TRYPANOTHIONE OVERPRODUCTION AND RESISTANCE TO ANTIMONIALS AND ARSENICALS IN LEISHMANIA

Leishmania resistant to arsenicals and anti-monials extrude arsenite. Previous results of arsenite uptake into plasma membrane-enriched vesi cles suggested that the transported species is a thiol adduct of arsen ite. In this paper, we demonstrate that promastigotes of arsenite-resi stant Leishmania tarentolae have increased levels of intracellular thi ols, High-pressure liquid chromatography of the total thiols showed th at a single peak of material was elevated almost 40-fold. The major sp ecies in this peak was identified by matrix-assisted laser desorption/ ionization mass spectrometry as N-1,N-8-bis-(glutathionyl)spermidine ( trypanothione). The trypanothione adduct of arsenite was effectively t ransported by the As-thiol pump, No difference in pump activity was ob served in wild type and mutants. A model for drug resistance is propos ed in which Sb(V)/As(V)-containing compounds, including the antileishm anial drug Pentostam, are reduced intracellularly to Sb(III)/As(III), conjugated to trypanothione, and extruded by the As-thiol pump. The ra te-limiting step in resistance is proposed to be formation of the meta lloid-thiol pump substrates, so that increased synthesis of trypanothi one produces resistance, Increased synthesis of the substrate rather t han an increase in the number of pump molecules is a novel mechanism f or drug resistance.