Research Article
Metabolic changes accompanying the loss of fumarate hydratase and malate–quinone oxidoreductase in the asexual blood stage of Plasmodium falciparum

https://doi.org/10.1016/j.jbc.2022.101897Get rights and content
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In the glucose-rich milieu of red blood cells, asexually replicating malarial parasites mainly rely on glycolysis for ATP production, with limited carbon flux through the mitochondrial tricarboxylic acid (TCA) cycle. By contrast, gametocytes and mosquito-stage parasites exhibit an increased dependence on the TCA cycle and oxidative phosphorylation for more economical energy generation. Prior genetic studies supported these stage-specific metabolic preferences by revealing that six of eight TCA cycle enzymes are completely dispensable during the asexual blood stages of Plasmodium falciparum, with only fumarate hydratase (FH) and malate–quinone oxidoreductase (MQO) being refractory to deletion. Several hypotheses have been put forth to explain the possible essentiality of FH and MQO, including their participation in a malate shuttle between the mitochondrial matrix and the cytosol. However, using newer genetic techniques like CRISPR and dimerizable Cre, we were able to generate deletion strains of FH and MQO in P. falciparum. We employed metabolomic analyses to characterize a double knockout mutant of FH and MQO (ΔFM) and identified changes in purine salvage and urea cycle metabolism that may help to limit fumarate accumulation. Correspondingly, we found that the ΔFM mutant was more sensitive to exogenous fumarate, which is known to cause toxicity by modifying and inactivating proteins and metabolites. Overall, our data indicate that P. falciparum is able to adequately compensate for the loss of FH and MQO, rendering them unsuitable targets for drug development.

Keywords

malaria
Plasmodium falciparum
mitochondrion
tricarboxylic acid cycle
aspartate shuttle
DiCre
metabolomics
oxaloacetate

Abbreviations

AAT
aspartate aminotransferase
ADSS
adenylosuccinate synthase
ARSL
argininosuccinate lyase
aTC
anhydrotetracycline
CMA
Complete Medium with Albumax
DHODH
dihydroorotate dehydrogenase
diCre
dimerizable Cre
FC
fold change
FH
fumarate hydratase
ΔFM
double knockout mutant of FH and MQO
gRNA
guide RNA
HA
homology arm
IDC
intraerythrocytic developmental cycle
iRBC
infected red blood cell
mETC
mitochondrial electron transport chain
MQO
malate–quinone oxidoreductase
PEP
phosphoenolpyruvate
PEPC
phosphoenolpyruvate carboxylase
RBC
red blood cell
RT
room temperature
SFG
superfolder GFP
S7P
sedoheptulose 7-phosphate
TCA
tricarboxylic acid
uRBC
uninfected red blood cell

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