The investigators propose a five-year research program to characterize the role of the cytoplasmic, juxtamembrane (JM) domain of the platelet-derived growth factor (PDGF) receptor tyrosine kinase. They previously reported that missense mutations in this region of the PDGF receptor and related receptors caused constitutive receptor activation. Similar mutations have been described in the stem cell factor receptor in some human tumors. In addition, they have presented sequence and biochemical evidence that this region of the receptors contains a WW-like domain, a protein-protein interaction domain present in numerous signaling molecules. They will first systematically construct mutations in the PDGF receptor JM domain, express the mutant receptors in mammalian cells and determine which are constitutively activated. Second, they will conduct a number of in vitro and in vivo assays to investigate the basis of the constitutively active phenotype. Third, they propose a number of approaches to identify proteins that bind the PDGF receptor JM region. Finally, they will confirm that binding of these proteins to the PDGF receptor occurs in intact cells and use their panel of mutants to explore the role of these proteins in receptor activation. These experiments should provide new insights into the function and regulation of receptor tyrosine kinases and into human tumorigenesis, and they should elucidate the binding properties of a new family of WW domains.
