Further evidence for a phospholipase C-coupled G protein in hamster fibroblasts. Induction of inositol phosphate formation by fluoroaluminate and vanadate and inhibition by pertussis toxin.
We have previously reported that alpha-thrombin induces in resting hamster fibroblasts (CCL39) the formation of inositol phosphates (IP) by activating a GTP-binding protein (G protein) sensitive to pertussis toxin (Paris, S., and Pouysségur, J. (1986) EMBO J. 5, 55-60). Here we show that IP formation in CCL39 cells can also be induced by NaF with AlCl3 and by vanadate. In the presence of Li+, IP accumulation is linear over 30 min with no detectable lag and is concentration-dependent. NaF alone is slightly stimulatory, but a marked potentiation is observed in the presence of AlCl3, by itself without effect. Maximal stimulation is obtained with 10 mM NaF and 3 microM AlCl3, and with vanadate half-maximal effect is achieved at 0.3 mM. Both stimulations are markedly inhibited (up to 80%) by pertussis toxin (half-maximal inhibition at 1-2 ng/ml). We therefore conclude that phospholipase C is stimulated by NaF plus AlCl3 (presumably acting as AlF-4) and by vanadate by direct activation of the regulatory G protein. In addition, NaF inhibits the inositol-1-phosphatase, but this effect is not potentiated by AlCl3. Similarly, vanadate inhibits inositol trisphosphate degradation. Maximal stimulations of phospholipase C by AlF-4 and vanadate are not additive, whereas they are both additive with thrombin effects. Pretreatment of cells for 15 min with the phorbol ester 12-O-tetradecanoylphorbol-13-acetate nearly completely abolishes induction of IP formation by AlF-4 and vanadate, suggesting that protein kinase C exerts a feedback negative control either on the G protein or on phospholipase C itself. An increase in cellular cyclic AMP similarly results in a marked attenuation of AlF-4-induced IP formation, indicating that activation of phospholipase C can be controlled also by cyclic AMP. However, the stimulatory effect of AlF-4 on phospholipase C is clearly dissociated from its effect on the adenylate cyclase system.
D037361:Adenylate Cyclase Toxin; D000527:Alprostadil / Q000494:pharmacology; D000535:Aluminum / Q000494:pharmacology*; D000077410:Aluminum Chloride; D017607:Aluminum Compounds; D000818:Animals; D002460:Cell Line; D002712:Chlorides / Q000494:pharmacology; D006224:Cricetinae; D003412:Cricetulus; D000242:Cyclic AMP / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004357:Drug Synergism; D005347:Fibroblasts / Q000201:enzymology*; D005459:Fluorides; D005461:Fluorine / Q000494:pharmacology*; D019204:GTP-Binding Proteins / Q000378:metabolism*; D007295:Inositol Phosphates / Q000096:biosynthesis*; D008094:Lithium / Q000494:pharmacology; D037342:Pertussis Toxin; D012969:Sodium Fluoride / Q000494:pharmacology; D013403:Sugar Phosphates / Q000096:biosynthesis*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D013917:Thrombin / Q000494:pharmacology; D010738:Type C Phospholipases / Q000378:metabolism*; D014638:Vanadates; D014639:Vanadium / Q000494:pharmacology*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology*
D037361:Adenylate Cyclase Toxin; D017607:Aluminum Compounds; D002712:Chlorides; D007295:Inositol Phosphates; D013403:Sugar Phosphates; D010566:Virulence Factors, Bordetella; D014639:Vanadium; C050992:tetrafluoroaluminate; D005461:Fluorine; D000077410:Aluminum Chloride; D014638:Vanadates; D012969:Sodium Fluoride; D008094:Lithium; D000535:Aluminum; D000242:Cyclic AMP; D037342:Pertussis Toxin; D010738:Type C Phospholipases; D013917:Thrombin; D019204:GTP-Binding Proteins; D000527:Alprostadil; D013755:Tetradecanoylphorbol Acetate; D005459:Fluorides