Abstract
The apicoplast of Plasmodium falciparum parasites is believed to rely on the import of three-carbon phosphate compounds for use in organelle anabolic pathways, in addition to the generation of energy and reducing power within the organelle. We generated a series of genetic deletions in an apicoplast metabolic bypass line to determine which genes involved in apicoplast carbon metabolism are required for blood-stage parasite survival and organelle maintenance. We found that pyruvate kinase II (PyrKII) is essential for organelle maintenance, but that production of pyruvate by PyrKII is not responsible for this phenomenon. Enzymatic characterization of PyrKII revealed activity against all NDPs and dNDPs tested, suggesting that it may be capable of generating a broad range of nucleotide triphosphates. Conditional mislocalization of PyrKII resulted in decreased transcript levels within the apicoplast that preceded organelle disruption, suggesting that PyrKII is required for organelle maintenance due to its role in nucleotide triphosphate generation.
Highlights
With increasing resistance to current front-line antimalarials, there is a crucial need to find new therapeutic interventions with novel mechanisms of action (Dondorp et al, 2009; Trape, 2001)
Upon removal of mevalonate both deletion lines failed to grow (Figure 2d). These results demonstrate that both outer triose phosphate transporter (oTPT) and inner triose phosphate transporter (iTPT) are required for apicoplast maintenance in addition to blood-stage parasite survival
While these results are consistent with what was found in P. berghei for oTPT they are notably in conflict with what was found for iTPT, which did not cause a significant growth defect in P. berghei when deleted (Banerjee et al, 2012)
Summary
With increasing resistance to current front-line antimalarials, there is a crucial need to find new therapeutic interventions with novel mechanisms of action (Dondorp et al, 2009; Trape, 2001). The apicoplast organelle within the parasite has often been considered as a source of new drug targets since it is required for blood-stage survival, in addition to possessing evolutionarily distinct biochemical pathways that are not present in the human host (Goodman and McFadden, 2013; Janouskovec et al, 2010; Mukherjee and Sadhukhan, 2016). It is hypothesized that the apicoplast meets these needs primarily through the import and metabolism of three-carbon phosphates generated by parasite glycolysis (Lim et al, 2010; Ralph et al, 2004; Zocher et al, 2012). These three-carbon phosphate compounds include dihydroxyacetone phosphate (DHAP) and phosphoenolpyruvate (PEP), which are imported into the organelle through the outer triose phosphate transporter (oTPT) and inner triose phosphate transporter (iTPT) embedded in the outer and inner apicoplast membranes, respectively (Figure 1; Lim et al, 2010; Lim et al, 2016; Mullin et al, 2006)
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