title: Phosphorylation of Eukaryotic Initiation Factor-2α during Stress and Encystation in Entamoeba Species.
abstract: Entamoeba histolytica is an enteric pathogen responsible for amoebic dysentery and liver abscess. It alternates between the host-restricted trophozoite form and the infective environmentally-stable cyst stage. Throughout its lifecycle E. histolytica experiences stress, in part, from host immune pressure. Conversion to cysts is presumed to be a stress-response. In other systems, stress induces phosphorylation of a serine residue on eukaryotic translation initiation factor-2α (eIF2α). This inhibits eIF2α activity resulting in a general decline in protein synthesis. Genomic data reveal that E. histolytica possesses eIF2α (EheIF2α) with a conserved phosphorylatable serine at position 59 (Ser59). Thus, this pathogen may have the machinery for stress-induced translational control. To test this, we exposed cells to different stress conditions and measured the level of total and phospho-EheIF2α. Long-term serum starvation, long-term heat shock, and oxidative stress induced an increase in the level of phospho-EheIF2α, while short-term serum starvation, short-term heat shock, or glucose deprivation did not. Long-term serum starvation also caused a decrease in polyribosome abundance, which is in accordance with the observation that this condition induces phosphorylation of EheIF2α. We generated transgenic cells that overexpress wildtype EheIF2α, a non-phosphorylatable variant of eIF2α in which Ser59 was mutated to alanine (EheIF2α-S59A), or a phosphomimetic variant of eIF2α in which Ser59 was mutated to aspartic acid (EheIF2α-S59D). Consistent with the known functions of eIF2α, cells expressing wildtype or EheIF2α-S59D exhibited increased or decreased translation, respectively. Surprisingly, cells expressing EheIF2α-S59A also exhibited reduced translation. Cells expressing EheIF2α-S59D were more resistant to long-term serum starvation underscoring the significance of EheIF2α phosphorylation in managing stress. Finally, phospho-eIF2α accumulated during encystation in E. invadens, a model encystation system. Together, these data demonstrate that the eIF2α-dependent stress response system is operational in Entamoeba species.
authors.affiliation: Department of Biological Sciences, Clemson University Clemson, South Carolina, United States of America.|Eukaryotic Pathogens Innovation Center (EPIC) Clemson University Clemson, South Carolina, United States of America.
authors.forename: Holland M
authors.initials: HM
authors.lastname: Hendrick
authors.affiliation: Department of Biological Sciences, Clemson University Clemson, South Carolina, United States of America.|Eukaryotic Pathogens Innovation Center (EPIC) Clemson University Clemson, South Carolina, United States of America.
authors.forename: Brenda H
authors.initials: BH
authors.lastname: Welter
authors.affiliation: Department of Biological Sciences, Clemson University Clemson, South Carolina, United States of America.|Eukaryotic Pathogens Innovation Center (EPIC) Clemson University Clemson, South Carolina, United States of America.
authors.forename: Matthew A
authors.identifier: http://orcid.org/0000-0002-1904-7226
authors.initials: MA
authors.lastname: Hapstack
authors.affiliation: Department of Biological Sciences, Clemson University Clemson, South Carolina, United States of America.|Eukaryotic Pathogens Innovation Center (EPIC) Clemson University Clemson, South Carolina, United States of America.
authors.forename: Steven E
authors.initials: SE
authors.lastname: Sykes
authors.affiliation: Department of Pharmacology and Toxicology Indiana University School of Medicine Indianaplois, IN United States of America.|Department of Microbiology and Immunology Indiana University School of Medicine Indianapolis, IN United States of America.
authors.forename: William J
authors.initials: WJ
authors.lastname: Sullivan
authors.affiliation: Department of Biological Sciences, Clemson University Clemson, South Carolina, United States of America.|Eukaryotic Pathogens Innovation Center (EPIC) Clemson University Clemson, South Carolina, United States of America.
authors.forename: Lesly A
authors.initials: LA
authors.lastname: Temesvari
mesh_terms: D015153:Blotting, Western; D004747:Entamoeba / Q000502:physiology*; D015852:Eukaryotic Initiation Factor-2 / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D030781:Organisms, Genetically Modified; D000069289:Parasite Encystment / Q000502:physiology*; D010766:Phosphorylation; D016133:Polymerase Chain Reaction; D013312:Stress, Physiological / Q000502:physiology*
chemical_list: D015852:Eukaryotic Initiation Factor-2