Disruption of the trypanothione reductase gene of Leishmania decreases its ability to survive oxidative stress in macrophages

Carole Dumas, Marc Ouellette, Jorge Tovar, Mark L. Cunningham, Alan H. Fairlamb, Samira Tamar, Martin Olivier, Barbara Papadopoulou (Lead / Corresponding author)

    Research output: Contribution to journalArticlepeer-review

    290 Citations (Scopus)

    Abstract

    Parasitic protozoa belonging to the order Kinetoplastida contain trypanothione as their major thiol. Trypanothione reductase (TR), the enzyme responsible for maintaining trypanothione in its reduced form, is thought to be central to the redox defence systems of trypanosomatids. To investigate further the physiological role of TR in Leishmania, we attempted to create TR-knockout mutants by gene disruption in L. donovani and L. major strains using the selectable markers neomycin and hygromycin phosphotransferases. TR is likely to be an important gene for parasite survival since all our attempts to obtain a TR null mutant in L. dottovani failed. Instead, we obtained mutants with a partial trisomy for the TR locus where, despite the successful disruption of two TR alleles by gene targeting, a third TR copy was generated as a result of genomic rearrangements involving the translocation of a TR-containing region to a larger chromosome. Mutants of L. donovani and L. major possessing only one wild-type TR allele express less TR mRNA and have lower TR activity compared with wild-type cells carrying two copies of the TR gene. Significantly, these mutants show attenuated infectivity with a markedly decreased capacity to survive intracellularly within macrophages, provided that the latter are producing reactive oxygen intermediates.

    Original languageEnglish
    Pages (from-to)2590-2598
    Number of pages9
    JournalEMBO Journal
    Volume16
    Issue number10
    DOIs
    Publication statusPublished - 15 May 1997

    Keywords

    • Gene targeting
    • Intracellular survival
    • Leishmania
    • Oxidative stress
    • Trypanothione reductase

    ASJC Scopus subject areas

    • General Neuroscience
    • Molecular Biology
    • General Biochemistry,Genetics and Molecular Biology
    • General Immunology and Microbiology

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