Metalloproteolytic release of endothelial cell protein C receptor.
Previous studies observed that there is about 100 ng/ml soluble endothelial cell protein C receptor (EPCR) in human plasma and that the levels increase in inflammatory diseases. In this study we examine the potential mechanisms involved in release of EPCR from cells. We find that EPCR is released from the surface of endothelium and transfected 293 cells by a metalloprotease in a constitutive fashion. The mass of soluble EPCR is 4 kDa less than intact EPCR. Release is blocked by either the hydroxamic acid based inhibitor, KD-IX-73-4 or by 1,10-phenanthroline, but not by matrix metalloprotease inhibitors. Release is stimulated by phorbol 12-myristate 13-acetate, thrombin, interleukin-1beta, and hydrogen peroxide. Stimulation with these agents reduces EPCR expression levels sufficiently to decrease the rate of protein C activation to a limited extent. The influence of phorbol 12-myristate 13-acetate on both EPCR release and inhibition of protein C activation are enhanced by microtubule disruption with nocodazole. EPCR release is augmented by transfection of EPCR expressing 293 cells with caveolin, suggesting that release is caveolae dependent. These studies indicate that metalloproteolytic release of EPCR is a highly regulated process that is sensitive to both coagulation factors and inflammatory mediators.
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Howard Hughes Medical Institute, Oklahoma Medical Research Foundation, Oklahoma City, Oklahoma 73104, USA.
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D001779:Blood Coagulation Factors; D051242:Caveolin 1; D022461:Caveolins; D002460:Cell Line; D018076:DNA, Complementary / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000819:agonists; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D007375:Interleukin-1 / Q000494:pharmacology; D013058:Mass Spectrometry; D008565:Membrane Proteins / Q000494:pharmacology; D008667:Metalloproteins / Q000378:metabolism*; D015739:Nocodazole / Q000494:pharmacology; D010455:Peptides / Q000494:pharmacology; D011486:Protein C / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000302:isolation & purification / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013917:Thrombin / Q000494:pharmacology; D013997:Time Factors; D014157:Transcription Factors / Q000819:agonists; D014471:Umbilical Veins / Q000187:drug effects / Q000378:metabolism
D001779:Blood Coagulation Factors; C495540:CAV1 protein, human; D051242:Caveolin 1; D022461:Caveolins; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D007375:Interleukin-1; D008565:Membrane Proteins; D008667:Metalloproteins; D010455:Peptides; D011486:Protein C; D011956:Receptors, Cell Surface; D029701:Saccharomyces cerevisiae Proteins; D014157:Transcription Factors; C086677:YAP1 protein, S cerevisiae; C091963:activated protein C receptor; D006861:Hydrogen Peroxide; D013917:Thrombin; D013755:Tetradecanoylphorbol Acetate; D015739:Nocodazole