The 230 kDa mature form of KDR/Flk-1 (VEGF receptor-2) activates the PLC-gamma pathway and partially induces mitotic signals in NIH3T3 fibroblasts.
KDR/Flk-1 tyrosine kinase, one of the two receptors for Vascular Endothelial Growth Factor (VEGF) has been shown to generate the major part of mitotic signals in endothelial cells, although the mechanisms are poorly understood. Here we examined the processing and signal transduction of KDR/Flk-1. Both in endothelial cells and in NIH3T3 cells expressing KDR/Flk-1, an immature form of KDR/Flk-1 with a molecular mass of about 150 kDa was glycosylated to create a 200 kDa intermediate, and after further glycosylation a mature 230 kDa was expressed on the cell surface. Only this 230 kDa form was rapidly and transiently phosphorylated on tyrosine residues in the presence of VEGF. As a major substrate of KDR/Flk-1, PLC-gamma was found to be rapidly tyrosine-phosphorylated and associated with KDR/Flk-1 both in endothelial cells and NIH3T3 cells. Interestingly, however, a prompt activation of MAP kinase and subsequent strong mitotic signaling were generated only in the endothelial cell background. Activation of MAP kinase in NIH3T3 cells overexpressing KDR/Flk-1 showed a slower response as maximum levels were only attained after 20 min compared to 5 min in sinusoidal endothelial cells. These results suggest that the KDR/Flk-1 utilizes cell type-specific signal transduction pathway(s) for MAP kinase activation and the mitotic response in endothelial cells.
Takahashi
T
T
Department of Genetics, Institute of Medical Science, University of Tokyo, Minato-ku, Japan.
Shibuya
M
M
D016475:3T3 Cells; D000818:Animals; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D016228:Endothelial Growth Factors / Q000494:pharmacology*; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D006031:Glycosylation / Q000187:drug effects; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism*; D008099:Liver / Q000098:blood supply; D008222:Lymphokines / Q000494:pharmacology*; D051379:Mice; D008938:Mitosis; D008970:Molecular Weight; D009928:Organ Specificity; D051966:Phospholipase C gamma; D010766:Phosphorylation / Q000187:drug effects; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D011817:Rabbits; D051381:Rats; D020794:Receptor Protein-Tyrosine Kinases / Q000187:drug effects / Q000502:physiology*; D017978:Receptors, Growth Factor / Q000187:drug effects / Q000502:physiology*; D040262:Receptors, Vascular Endothelial Growth Factor; D011994:Recombinant Proteins / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects; D010738:Type C Phospholipases / Q000378:metabolism*; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors
D016228:Endothelial Growth Factors; D007527:Isoenzymes; D008222:Lymphokines; D017978:Receptors, Growth Factor; D011994:Recombinant Proteins; D042461:Vascular Endothelial Growth Factor A; D042442:Vascular Endothelial Growth Factors; D020794:Receptor Protein-Tyrosine Kinases; D040262:Receptors, Vascular Endothelial Growth Factor; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D010738:Type C Phospholipases; D051966:Phospholipase C gamma