Protease nexin-1, a thrombin inhibitor, is regulated by interleukin-1 and 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, proliferation, and differentiation. Protease nexin-1 (PN-1) is a potent thrombin inhibitor in the extracellular environment. Since injury-related factors are known to regulate the synthesis and secretion of PN-1, the inhibitor may serve to modulate the actions of thrombin during injury. Here we report the molecular mechanisms that underlie this regulation. 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 with both cycloheximide and IL-1 reduced the levels of PN-1 mRNA. Nuclear 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 contrast, 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 functioned 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 the level of transcription, IL-1 in the presence of ongoing protein synthesis regulates PN-1 production predominantly in a post-transcriptional fashion by increasing the half-life of the PN-1 mRNA.
Guttridge
D C
DC
Department of Microbiology, College of Medicine, University of California, Irvine 92717.
Lau
A L
AL
Cunningham
D D
DD
D016564:Amyloid beta-Protein Precursor; D002352:Carrier Proteins / Q000235:genetics* / Q000378:metabolism / Q000494:pharmacology; D003513:Cycloheximide / Q000494:pharmacology; D016207:Cytokines / Q000494:pharmacology; D003609:Dactinomycin / Q000494:pharmacology; D003907:Dexamethasone / Q000494:pharmacology*; D004355:Drug Stability; D005109:Extracellular Matrix / Q000378:metabolism; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism*; D005786:Gene Expression Regulation; D006801:Humans; D007375:Interleukin-1 / Q000494:pharmacology*; D058255:Protease Nexins; D012333:RNA, Messenger / Q000378:metabolism; D011956:Receptors, Cell Surface; D058258:Serpin E2; D013917:Thrombin / Q000037:antagonists & inhibitors* / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects
D016564:Amyloid beta-Protein Precursor; D002352:Carrier Proteins; D016207:Cytokines; D007375:Interleukin-1; D058255:Protease Nexins; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; C545486:SERPINE2 protein, human; D058258:Serpin E2; D003609:Dactinomycin; D003907:Dexamethasone; D003513:Cycloheximide; D013917:Thrombin