Tethering of the platelet-derived growth factor beta receptor to G-protein-coupled receptors. A novel platform for integrative signaling by these receptor classes in mammalian cells.
Here we provide evidence to show that the platelet-derived growth factor beta receptor is tethered to endogenous G-protein-coupled receptor(s) in human embryonic kidney 293 cells. The tethered receptor complex provides a platform on which receptor tyrosine kinase and G-protein-coupled receptor signals can be integrated to produce more efficient stimulation of the p42/p44 mitogen-activated protein kinase pathway. This was based on several lines of evidence. First, we have shown that pertussis toxin (which uncouples G-protein-coupled receptors from inhibitory G-proteins) reduced the platelet-derived growth factor stimulation of p42/p44 mitogen-activated protein kinase. Second, transfection of cells with inhibitory G-protein alpha subunit increased the activation of p42/p44 mitogen-activated protein kinase by platelet-derived growth factor. Third, platelet-derived growth factor stimulated the tyrosine phosphorylation of the inhibitory G-protein alpha subunit, which was blocked by the platelet-derived growth factor kinase inhibitor, tyrphostin AG 1296. We have also shown that the platelet-derived growth factor beta receptor forms a tethered complex with Myc-tagged endothelial differentiation gene 1 (a G-protein-coupled receptor whose agonist is sphingosine 1-phosphate) in cells co-transfected with these receptors. This facilitates platelet-derived growth factor-stimulated tyrosine phosphorylation of the inhibitory G-protein alpha subunit and increases p42/p44 mitogen-activated protein kinase activation. In addition, we found that G-protein-coupled receptor kinase 2 and beta-arrestin I can associate with the platelet-derived growth factor beta receptor. These proteins play an important role in regulating endocytosis of G-protein-coupled receptor signal complexes, which is required for activation of p42/p44 mitogen-activated protein kinase. Thus, platelet-derived growth factor beta receptor signaling may be initiated by G-protein-coupled receptor kinase 2/beta-arrestin I that has been recruited to the platelet-derived growth factor beta receptor by its tethering to a G-protein-coupled receptor(s). These results provide a model that may account for the co-mitogenic effect of certain G-protein-coupled receptor agonists with platelet-derived growth factor on DNA synthesis.
Alderton
F
F
Department of Physiology and Pharmacology, Strathclyde Institute for Biomedical Sciences, University of Strathclyde, 27 Taylor St., Glasgow, G4 0NR, Scotland, United Kingdom.
Rakhit
S
S
Kong
K C
KC
Palmer
T
T
Sambi
B
B
Pyne
S
S
Pyne
N J
NJ
D002460:Cell Line; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D008246:Lysophospholipids; D019950:Mitogen-Activated Protein Kinase 1; D037342:Pertussis Toxin; D010766:Phosphorylation; D010982:Platelet-Derived Growth Factor / Q000378:metabolism; D011233:Precipitin Tests; D011485:Protein Binding; D020797:Receptor, Platelet-Derived Growth Factor beta / Q000737:chemistry* / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000737:chemistry*; D011994:Recombinant Proteins / Q000378:metabolism; D015398:Signal Transduction; D013110:Sphingosine / Q000031:analogs & derivatives / Q000378:metabolism; D014162:Transfection; D020032:Tyrphostins / Q000494:pharmacology; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D004791:Enzyme Inhibitors; D008246:Lysophospholipids; D010982:Platelet-Derived Growth Factor; D011956:Receptors, Cell Surface; D011994:Recombinant Proteins; D020032:Tyrphostins; D010566:Virulence Factors, Bordetella; C106250:6,7-dimethoxy-3-phenylquinoxaline; C060506:sphingosine 1-phosphate; D037342:Pertussis Toxin; D020797:Receptor, Platelet-Derived Growth Factor beta; D019950:Mitogen-Activated Protein Kinase 1; D013110:Sphingosine