Down-regulation of Leishmania donovani trypanothione reductase by heterologous expression of a trans-dominant mutant homologue: Effect on parasite intracellular survival

TY - JOUR

T1 - Down-regulation of Leishmania donovani trypanothione reductase by heterologous expression of a trans-dominant mutant homologue

T2 - Effect on parasite intracellular survival

AU - Tovar, Jorge

AU - Cunningham, M.L.

AU - Smith, Aden C.

AU - Croft, Simon L.

AU - Fairlamb, Alan H.

N1 - Medline is the source for the MeSH terms of this document.

PY - 1998/4/28

Y1 - 1998/4/28

N2 - A trans-dominant mutational strategy was used to down-regulate trypanothione reductase (TR) activity levels in Leishmania donovani, the causative agent of visceral leishmaniasis in humans. TR, regarded as an ideal drug target against trypanosomatid infections, is a homodimeric flavoprotein oxidoreductase unique to these organisms that plays a central role in the enzymatic regeneration of the thiol pool. Extrachromosomal, heterologous expression of a transdominant mutant version of the Trypanosoma cruzi enzyme in L. donovani resulted in the formation of inactive cross-species heterodimers and in a dramatic decrease of endogenous TR activity levels. Recombinant cells depleted of up to 85% of TR activity were significantly impaired in their ability to regenerate dihydrotrypanothione from trypanothione disulfide following oxidation with diamide. Nonetheless trans- dominant mutant recombinants were still capable of maintaining a reduced intracellular environment during cell growth in culture and were able to metabolize hydrogen peroxide at wildtype rates in vitro. Importantly, however, cells expressing the trans-dominant mutant enzyme displayed a decreased ability to survive inside activated macrophages in a murine model of Leishmania infection. The apparent inability of Leishmania to modulate the expression of active TR homodimers in response to the expression of trans- dominant mutant protein suggests that specific inhibitors of this enzyme should be useful anti-leishmanial agents.

AB - A trans-dominant mutational strategy was used to down-regulate trypanothione reductase (TR) activity levels in Leishmania donovani, the causative agent of visceral leishmaniasis in humans. TR, regarded as an ideal drug target against trypanosomatid infections, is a homodimeric flavoprotein oxidoreductase unique to these organisms that plays a central role in the enzymatic regeneration of the thiol pool. Extrachromosomal, heterologous expression of a transdominant mutant version of the Trypanosoma cruzi enzyme in L. donovani resulted in the formation of inactive cross-species heterodimers and in a dramatic decrease of endogenous TR activity levels. Recombinant cells depleted of up to 85% of TR activity were significantly impaired in their ability to regenerate dihydrotrypanothione from trypanothione disulfide following oxidation with diamide. Nonetheless trans- dominant mutant recombinants were still capable of maintaining a reduced intracellular environment during cell growth in culture and were able to metabolize hydrogen peroxide at wildtype rates in vitro. Importantly, however, cells expressing the trans-dominant mutant enzyme displayed a decreased ability to survive inside activated macrophages in a murine model of Leishmania infection. The apparent inability of Leishmania to modulate the expression of active TR homodimers in response to the expression of trans- dominant mutant protein suggests that specific inhibitors of this enzyme should be useful anti-leishmanial agents.

UR - https://www.scopus.com/pages/publications/0032574808

U2 - 10.1073/pnas.95.9.5311

DO - 10.1073/pnas.95.9.5311

M3 - Article

AN - SCOPUS:0032574808

SN - 0027-8424

VL - 95

SP - 5311

EP - 5316

JO - Proceedings of the National Academy of Sciences of the United States of America

JF - Proceedings of the National Academy of Sciences of the United States of America

IS - 9

ER -