Life cycle studies of the hexose transporter ofPlasmodiumspecies and genetic validation of their essentiality

Abstract

A Plasmodium falciparumhexose transporter (PfHT) has previously been shown to be a facilitative glucose and fructose transporter. Its expression in Xenopus laevisoocytes and the use of a glucose analogue inhibitor permitted chemical validation of PfHT as a novel drug target. Following recent re-annotations of the P. falciparum genome, other putative sugar transporters have been identified. To investigate further if PfHT is the key supplier of hexose to P. falciparum and to extend studies to different stages of Plasmodium spp., we functionally analysed the hexose transporters of both the human parasite P. falciparum and the rodent parasite Plasmodium berghei using gene targeting strategies. We show here the essential function of pfht for the erythrocytic parasite growth as it was not possible to knockout pfht unless the gene was complemented by an episomal construct. Also, we show that parasites are rescued from the toxic effect of a glucose analogue inhibitor when pfht is overexpressed in these transfectants. We found that the rodent malaria parasite orthologue, P. berghei hexose transporter (PbHT) gene, was similarly refractory to knockout attempts. However, using a single cross-over transfection strategy, we generated transgenic P. berghei parasites expressing a PbHT–GFP fusion protein suggesting that locus is amenable for gene targeting. Analysis of pbht-gfp transgenic parasites showed that PbHT is constitutively expressed through all the stages in the mosquito host in addition to asexual stages. These results provide genetic support for prioritizing PfHT as a target for novel antimalarials that can inhibit glucose uptake and kill parasites, as well as unveiling the expression of this hexose transporter in mosquito stages of the parasite, where it is also likely to be critical for survival.

Glucose‐6‐phosphate metabolism in Plasmodium falciparum

“…Moreover, the glucosederived inhibitors show an inhibition of asexual blood stage P. falciparum growth in vitro and of different P. berghei life cycle stages in vivo, thereby validating PfHT as a valuable antimalarial drug target (46). This assumption was further supported by a study showing that PfHT is essential for both P. falciparum and P. berghei (47). Additionally, three other putative sugar transporters have been identified in the genome of P. falciparum.…”

Impact of trehalose transporter knockdown onAnopheles gambiaestress adaptation and susceptibility toPlasmodium falciparuminfection

“…So far, orthologs of these genes have not been identified in Plasmodium spp. A hexose transporter was cloned from P. falciparum, but targeted gene deletion proved lethal during the erythrocytic stages of the parasite (29). Therefore, it is likely that P. falciparum propagation during vector stages depletes glucose from the mosquito by scavenging uptake into the parasite cell through membrane transporters.…”

Apicomplexan Energy Metabolism: Carbon Source Promiscuity and the Quiescence Hyperbole

“…Up to 93% of glucose is converted directly to lactate in asexual stages [26][ 4 _ T D $ D I F F ] and is ultimately excreted into the surrounding host cell. Perturbation of glucose uptake is detrimental to parasite growth [27,28], suggesting that glycolysis is an important part of the parasite's strategy for rapid proliferation. In a manner analogous to the Warburg effect observed in highly-proliferative cancer lines and other rapidly-growing cells (e.g., yeast and bloodstream Trypanosoma spp.…”