title: A pyrophosphatase regulating polyphosphate metabolism in acidocalcisomes is essential for Trypanosoma brucei virulence in mice.
abstract: We report the functional characterization of a soluble pyrophosphatase (TbVSP1), which localizes to acidocalcisomes, a vesicular acidic compartment of Trypanosoma brucei. Depending on the pH and the cofactors Mg(2+) or Zn(2+), both present in the compartment, the enzyme hydrolyzes either inorganic pyrophosphate (PP(i)) (k(cat) = 385 s(-1)) or tripolyP (polyP(3)) and polyphosphate (polyP) of 28 residues (polyP(28)) with k(cat) values of 52 and 3.5 s(-1), respectively. An unusual N-terminal domain of 160 amino acids, containing a putative calcium EF-hand-binding domain, is involved in protein oligomerization. Using double-stranded RNA interference methodology, we produced an inducible bloodstream form (BF) deficient in the TbVSP1 protein (BFiVSP1). The long-chain polyP levels of these mutants were reduced by 60%. Their phenotypes revealed a deficient polyP metabolism, as indicated by their defective response to phosphate starvation and hyposmotic stress. BFiVSP1 did not cause acute virulent infection in mice, demonstrating that TbVSP1 is essential for growth of bloodstream forms in the mammalian host.
authors: Laboratoire de Genomique Fonctionnelle des Trypanosomatides, UMR-CNRS 5162, 146, rue Leo Saignat, 33076 Bordeaux, France.
authors: Guillaume
authors: G
authors: Lemercier
authors: Benoit
authors: B
authors: Espiau
authors: Felix A
authors: FA
authors: Ruiz
authors: Mauricio
authors: M
authors: Vieira
authors: Shuhong
authors: S
authors: Luo
authors: Theo
authors: T
authors: Baltz
authors: Roberto
authors: R
authors: Docampo
authors: Norbert
authors: N
authors: Bakalara
mesh_terms: D017766:Acid Anhydride Hydrolases / Q000737:chemistry; D000251:Adenosine Triphosphatases / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D015153:Blotting, Western; D002118:Calcium / Q000737:chemistry; D002845:Chromatography; D002850:Chromatography, Gel; D003001:Cloning, Molecular; D003360:Cosmids; D004305:Dose-Response Relationship, Drug; D010450:Endopeptidases / Q000502:physiology*; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D015151:Immunoblotting; D007700:Kinetics; D008274:Magnesium / Q000494:pharmacology; D051379:Mice; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D009995:Osmosis; D010641:Phenotype; D010710:Phosphates; D011122:Polyphosphates / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D015800:Protozoan Proteins; D011755:Pyrophosphatases / Q000737:chemistry* / Q000378:metabolism*; D034622:RNA Interference; D012330:RNA, Double-Stranded / Q000737:chemistry; D011994:Recombinant Proteins / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D013997:Time Factors; D014162:Transfection; D014346:Trypanosoma brucei brucei / Q000472:pathogenicity*; D014774:Virulence; D015032:Zinc / Q000494:pharmacology
chemical_list: D010710:Phosphates; D011122:Polyphosphates; D015800:Protozoan Proteins; D012330:RNA, Double-Stranded; D011994:Recombinant Proteins; D010450:Endopeptidases; D017766:Acid Anhydride Hydrolases; D000251:Adenosine Triphosphatases; D011755:Pyrophosphatases; C482344:VSP1 protein, Trypanosoma brucei; C020137:exopolyphosphatase; D008274:Magnesium; D015032:Zinc; D002118:Calcium