Thrombin exerts a dual effect on stimulated adenylate cyclase in hamster fibroblasts, an inhibition via a GTP-binding protein and a potentiation via activation of protein kinase C.
Previous studies in Chinese-hamster fibroblasts (CCL39 line) indicate that an important signalling pathway involved in thrombin's mitogenicity is the activation of a phosphoinositide-specific phospholipase C, mediated by a pertussis-toxin-sensitive GTP-binding protein (Gp). The present studies examine the effects of thrombin on the adenylate cyclase system and the interactions between the two signal transduction pathways. We report that thrombin exerts two opposite effects on cyclic AMP accumulation stimulated by cholera toxin, forskolin or prostaglandin E1. (1) Low thrombin concentrations (below 0.1 nM) decrease cyclic AMP formation. A similar inhibition is induced by A1F4-, and both thrombin- and A1F4- -induced inhibitions are abolished by pertussis toxin. (2) Increasing thrombin concentration from 0.1 to 10 nM results in a progressive suppression of adenylate cyclase inhibition and in a marked enhancement of cyclic AMP formation in pertussis-toxin-treated cells. A similar stimulation is induced by an active phorbol ester, and thrombin-induced potentiation of adenylate cyclase is suppressed by down-regulation of protein kinase C. Therefore, we conclude that (1) the inhibitory effect of thrombin on adenylate cyclase is the direct consequence of the activation of a pertussis-toxin-sensitive inhibitory GTP-binding protein (Gi) possibly identical with Gp, and (2) the potentiating effect of thrombin on cyclic AMP formation is due to stimulation of protein kinase C, as an indirect consequence of Gp activation. Our results suggest that the target of protein kinase C is an element of the adenylate cyclase-stimulatory GTP-binding protein (Gs) complex. At low thrombin concentrations, activation of phospholipase C is greatly attenuated by increased cyclic AMP, leading to predominance of the Gi-mediated inhibition.
Magnaldo
I
I
Centre de Biochimie, C.N.R.S., Université de Nice, Faculté des Sciences, Parc Valrose, France.
Pouysségur
J
J
Paris
S
S
D037361:Adenylate Cyclase Toxin; D000262:Adenylyl Cyclases / Q000096:biosynthesis*; D000527:Alprostadil / Q000494:pharmacology; D000535:Aluminum / Q000494:pharmacology; D017607:Aluminum Compounds; D000818:Animals; D002460:Cell Line; D002772:Cholera Toxin / Q000494:pharmacology; D006224:Cricetinae; D003412:Cricetulus; D000242:Cyclic AMP / Q000378:metabolism; D004357:Drug Synergism; D004789:Enzyme Activation / Q000187:drug effects; D004790:Enzyme Induction / Q000187:drug effects; D005347:Fibroblasts / Q000187:drug effects / Q000201:enzymology*; D005459:Fluorides / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000378:metabolism*; D037342:Pertussis Toxin; D015240:Phorbol 12,13-Dibutyrate; D010716:Phosphatidylinositols / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism*; D013917:Thrombin / Q000494:pharmacology*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D037361:Adenylate Cyclase Toxin; D017607:Aluminum Compounds; D010716:Phosphatidylinositols; D010566:Virulence Factors, Bordetella; D015240:Phorbol 12,13-Dibutyrate; D002772:Cholera Toxin; D000535:Aluminum; D000242:Cyclic AMP; D037342:Pertussis Toxin; D011493:Protein Kinase C; D013917:Thrombin; D019204:GTP-Binding Proteins; D000262:Adenylyl Cyclases; D000527:Alprostadil; D005459:Fluorides; C032311:aluminum fluoride