Abstract

ABC transporters of the mitochondria (ATMs) are conserved in many different species. In the model plant Arabidopsis thaliana, ATM3 is required for cytosolic assembly of Fe-S clusters and is thought to provide a so far unknown precursor for the cytosolic Fe-S cluster assembly pathway. Previous studies in Arabidopsis suggested that the transported compound contains sulphur (but not iron) in an unknown chemical form (1). Additionally it was shown that glutathione (GSH) plays a role in the function of the yeast orthologue Atm1 (2, 3).

The aim of our study is to characterize the substrate specificity of plant AtATM3 and its functional orthologue yeast Atm1. Using buthionine sulfoximine (BSO), an inhibitor of the first step of glutathione synthesis, we assessed the effect of GSH depletion on Arabidopsis atm3 mutants. We observed reduction of root growth in atm3 mutants in presence of low concentrations of BSO suggesting a role of GSH in ATM function. To investigate the substrate specificity of ATMs further we have established an expression system in the gram-positive bacterium Lactococcus lactis, and have purified ATMs in their functional form by affinity purification. Using purified protein we screened for potential substrates using ATP hydrolysis measurements and tested transport in inside-out vesicles. We also found a genetic link between ATM3 and the mitochondrial sulphur dioxygenase ETHE1 suggesting the persulfide of glutathione is of physiological relevance in the ATM transport reaction.