Ornithine decarboxylase gene deletion mutants of Leishmania donovani

Yuqui Jiang, Sigrid C. Roberts, Armande Jardim, Nicola S. Carter, Sarah Shin, Mark Ariyanayagam, Alan H. Fairlamb, Buddy Ullman

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Abstract

A knockout strain of Leishmania donovani lacking both ornithine decarboxylase (ODC) alleles has been created by targeted gene replacement. Growth of Δodc cells in polyamine-deficient medium resulted in a rapid and profound depletion of cellular putrescine pools, although levels of spermidine were relatively unaffected. Concentrations of trypanothione, a spermidine conjugate, were also reduced, whereas glutathione concentrations were augmented. The Δodc L. donovani exhibited an auxotrophy for polyamines that could be circumvented by the addition of the naturally occurring polyamines, putrescine or spermidine, to the culture medium. Whereas putrescine supplementation restored intracellular pools of both putrescine and spermidine, exogenous spermidine was not converted back to putrescine, indicating that spermidine alone is sufficient to meet the polyamine requirement, and that L. donovani does not express the enzymatic machinery for polyamine degradation. The lack of a polyamine catabolic pathway in intact parasites was confirmed radiometrically. In addition, the Δodc strain could grow in medium supplemented with either 1,3-diaminopropane or 1,5diaminopentane (cadaverine), but polyamine auxotrophy could not be overcome by other aliphatic diamines or spermine. These data establish genetically that ODC is an essential gene in L. donovani, define the polyamine requirements of the parasite, and reveal the absence of a polyamine-degradative pathway.

Original languageEnglish (US)
Pages (from-to)3781-3788
Number of pages8
JournalJournal of Biological Chemistry
Volume274
Issue number6
DOIs
StatePublished - Feb 5 1999

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ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Jiang, Y., Roberts, S. C., Jardim, A., Carter, N. S., Shin, S., Ariyanayagam, M., Fairlamb, A. H., & Ullman, B. (1999). Ornithine decarboxylase gene deletion mutants of Leishmania donovani. Journal of Biological Chemistry, 274(6), 3781-3788. https://doi.org/10.1074/jbc.274.6.3781