Exported Epoxide Hydrolases Modulate Erythrocyte Vasoactive Lipids during Plasmodium falciparum Infection

Abstract

Erythrocytes are reservoirs of important epoxide-containing lipid signaling molecules, including epoxyeicosatrienoic acids (EETs). EETs function as vasodilators and anti-inflammatory modulators in the bloodstream. Bioactive EETs are hydrolyzed to less active diols (dihydroxyeicosatrienoic acids) by epoxide hydrolases (EHs). The malaria parasite Plasmodium falciparum infects host red blood cells (RBCs) and exports hundreds of proteins into the RBC compartment. In this study, we show that two parasite epoxide hydrolases, P. falciparum epoxide hydrolases 1 (PfEH1) and 2 (PfEH2), both with noncanonical serine nucleophiles, are exported to the periphery of infected RBCs. PfEH1 and PfEH2 were successfully expressed in Escherichia coli, and they hydrolyzed physiologically relevant erythrocyte EETs. Mutations in active site residues of PfEH1 ablated the ability of the enzyme to hydrolyze an epoxide substrate. Overexpression of PfEH1 or PfEH2 in parasite-infected RBCs resulted in a significant alteration in the epoxide fatty acids stored in RBC phospholipids. We hypothesize that the parasite disruption of epoxide-containing signaling lipids leads to perturbed vascular function, creating favorable conditions for binding and sequestration of infected RBCs to the microvascular endothelium.

The chaperonin TRiC forms an oligomeric complex in the malaria parasite cytosol

“…, ; Mesen‐Ramirez et al . , ; Spillman, Dalmia, & Goldberg, ), due to endocytosis of exported proteins during residence in the PV and proteolysis down to the tightly folded GFP/tag in the digestive vacuole.…”

“…Consistent with this, evaluation of the full-length immunoblot from Figure S2) failed to reveal ã 27 kDa GFP "core". Exported proteins often display a proteolysis product (Boddey et al, 2016;Mesen-Ramirez et al, 2016;Spillman, Dalmia, & Goldberg, 2016), due to endocytosis of exported proteins during residence in the PV and proteolysis down to the tightly folded GFP/tag in the digestive vacuole.…”

Active-Site Flexibility and Substrate Specificity in a Bacterial Virulence Factor: Crystallographic Snapshots of an Epoxide Hydrolase

“…This may be especially pertinent for other EHs deployed by pathogens, aiding in the identification of a more complete set of substrate targets (Morisseau, 2013; Spillman et al, 2016). As we have shown, this approach can reveal unexpected active-site rearrangements and their effects on substrate selectivity.…”

The metabolic repair enzyme phosphoglycolate phosphatase regulates central carbon metabolism and fosmidomycin sensitivity inPlasmodium falciparum

“…For knock-out constructs, NF54 parasites were transfected at ring stage using 75 μg of each plasmid (pL7 and pUF-1-Cas9). For overexpression constructs, NF54 parasites were transfected at ring stage using 100 μg of pTEOE and 50 μg of pHTH as previously described (37). DNA/Cytomix were electroporated into red blood cells at 310V, 950 uF as previously described (38).…”