Platelet-derived growth factor-induced signal transduction
Abstract: Platelet-derived growth factor (PDGF), a potent mitogen for mesenchymal cells, elicits its effects by binding to cell surface tyrosine kinase receptors, denoted α- and β-receptors. This thesis describes the mechanism of interaction between stimulated PDGF receptors and various intracellular molecules.PDGF-B is homologous to the transforming protein v-sis encoded by the simian sarcoma virus (SSV). Transformation of cells by the v-sis oncogene occurs through an autocrine mechanism, whereby the v-Sis protein produced by a cell activates the cells own PDGF receptors. We investigated the subcellular location of the autocrine signal and found that in c-Sis/B-chain transformed cells, autophosphorylated intracellular receptors initiated activation of phosphatidylinositol 3' kinase, and tyrosine phosphorylation of phospholipase C-γ (PLC-γ) and Ras GTPase-activating protein (RasGAP). These signals were, however, not sufficient for transformation of the cells.Receptor-association with Grb2 and SHP-2, phosphorylation of Shc and SHP-2 and activation of Raf and Src kinases, on the other hand, correlated with the transformed phenotype. Intermediate filaments (IFs) are one of three major components of the cytoskeleton. We observed a marked reorganization of vimentin containing IFs within 30 minutes of stimulation with PDGF, in which the fine fibrillar vimentin network aggregated into a dense coil around the nucleus. Concomitantly, the amount of soluble vimentin in the cytoplasm was decreased The reorganization of vimentin was dependent on intact organization of actin-containing microfilaments. PDGF stimulation also resulted in increased tyrosine phosphorylation of vimentin, a reaction which appears to involve Src kinase activity.Activation of Stat and Jak proteins in response to PDGF was also investigated. Stat1, Stat3 and Stat5 were tyrosine phosphorylated and activated in response to PDGF. Ligand-stimulated PDGF β-receptor associated with Jak1, Jak2 and Tyk2, and Jak1 was tyrosine phosphorylated in response to PDGF. Activation of both Stats and Jaks was more efficiently mediated by the PDGF β-receptor than the α-receptor. The interaction between β-receptor and Stat5 was investigated further and tyrosine residues 579, 581 and 775 in the β-receptor were found to be important for PDGF-induced phosphorylation and activation of Stat5. In addition, the MAP kinase cascade was found to negatively modulate Stat5 activity in PDGF-stimulated cells, as revealed by use of a MEK specific inhibitor (PD98059). This negative effect by MEK appears not to be exerted through a direct phosphorylation mechanism, since serine phosphorylation of Stat5 was not affected by the presence of the MEK inhibitor.
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