title: An H(+)-ATPase regulates cytoplasmic pH in Pneumocystis carinii trophozoites.
abstract: Pneumocystis carinii is an opportunistic fungus which causes interstitial pneumonia in patients with acquired immunodeficiency syndrome (AIDS). Cytoplasmic pH (pHi) regulation in short-term-cultured P. carinii trophozoites was studied using the fluorescent dye 2',7'-bis-(2-carboxyethyl)-5-(-6)-carboxyfluorescein. With an extracellular pH of 7.4, the mean baseline pHi of P. carinii trophozoites was 7.40 +/- 0.10 (n = 8). This steady-state pHi was not significantly affected in the absence of extracellular Na+ or K+. Moreover, steady-state pHi was maintained in the nominal absence of HCO3- and was not affected by the Cl-/HCO(3-)-exchanger inhibitor 4, 4'-di-isothiocyanato-dihydrostilbene-2, 2'-disulphonic acid (100 microM), or the Na+/H(+)-exchanger inhibitor N-ethyl-N-isopropylamiloride (100 microM). In contrast, the general inhibitors of ATPases, N-ethylmaleimide (1 mM), and dicyclohexylcarbodi-imide (100 microM), and the inhibitor of yeast H(+)-ATPase, diethylstilbestrol (12.5-100 microM), decreased pHi, while the K+/H(+)-ATPase inhibitor omeprazole (50-400 microM), and the vacuolar-type H(+)-ATPase inhibitor bafilomycin A1 (1-5 microM) only produced a dose-dependent acidification of the cells when used at high concentrations. In addition, steady-state pHi depended on the availability of cellular ATP, since it was decreased by the ATP synthase inhibitors oligomycin (1 microgram/ml) and sodium azide (1 mM), and by the uncoupler of oxidative phosphorylation carbonyl cyanide p-trifluorophenylhydrazone (1 microM), agents that were able to deplete significantly the intracellular ATP levels. Taken together, these results are consistent with an important role of an H(+)-ATPase similar to those found in other fungi in the regulation of pHi homoeostasis in P. carinii trophozoites.
authors.affiliation: Department of Veterinary Pathobiology, University of Illinois at Urbana-Champaign 61801, USA.
authors.forename: R
authors.initials: R
authors.lastname: Docampo
authors.forename: N M
authors.initials: NM
authors.lastname: Vanderheyden
authors.forename: M M
authors.initials: MM
authors.lastname: Shaw
authors.forename: P J
authors.initials: PJ
authors.lastname: Durant
authors.forename: M S
authors.initials: MS
authors.lastname: Bartlett
authors.forename: J W
authors.initials: JW
authors.lastname: Smith
authors.forename: G L
authors.initials: GL
authors.lastname: McLaughlin
mesh_terms: D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D001386:Azides / Q000494:pharmacology; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone / Q000494:pharmacology; D002460:Cell Line; D003593:Cytoplasm / Q000378:metabolism*; D004024:Dicyclohexylcarbodiimide / Q000494:pharmacology; D004054:Diethylstilbestrol / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005033:Ethylmaleimide / Q000494:pharmacology; D005452:Fluoresceins / Q000378:metabolism; D005456:Fluorescent Dyes / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007476:Ionophores / Q000494:pharmacology; D008168:Lung / Q000382:microbiology; D011010:Pneumocystis / Q000201:enzymology*; D011188:Potassium / Q000494:pharmacology; D006180:Proton-Translocating ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D012964:Sodium / Q000494:pharmacology; D019810:Sodium Azide; D013439:Sulfhydryl Reagents / Q000494:pharmacology
chemical_list: D001386:Azides; D004791:Enzyme Inhibitors; D005452:Fluoresceins; D005456:Fluorescent Dyes; D007476:Ionophores; D013439:Sulfhydryl Reagents; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; D004024:Dicyclohexylcarbodiimide; D004054:Diethylstilbestrol; C043829:2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein; D019810:Sodium Azide; D012964:Sodium; D006180:Proton-Translocating ATPases; D005033:Ethylmaleimide; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D011188:Potassium