RK-33 activates multiple ISR kinases to induce stress granules. (A). HapI cells were treated with 10 μM RK-33 for 1 h before lysis and western blot for indicated proteins. Representative blot shown. (B) U2OS cells were treated with RK-33 for 1 h prior to fixation and immunofluorescence for DDX3X and PABP. Representative images shown, one independent experiment. (C) U2OS cells were pretreated with 2BAct for 1.5 h, followed by 10 μM RK-33 ±2Bact for 1 h prior to fixation and immunofluorescence for DDX3X and PABP. The percentage of cells with stress granules were determined. Representative image shown. N > 100 cells in each condition were quantified from three independent experiments. Statistical significance: (ns) not significant, (*) P ≤ 0.05, (**) P ≤ 0.01, (***) P ≤ 0.001. Tests used: unpaired t-test with Welch correction (C). Error bars represent mean ± SEM.
