A nonmitochondrial hydrogen production in Naegleria gruberi

Naegleria gruberi is a free-living heterotrophic aerobic amoeba well known for its ability to transform from an amoeba to a flagellate form. The genome of N. gruberi has been recently published, and in silico predictions demonstrated that Naegleria has the capacity for both aerobic respiration and anaerobic biochemistry to produce molecular hydrogen in its mitochondria. This finding was considered to have fundamental implications on the evolution of mitochondrial metabolism and of the last eukaryotic common ancestor. However, no actual experimental data have been shown to support this hypothesis. For this reason, we have decided to investigate the anaerobic metabolism of the mitochondrion of N. gruberi. Using in vivo biochemical assays, we have demonstrated that N. gruberi has indeed a functional [FeFe]-hydrogenase, an enzyme that is attributed to anaerobic organisms. Surprisingly, in contrast to the published predictions, we have demonstrated that hydrogenase is localized exclusively in the cytosol, while no hydrogenase activity was associated with mitochondria of the organism. In addition, cytosolic localization displayed for HydE, a marker component of hydrogenase maturases. Naegleria gruberi, an obligate aerobic organism and one of the earliest eukaryotes, is producing hydrogen, a function that raises questions on the purpose of this pathway for the lifestyle of the organism and potentially on the evolution of eukaryotes

Inhibition of Fatty Acid Oxidation as a New Target To Treat Primary Amoebic Meningoencephalitis

Primary amoebic meningoencephalitis (PAM) is a rapidly fatal infection caused by the free-living amoeba Naegleria fowleri The amoeba migrates along the olfactory nerve to the brain, resulting in seizures, coma, and, eventually, death. Previous research has shown that Naegleria gruberi, a close relative of N. fowleri, prefers lipids over glucose as an energy source. Therefore, we tested several already-approved inhibitors of fatty acid oxidation alongside the currently used drugs amphotericin B and miltefosine. Our data demonstrate that etomoxir, orlistat, perhexiline, thioridazine, and valproic acid inhibited growth of N. gruberi We then tested these compounds on N. fowleri and found etomoxir, perhexiline, and thioridazine to be effective growth inhibitors. Hence, not only are lipids the preferred food source for N. gruberi, but also oxidation of fatty acids seems to be essential for growth of N. fowleri Inhibition of fatty acid oxidation could result in new treatment options, as thioridazine inhibits N. fowleri growth in concentrations that can be reached at the site of infection. It could also potentiate currently used therapy, as checkerboard assays revealed synergy between miltefosine and etomoxir. Animal testing should be performed to confirm the added value of these inhibitors. Although the development of new drugs and randomized controlled trials for this rare disease are nearly impossible, inhibition of fatty acid oxidation seems a promising strategy as we showed effectivity of several drugs that are or have been in use and that thus could be repurposed to treat PAM in the future

Iron Uptake Mechanisms in Marine Phytoplankton

International audienceOceanic phytoplankton species have highly efficient mechanisms of iron acquisition, as they can take up iron from environments in which it is present at subnanomolar concentrations. In eukaryotes, three main models were proposed for iron transport into the cells by first studying the kinetics of iron uptake in different algal species and then, more recently, by using modern biological techniques on the model diatom Phaeodactylum tricornutum. In the first model, the rate of uptake is dependent on the concentration of unchelated Fe species, and is thus limited thermodynamically. Iron is transported by endocytosis after carbonate-dependent binding of Fe(III)' (inorganic soluble ferric species) to phytotransferrin at the cell surface. In this strategy the cells are able to take up iron from very low iron concentration. In an alternative model, kinetically limited for iron acquisition, the extracellular reduction of all iron species (including Fe') is a prerequisite for iron acquisition. This strategy allows the cells to take up iron from a great variety of ferric species. In a third model, hydroxamate siderophores can be transported by endocytosis (dependent on ISIP1) after binding to the FBP1 protein, and iron is released from the siderophores by FRE2-dependent reduction. In prokaryotes, one mechanism of iron uptake is based on the use of siderophores excreted by the cells. Iron-loaded siderophores are transported across the cell outer membrane via a TonB-dependent transporter (TBDT), and are then transported into the cells by an ABC transporter. Open ocean cyanobacteria do not excrete siderophores but can probably use siderophores produced by other organisms. In an alternative model, inorganic ferric species are transported through the outer membrane by TBDT or by porins, and are taken up by the ABC transporter system FutABC. Alternatively, ferric iron of the periplasmic space can be reduced by the alternative respiratory terminal oxidase (ARTO) and the ferrous ions can be transported by divalent metal transporters (FeoB or ZIP). After reoxidation, iron can be taken up by the high-affinity permease Ftr1

Crusade for iron: iron uptake in unicellular eukaryotes and its significance for virulence.

International audienceThe effective acquisition of iron is a pre-requisite for survival of all organisms, especially parasites that have a high iron requirement. In mammals, iron homeostasis is meticulously regulated; extracellular free iron is essentially unavailable and host iron availability has a crucial role in the host-pathogen relationship. Therefore, pathogens use specialized and effective mechanisms to acquire iron. In this review, we summarize the iron-uptake systems in eukaryotic unicellular organisms with particular focus on the pathogenic species: Candida albicans, Tritrichomonas foetus, Trypanosoma brucei and Leishmania spp. We describe the diversity of their iron-uptake mechanisms and highlight the importance of the process for virulence

Nonreductive Iron Uptake Mechanism in the Marine Alveolate Chromera velia1[W]

Chromera velia is a newly cultured photosynthetic marine alveolate. This microalga has a high iron requirement for respiration and photosynthesis, although its natural environment contains less than 1 nm of this metal. We found that this organism uses a novel mechanism of iron uptake, differing from the classic reductive and siderophore-mediated iron uptake systems characterized in the model yeast Saccharomyces cerevisiae and present in most yeasts and terrestrial plants. C. velia has no trans-plasma membrane electron transfer system, and thus cannot reduce extracellular ferric chelates. It is also unable to use hydroxamate siderophores as iron sources. Iron uptake from ferric citrate by C. velia is not inhibited by a ferrous chelator, but the rate of uptake is strongly decreased by increasing the ferric ligand (citrate) concentration. The cell wall contains a large number of iron binding sites, allowing the cells to concentrate iron in the vicinity of the transport sites. We describe a model of iron uptake in which aqueous ferric ions are first concentrated in the cell wall before being taken up by the cells without prior reduction. We discuss our results in relation to the strategies used by the phytoplankton to take up iron in the oceans