title: The Entamoeba histolytica Dnmt2 homolog (Ehmeth) confers resistance to nitrosative stress.
abstract: Nitric oxide (NO) has antimicrobial properties against many pathogens due to its reactivity as an S-nitrosylating agent. It inhibits many of the key enzymes that are involved in the metabolism and virulence of the parasite Entamoeba histolytica through S-nitrosylation of essential cysteine residues. Very little information is available on the mechanism of resistance to NO by pathogens in general and by this parasite in particular. Here, we report that exposure of the parasites to S-nitrosoglutathione (GSNO), an NO donor molecule, strongly reduces their viability and protein synthesis. However, the deleterious effects of NO were significantly reduced in trophozoites overexpressing Ehmeth, the cytosine-5 methyltransferase of the Dnmt2 family. Since these trophozoites also exhibited high levels of tRNA(Asp) methylation, the high levels suggested that Ehmeth-mediated tRNA(Asp) methylation is part of the resistance mechanism to NO. We previously reported that enolase, another glycolytic enzyme, binds to Ehmeth and inhibits its activity. We observed that the amount of Ehmeth-enolase complex was significantly reduced in GSNO-treated E. histolytica, which explains the aforementioned increase of tRNA methylation. Specifically, we demonstrated via site-directed mutagenesis that cysteine residues 228 and 229 of Ehmeth are susceptible to S-nitrosylation and are crucial for Ehmeth binding to enolase and for Ehmeth-mediated resistance to NO. These results indicate that Ehmeth has a central role in the response of the parasite to NO, and they contribute to the growing evidence that NO is a regulator of epigenetic mechanisms.
authors.affiliation: Department of Molecular Microbiology, Bruce Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, Haifa, Israel.
authors.forename: Rivi
authors.initials: R
authors.lastname: Hertz
authors.forename: Ayala
authors.initials: A
authors.lastname: Tovy
authors.forename: Michael
authors.initials: M
authors.lastname: Kirschenbaum
authors.forename: Meirav
authors.initials: M
authors.lastname: Geffen
authors.forename: Tomoyoshi
authors.initials: T
authors.lastname: Nozaki
authors.forename: Noam
authors.initials: N
authors.lastname: Adir
authors.forename: Serge
authors.initials: S
authors.lastname: Ankri
mesh_terms: D003545:Cysteine / Q000737:chemistry / Q000378:metabolism; D004248:DNA (Cytosine-5-)-Methyltransferases / Q000235:genetics* / Q000378:metabolism; D004351:Drug Resistance / Q000235:genetics*; D004748:Entamoeba histolytica / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D044127:Epigenesis, Genetic; D008745:Methylation; D016297:Mutagenesis, Site-Directed; D009569:Nitric Oxide / Q000378:metabolism / Q000494:pharmacology; D020030:Nitric Oxide Donors / Q000378:metabolism / Q000494:pharmacology*; D010751:Phosphopyruvate Hydratase / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D014176:Protein Biosynthesis / Q000187:drug effects; D015800:Protozoan Proteins / Q000235:genetics* / Q000378:metabolism; D012343:RNA, Transfer / Q000378:metabolism; D026422:S-Nitrosoglutathione / Q000378:metabolism / Q000494:pharmacology*; D015398:Signal Transduction; D013312:Stress, Physiological; D053058:Trophozoites / Q000187:drug effects / Q000254:growth & development / Q000378:metabolism
chemical_list: D020030:Nitric Oxide Donors; D015800:Protozoan Proteins; D009569:Nitric Oxide; D026422:S-Nitrosoglutathione; D012343:RNA, Transfer; D004248:DNA (Cytosine-5-)-Methyltransferases; D010751:Phosphopyruvate Hydratase; D003545:Cysteine