Parasitology Research

, Volume 100, Issue 4, pp 887–892

Cofactor-independent phosphoglycerate mutase is an essential gene in procyclic form Trypanosoma brucei

  • Appolinaire Djikeng
  • Sylvine Raverdy
  • Jeremy Foster
  • Daniella Bartholomeu
  • Yinhua Zhang
  • Najib M. El-Sayed
  • Clotilde Carlow
Short Communication

Abstract

Glycolysis and gluconeogenesis are, in part, driven by the interconversion of 3- and 2-phosphoglycerate (3-PG and 2-PG) which is performed by phosphoglycerate mutases (PGAMs) which can be cofactor dependant (dPGAM) or cofactor independent (iPGAM). The African trypanosome, Trypanosoma brucei, possesses the iPGAM form which is thought to play an important role in glycolysis. Here, we report on the use of RNA interference to down-regulate the T. brucei iPGAM in procyclic form T. brucei and evaluation of the resulting phenotype. We first demonstrated biochemically that depletion of the steady state levels of iPGM mRNA correlates with a marked reduction of enzyme activity. We further show that iPGAM is required for cell growth in procyclic T. brucei.

References

  1. Albert MA, Haanstra JR, Hannaert V, Van Roy J, Opperdoes FR, Bakker BM, Michels PA (2005) Experimental and in silico analyses of glycolytic flux control in bloodstream form Trypanosoma brucei. J Biol Chem 280:28306–28315PubMedCrossRefGoogle Scholar
  2. Barrett MP, Burchmore RJ, Stich A, Lazzari JO, Frasch AC, Cazzulo JJ, Krishna S (2003) The trypanosomiases. Lancet 362:1469–1480PubMedCrossRefGoogle Scholar
  3. Berriman M, Ghedin E, Hertz-Fowler C, Blandin G, Renauld H, Bartholomeu DC, Lennard NJ et al. (2005) The genome of the African trypanosome Trypanosoma brucei. Science 309:416–422PubMedCrossRefGoogle Scholar
  4. Besteiro S, Barrett MP, Riviere L, Bringaud F (2005) Energy generation in insect stages of Trypanosoma brucei: metabolism in flux. Trends Parasitol 21:185–191PubMedCrossRefGoogle Scholar
  5. Chevalier N, Rigden DJ, Van Roy J, Opperdoes FR, Michels PA (2000) Trypanosoma brucei contains a 2,3-bisphosphoglycerate independent phosphoglycerate mutase. Eur J Biochem 267:1464–1472PubMedCrossRefGoogle Scholar
  6. Collet JF, Stroobant V, Van Schaftingen E (2001) The 2,3-bisphosphoglycerate-independent phosphoglycerate mutase from Trypanosoma brucei: metal-ion dependency and phosphoenzyme formation. FEMS Microbiol Lett 204:39–44PubMedCrossRefGoogle Scholar
  7. Coustou V, Besteiro S, Biran M, Diolez P, Bouchaud V, Voisin P,Michels PA et al. (2003) ATP generation in the Trypanosoma brucei procyclic form: cytosolic substrate level is essential, but not oxidative phosphorylation. J Biol Chem 278:49625–49635PubMedCrossRefGoogle Scholar
  8. Cross GA, Klein RA, Linstead DJ (1975) Utilization of amino acids by Trypanosoma brucei in culture: l-threonine as a precursor for acetate. Parasitology 71:311–326PubMedCrossRefGoogle Scholar
  9. Cunningham I (1977) New culture medium for maintenance of tsetse tissues and growth of trypanosomatids. J Protozool 24:325–329Google Scholar
  10. Djikeng A, Shen S, Tschudi C, Ullu E (2004) Analysis of gene function in Trypanosoma brucei using RNA interference. Methods Mol Biol 270:287–298PubMedGoogle Scholar
  11. Drew ME, Morris JC, Wang Z, Wells L, Sanchez M, Landfear SM, Englund PT (2003) The adenosine analog tubercidin inhibits glycolysis in Trypanosoma brucei as revealed by an RNA interference library. J Biol Chem 278:46596–46600PubMedCrossRefGoogle Scholar
  12. El-Sayed NM, Myler PJ, Bartholomeu DC, Nilsson D, Aggarwal G, Tran AN, Ghedin E et al. (2005a) The genome sequence of Trypanosoma cruzi, etiologic agent of Chagas disease. Science 309:409–415PubMedCrossRefGoogle Scholar
  13. El-Sayed NM, Myler PJ, Blandin G, Berriman M, Crabtree J, Aggarwal G, Caler E et al. (2005b) Comparative genomics of trypanosomatid parasitic protozoa. Science 309:404–409PubMedCrossRefGoogle Scholar
  14. Guerra DG, Vertommen D, Fothergill-Gilmore LA, Opperdoes FR, Michels PA (2004) Characterization of the cofactor-independent phosphoglycerate mutase from Leishmania mexicana mexicana. Histidines that coordinate the two metal ions in the active site show different susceptibilities to irreversible chemical modification. Eur J Biochem 271:1798–1810PubMedCrossRefGoogle Scholar
