Platelet derived growth factor (PDGF)--the product of c-sis proto-oncogene (p28sis), initiates cellular replication and transformation by binding to a high affinity cell surface receptor. Recently I purified the human PDGF-receptor and demonstrated for the first time the presence of PDGF-stimulatable tyrosine kinase activity in the solubilized and purified receptor preparation. One of the objectives of this proposal is to purify the receptor to homogeneity and to test rigorously whether the tyrosine kinase is intrinsic to the receptor.
Other aims relate to studies on biosynthesis, processing, subcellular localization and endocytic fate of the receptor. My specific objectives are as follows:
Aim 1. Further purification of human and murine PDGF-receptor to homogeneity and structural/functional characterization of the purified receptor. The purified receptor will be tested for its ability to be convalently labeled by ATP analogues. The domain sub-structure of the receptor will be studied by limited proteolysis. The receptor fragment representing the putative tyrosine kinase domain will be identified by autophosphorylation assay and by affinity labeling with a radiolabeled ATP analogue. The fragment representing the growth factor binding domain will be identified by 125I-PDGF binding and by chemical cross-linking to 125I-PDGF.
Aim 2. Generation of polyclonal and monoclonal antibodies to different functional sites of the receptor. The antibodies will be classified according to their specificity for various sites, such as PDGF-binding site, tyrosine kinase site, etc. All the antibodies will be tested for PDGF-like mitogenic activity.
Aim 3. Use the antibodies to investigate biosynthesis, turnover, glycosylation and maturation of the PDGF-receptor.
Aim 4. Studies on endocytic fate and recycling behavior of the PDGF-receptor in response to PDGF. These studies will involve indirect immunofluorescent staining and immunogold electron microscopy of cells using appropriate anti-receptor antibodies. Recycling behaviour of the internalized receptor will be examined by both immunocythchemical techniques and by 125I-PDGF binding assay. In addition, I plan to study localization of the receptor in simian sarcoma virus (SSV)-transformed cells.

National Institute of Health (NIH)
National Cancer Institute (NCI)
First Independent Research Support & Transition (FIRST) Awards (R29)
Project #
Application #
Study Section
Molecular Cytology Study Section (CTY)
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
Children's Hospital of Philadelphia
United States
Zip Code
Bishayee, S (1997) A novel 200 kDa plasma membrane glycoprotein with high basal tyrosine kinase activity in tumour cells. Indian J Biochem Biophys 34:18-24
Panneerselvam, K; Kanakaraj, P; Raj, S et al. (1995) Characterization of a novel epidermal-growth-factor-receptor-related 200-kDa tyrosine kinase in tumor cells. Eur J Biochem 230:951-7
Raj, S; Kanakaraj, P; Khan, S A et al. (1992) Type-specific antibodies to the platelet-derived growth factor receptors: role in elucidating the structural and functional characteristics of receptor types. Biochemistry 31:1774-9
Kanakaraj, P; Raj, S; Khan, S A et al. (1991) Ligand-induced interaction between alpha- and beta-type platelet-derived growth factor (PDGF) receptors: role of receptor heterodimers in kinase activation. Biochemistry 30:1761-7
Basu, A; Raghunath, M; Bishayee, S et al. (1989) Inhibition of tyrosine kinase activity of the epidermal growth factor (EGF) receptor by a truncated receptor form that binds to EGF: role for interreceptor interaction in kinase regulation. Mol Cell Biol 9:671-7
Bishayee, S; Majumdar, S; Khire, J et al. (1989) Ligand-induced dimerization of the platelet-derived growth factor receptor. Monomer-dimer interconversion occurs independent of receptor phosphorylation. J Biol Chem 264:11699-705
Grundy, P; Bishayee, S; Disa, S et al. (1989) Modulation of platelet-derived growth factor receptor function in BP3T3, a chemically transformed BALB/c-3T3 cell line. Cancer Res 49:3581-6
Bishayee, S; Majumdar, S; Scher, C D et al. (1988) Characterization of a novel anti-peptide antibody that recognizes a specific conformation of the platelet-derived growth factor receptor. Mol Cell Biol 8:3696-702