Thrombin promotes actin polymerization in U937 human monocyte-macrophage cells. Analysis of the signalling mechanisms mediating actin polymerization.
The U937 human monocyte-macrophage cell line was used to examine the effect of thrombin, an ill-defined chemoattractant, on the polymerization of actin, a process essential for cell motility. In differentiated macrophage-like U937 cells, thrombin (0.5-50 units/ml) caused a rapid dose-dependent increase in the formation of filamentous (F-) actin, detected by the staining of F-actin with the fluorescent toxin, 7-nitrobenz-2-oxa-1,3-diazole-phallacidin. In contrast with other chemoattractants such as N-formylmethionyl-leucylphenylalanine or C5a, actin polymerization in response to thrombin occurred via a pertussis-toxin-insensitive G1-(inhibitory G-protein) independent signalling pathway. Further, this response was not affected by the Ca2+ chelator EGTA or by the specific protein kinase C (PKC) inhibitor RO-31-8220. The response to thrombin was not mimicked by the Ca2+ ionophore ionomycin or by the direct PKC activator phorbol 12-myristate 13-acetate. The thrombin response was, however, inhibited by the non-specific protein kinase inhibitor staurosporine. The present results suggest that in U937 cells thrombin stimulates the formation of F-actin via a signalling pathway independent of (i) the activation of PKC, (ii) the mobilization of intracellular Ca2+ and (iii) the activation of Ca(2+)-dependent protein kinases, but dependent on the activation of an undefined staurosporine-sensitive protein kinase.
Joseph
S
S
Department of Clinical Pharmacology, Royal Postgraduate Medical School, London, U.K.
MacDermot
J
J
D000199:Actins / Q000378:metabolism*; D000470:Alkaloids / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism; D002413:Cations, Divalent; D002460:Cell Line; D002634:Chemotaxis, Leukocyte; D019204:GTP-Binding Proteins / Q000378:metabolism; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D015759:Ionomycin / Q000494:pharmacology; D008264:Macrophages / Q000187:drug effects / Q000378:metabolism*; D009000:Monocytes / Q000187:drug effects / Q000378:metabolism*; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D037342:Pertussis Toxin; D010972:Platelet Activating Factor / Q000494:pharmacology; D011108:Polymers; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D015398:Signal Transduction; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013917:Thrombin / Q000378:metabolism*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D000199:Actins; D000470:Alkaloids; D002413:Cations, Divalent; D007211:Indoles; D010972:Platelet Activating Factor; D011108:Polymers; D010566:Virulence Factors, Bordetella; D015759:Ionomycin; D009240:N-Formylmethionine Leucyl-Phenylalanine; D037342:Pertussis Toxin; D011493:Protein Kinase C; D013917:Thrombin; D019204:GTP-Binding Proteins; D019311:Staurosporine; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium; C064758:Ro 31-8220