UDP-galactose and acetyl-CoA transporters as Plasmodium multidrug resistance genes.

A molecular understanding of drug resistance mechanisms enables surveillance of the effectiveness of new antimicrobial therapies during development and deployment in the field. We used conventional drug resistance selection as well as a regime of limiting dilution at early stages of drug treatment to probe two antimalarial imidazolopiperazines, KAF156 and GNF179. The latter approach permits the isolation of low-fitness mutants that might otherwise be out-competed during selection. Whole-genome sequencing of 24 independently derived resistant Plasmodium falciparum clones revealed four parasites with mutations in the known cyclic amine resistance locus (pfcarl) and a further 20 with mutations in two previously unreported P. falciparum drug resistance genes, an acetyl-CoA transporter (pfact) and a UDP-galactose transporter (pfugt). Mutations were validated both in vitro by CRISPR editing in P. falciparum and in vivo by evolution of resistant Plasmodium berghei mutants. Both PfACT and PfUGT were localized to the endoplasmic reticulum by fluorescence microscopy. As mutations in pfact and pfugt conveyed resistance against additional unrelated chemical scaffolds, these genes are probably involved in broad mechanisms of antimalarial drug resistance.

Targeting of a Transporter to the Outer Apicoplast Membrane in the Human Malaria Parasite Plasmodium falciparum.

Apicoplasts are vestigial plastids in apicomplexan parasites like Plasmodium, the causative agent of malaria. Apicomplexan parasites are dependant on their apicoplasts for synthesis of various molecules that they are unable to scavenge in sufficient quantity from their host, which makes apicoplasts attractive drug targets. Proteins known as plastid phosphate translocators (pPTs) are embedded in the outer apicoplast membrane and are responsible for the import of carbon, energy and reducing power to drive anabolic synthesis in the organelle. We investigated how a pPT is targeted into the outer apicoplast membrane of the human malaria parasite P. falciparum. We showed that a transmembrane domain is likely to act as a recessed signal anchor to direct the protein into the endomembrane system, and that a tyrosine in the cytosolic N-terminus of the protein is essential for targeting, but one or more, as yet unidentified, factors are also essential to direct the protein into the outer apicoplast membrane.

The evolution, metabolism and functions of the apicoplast.

The malaria parasite, Plasmodium falciparum, harbours a relict plastid known as the 'apicoplast'. The discovery of the apicoplast ushered in an exciting new prospect for drug development against the parasite. The eubacterial ancestry of the organelle offers a wealth of opportunities for the development of therapeutic interventions. Morphological, biochemical and bioinformatic studies of the apicoplast have further reinforced its 'plant-like' characteristics and potential as a drug target. However, we are still not sure why the apicoplast is essential for the parasite's survival. This review explores the origins and metabolic functions of the apicoplast. In an attempt to decipher the role of the organelle within the parasite we also take a closer look at the transporters decorating the plastid to better understand the metabolic exchanges between the apicoplast and the rest of the parasite cell.

The carbon and energy sources of the non-photosynthetic plastid in the malaria parasite.

The malaria parasite harbours an indispensable plastid known as the 'apicoplast'. The apicoplast's exact role remains uncertain, but it houses components involved in fatty acid, isoprenoid and haem biosyntheses. These pathways offer opportunities to develop anti-malarials. In the absence of photosynthesis, how apicoplast anabolism is fuelled is unclear. Here we investigated plant-like transporters of the apicoplast and measured their substrate preferences using a novel cell-free assay system to explore the carbon and energy sources of the apicoplast. The transporters exchange triose phosphate and phosphoenolpyruvate for inorganic phosphate, demonstrating that the apicoplast taps into host-derived glucose to fuel its metabolism.

New proteins in the apicoplast membranes: time to rethink apicoplast protein targeting.

Several apicomplexan parasites harbour an essential plastid known as the apicoplast. Apicoplasts import proteins and metabolites for several biological functions, but how import is achieved is largely unknown. Two recent reports have identified novel proteins in the apicoplast membranes, providing new perspectives on how proteins traffic to this organelle. The first report contributes to a newly recognized apicoplast-targeting pathway for membrane proteins, and the second identifies the first member of the protein-translocation complex in apicoplasts.

Membrane transporters in the relict plastid of malaria parasites.

Malaria parasites contain a nonphotosynthetic plastid homologous to chloroplasts of plants. The parasite plastid synthesizes fatty acids, heme, iron sulfur clusters and isoprenoid precursors and is indispensable, making it an attractive target for antiparasite drugs. How parasite plastid biosynthetic pathways are fuelled in the absence of photosynthetic capture of energy and carbon was not clear. Here, we describe a pair of parasite transporter proteins, PfiTPT and PfoTPT, that are homologues of plant chloroplast innermost membrane transporters responsible for moving phosphorylated C3, C5, and C6 compounds across the plant chloroplast envelope. PfiTPT is shown to be localized in the innermost membrane of the parasite plastid courtesy of a cleavable N-terminal targeting sequence. PfoTPT lacks such a targeting sequence, but is shown to localize in the outermost parasite plastid membrane with its termini projecting into the cytosol. We have identified these membrane proteins in the parasite plastid and determined membrane orientation for PfoTPT. PfiTPT and PfoTPT are proposed to act in tandem to transport phosphorylated C3 compounds from the parasite cytosol into the plastid. Thus, the transporters could shunt glycolytic derivatives of glucose scavenged from the host into the plastid providing carbon, reducing equivalents and ATP to power the organelle.