Previous studies in Trypanosoma cruzi, the etiologic agent of Chagas disease, have resulted in the cloning and sequencing of a pair of tandemly linked genes (TcHA1 and TcHA2) that encode P (phospho-intermediate form)-type H+-ATPases with homology to fungal and plant proton-pumping ATPases. In the present study, we demonstrate that these pumps are present in the plasma membrane and intracellular compartments of three different stages of T. cruzi. The main intracellular compartment containing these ATPases in epimastigotes was identified as the reservosome. This identification was achieved by immunofluorescence assays and immunoelectron microscopy showing their co-localization with cruzipain, and by subcellular fractionation and detection of their activity. ATP-dependent proton transport by isolated reservosomes was sensitive to vanadate and insensitive to bafilomycin A1, which is in agreement with the localization of P-type H+-ATPases in these organelles. Analysis by confocal immunofluorescence microscopy revealed that epitope–tagged TcHA1-Ty1 and TcHA2-Ty1 gene products are localized in the reservosomes, whereas the TcHA1-Ty1 gene product is additionally present in the plasma membrane. Immunogold electron microscopy showed the presence of the H+-ATPases in other compartments of the endocytic pathway such as the cytostome and endosomal vesicles, suggesting that in contrast with most cells investigated until now, the endocytic pathway of T. cruzi is acidified by a P-type H+-ATPase.
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December 2005
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Research Article|
December 06 2005
Role for a P-type H+-ATPase in the acidification of the endocytic pathway of Trypanosoma cruzi
Mauricio Vieira;
Mauricio Vieira
*Laboratory of Molecular Parasitology, Department of Pathobiology and Center for Zoonoses Research, University of Illinois at Urbana–Champaign, Urbana, IL 61802, U.S.A.
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Peter Rohloff;
Peter Rohloff
*Laboratory of Molecular Parasitology, Department of Pathobiology and Center for Zoonoses Research, University of Illinois at Urbana–Champaign, Urbana, IL 61802, U.S.A.
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Shuhong Luo;
Shuhong Luo
*Laboratory of Molecular Parasitology, Department of Pathobiology and Center for Zoonoses Research, University of Illinois at Urbana–Champaign, Urbana, IL 61802, U.S.A.
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Narcisa L. Cunha-E-Silva;
Narcisa L. Cunha-E-Silva
†Instituto de Biofisica Carlos Chagas Filho (IBCCF), Universidade Federal do Rio de Janeiro, Rio de Janeiro, 21941 RJ, Brazil
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Wanderley De Souza;
Wanderley De Souza
†Instituto de Biofisica Carlos Chagas Filho (IBCCF), Universidade Federal do Rio de Janeiro, Rio de Janeiro, 21941 RJ, Brazil
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Roberto Docampo
Roberto Docampo
1
*Laboratory of Molecular Parasitology, Department of Pathobiology and Center for Zoonoses Research, University of Illinois at Urbana–Champaign, Urbana, IL 61802, U.S.A.
‡Department of Cellular Biology and Center for Tropical and Global Emerging Diseases, University of Georgia, 30602 Athens, U.S.A.
1To whom correspondence should be addressed (email [email protected]).
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Publisher: Portland Press Ltd
Received:
August 12 2005
Revision Received:
September 07 2005
Accepted:
September 08 2005
Accepted Manuscript online:
September 08 2005
Online ISSN: 1470-8728
Print ISSN: 0264-6021
The Biochemical Society, London
2005
Biochem J (2005) 392 (3): 467–474.
Article history
Received:
August 12 2005
Revision Received:
September 07 2005
Accepted:
September 08 2005
Accepted Manuscript online:
September 08 2005
Citation
Mauricio Vieira, Peter Rohloff, Shuhong Luo, Narcisa L. Cunha-E-Silva, Wanderley De Souza, Roberto Docampo; Role for a P-type H+-ATPase in the acidification of the endocytic pathway of Trypanosoma cruzi. Biochem J 15 December 2005; 392 (3): 467–474. doi: https://doi.org/10.1042/BJ20051319
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