Phosphorylation of Eukaryotic Initiation Factor-2α during Stress and Encystation in Entamoeba Species
Fig 9
Phospho-eIF2α accumulates during encystation in E. invadens.
(A) Western blot analysis of phospho-eIF2α and total eIF2α in E. invadens trophozoites (T) or trophozoites induced to encyst for 24 h. (B) Western blot analysis of phospho-eIF2α and total eIF2α in E. invadens trophozoites (T) induced to encyst for 72 h. The 72 h population was probed before (72) and after (72C) treatment with sarkosyl detergent, which eliminates un-encysted trophozoites from the population. (C) The ratio of phospho-eIF2α to total eIF2α was determined by scanning densitometry and image analysis (Image J, NCBI). The data represent the mean (± standard error) of at least three separate trials. All densitometry values were corrected for load using a single Coomassie stained band and the ratio of phospho-eIF2α to total eIF2α in control unencysted cells was arbitrarily set to 1.0. Representative Western blots are shown. There was significant accumulation of phospho-EieIF2α in E. invadens after 24 hours (24, *P<0.05) and 72 hours (72; **P<0.01) into encystation as well as in detergent-purified cysts (72C; ***P<0.001) (B). Coomassie blue staining of SDS-PAGE gels revealed equal protein loading. The lanes with the molecular weight markers (M) are indicated.