Aims: To determine the role of the kynurenine pathway in rhodoquinone and de novo NAD+ biosynthesis and whether NAD+ rescue pathways are essential in parasitic worms (helminths). Results: We demonstrate that rhodoquinone, the key electron transporter used by helminths under hypoxia, derives from the tryptophan catabolism even in the presence of a minimal kynurenine pathway. We show that of the kynurenine pathway genes only the kynureninase and tryptophan/indoleamine dioxygenases are essential for rhodoquinone biosynthesis. Metabolic labeling with tryptophan revealed that the lack of the formamidase and kynurenine monooxygenase genes did not preclude rhodoquinone biosynthesis in the flatworm Mesocestoides corti. In contrast, a minimal kynurenine pathway prevented de novo NAD+ biosynthesis, as revealed by metabolic labeling in M. corti, which also lacks the 3-hydroxyanthranilate 3,4-dioxygenase gene. Our results indicate that most helminths depend solely on NAD+ rescue pathways, and some lineages rely exclusively on the nicotinamide salvage pathway. Importantly, the inhibition of the NAD+ recycling enzyme nicotinamide phosphoribosyltransferase with FK866 led cultured M. corti to death. Innovation: We use comparative genomics of more than 100 hundred helminth genomes, metabolic labeling, HPLC-MS targeted metabolomics, and enzyme inhibitors to define pathways that lead to rhodoquinone and NAD+ biosynthesis in helminths. We identified the essential enzymes of these pathways in helminth lineages, revealing new potential pharmacological targets for helminthiasis. Conclusions: Our results demonstrate that a minimal kynurenine pathway was evolutionary maintained for rhodoquinone and not for de novo NAD+ biosynthesis in helminths, and shed light on the essentiality of NAD+ rescue pathways in helminths.

Selenoproteins are a diverse group of proteins containing selenocysteine (Sec)—the twenty-first amino acid—incorporated during translation via a unique recoding mechanism 1,2 . Selenoproteins fulfil essential roles in many organisms ¹ , yet are not ubiquitous across the tree of life 3–7 . In particular, fungi were deemed devoid of selenoproteins 4,5,8 . However, we show here that Sec is utilized by nine species belonging to diverse early-branching fungal phyla, as evidenced by the genomic presence of both Sec machinery and selenoproteins. Most fungal selenoproteins lack consensus Sec recoding signals (SECIS elements ⁹ ) but exhibit other RNA structures, suggesting altered mechanisms of Sec insertion in fungi. Phylogenetic analyses support a scenario of vertical inheritance of the Sec trait within eukaryotes and fungi. Sec was then lost in numerous independent events in various fungal lineages. Notably, Sec was lost at the base of Dikarya, resulting in the absence of selenoproteins in Saccharomyces cerevisiae and other well-studied fungi. Our results indicate that, despite scattered occurrence, selenoproteins are found in all kingdoms of life. © 2019, The Author(s), under exclusive licence to Springer Nature Limited.

Searching for prospective vanadium-based agents against Trypanosoma cruzi, the parasite causing Chagas disease, four new [VVO(8HQ–H)(L–2H)] compounds, where 8HQ is 8-hydroxyquinoline and L are tridentate salicylaldehyde semicarbazone derivatives L1–L4, were synthesized and characterized in the solid state and in solution. The compounds were evaluated on T. cruzi epimastigotes (CL Brener) as well as on VERO cells, as mammalian cell model. Compounds showed activity against T. cruzi (IC50 6.2–10.5 μM) of the same order than Nifurtimox and 8HQ, and a four- to sevenfold activity increase with respect to the free semicarbazones. For comparison, [VVO2(L–H)] series was prepared and the new [VVO2(L3–H)] was fully characterized. They showed negligible activity and low selectivity towards the parasite. The inclusion of 8HQ as ligand in [VVO(8HQ–H)(L–2H)] compounds led to good activities and increased selectivity towards the parasite with respect to 8HQ. ⁵¹V NMR experiments, performed to get insight into the nature of the active species, suggested partial decomposition of the compounds in solution to [VVO2(L–H)] and 8HQ. Depending on the dose, the compounds act as trypanocide or trypanostatic. A high uptake of vanadium in the parasites (58.51–88.9% depending on dose) and a preferential accumulation in the soluble protein fraction of the parasite was determined. Treated parasites do not seem to show a late apoptotic/necrotic phenotype suggesting a different cell death mechanism. In vivo toxicity study on zebrafish model showed no toxicity up to a 25 µM concentration of [VVO(8HQ–H)(L1–2H)]. These compounds could be considered prospective anti-T. cruzi agents that deserve further research.