Binding of platelet-derived growth factor (PDGF) to its cell surface receptors stimulates a variety of biochemical and biological responses. Two distinct PDGF receptors have been identified and each exhibits different binding affinities for the variant PDGF isoforms. The alpha PDGF receptor binds PDGF AA and PDGF BB, whereas the beta PDGF receptor binds only PDGF BB. To determine the region of the alpha PDGF receptor responsible for PDGF AA binding, chimeric molecules between the alpha and beta PDGF receptors were constructed and transfected into the PDGF receptor-negative cell line, 32D. Binding analysis and mitogenic assays revealed that amino acids 1-340 of the alpha PDGF receptor comprised the region of the receptor responsible for PDGF AA binding specificity. This region corresponds to immunoglobulin-like subdomains 1, 2 and 3 of the extracellular portion of the receptor. The mechanism of receptor activation was also studied utilizing recombinant PDGF receptors in the baculovirus expression system. In these experiments, intact Sf9 cells expressing beta PDGF receptors were stimulated with PDGF ligand. treated with a cross-linking agent, lysed, and submitted to western blot analysis. These experiments revealed that at early time points of infection, the receptor is phosphorylated and forms receptor dimers with ligand stimulation. In contrast, at late time points of infection, receptor phosphorylation was not ligand-dependent and appeared to be a function of increasing receptor concentration. Baculovirus-produced recombinant PDGF receptors were also utilized to screen hybridoma supernatants for antireceptor monoclonal antibodies, and a monoclonal antibody specific for human alpha PDGF receptor was identified. This antibody is reactive in ELISA assays, immunoprecipitations, and in western blotting of nonreduced receptor.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Intramural Research (Z01)
Project #
1Z01CP005634-02
Application #
3853538
Study Section
Project Start
Project End
Budget Start
Budget End
Support Year
2
Fiscal Year
1991
Total Cost
Indirect Cost
Name
Division of Cancer Epidemiology and Genetics
Department
Type
DUNS #
City
State
Country
United States
Zip Code