Mechanism of cAMP-induced H(+)-efflux of Dictyostelium cells: a role for fatty acids.
Aggregating Dictyostelium cells release protons when stimulated with cAMP. To find out whether the protons are generated by acidic vesicles or in the cytosol, we permeabilized the cells and found that this did not alter the cAMP-response. Proton efflux in intact cells was inhibited by preincubation with the V-type H(+) ATPase inhibitor concanamycin A and with the plasma membrane H(+) ATPase blocker miconazole. Surprisingly, miconazole also inhibited efflux in permeabilized cells, indicating that this type of H(+) ATPase is present on intracellular vesicles as well. Vesicular acidification was inhibited by miconazole and by concanamycin A, suggesting that the acidic vesicles contain both V-type and P-type H(+) ATPases. Moreover, concanamycin A and miconazole acted in concert, both in intact cells and in vesicles. The mechanism of cAMP-induced Ca2(+)-fluxes involves phospholipase A2 activity. Fatty acids circumvent the plasma membrane and stimulate vesicular Ca2(+)-efflux. Here we show that arachidonic acid elicited H(+)-efflux not only from intact cells but also from acidic vesicles. The target of regulation by arachidonic acid seemed to be the vesicular Ca2(+)-release channel.
Flaadt
H
H
Faculty of Biology, University of Konstanz, D-78457 Konstanz, Germany.
Schaloske
R
R
Malchow
D
D
D009327:4-Chloro-7-nitrobenzofurazan / Q000494:pharmacology; D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D016718:Arachidonic Acid / Q000494:pharmacology; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology*; D000242:Cyclic AMP / Q000502:physiology*; D004023:Dictyostelium / Q000166:cytology / Q000378:metabolism*; D005227:Fatty Acids / Q000502:physiology*; D005372:Filipin / Q000494:pharmacology; D006859:Hydrogen / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D017136:Ion Transport / Q000187:drug effects; D018942:Macrolides; D008565:Membrane Proteins / Q000037:antagonists & inhibitors / Q000502:physiology*; D008825:Miconazole / Q000494:pharmacology; D008954:Models, Biological; D015388:Organelles / Q000187:drug effects / Q000378:metabolism*; D010741:Phospholipases A / Q000502:physiology; D054467:Phospholipases A2; D006180:Proton-Translocating ATPases / Q000037:antagonists & inhibitors / Q000502:physiology*; D011522:Protons
D000900:Anti-Bacterial Agents; D005227:Fatty Acids; D018942:Macrolides; D008565:Membrane Proteins; D011522:Protons; D016718:Arachidonic Acid; D008825:Miconazole; D006859:Hydrogen; C036978:concanamycin A; D005372:Filipin; D000242:Cyclic AMP; D010741:Phospholipases A; D054467:Phospholipases A2; D006180:Proton-Translocating ATPases; D009327:4-Chloro-7-nitrobenzofurazan