Skip to main content
Biochemical Journal logoLink to Biochemical Journal
. 1996 Aug 15;318(Pt 1):103–109. doi: 10.1042/bj3180103

The role of a H(+)-ATPase in the regulation of cytoplasmic pH in Trypanosoma cruzi epimastigotes.

N Vanderheyden 1, G Benaim 1, R Docampo 1
PMCID: PMC1217594  PMID: 8761458

Abstract

Cytoplasmic pH (pHi) regulation was studied in Trypanosoma cruzi epimastigotes using fluorescent probes. Steady-state pHi was maintained even in the absence of extracellular Na+ or K+, but was significantly decreased in the absence of Cl-. Acid-loaded epimastigotes regained normal pHi by a process that was ATP-dependent and sensitive to N-ethylmaleimide, dicyclohexyl-carbodi-imide and diethylstiboestrol, suggesting involvement of a H(+)-pumping ATPase. Recovery from an acid load was independent of extracellular Na+ or K+ and insensitive to omeprazole, vanadate and low concentrations of bafilomycin A1. Using the fluorescent probe bisoxonol to measure the membrane potential of intact cells, acid loading of epimastigotes was shown to result in a dicyclohexylcarbodi-imide-sensitive hyperpolarization, which suggests electrogenic pumping of protons across the plasma membrane. Addition of glucose, but not of 6-deoxyglucose, produced a transient cellular acidification of possible metabolic origin, and increased the rate of recovery from an acid load. Taken together, these results are consistent with an important role of a H(+)-ATPase in the regulation of pHi homoeostasis in T. cruzi.

