A vacuolar-type H+-pyrophosphatase governs maintenance of functional acidocalcisomes and growth of the insect and mammalian forms of Trypanosoma brucei

Guillaume Lemercier, Sandrine Dutoya, Shuhong Luo, Felix A. Ruiz, Claudia D Rodrigues, Théo Baltz, Roberto Docampo, Norbert Bakalara

Research output: Contribution to journalArticle

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Abstract

Vacuolar proton pyrophosphatases (V-H+-PPases) are electrogenic proton pumps found in many organisms of considerable industrial, environmental, and clinical importance. V-H+-PPases of several parasites were shown to be associated with acidic vacuoles named acidocalcisomes, which contain polyphosphate and calcium. In this work we functionally characterized a Trypanosoma brucei V-H+-PPase gene by using double-stranded RNA interference methodology to produce inducible V-H+-PPase-deficient strains of procyclic and bloodstream forms (PFiVP1 and BFiVP1). Acidocalcisomes of these mutated parasites lost acidity and contained 90% less polyphosphate. PFiVP1 did not release calcium after the addition of nigericin, and its total acidity was reduced by 70%. This mutant also failed to stabilize its intracellular pH on exposure to external basic pH >7.4 and recovered from intracellular acidification at a slower rate and to a more acidic final intracellular pH. In the absence of T. brucei V-H+-PPase expression, PFiVP1 and BFiVP1 grew at a slower rate with doubling times of 27 h instead of 15 h, and 10 h instead of 7.5 h, respectively. Moreover, BFiVP1 could not grow over 5 × 105 cells/ml corresponding to a cell density reduction of five times for bloodstream form stationary phase growth.

Original languageEnglish
Pages (from-to)37369-37376
Number of pages8
JournalJournal of Biological Chemistry
Volume277
Issue number40
DOIs
StatePublished - Oct 4 2002
Externally publishedYes

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Inorganic Pyrophosphatase
Trypanosoma brucei brucei
Insects
Maintenance
Growth
Polyphosphates
Acidity
Parasites
Nigericin
Pyrophosphatases
Calcium
Proton Pumps
Double-Stranded RNA
Acidification
RNA Interference
Vacuoles
Protons
Cell Count
Genes

ASJC Scopus subject areas

  • Biochemistry

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A vacuolar-type H+-pyrophosphatase governs maintenance of functional acidocalcisomes and growth of the insect and mammalian forms of Trypanosoma brucei. / Lemercier, Guillaume; Dutoya, Sandrine; Luo, Shuhong; Ruiz, Felix A.; Rodrigues, Claudia D; Baltz, Théo; Docampo, Roberto; Bakalara, Norbert.

In: Journal of Biological Chemistry, Vol. 277, No. 40, 04.10.2002, p. 37369-37376.

Research output: Contribution to journalArticle

Lemercier, Guillaume ; Dutoya, Sandrine ; Luo, Shuhong ; Ruiz, Felix A. ; Rodrigues, Claudia D ; Baltz, Théo ; Docampo, Roberto ; Bakalara, Norbert. / A vacuolar-type H+-pyrophosphatase governs maintenance of functional acidocalcisomes and growth of the insect and mammalian forms of Trypanosoma brucei. In: Journal of Biological Chemistry. 2002 ; Vol. 277, No. 40. pp. 37369-37376.
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