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 O. Rodrigues, Théo Baltz, Roberto Docampo, Norbert Bakalara

Research output: Contribution to journalArticle

68 Scopus citations

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

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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