Ca2+/H+ exchange in acidic vacuoles of Trypanosoma brucei.
The use of digitonin to permeabilize the plasma membrane of Trypanosoma brucei procyclic and bloodstream trypomastigotes allowed the identification of a non-mitochondrial nigericin-sensitive Ca2+ compartment. The proton ionophore carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP) was able to cause Ca2+ release from this compartment, which was also sensitive to sodium orthovanadate. Preincubation of the cells with the vacuolar H(+)-ATPase inhibitor bafilomycin A1 greatly reduced the nigericin-sensitive Ca2+ compartment. Bafilomycin A1 inhibited the initial rate of ATP-dependent non-mitochondrial Ca2+ uptake and stimulated the initial rate of nigericin-induced Ca2+ release by permeabilized procyclic trypomastigotes. ATP-dependent and bafilomycin A1- and 7-chloro-4-nitrobenz-2-oxa-1,3-diazole (NBD-Cl)-sensitive Acridine Orange uptake was demonstrated in permeabilized cells. Under these conditions Acridine Orange was concentrated in abundant cytoplasmic round vacuoles by a process inhibited by bafilomycin A1, NBD-Cl, nigericin, and Ca2+. Vanadate or EGTA significantly increased Acridine Orange uptake, while Ca2+ released Acridine Orange from these preparations, thus suggesting that the dye and Ca2+ were being accumulated in the same acidic vacuole. Acridine Orange uptake was reversed by nigericin, bafilomycin A1 and NH4Cl. The results are consistent with the presence of a Ca2+/H(+)-ATPase system pumping Ca2+ into an acidic vacuole, that we tentatively named the acidocalcisome.
Vercesi
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Department of Veterinary Pathobiology, University of Illinois at Urbana-Champaign 61801.
Moreno
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Docampo
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D000165:Acridine Orange / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone / Q000494:pharmacology; D006859:Hydrogen / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D018942:Macrolides; D017494:Proton Pumps / Q000378:metabolism; D014346:Trypanosoma brucei brucei / Q000378:metabolism* / Q000648:ultrastructure; D014617:Vacuoles / Q000378:metabolism
D000900:Anti-Bacterial Agents; D018942:Macrolides; D017494:Proton Pumps; C057620:bafilomycin A; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; D006859:Hydrogen; D000255:Adenosine Triphosphate; D000165:Acridine Orange; D002118:Calcium