Understanding the molecular basis of cancer has been a major goal of the biological sciences for many years. This work was accelerated with the discovery that the retroviral oncogenes represented normal cellular genes that were either mutated or expressed inappropriately. However, for the majority of these genes, the determination of their actual function has proved elusive. The discovery that the v-sis oncogene was homologous to the B-chain of the heterodimeric platelet-derived growth factor (PDGF) was one of the first findings of a direct link between these genes and molecules that controlled the cell cycle. The study of PDGF has given and will continue to yield extensive insights into the control of the cell cycle and the process of tumorigenesis. The long-term objective of this application is to identify some of the proteins that are necessary for PDGF to induce transformation and eventually relate these to the process of tumorigenesis. The physiological action of PDGF is controlled by the interaction of the PDGF with the two receptors, alpha and beta. The B-chain binds to both types while the A-chain binds only to the type alpha receptor. The ability of the A-chain to bind to and signal through the type alpha receptor appears to be highly correlated with its ability to induce foci of cells in cultaure. However the secondary signaling mechanisms necessary for transformation which are used by the type beta receptor but not the type alpha receptor are not understood or known. The only differences that are known to date about the autophosphorylated tyrosine residues which are used to control the interactions of the receptors and the secondary signaling molecules are that two residues are unique to the type beta receptor. The hypothesis of this grant is that at least one of these residues accounts for the difference in the ability of the type beta receptor to transform cells.
The specific aims of this project are to create receptors that consist primarily of the type alpha receptor but also have the unique tyrosines from the type beta receptor substituted at the appropriate positions. The ability of these receptors to allow the A-chain to transform cells will be assessed. These receptors will be created by PCR and standard cloning techniques. The Syp and GRB2 proteins, which are known to associate with these unique tyrosine residues, will also be mutagenized by PCR to create proteins that will interfere with the normal functioning of the endogenous protein (dominant negative mutants). Furthermore, as these proteins are known to affect wither ras and/or the activation of the MAP kinases, the activity of the MAP kinases will also be determined. These experiments will add significantly to our understanding of the mechanisms involved in PDGF transformation and tumorigenesis.
| Shimamura, Takeshi; Hsu, Tin-Chen; Colburn, Nancy H et al. (2002) Activation of NF-kappaB is required for PDGF-B chain to transform NIH3T3 cells. Exp Cell Res 274:157-67 |
| Murphy, K; Shimamura, T; Bejcek, B E (2001) Use of fluorescently labeled DNA and a scanner for electrophoretic mobility shift assays. Biotechniques 30:504-6, 508 |
| Jazayeri, A; McGee, J; Shimamura, T et al. (2000) SHP-2 can suppress transformation induced by platelet-derived growth factor. Exp Cell Res 254:197-203 |
