title: Gene induction by coagulation factor Xa is mediated by activation of protease-activated receptor 1.
abstract: Cell signaling by coagulation factor Xa (Xa) contributes to pro-inflammatory responses in vivo. This study characterizes the signaling mechanism of Xa in a HeLa cell line that expresses protease-activated receptor 1 (PAR-1) but not PAR-2, -3, or -4. Xa induced NF-kappaB in HeLa cells efficiently but with delayed kinetics compared to thrombin. This delay caused no difference in gene expression patterns, as determined by high-density microarray analysis. Both proteases prominently induced the angiogenesis-promoting gene Cyr61 and connective tissue growth factor. Inhibition of PAR-1 cleavage abolished MAP kinase phosphorylation and gene induction by Xa, demonstrating that Xa signals through PAR-1 and not through a novel member of the PAR family. Activation of cell surface prothrombin with the snake venom enzyme Ecarin also produced PAR-1-dependent signaling. However, though the response to Ecarin was completely blocked by the thrombin inhibitor hirudin, the response to Xa was not. This suggests that the Xa response is not mediated by locally generated thrombin. The concentration dependence of Xa for PAR-1 activation is consistent with previously characterized Xa-mediated PAR-2 signaling, suggesting that local concentration of Xa on the cell surface, rather than sequence-specific recognition of the PAR scissile bond, determines receptor cleavage. This study demonstrates that PAR-1 cleavage by Xa can elicit the same cellular response as thrombin, but mechanistic differences in receptor recognition may be crucial for specific roles for Xa in signaling during spatial or temporal separation from thrombin generation.
authors.affiliation: Department of Immunology and Vascular Biology, The Scripps Research Institute, La Jolla, CA, USA.
authors.forename: M
authors.initials: M
authors.lastname: Riewald
authors.forename: V V
authors.initials: VV
authors.lastname: Kravchenko
authors.forename: R J
authors.initials: RJ
authors.lastname: Petrovan
authors.forename: P J
authors.initials: PJ
authors.lastname: O'Brien
authors.forename: L F
authors.initials: LF
authors.lastname: Brass
authors.forename: R J
authors.initials: RJ
authors.lastname: Ulevitch
authors.forename: W
authors.initials: W
authors.lastname: Ruf
mesh_terms: D000991:Antithrombins / Q000494:pharmacology; D002460:Cell Line; D055513:Connective Tissue Growth Factor; D055515:Cysteine-Rich Protein 61; D004730:Endothelium, Vascular; D004789:Enzyme Activation / Q000187:drug effects; D015951:Factor Xa / Q000494:pharmacology*; D015870:Gene Expression; D006133:Growth Substances / Q000235:genetics; D006367:HeLa Cells; D006629:Hirudins / Q000494:pharmacology; D006801:Humans; D017874:Immediate-Early Proteins / Q000235:genetics; D036341:Intercellular Signaling Peptides and Proteins; D007700:Kinetics; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D016328:NF-kappa B / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D010766:Phosphorylation; D044463:Receptor, PAR-1; D018179:Receptors, Thrombin / Q000235:genetics*; D015398:Signal Transduction; D013917:Thrombin / Q000378:metabolism / Q000494:pharmacology; D014471:Umbilical Veins
chemical_list: D000991:Antithrombins; C501848:CCN1 protein, human; C526317:CCN2 protein, human; D055515:Cysteine-Rich Protein 61; D006133:Growth Substances; D006629:Hirudins; D017874:Immediate-Early Proteins; D036341:Intercellular Signaling Peptides and Proteins; D016328:NF-kappa B; D044463:Receptor, PAR-1; D018179:Receptors, Thrombin; D055513:Connective Tissue Growth Factor; D020928:Mitogen-Activated Protein Kinases; D013917:Thrombin; D015951:Factor Xa