Fatty acids induce release of Ca2+ from acidosomal stores and activate capacitative Ca2+ entry in Dictyostelium discoideum.
cAMP-induced Ca2+ fluxes in Dictyostelium discoideum largely depend on phospholipase A2 activity generating non-esterified fatty acids [Schaloske and Malchow (1997) Biochem. J. 327, 233-238]. In the present study the effect of fatty acids on Ca2+ homoeostasis in D. discoideum was investigated. Cytosolic free Ca2+ concentration ([Ca2+]i) was analysed by digital imaging of single fura2-dextran-loaded cells. Arachidonic acid and linoleic acid induced a transient increase in [Ca2+]i. The concentration of arachidonic acid determined the percentage of responding cells, with the mean height of the increase being dose-independent. In nominally Ca2+-free medium or in the presence of bis-(o-aminophenoxy)ethane-N, N,N',N'-tetra-acetic acid (BAPTA), no [Ca2+]i transient was detectable. In spite of this, we found that (1) arachidonic acid induced Ca2+ release from permeabilized cells and from vesicular fractions at concentrations that elicited Ca2+ influx in intact cells and (2) Ca2+ entry was inhibited by inhibitors of Ca2+-transport ATPases and V-type H+-ATPase, indicating that intracellular Ca2+ release precedes Ca2+ entry. Inhibition studies and mutant analysis point to the acidosomal Ca2+ stores as a target of fatty acids. Although fatty acids can substitute fully for cAMP with respect to Ca2+ influx in wild-type cells, experiments with a mutant strain revealed that cAMP also sensitizes the Ca2+-entry mechanism: cAMP-induced Ca2+ influx was normal in a phospholipase C knockout mutant but influx was fairly insensitive to arachidonic acid in this strain. This defect could be overcome by higher doses of arachidonic acid which cause sufficient Ca2+ to be released from the stores to trigger extracellular Ca2+ entry.
Schaloske
R
R
Fakultät für Biologie, Universität Konstanz, D-78457 Konstanz, Germany.
Sonnemann
J
J
Malchow
D
D
Schlatterer
C
C
D000251:Adenosine Triphosphatases / Q000037:antagonists & inhibitors; D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D016718:Arachidonic Acid / Q000494:pharmacology; D002118:Calcium / Q000493:pharmacokinetics*; D000252:Calcium-Transporting ATPases / Q000378:metabolism; D000242:Cyclic AMP / Q000494:pharmacology; D004023:Dictyostelium / Q000502:physiology*; D005227:Fatty Acids / Q000494:pharmacology*; D005456:Fluorescent Dyes / Q000378:metabolism; D016257:Fura-2 / Q000378:metabolism; D006873:Hydroquinones / Q000494:pharmacology; D019787:Linoleic Acid / Q000494:pharmacology; D018942:Macrolides; D006180:Proton-Translocating ATPases / Q000378:metabolism; D010738:Type C Phospholipases / Q000378:metabolism
D000900:Anti-Bacterial Agents; D005227:Fatty Acids; D005456:Fluorescent Dyes; D006873:Hydroquinones; D018942:Macrolides; C019359:2,5-di-tert-butylhydroquinone; D016718:Arachidonic Acid; C036978:concanamycin A; D019787:Linoleic Acid; D000242:Cyclic AMP; D010738:Type C Phospholipases; D000251:Adenosine Triphosphatases; D006180:Proton-Translocating ATPases; D000252:Calcium-Transporting ATPases; D002118:Calcium; D016257:Fura-2