The long term goals of the studies outlined in this proposal are to understand how tyrosine kinase onco-proteins (e.g., pp60src) and effectors of normal mitogenic responses (e.g., growth factors and their receptors) function together to influence cellular growth regulation. Oncogenic transformation mediated by pp60src induces the expression of a novel growth factor receptor, c-sea. C-sea expression is also induced in response to serum stimulation of quiescent fibroblasts, suggesting that c-sea expression may be linked to mitogen induced proliferation. We propose to study the molecular events linking pp60src expression, mitogen stimulation and expression of the c-sea growth factor receptor.
In Aim 1 we will identify and characterize the c-sea gene product. The sequence analysis of chicken c-sea and cloning and sequencing of the human homologue of c-sea will be carried out. Antibodies to both extra-cellular and cytoplasmic domains of the c-sea protein will be used to identify and characterize the c-sea protein. We will use both c-sea cDNA probes and antibodies to determine the tissue and cell type distribution of c-sea expression, and investigate the cell cycle dependent expression of c-sea.
In Aim 2 we will characterize the c-sea protein and assess its possible function as a receptor-like tyrosine protein kinase. We will isolate cell lines over- expressing the complete c-sea gene and various structurally permuted variants of c-sea. These cell lines will be analyzed with respect to morphological phenotype, growth factor responses and tyrosine phosphorylation of the c-sea protein and tyrosine phosphorylation of putative cellular targets for c-sea. We will attempt to activate the over- expressed c-sea receptor in a ligand-dependent manner and initiate experiments to identify the natural ligand for the c-sea receptor.
In Aim 3 we will determine the role of c-sea expression in contributing to and/or mediating the transformed phenotype of src transformed cells. We will attempt to block functional expression of c-sea in src transformed cells and assess the phenotypic consequences of such a block and we will carry out studies to identify the src induced signalling pathways that lead to c- sea induction. The analysis of the c-sea receptor may provide insights into the possible altered expression of c-sea in human and animal tumors and provide information regarding the mechanisms of c-sea gene expression during the cell cycle.

National Institute of Health (NIH)
National Cancer Institute (NCI)
Research Project (R01)
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Virology Study Section (VR)
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University of Virginia
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Huff, J L; Jelinek, M A; Jamieson, T A et al. (1996) Expression and maturation of the cellular sea receptor, a member of the hepatocyte growth factor (HGF) receptor family of protein tyrosine kinases. Oncogene 12:299-307
Huff, J L; Jelinek, M A; Borgman, C A et al. (1993) The protooncogene c-sea encodes a transmembrane protein-tyrosine kinase related to the Met/hepatocyte growth factor/scatter factor receptor. Proc Natl Acad Sci U S A 90:6140-4
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Heaney, M L; Pierce, J; Parsons, J T (1986) Site-directed mutagenesis of the gag-myc gene of avian myelocytomatosis virus 29: biological activity and intracellular localization of structurally altered proteins. J Virol 60:167-76
Wilkerson, V W; Bryant, D L; Parsons, J T (1985) Rous sarcoma virus variants that encode src proteins with an altered carboxy terminus are defective for cellular transformation. J Virol 55:314-21
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