Characterisation of the SUF FeS cluster machinery in the amitochondriate eukaryoteMonocercomonoides exilis

Abstract

AbstractMonocercomonoides exilisis the first eukaryotic organism described as a complete amitochondriate, yet it shares common features with heterotrophic anaerobic/microaerophilic protists, some of which bear divergent mitochondrion-related organelles or MROs. It has been postulated that the retention of these organelles stems from their involvement in the assembly of essential cytosolic and nuclear FeS proteins, whose maturation requires the evolutionarily conserved mitochondrial ISC and cytosolic CIA machineries. The amitochondriateM. exilislacks genes encoding the ISC machinery yet contains a bacteria-derived SUF system (MeSuf), composed of the cysteine desulphurase SufS fused to SufD and SufU, as well as the FeS scaffolding components MeSufB and MeSufC. Here, we show that expression of the M. exilis SUFgenes, either individually or in tandem, can restore the maturation of the FeS protein IscR in theEscherichia colidouble mutants of ΔsufSΔiscSand ΔsufBΔiscUA. In vivoandin vitrostudies indicate that purified MeSufB, MeSufC and MeSufDSU proteins interact suggesting that they act as a complex in the protist. MeSufBC can undergo conformational changes in the presence of ATP and assemble FeS clusters under anaerobic conditions in presence and absence of ATPin vitro. Altogether, these results indicate that the dynamically interacting MeSufDSUBC proteins may function as an FeS cluster assembly complex inM. exilisthereby being capable of replacing the organelle-enclosed ISC system of canonical eukaryotes.