Full Text

The Full Text of this article is available as a PDF (565.2 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Anderson S. A., Jiang S., Mukkada A. J. The beta-aspartyl phosphate intermediate in a Leishmania donovani promastigote plasma membrane P-type ATPase. Biochim Biophys Acta. 1994 Oct 12;1195(1):81–88. doi: 10.1016/0005-2736(94)90012-4. [DOI] [PubMed] [Google Scholar]
  2. Anderson S. A., Mukkada A. J. Biochemical and immunochemical characterization of a P-type ATPase from Leishmania donovani promastigote plasma membrane. Biochim Biophys Acta. 1994 Oct 12;1195(1):71–80. doi: 10.1016/0005-2736(94)90011-6. [DOI] [PubMed] [Google Scholar]
  3. Aronson P. S. Kinetic properties of the plasma membrane Na+-H+ exchanger. Annu Rev Physiol. 1985;47:545–560. doi: 10.1146/annurev.ph.47.030185.002553. [DOI] [PubMed] [Google Scholar]
  4. Benaim G., Losada S., Gadelha F. R., Docampo R. A calmodulin-activated (Ca(2+)-Mg2+)-ATPase is involved in Ca2+ transport by plasma membrane vesicles from Trypanosoma cruzi. Biochem J. 1991 Dec 15;280(Pt 3):715–720. doi: 10.1042/bj2800715. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Benaim G., Moreno S. N., Hutchinson G., Cervino V., Hermoso T., Romero P. J., Ruiz F., de Souza W., Docampo R. Characterization of the plasma-membrane calcium pump from Trypanosoma cruzi. Biochem J. 1995 Feb 15;306(Pt 1):299–303. doi: 10.1042/bj3060299. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Boron W. F. Transport of H+ and of ionic weak acids and bases. J Membr Biol. 1983;72(1-2):1–16. doi: 10.1007/BF01870311. [DOI] [PubMed] [Google Scholar]
  7. Bowman E. J., Siebers A., Altendorf K. Bafilomycins: a class of inhibitors of membrane ATPases from microorganisms, animal cells, and plant cells. Proc Natl Acad Sci U S A. 1988 Nov;85(21):7972–7976. doi: 10.1073/pnas.85.21.7972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Cabantchik Z. I., Knauf P. A., Rothstein A. The anion transport system of the red blood cell. The role of membrane protein evaluated by the use of 'probes'. Biochim Biophys Acta. 1978 Sep 29;515(3):239–302. doi: 10.1016/0304-4157(78)90016-3. [DOI] [PubMed] [Google Scholar]
  9. Cantley L. C., Jr, Aisen P. The fate of cytoplasmic vanadium. Implications on (NA,K)-ATPase inhibition. J Biol Chem. 1979 Mar 25;254(6):1781–1784. [PubMed] [Google Scholar]
  10. Docampo R., Scott D. A., Vercesi A. E., Moreno S. N. Intracellular Ca2+ storage in acidocalcisomes of Trypanosoma cruzi. Biochem J. 1995 Sep 15;310(Pt 3):1005–1012. doi: 10.1042/bj3101005. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Docampo R., Vanderheyden N. M., Shaw M. M., Durant P. J., Bartlett M. S., Smith J. W., McLaughlin G. L. An H(+)-ATPase regulates cytoplasmic pH in Pneumocystis carinii trophozoites. Biochem J. 1996 Jun 1;316(Pt 2):681–684. doi: 10.1042/bj3160681. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Glaser T. A., Baatz J. E., Kreishman G. P., Mukkada A. J. pH homeostasis in Leishmania donovani amastigotes and promastigotes. Proc Natl Acad Sci U S A. 1988 Oct;85(20):7602–7606. doi: 10.1073/pnas.85.20.7602. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Hersey S. J., Sachs G. Gastric acid secretion. Physiol Rev. 1995 Jan;75(1):155–189. doi: 10.1152/physrev.1995.75.1.155. [DOI] [PubMed] [Google Scholar]
  14. Jiang S., Anderson S. A., Winget G. D., Mukkada A. J. Plasma membrane K+/H(+)-ATPase from Leishmania donovani. J Cell Physiol. 1994 Apr;159(1):60–66. doi: 10.1002/jcp.1041590109. [DOI] [PubMed] [Google Scholar]
  15. Kleyman T. R., Cragoe E. J., Jr Amiloride and its analogs as tools in the study of ion transport. J Membr Biol. 1988 Oct;105(1):1–21. doi: 10.1007/BF01871102. [DOI] [PubMed] [Google Scholar]
  16. Law S. S., Mukkada A. J. Transport of L-proline and its regulation in Leishmania tropica promastigotes. J Protozool. 1979 May;26(2):295–301. doi: 10.1111/j.1550-7408.1979.tb02784.x. [DOI] [PubMed] [Google Scholar]
  17. Liveanu V., Webster P., Zilberstein D. Localization of the plasma membrane and mitochondrial H(+)-ATPases in Leishmania donovani promastigotes. Eur J Cell Biol. 1991 Feb;54(1):95–101. [PubMed] [Google Scholar]
  18. Martinez-Zaguilan R., Lynch R. M., Martinez G. M., Gillies R. J. Vacuolar-type H(+)-ATPases are functionally expressed in plasma membranes of human tumor cells. Am J Physiol. 1993 Oct;265(4 Pt 1):C1015–C1029. doi: 10.1152/ajpcell.1993.265.4.C1015. [DOI] [PubMed] [Google Scholar]
  19. Meade J. C., Hudson K. M., Stringer S. L., Stringer J. R. A tandem pair of Leishmania donovani cation transporting ATPase genes encode isoforms that are differentially expressed. Mol Biochem Parasitol. 1989 Feb;33(1):81–91. doi: 10.1016/0166-6851(89)90045-5. [DOI] [PubMed] [Google Scholar]
  20. Meade J. C., Shaw J., Lemaster S., Gallagher G., Stringer J. R. Structure and expression of a tandem gene pair in Leishmania donovani that encodes a protein structurally homologous to eucaryotic cation-transporting ATPases. Mol Cell Biol. 1987 Nov;7(11):3937–3946. doi: 10.1128/mcb.7.11.3937. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Negulescu P. A., Machen T. E. Intracellular ion activities and membrane transport in parietal cells measured with fluorescent dyes. Methods Enzymol. 1990;192:38–81. doi: 10.1016/0076-6879(90)92062-i. [DOI] [PubMed] [Google Scholar]
