Characterization of the receptors for vascular endothelial growth factor.
Vascular endothelial growth factor (vEGF) is a recently discovered mitogen for endothelial cells. It is also a potent angiogenic factor. We have characterized the vEGF receptors of endothelial cells using both binding and cross-linking techniques. Scatchard analysis of equilibrium binding experiments revealed two types of high-affinity binding sites on the cell surfaces of bovine endothelial cells. One of the sites has a dissociation constant of 10(-12) M and is present at a density of 3 x 10(3) receptors/cell. The other has a dissociation constant of 10(-11) M, with 4 x 10(4) receptors/cell. A high molecular weight complex containing 125I-vEGF is formed when 125I-vEGF is cross-linked to bovine endothelial cells. This complex has an apparent molecular mass of 225 kDa. Two other faintly labeled complexes with apparent molecular masses of 170 and 195 kDa also are detected. Reduction in the presence of dithiothreitol causes a substantial increase in the labeling intensity of the 170- and 195-kDa complexes, suggesting that these complexes are derived from the 225-kDa complex by reduction of disulfide bonds. The labeling of the vEGF receptors was inhibited by an excess of unlabeled vEGF but not by high concentrations of several other growth factors. Suramin and protamine, as well as several species of lectins, inhibited the binding. The expression of functional vEGF receptors was inhibited when the cells were preincubated with tunicamycin, indicating that glycosylation of the receptor is important for the expression of functional vEGF receptors. Pretreatment with swainsonine on the other hand, did not prevent formation of functional receptors. However, the mass of the 225-kDa complex is decreased by 20 kDa when 125I-vEGF is cross-linked to swainsonine-treated endothelial cells.
Vaisman
N
N
Department of Biology, Israel Institute of Technology, Technion City, Haifa.
Gospodarowicz
D
D
Neufeld
G
G
D000818:Animals; D002417:Cattle; D002460:Cell Line; D006224:Cricetinae; D003432:Cross-Linking Reagents; D004220:Disulfides / Q000378:metabolism; D004229:Dithiothreitol / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D016228:Endothelial Growth Factors / Q000378:metabolism*; D004730:Endothelium, Vascular / Q000378:metabolism*; D006031:Glycosylation; D051379:Mice; D008807:Mice, Inbred BALB C; D008970:Molecular Weight; D011479:Protamines / Q000494:pharmacology; D011975:Receptors, Mitogen / Q000037:antagonists & inhibitors / Q000378:metabolism*; D040262:Receptors, Vascular Endothelial Growth Factor; D013498:Suramin / Q000494:pharmacology; D014415:Tunicamycin / Q000494:pharmacology
D003432:Cross-Linking Reagents; D004220:Disulfides; D016228:Endothelial Growth Factors; D011479:Protamines; D011975:Receptors, Mitogen; D014415:Tunicamycin; D013498:Suramin; D040262:Receptors, Vascular Endothelial Growth Factor; D004229:Dithiothreitol