The long-term objective of this project is a combined biochemical and genetic analysis of the hormonal regulation of cell membrane characteristics relevant to neoplasia. These studies will focus on the glucocorticoid and cyclic nucleotide regulation of the serine protease, plasminogen activator in HTC rat hepatoma cells in culture. We have demonstrated that the glucocorticoid dexamethasone rapidly inhibits plasminogen activator activity, and that this inhibition is secondary to the induction of a 50 kilodalton inhibitor specific for plasminogen activation rather than to a decrease in the amount of activator. We have isolated a unique set of variant HTC cell lines selectively resistant to the dexamethasone inhibition in which glucocorticoids fail to induce the inhibitor. In the third year of this project, the major effort will be aimed at the purification of the dexamethasone-induced inhibitor of plasminogen activator (PAI). We will utilize immunoaffinity chromatography with immobilized antibodies directed against the PA inhibitor isolated from bovine aortic endothelial cells. This antibody cross-reacts with HTC PAI. We will the prepare antibodies to the HTC PAI in order to study its hormonal regulation. Limited proteolysis mediated by the plasminogen activator-plasmin cascade plays a significant role in the behavior of transformed cells as well as in many normal processes. These studies should increase our understanding of the hormonal regulation of this important cell membrane property and of the membrane biology of neoplastic cells. (A)
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