  22. Ramos S., Balbín M., Raposo M., Valle E., Pardo L. A. The mechanism of intracellular acidification induced by glucose in Saccharomyces cerevisiae. J Gen Microbiol. 1989 Sep;135(9):2413–2422. doi: 10.1099/00221287-135-9-2413. [DOI] [PubMed] [Google Scholar]
  23. Rink T. J., Montecucco C., Hesketh T. R., Tsien R. Y. Lymphocyte membrane potential assessed with fluorescent probes. Biochim Biophys Acta. 1980;595(1):15–30. doi: 10.1016/0005-2736(80)90243-6. [DOI] [PubMed] [Google Scholar]
  24. Roos A., Boron W. F. Intracellular pH. Physiol Rev. 1981 Apr;61(2):296–434. doi: 10.1152/physrev.1981.61.2.296. [DOI] [PubMed] [Google Scholar]
  25. Schenkman S., Robbins E. S., Nussenzweig V. Attachment of Trypanosoma cruzi to mammalian cells requires parasite energy, and invasion can be independent of the target cell cytoskeleton. Infect Immun. 1991 Feb;59(2):645–654. doi: 10.1128/iai.59.2.645-654.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Schuster V. L., Fejes-Tóth G., Naray-Fejes-Tóth A., Gluck S. Colocalization of H(+)-ATPase and band 3 anion exchanger in rabbit collecting duct intercalated cells. Am J Physiol. 1991 Apr;260(4 Pt 2):F506–F517. doi: 10.1152/ajprenal.1991.260.4.F506. [DOI] [PubMed] [Google Scholar]
  27. Serrano R. Structure and function of proton translocating ATPase in plasma membranes of plants and fungi. Biochim Biophys Acta. 1988 Feb 24;947(1):1–28. doi: 10.1016/0304-4157(88)90017-2. [DOI] [PubMed] [Google Scholar]
  28. Swallow C. J., Grinstein S., Rotstein O. D. A vacuolar type H(+)-ATPase regulates cytoplasmic pH in murine macrophages. J Biol Chem. 1990 May 5;265(13):7645–7654. [PubMed] [Google Scholar]
  29. Swallow C. J., Grinstein S., Rotstein O. D. Cytoplasmic pH regulation in macrophages by an ATP-dependent and N,N'-dicyclohexylcarbodiimide-sensitive mechanism. Possible involvement of a plasma membrane proton pump. J Biol Chem. 1988 Dec 25;263(36):19558–19563. [PubMed] [Google Scholar]
  30. Tardieux I., Webster P., Ravesloot J., Boron W., Lunn J. A., Heuser J. E., Andrews N. W. Lysosome recruitment and fusion are early events required for trypanosome invasion of mammalian cells. Cell. 1992 Dec 24;71(7):1117–1130. doi: 10.1016/s0092-8674(05)80061-3. [DOI] [PubMed] [Google Scholar]
  31. Thissen J. A., Wang C. C. Maintenance of internal pH and an electrochemical gradient in Trypanosoma brucei. Exp Parasitol. 1991 Apr;72(3):243–251. doi: 10.1016/0014-4894(91)90143-k. [DOI] [PubMed] [Google Scholar]
  32. Tomlinson S., Vandekerckhove F., Frevert U., Nussenzweig V. The induction of Trypanosoma cruzi trypomastigote to amastigote transformation by low pH. Parasitology. 1995 Jun;110(Pt 5):547–554. doi: 10.1017/s0031182000065264. [DOI] [PubMed] [Google Scholar]
  33. Trudel S., Downey G. P., Grinstein S., Pâquet M. R. Activation of permeabilized HL60 cells by vanadate. Evidence for divergent signalling pathways. Biochem J. 1990 Jul 1;269(1):127–131. doi: 10.1042/bj2690127. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Turrini F., Sabolić I., Zimolo Z., Moewes B., Burckhardt G. Relation of ATPases in rat renal brush-border membranes to ATP-driven H+ secretion. J Membr Biol. 1989 Jan;107(1):1–12. doi: 10.1007/BF01871078. [DOI] [PubMed] [Google Scholar]
  35. Vieira L., Lavan A., Dagger F., Cabantchik Z. I. The role of anions in pH regulation of Leishmania major promastigotes. J Biol Chem. 1994 Jun 10;269(23):16254–16259. [PubMed] [Google Scholar]
  36. Vieira L., Slotki I., Cabantchik Z. I. Chloride conductive pathways which support electrogenic H+ pumping by Leishmania major promastigotes. J Biol Chem. 1995 Mar 10;270(10):5299–5304. doi: 10.1074/jbc.270.10.5299. [DOI] [PubMed] [Google Scholar]
  37. Vänänen H. K., Karhukorpi E. K., Sundquist K., Wallmark B., Roininen I., Hentunen T., Tuukkanen J., Lakkakorpi P. Evidence for the presence of a proton pump of the vacuolar H(+)-ATPase type in the ruffled borders of osteoclasts. J Cell Biol. 1990 Sep;111(3):1305–1311. doi: 10.1083/jcb.111.3.1305. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Zilberstein D., Dwyer D. M. Protonmotive force-driven active transport of D-glucose and L-proline in the protozoan parasite Leishmania donovani. Proc Natl Acad Sci U S A. 1985 Mar;82(6):1716–1720. doi: 10.1073/pnas.82.6.1716. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Zilberstein D., Philosoph H., Gepstein A. Maintenance of cytoplasmic pH and proton motive force in promastigotes of Leishmania donovani. Mol Biochem Parasitol. 1989 Sep;36(2):109–117. doi: 10.1016/0166-6851(89)90183-7. [DOI] [PubMed] [Google Scholar]
  40. van Adelsberg J., Al-Awqati Q. Regulation of cell pH by Ca+2-mediated exocytotic insertion of H+-ATPases. J Cell Biol. 1986 May;102(5):1638–1645. doi: 10.1083/jcb.102.5.1638. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES