Mechanism of tissue factor activation on HL-60 cells.
Tissue factor (TF) procoagulant activity (PCA) on the surface of intact HL-60 cells is encrypted. This latent TF PCA was activated by exposing the cells to ionomycin, a calcium ionophore. Within seconds an increase in TF PCA of greater than 100-fold was observed. The ionomycin effect was blocked by pretreating the cells with calmidazolium, a calmodulin inhibitor. Changes in TF structure and function, coincident with the ionophore-induced increase in TF PCA, were identified. TF-factor VIIa complexes formed on both untreated and ionophore-treated cells, but pseudosubstrate inhibitors only bound to TF-factor VIIa on the ionophore-treated cells. TF PCA was inhibited by reacting cells with sulfosuccinimidyl-6-(biotinamido)hexanoate, and the rate of this reaction increased twofold after cells were exposed to ionomycin. When proteins on the surface of untreated cells, expressing minimal TF PCA, were cross-linked with 3-3'-dithiobis(sulfosuccinimidylpropionate), cross-linked TF dimers were produced. TF cross-linking was prevented by first treating the cells with ionomycin. These results suggest a mechanism for the ionomycin-induced increase in TF PCA. TF activation appears to be a calmodulin-dependent process, which exposes an essential macromolecular substrate binding site on TF, possibly as the result of a change in TF quaternary structure.
Bach
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Veterans Affairs Medical Center, Research Service (151), Minneapolis, MN 55417, USA.
Moldow
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CF
D000925:Anticoagulants / Q000494:pharmacology; D002147:Calmodulin / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism*; D003432:Cross-Linking Reagents; D004791:Enzyme Inhibitors / Q000494:pharmacology; D015942:Factor VIIa / Q000378:metabolism; D015951:Factor Xa / Q000378:metabolism; D015870:Gene Expression / Q000187:drug effects; D018922:HL-60 Cells; D006801:Humans; D007093:Imidazoles / Q000494:pharmacology; D015759:Ionomycin / Q000494:pharmacology; D007700:Kinetics; D008074:Lipoproteins / Q000494:pharmacology; D046911:Macromolecular Substances; D013388:Succinimides / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013925:Thromboplastin / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism*
D000925:Anticoagulants; D002147:Calmodulin; D003432:Cross-Linking Reagents; D004791:Enzyme Inhibitors; D007093:Imidazoles; D008074:Lipoproteins; D046911:Macromolecular Substances; D013388:Succinimides; C051928:lipoprotein-associated coagulation inhibitor; C031938:calmidazolium; D015759:Ionomycin; D013925:Thromboplastin; D015942:Factor VIIa; D015951:Factor Xa; C011240:dithiobis(succinimidylpropionate); D013755:Tetradecanoylphorbol Acetate