FIG. 5.

OGFOD1 regulates the phosphorylation of eIF2α during recovery from arsenite-induced stress. (A) Abundance of phosphorylated eIF2α decreases upon knockdown of OGFOD1 or G3BP1. Cells were treated with a negative-control siRNA (neg. cntrl.) and siRNAs against G3BP1 and OGFOD1 72 h prior to harvest. Extracts were prepared from unstressed cells (untreated), arsenite-treated cells, and cells recovering from arsenite-induced stress. Proteins were analyzed by Western blotting. (B) Abundance of phosphorylated eIF2α increases after overexpression of OGFOD1 or G3BP1. Extracts were prepared as described above from HeLa cells transiently transfected with an empty DNA vector (neg. cntrl.) or with plasmids expressing G3BP1 and OGFOD1 cDNAs and subjected to Western blot analysis. (C) Abundance of phosphorylated eIF2α in cells recovering from arsenite-induced stress changes upon knockdown or overexpression of OGFOD1 and G3BP1. Levels of phosphorylated eIF2α and total eIF2α were quantitated for three independent experiments, using Image J software. Representative Western blots are shown in panels A and B. Percent changes in phosphorylated eIF2α were calculated relative to total eIF2α levels and normalized to the negative control. Error bars represent the standard errors of the means.