PLASMINOGEN ACTIVATOR INHIBITOR TYPE 1: STUDIES ON STRUCTURE AND REGULATION

Abstract

Human plasminogen activator inhibitor type-1 is an Mr∼54,000 protein which specifically inhibits urokinase-type (u-PA) and tissue-type (t-PA) plasminogen activators. During inhibition, u-PA and t-PA convert PAI-1 to an inactive form with Mr∼50,000. We have determined the amino-terminal amino acid sequence of native and converted PAI-1, and isolated and partly sequenced PAI-1 cDNA. The data show that the conversion of PAI-1 consists of cleavage of an Arg-Met bond 33 residues from the carboxy-terminus, thus localizing the reactive center of the inhibitor to that position, and identifying PAI-1 as an "arg-serpin". PAI-1 activity is known to be influenced by a number of agents; we have studied the mechanisms of the stimulation of PAI-1 activity by transforming growth factor-β (TGF-β) and the synthetic glucocorticoid dexame-thasone in human WI-38 lung fibroblasts and HT-1080 fibrosarcoma cells. Bytheuse of PAI-1 cDNA, TGF-β was found to course a rapid increase in PAI-1 mRNA level in WI-38 cells, reaching a maximal 50-fold enhancement after 8 hours. Dexamethasone caused a 10-fold increase in PAI-1 mRNA in HT-1080 cells, which was detectable after 4 hours and became maximal after 16 hours. In both cases, the 3.4 as well as the 2.4 Kb-PAI-1-mRNA species were increased. Quantitative studies on the effect of these agents on PAI-1 protein levels in cell extracts and culture media by ELISA gave results consistent with the effects on PAI-1 mRNA. These studies suggest that TGF-β and glucocorticoids may exert important controls over plasminogen activation-mediated extracellular proteolysis through an enhancement of PAI-1 gene transcription.