A Pyrophosphatase Regulating Polyphosphate Metabolism in Acidocalcisomes Is Essential for Trypanosoma brucei Virulence in Mice

Lemercier, Guillaume; Espiau, Benoît; Ruiz, Félix A.; Vieira, Marcelo Bernardes; Luo, Shuhong; Baltz, Théo; Docampo, Roberto; Bakalara, Norbert

doi:10.1074/jbc.m309974200

Cited by 77 publications

(124 citation statements)

References 41 publications

(50 reference statements)

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“…Therefore, loss of PPX was expected to lead to an overall increase in levels of cellular polyP unless there was compensatory up-regulation of the expression of PPK as in B. cereus (32). However, there was no change in ppk mRNA levels following loss of ppx, and total cellular polyP levels were elevated compared with the wild type.…”

Section: Discussionmentioning

confidence: 90%

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The Role of the Exopolyphosphatase PPX in Avoidance by Neisseria meningitidis of Complement-mediated Killing

Zhang

Tang

2010

Journal of Biological Chemistry

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The complement system is critical for immunity against the important human pathogen Neisseria meningitidis. We describe the isolation of a meningococcal mutant lacking PPX, an exopolyphosphatase responsible for cleaving cellular polyphosphate, a polymer of tens to hundreds of orthophosphate residues found in virtually all living cells. Bacteria lacking PPX exhibit increased resistance to complement-mediated killing. By site directed mutagenesis, we define amino acids necessary for the biochemical activity of meningococcal PPX, including a conserved glutamate (Glu 117 ) and residues in the Walker B box predicted to be involved in binding to phosphate. We show that the biochemical activity of PPX is necessary for interactions with the complement. The relative resistance of the ppx mutant does not result from changes in structures (such as capsule, lipopolysaccharide, and factor H-binding protein), which are known to be required for evasion of this key aspect of host immunity. Instead, expression of PPX modifies the interaction of N. meningitidis with the alternative pathway of complement activation.

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Section: Discussionmentioning

confidence: 90%

“…In Bacillus cereus, loss of PPX is compensated for by a reduction in expression of PPK, the enzyme responsible for polyP synthesis (32). Therefore, we determined whether levels of ppk mRNA are influenced by the presence of PPX in N. meningitidis.…”

Section: Loss Of Nmb1467 Enhances Survival Of N Meningitidis Inmentioning

confidence: 99%

The Role of the Exopolyphosphatase PPX in Avoidance by Neisseria meningitidis of Complement-mediated Killing

Zhang

Tang

2010

Journal of Biological Chemistry

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“…Labeling occurred predominantly in the membrane of the organelle, despite the presence of gold particles in the matrix of some acidocalcisomes (Fig. 8B), which might be explained by the presence of soluble pyrophosphatases in these organelles, as occurs in T. brucei (Lemercier et al 2003). Because a polyclonal antibody was used, it is possible that it has recognized specific sites of the soluble pyrophosphatases that are conserved in the vacuolar-type pyrophosphatase (i.e., PPi binding sites).…”

Section: Localization Of the V-h + -Ppase In Acidocalcisomes And Its mentioning

confidence: 99%

Dynamics of polymorphism of acidocalcisomes in Leishmania parasites

Miranda

Docampo

Grillo

et al. 2004

Histochemistry and Cell Biology

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Growth of Leishmania mexicana amazonensis promastigotes in different culture media resulted in structurally and chemically different acidocalcisomes. When grown in SDM-79 medium, the promastigotes showed large spherical acidocalcisomes of up to 1.2 mm diameter distributed throughout the cell. X-ray microanalysis and elemental mapping of the organelles showed large amounts of oxygen, phosphorus, sodium, potassium, magnesium, calcium, and zinc. Immunofluorescence microscopy using antisera raised against a peptide sequence of the vacuolar-type proton pyrophosphatase of Arabidopsis thaliana that is conserved in the Leishmania enzyme, indicated localization in acidocalcisomes. When cells were transferred to Warren's medium, the acidocalcisomes transformed from spherical into branched tubular organelles. The labeling pattern of the vacuolar proton-pyrophosphatase, considered as a marker for the organelle, changed accompanying the structural changes of the acidocalcisomes, and the enzyme showed an apparently lower proton-transporting activity when measured in digitonin-permeabilized promastigotes. X-ray microanalysis and elemental mapping of these structures revealed the additional presence of iron. Together, the results reveal that the morphology and composition of acidocalcisomes are greatly influenced by the culture con-ditions.

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“…The soluble acidocalcisomal protein VSP1 is found in acidocalcisomes of ⌬ap3␦ promastigotes VSP1 is a soluble pyrophosphatase found in T. brucei (Lemercier et al, 2004) and L. amazonensis (Espiau et al, 2006) acidocalcisomes. The ~50-kDa VSP1 was detected in L. major wildtype, AP3␦-null mutant and AP3␦-re-expressing cell extracts with an anti-VSP1 antibody (Fig.…”

Section: ⌬Ap3␦ Promastigotes Lack Acidocalcisomal Membrane Proteinsmentioning

confidence: 99%

“…Western blot analysis was performed on cell fractions or total cell extracts (from 10 7 promastigotes) with the anti-TbVP1 rabbit antibody (Lemercier et al, 2002) and rabbit anti-TbVSP1 (Espiau et al, 2006;Lemercier et al, 2004) diluted at 1:1000. Other western blot analyses involved the use of rabbit anti-HASP antibody (Flinn et al, 1994) at a 1:1000 dilution and mouse anti-TbEF1␣ antibody (Upstate) at 1:10,000.…”

Section: Western Blottingmentioning

confidence: 99%

The AP3 adaptor is involved in the transport of membrane proteins to acidocalcisomes of Leishmania

Besteiro

Tonn

Tetley

et al. 2008

Journal of Cell Science

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Lysosomal function is crucial for the differentiation and infectivity of the parasitic protozoon Leishmania major. To study lysosomal biogenesis, an L. major mutant deficient in the ␦ subunit of the adaptor protein 3 (AP3 ␦) complex was generated. Structure and proteolytic capacity of the lysosomal compartment were apparently unaffected in the AP3-deficient mutant; however, defects were identified in its acidocalcisomes. These are acidic organelles enriched in calcium and phosphorus, conserved from bacteria to eukaryotes, whose function remains enigmatic. The acidocalcisomes of the L. major mutant lacked membrane-bound proton pumps (notably V-H + -PPase), were less acidic than normal acidocalcisomes and devoid of polyphosphate, but contained a soluble pyrophosphatase. The mutant parasites were viable in vitro, but were unable to establish an infection in mice, which indicates a role for AP3 in determining -possibly through an acidocalcisome-related function -the virulence of the parasite. AP3 transport function has been linked previously to lysosome-related organelles such as platelet dense granules, which appear to share several features with acidocalcisomes. Our findings, implicating that AP3 has a role in transport to acidocalcisomes, thus provide further evidence that biogenesis of acidocalcisomes resembles that of lysosome-related organelles, and that both may have conserved origins. Supplementary material available online at

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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.

Cited by 77 publications

References 41 publications

The Role of the Exopolyphosphatase PPX in Avoidance by Neisseria meningitidis of Complement-mediated Killing

The Role of the Exopolyphosphatase PPX in Avoidance by Neisseria meningitidis of Complement-mediated Killing

Dynamics of polymorphism of acidocalcisomes in Leishmania parasites

The AP3 adaptor is involved in the transport of membrane proteins to acidocalcisomes of Leishmania

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