A key cytosolic iron–sulfur cluster synthesis protein localizes to the mitochondrion of Toxoplasma gondii

Abstract

Iron–sulfur (Fe‐S) clusters are prosthetic groups on proteins that function in a range of enzymatic and electron transfer reactions. Fe‐S cluster synthesis is essential for the survival of all eukaryotes. Independent Fe‐S cluster biosynthesis pathways occur in the mitochondrion, plastid, and cytosolic compartments of eukaryotic cells. Little is known about the cytosolic Fe‐S cluster biosynthesis in apicomplexan parasites, the causative agents of diseases such as malaria and toxoplasmosis. NBP35 serves as a key scaffold protein on which cytosolic Fe‐S clusters assemble, and has a cytosolic localization in most eukaryotes studied thus far. Unexpectedly, we found that the NBP35 homolog of the apicomplexan Toxoplasma gondii (TgNBP35) localizes to the outer mitochondrial membrane, with mitochondrial targeting mediated by an N‐terminal transmembrane domain. We demonstrate that TgNBP35 is critical for parasite proliferation, but that, despite its mitochondrial localization, it is not required for Fe‐S cluster synthesis in the mitochondrion. Instead, we establish that TgNBP35 is important for the biogenesis of cytosolic Fe‐S proteins. Our data are consistent with TgNBP35 playing a central and specific role in cytosolic Fe‐S cluster biosynthesis, and imply that the assembly of cytosolic Fe‐S clusters occurs on the cytosolic face of the outer mitochondrial membrane in these parasites.

Differential contribution of two organelles of endosymbiotic origin to iron-sulfur cluster synthesis and overall fitness in Toxoplasma

“…It should be noted that although we believe the drop in mitochondrial membrane potential is likely due to a specific alteration of the respiratory chain, it may also be due to a loss of parasite viability. Reassuringly, our results are in line with the recent findings obtained by Aw et al, who generated a mutant of mitochondrial Fe-S cluster synthesis (by depleting TgNFS1), and observed a sharp decrease in TgSDHB abundance and a clear drop in mitochondrial O 2 consumption rate [25].…”

“…Accordingly, perhaps the most obvious consequence of disrupting the ISC pathway was the profound impact on the mitochondrial respiratory capacity, as evidenced experimentally by measuring the mitochondrial membrane potential (Figure 8D), and supported by quantitative analyses showing a clear drop in expression of many respiratory complex proteins (Figure 8A, B, and C, Table S4). This is also in line with the recent description of another mitochondrial Fe-S cluster synthesis mutant that showed a marked alteration of its respiratory capacity [25]. Although the mitochondrion, through the TCA cycle and the respiratory chain/oxidative phosphorylation, contributes to energy production in tachyzoites [91], the glycolytic flux is also believed to be a major source of carbon and energy for these parasites [92].…”

“…Our quantitative proteomics data suggests it is also the case in T. gondii , as several putative nuclear Fe-S proteins involved in gene transcription (such as DNA-dependent RNA polymerases) or DNA repair (like DNA endonulease III) were found to be impacted by TgISU1 depletion. The CIA pathway has recently been shown to be important for tachyzoite proliferation [25], and several of the cytoplasmic or nuclear Fe-S cluster-containing proteins are likely essential for parasite viability. It is thus possible that, in spite of their conversion to a stress-resistant form, the long-term viability of TgISU1 parasites could be affected beyond recovery.…”

“…Like in plants and algae, apicoplast-containing Apicomplexa seem to harbour the three (ISC, SUF and CIA) Fe-S cluster synthesis pathways. Although the CIA pathway was recently shown to be important for Toxoplasma fitness [25], investigations in apicomplexan parasites have been so far almost exclusively focused on the apicoplast-located SUF pathway [26–30] and mostly in Plasmodium species. The SUF pathway was shown to be essential for the viability of malaria parasites during both the erythrocytic and sexual stages of development, and has thus been recognized as a putative avenue for discovering new antiparasitic drug targets (reviewed in [31]).…”

The vacuolar iron transporter mediates iron detoxification inToxoplasma gondii

“…In common with most eukaryotes, elemental analysis found that around 5% of the atoms of T. gondii atoms are iron (Al-sandaqchi et al, 2018). This iron is required for a number of essential cellular processes, including heme biosynthesis (Bergmann et al, 2020;Kloehn et al, 2020), iron sulphur (Fe-S) cluster biogenesis (Aw et al, 2020), and as a cofactor in several important proteins such as catalase (Kwok et al, 2004;Odberg-Ferragut et al, 2000), prolyl hydroxylases (Florimond et al, 2019) and the deoxyribonucleotide synthesis enzyme ribonucleotide reductase. Iron acquisition and use have been studied in various pathogenic parasites such as Trichomonas (Sehgal et al, 2012) and in some detail in kinetoplastids such as Leishmania (Zaidi et al, 2017) and Trypanosoma (Carbajo et al, 2021).…”