Trypanothione overproduction and resistance to antimonials and arsenicals in Leishmania |
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Authors: | R Mukhopadhyay S Dey N Xu D Gage J Lightbody M Ouellette BP Rosen |
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Affiliation: | Department of Biochemistry and Molecular Biology, Wayne State University School of Medicine, Detroit, MI 48201, USA. |
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Abstract: | Leishmania resistant to arsenicals and antimonials extrude arsenite. Previous results of arsenite uptake into plasma membrane-enriched vesicles suggested that the transported species is a thiol adduct of arsenite. In this paper, we demonstrate that promastigotes of arsenite-resistant Leishmania tarentolae have increased levels of intracellular thiols. High-pressure liquid chromatography of the total thiols showed that a single peak of material was elevated almost 40-fold. The major species in this peak was identified by matrix-assisted laser desorption/ionization mass spectrometry as N1,N8-bis-(glutathionyl)spermidine (trypanothione). The trypanothione adduct of arsenite was effectively transported by the As-thiol pump. No difference in pump activity was observed in wild type and mutants. A model for drug resistance is proposed in which Sb(V)/As(V)-containing compounds, including the antileishmanial drug Pentostam, are reduced intracellularly to Sb(III)/As(III), conjugated to trypanothione, and extruded by the As-thiol pump. The rate-limiting step in resistance is proposed to be formation of the metalloid-thiol pump substrates, so that increased synthesis of trypanothione produces resistance. Increased synthesis of the substrate rather than an increase in the number of pump molecules is a novel mechanism for drug resistance. |
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