Novel approaches to kill Toxoplasma by exploiting the uncontrolled uptake of unsaturated fatty acids and vulnerability to lipid storage inhibition of the parasite [PublishAheadOfPrint]

Toxoplasma gondii, an obligate intracellular parasite replicating in mammalian cells within a parasitophorous vacuole (PV), is an avid scavenger of lipids retrieved from the host cell. Following lipid uptake, this parasite stores excess lipids into lipid droplets (LD). Here, we examined the lipid storage capacities of Toxoplasma upon supplementation of the culture medium with various fatty acids at physiological concentrations. Supplemental unsaturated fatty acids (oleate (OA), palmitoleate, linoleate) accumulate in large LD and impair parasite replication whereas saturated fatty acids (palmitate, stearate) neither stimulate LD formation nor impact growth. Examination of parasite growth defects with 0.4 mM OA reveals massive lipid deposits outside LD, indicating enzymatic inadequacies for storing neutral lipids in LD in response to the copious salvage of OA. Toxoplasma exposure to 0.5 mM OA leads to irreversible growth arrest and lipid-induced damage, confirming a major disconnect between fatty acid uptake and the parasite's cellular lipid requirements. The importance of neutral lipid synthesis and storage to avoid lipotoxicity is further highlighted by the selective vulnerability of Toxoplasma, both the proliferative and the encysted forms, to subtoxic concentrations of the acyl-CoA:diacylglycerol acyltransferase 1 (DGAT1) pharmacological inhibitor T863. T863-treated parasites do not form LD, but instead build up large membranous structures within the cytoplasm, which suggests improper channeling and management of lipid excess. Dual addition of OA and T863 to infected cells intensifies the deterioration of the parasite. Overall, our data pinpoint Toxoplasma DGAT as a promising drug target against toxoplasmosis, without incurring risk of toxicity for mammalian cells.

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