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Metabolome Analysis Revealed Increase in S-Methylcysteine and Phosphatidylisopropanolamine Synthesis upon l-Cysteine Deprivation in the Anaerobic Protozoan Parasite Entamoeba histolytica*

l-Cysteine is ubiquitous in all living organisms and is involved in a variety of functions, including the synthesis of iron-sulfur clusters and glutathione and the regulation of the structure, stability, and catalysis of proteins. In the protozoan parasite Entamoeba histolytica, the causative agent of amebiasis, l-cysteine plays an essential role in proliferation, adherence, and defense against oxidative stress; however, the essentiality of this amino acid in the pathways it regulates is not well understood. In the present study, we applied capillary electrophoresis time-of-flight mass spectrometry to quantitate charged metabolites modulated in response to l-cysteine deprivation in E. histolytica, which was selected as a model for examining the biological roles of l-cysteine. l-Cysteine deprivation had profound effects on glycolysis, amino acid, and phospholipid metabolism, with sharp decreases in the levels of l-cysteine, l-cystine, and S-adenosylmethionine and a dramatic accumulation of O-acetylserine and S-methylcysteine. We further demonstrated that S-methylcysteine is synthesized from methanethiol and O-acetylserine by cysteine synthase, which was previously considered to be involved in sulfur-assimilatory l-cysteine biosynthesis. In addition, l-cysteine depletion repressed glycolysis and energy generation, as it reduced acetyl-CoA, ethanol, and the major nucleotide di- and triphosphates, and led to the accumulation of glycolytic intermediates. Interestingly, l-cysteine depletion increased the synthesis of isopropanolamine and phosphatidylisopropanolamine, and it was confirmed that their increment was not a result of oxidative stress but was a specific response to l-cysteine depletion. We also identified a pathway in which isopropanolamine is synthesized from methylglyoxal via aminoacetone. To date, this study represents the first case where l-cysteine deprivation leads to drastic changes in core metabolic pathways, including energy, amino acid, and phospholipid metabolism.