The late ISC pathway interactome reveals mitosomal-cytoplasmic crosstalk in Giardia intestinalis

Abstract

Mitochondrial metabolism is entirely dependent on the biosynthesis of the [4Fe-4S] clusters, which are part of the subunits of the respiratory chain. The mitochondrial late ISC pathway mediates the formation of these clusters from simpler [2Fe-2S] molecules and transfers them to client proteins. Here, we characterized the late ISC pathway in one of the simplest mitochondria, mitosomes, of the anaerobic protist Giardia intestinalis that lost the respiratory chain and other hallmarks of mitochondria. Identification of the late ISC interactome revealed unexpected involvement of the aerobic marker protein BolA and specific interaction of IscA with the outer mitosomal membrane. Although we confirmed that the synthesis of the Fe-S cluster remained the only metabolic role of mitosomes, we also showed that mitosomes lack client proteins that require the [4Fe-4S] cluster. Instead, by knocking out the bolA gene from the G. intestinalis genome, we showed that, unlike aerobic mitochondria, the late ISC mitosomal pathway is involved in the assembly of cytosolic [4Fe-4S] clusters. Thus, this work reveals an unexpected link between the formation of mitochondrial and cytosolic [4Fe- 4S] clusters. This may either be a consequence of mitochondrial adaptation to life without oxygen, or it represents a general metabolic coupling that has not been previously observed in the complex mitochondrial metabolism of aerobes.

Extreme mitochondrial reduction in a novel group of free-living metamonads

“…Surprisingly, with the exceptions of mitochondrial Hsp70 (discussed further below) and ferredoxin, no other genes encoding ISC pathway components were detected in the barthelonids or any other of the BaSks (see Supplementary Table 1 for ISC proteins queried). The absence of some of these proteins, like Isd11 and ferredoxin reductase (Arh1), is not unexpected as they have not been detected in most metamonads (Motyčková et al, 2023). The absence of other early ISC pathway proteins implicated in [2Fe-2S] cluster synthesis (e.g.…”

“…The absence of other early ISC pathway proteins implicated in [2Fe-2S] cluster synthesis (e.g. frataxin, Jac1 (HscB) and Mge1 (GrpE)) is more puzzling as these components are usually detected in metamonads (Motyčková et al, 2023).…”

“…The absence of other early ISC pathway proteins implicated in [2Fe-2S] cluster synthesis (e.g. frataxin, Jac1 (HscB) and Mge1 (GrpE)) is more puzzling as these components are usually detected in metamonads (Motyčková et al, 2022). Similarly, some late ISC pathway components implicated in [4Fe-4S] cluster synthesis (Lill & Freibert, 2020) are found in other metamonads (e.g Grx5, Isa2, NFU1 and BolA; Motyčková et al, 2022), but were not detected in any BRCs (Supplementary Table 1).…”

“…frataxin, Jac1 (HscB) and Mge1 (GrpE)) is more puzzling as these components are usually detected in metamonads (Motyčková et al, 2022). Similarly, some late ISC pathway components implicated in [4Fe-4S] cluster synthesis (Lill & Freibert, 2020) are found in other metamonads (e.g Grx5, Isa2, NFU1 and BolA; Motyčková et al, 2022), but were not detected in any BRCs (Supplementary Table 1). This is strange given that the barthelonids possess mitochondrial NuoF subunits that typically house an [4Fe-4S] cluster (Ohnishi, 1998).…”