Tenascin, first characterized as a novel glioma-mesenchymal extracellular matrix (GMEM) glycoprotein (Bourdon et al., 1983;1985), is distinctive in its restricted and transient expression in fetal tissues and association with the neovasculature and stroma of tumors. We have shown that tenascin promotes cell attachment, which can be inhibited by peptides derived from tenascin. The interaction of cells with tenascin is mediated by one (TNR1) and perhaps two (TNR2) integrin cell surface receptor(s). In addition, we and others have found that tenascin has a heparin-binding site and likely interacts with matrix and cell surface proteoglycans. The highly selective spatial and temporal expression of tenascin during development, association with the tumor neovasculature, and cell receptor binding indicates a selective role in cell-matrix interactions important to tumor growth and angiogenesis. The role of tenascin will be examined through an analysis of the structure and function of tenascin and its receptor(s). We propose to: 1) complete the cloning and sequencing of tenascin cDNA; 2) make a detailed determination of the structural sites of cell receptor an heparin binding on tenascin. Synthetic peptides derived from tenascin will be examined for receptor or heparin binding activity. Bacterial and mammalian expression of tenascin cDNA-coded fusion proteins and mutants generated by site- mutagenesis and deletion will be used to localize receptor and heparin binding sites on tenascin; 3) characterize the TNR1 and TNR2 tenascin receptors. We will determine the identity and matrix ligand binding characteristics of these receptors though the use of receptor-specific antibodies, amino acid sequencing and ligand binding assays; 4) clone and sequence the novel alpha subunit of TNR1. The results will provide the structure of the receptor alpha subunit and perhaps insights into the function of the receptor. The cloning of the alpha subunit provides cDNA probes to examine receptor expression in cell lines and tissues and makes possible future studies into the role of the receptor; 5) examine the role of tenascin in cell motility, tumor growth, and angiogenesis in four successively more complex model systems (cell monolayers, tumor spheroids, tumor growth on chorio-allantoic membrane, nude mouse tumor xenografts). The goal of these studies is to gain an understanding of the role in tumor growth played by tenascin and its receptor(s) and the structural basis of tenascin-receptor interactions.
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