PROTEASE NEXIN-1, A THROMBIN INHIBITOR, IS REGULATED BY INTERLEUKIN-1AND DEXAMETHASONE IN NORMAL HUMAN FIBROBLASTS

Thrombin participates in several regulatory events following injury as a result of its effects on blood coagulation and cell migration, prol iferation, and differentiation. Protease nexin-1 (PN-1) is a potent th rombin inhibitor in the extracellular environment. Since injury-relate d factors are known to regulate the synthesis and secretion of PN-1, t he inhibitor may serve to modulate the actions of thrombin during inju ry. Here we report the molecular mechanisms that underlie this regulat ion. In normal human fibroblasts, interleukin-1 (IL-1) beta stimulated the synthesis and secretion of PN-1. The stimulation correlated with an increase in steady-state levels of PN-1 mRNA. Treatment of cells wi th both cycloheximide and IL-1 reduced the levels of PN-1 mRNA. Nuclea r run-on assays indicated that IL-1 modestly increased the rate of PN- 1 transcription. However, experiments with actinomycin D demonstrated that IL-1 significantly increased the half-life of the PN-1 mRNA. In c ontrast, dexamethasone (DXM) repressed the synthesis and secretion of PN-1 from fibroblasts. This effect correlated with a decrease in PN-1 mRNA. A sustained decrease in PN-1 mRNA was also seen when cells were treated with cycloheximide and DXM. In nuclear run-on assays, DXM func tioned as a transcriptional repressor of PN-1 synthesis. Treatment of cells with actinomycin D showed that DXM did not affect mRNA stability . Thus, our experiments demonstrate that IL-1 and DXM, which function biologically in different fashions, regulate the synthesis of PN-1 by separate molecular mechanisms. While DXM directly regulates PN-1 at th e level of transcription, IL-1 in the presence ot ongoing protein synt hesis regulates PN-1 production predominantly in a post-transcriptiona l fashion by increasing the half-life of the PN-1 mRNA.