  15. Guerra-Giraldez C, Quijada L, Clayton CE (2002) Compartmentation of enzymes in a microbody, the glycosome, is essential in Trypanosoma brucei. J Cell Sci 115:2651–2658PubMedGoogle Scholar
  16. Hannaert V, Bringaud F, Opperdoes FR, Michels PA (2003) Evolution of energy metabolism and its compartmentation in Kinetoplastida. Kinetoplastid Biol Dis 2:11PubMedCrossRefGoogle Scholar
  17. Ivens AC, Peacock CS, Worthey EA, Murphy L, Aggarwal G, Berriman M, Sisk E et al. (2005) The genome of the kinetoplastid parasite, Leishmania major. Science 309:436–442PubMedCrossRefGoogle Scholar
  18. Jannin J, Cattand P (2004) Treatment and control of human African trypanosomiasis. Curr Opin Infect Dis 17:565–571PubMedCrossRefGoogle Scholar
  19. Jedrzejas MJ, Chander M, Setlow P, Krishnasamy G (2000a) Mechanism of catalysis of the cofactor-independent phosphoglycerate mutase from Bacillus stearothermophilus. Crystal structure of the complex with 2-phosphoglycerate. J Biol Chem 275:23146–23153PubMedCrossRefGoogle Scholar
  20. Jedrzejas MJ, Chander M, Setlow P, Krishnasamy G (2000b) Structure and mechanism of action of a novel phosphoglycerate mutase from Bacillus stearothermophilus. EMBO J 19:1419–1431PubMedCrossRefGoogle Scholar
  21. Joshi PP, Shegokar VR, Powar RM, Herder S, Katti R, Salkar HR, Dani VS et al. (2005) Human trypanosomiasis caused by Trypanosoma evansi in India: the first case report. Am J Trop Med Hyg 73:491–495PubMedGoogle Scholar
  22. Kessler PS, Parsons M (2005) Probing the role of compartmentation of glycolysis in procyclic form Trypanosoma brucei: RNA interference studies of PEX14, hexokinase, and phosphofructokinase. J Biol Chem 280:9030–9036PubMedCrossRefGoogle Scholar
  23. Lamour N, Riviere L, Coustou V, Coombs GH, Barrett MP, Bringaud F (2005) Proline metabolism in procyclic Trypanosoma brucei is down-regulated in the presence of glucose. J Biol Chem 280:11902–11910PubMedCrossRefGoogle Scholar
  24. Leyva-Vazquez MA, Setlow P (1994) Cloning and nucleotide sequences of the genes encoding triose phosphate isomerase, phosphoglycerate mutase, and enolase from Bacillus subtilis. J Bacteriol 176:3903–3910PubMedGoogle Scholar
  25. Morris VL, Jackson DP, Grattan M, Hisnworth T, Cuppels DA (1995) Isolation and sequence analysis of the Pseudomonas syringae pv. tomato gene encoding a 2,3-diphosphoglycerate-independent phosphoglyceromutase. J Bacteriol 177:1727–1733PubMedGoogle Scholar
  26. Oduro KK, Flynn IW, Bowman IB (1980) Trypanosoma brucei: activities and subcellular distribution of glycolytic enzymes from differently disrupted cells. Exp Parasitol 50:123–135PubMedCrossRefGoogle Scholar
  27. Rigden DJ, Mello LV, Setlow P, Jedrzejas MJ (2002) Structure and mechanism of action of a cofactor-dependent phosphoglycerate mutase homolog from Bacillus stearothermophilus with broad specificity phosphatase activity. J Mol Biol 315:1129–1143PubMedCrossRefGoogle Scholar
  28. Wang Z, Morris JC, Drew ME, Englund PT (2000) Inhibition of Trypanosoma brucei gene expression by RNA interference using an integratable vector with opposing T7 promoters. J Biol Chem 275:40174–40179PubMedCrossRefGoogle Scholar
  29. Wirtz E, Leal S, Ochatt C, Cross GA (1999) A tightly regulated inducible expression system for conditional gene knock-outs and dominant-negative genetics in Trypanosoma brucei. Mol Biochem Parasitol 99:89–101PubMedCrossRefGoogle Scholar
  30. Zhang Y, Foster JM, Kumar S, Fougere M, Carlow CK (2004) Cofactor-independent phosphoglycerate mutase has an essential role in Caenorhabditis elegans and is conserved in parasitic nematodes. J Biol Chem 279:37185–37190PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2006

Authors and Affiliations

  • Appolinaire Djikeng
    • 1
  • Sylvine Raverdy
    • 2
  • Jeremy Foster
    • 2
  • Daniella Bartholomeu
    • 1
  • Yinhua Zhang
    • 2
  • Najib M. El-Sayed
    • 1
    • 3
  • Clotilde Carlow
    • 2
  1. 1.The Institute for Genomic Research (TIGR)RockvilleUSA
  2. 2.New England Biolabs (NEB)IpswichUSA
  3. 3.Department of Microbiology and Tropical MedicineGeorge Washington UniversityWashingtonUSA

Personalised recommendations