Stage-specific essentiality predictions of experimentally validated druggable targets and single-gene deletion experiments.

<p>Comprehensive map of experimentally tested treatment targets for <i>P</i>. <i>falciparum</i> with stage-specific model predictions projected (in color) projected on top of the map. Colored reaction pathways correspond to drug inhibition studies (Table C in <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1005895#pcbi.1005895.s002" target="_blank">S1 Tables</a>) and colored reaction names in rectangles correspond to single gene deletion experiments (Table B in <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1005895#pcbi.1005895.s002" target="_blank">S1 Tables</a>). The color legend inset corresponds to iAM-Pf480 predictions. Validated drug targets that are also predictive to reduce growth in proliferative as well as late gametocyte stages are of particular interest. This comprehensive assessment of the model and experimental results enables stratification of existing drugs, new drugs to target, as well as new areas of metabolism warranting further investigation. See <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1005895#pcbi.1005895.s004" target="_blank">S2 Fig</a> for the high resolution version of the figure.</p

Species-specific models provide mechanistic explanation for differences in drug response between human- and rodent-infecting malaria species.

<p><b>(a)</b> The core and pan metabolic content of 5 malaria species was identified based on the respective species-specific reconstructions. The core content, illustrated by the intersection of the Venn diagram, is shared by all species. The pan content represents the union of the content across all of the multi-species reconstructions. <b>(b)</b> 14 metabolic reactions differed in their presence across the 5 reconstructed Plasmodium species. <b>(c)</b> Thiamine pyrophosphokinase (TPK) and <b>(d)</b> Choline kinase (CK) were predicted by the models to be essential for the growth of the rodent-infecting species (<i>P</i>. <i>berghei</i>) while their deletion had no effect on the growth of human and non-human primate species. Differential essentiality of TPK is due to absence of phosphomethylpyrimidine kinase and thiamine-phosphate pyrophosphorylase the rodent-infecting species. In the case of CK, the differential essentiality is due to the absence of phosphoethanolamine N-methyltransferase. (See Table A in <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1005895#pcbi.1005895.s002" target="_blank">S1 Tables</a> for reactions abbreviations and gene-protein-reaction associations). <b>(e)</b> Pantothenate metabolism showed differences in essentiality between stage- and species-specific models. Tables indicate percentage in growth reduction compared to the WT upon deletion of the respective gene. ‘X’ indicates absence of a reaction from the respective reconstruction, ‘—‘ indicates no effect on growth upon deletion of the corresponding reaction and ‘%’ indicates the growth reduction percentage resulting from deletion of the corresponding gene. T: trophozoite, GII: early gametocyte stage, GV: late gametocyte stage, Ook: ookinete. See <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1005895#pcbi.1005895.s005" target="_blank">S3 Fig</a> for the high resolution version of the figure.</p

Life cycle stage specific models of <i>P</i>. <i>falciparum</i> predict gene targets that are essential for asexual, sexual and mosquito stages.

<p><b>(a)</b> The core and pan metabolic content of the genome-scale life cycle stage specific models of <i>P</i>. <i>falciparum</i> are 720 and 1002 reactions, respectively. <b>(b-c)</b> Parameters and AUC plots for performance evaluation of stage-specific pairwise differential gene expression comparisons following a similar approach to Nam, et al.[<a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1005895#pcbi.1005895.ref052" target="_blank">52</a>] (<a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1005895#pcbi.1005895.s004" target="_blank">S2 Fig</a>). AUROC: area under the receiver operator curve (ROC), DEG: differential expression analysis, T: trophozoite, GII: early gametocyte stage, GV: late gametocyte stage, Ook: ookinete.</p

Stage-specific central metabolic flux patterns in malaria.

<p><b>(a)</b> Correlated reaction sets for iAM-Pf480 were used to define stage and model specific pathways, which were analyzed and compared across different stages. Modularity indices (see <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1005895#sec012" target="_blank">methods</a>) were 0.023, 0.024, 0.026, 0.145, 0.022 for the T, Schizont, GII, GV, and Ook stages, respectively. In proliferative versus non-proliferative stages of malaria, there were changes in the patterns of central carbon metabolism, notably the non-oxidative PPP and glycolysis. <b>(b)</b> The direction of flux in the non-oxidative branch of PPP goes towards production of glycolytic intermediates in the Trophozoite, Schizont, GII, and GV stages but not the Ookinete stage. Reversal of non-oxidative pentose phosphate pathway fluxes in the Ook enables provision of ribose 5 phosphate (r5p) needed for the synthesis of nucleotide precursors of DNA. The non-oxidative branch in the schizont is colored in red indicating its coupling to growth rate in this stage (Table G in <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1005895#pcbi.1005895.s002" target="_blank">S1 Tables</a>). Both the oxidative and non-oxidative PPP branches were correlated in GII. Glycolysis was split into upper and lower branches in all stages except GV where the non-oxidative PPP branch was correlated with inositol metabolism. Arrows are omitted from the schizont pathway map to account for reduced flux values relative the other 4 stages. <b>(c)</b> Predicted sampled flux distribution are shown in the non-oxidative branch of PPP (Transketolase; TKT1) and inositol metabolism (myo-inositol-3-phosphate lyase; MI3PS) across all the stages showing increased involvement of inositol metabolism in the GV stage (see supplementary material for discussion and <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1005895#pcbi.1005895.s003" target="_blank">S1 Fig</a> for the high resolution version of the figure).</p