate