Figure 6. Jasplakinolide diminishes eIF2α phosphatase activity in vivo.
(A) Immunoblot for phosphorylated eIF2α (P-eIF2α), total eIF2α, and ATF4. Gcn2−/− MEFs were pre-treated with thapsigargin 300 nM for 30 min to induce eIF2α phosphorylation and ATF4 protein levels. GSK2606414A at 2 µM was then added for the indicated times. Protein lysates were analysed by SDS-PAGE and subjected to immunoblot. (B) Quantification of ‘A’ using ImageJ software. Mean ± SEM of n = 3 independent repeats. (C) Immunoblot for phosphorylated eIF2α (P-eIF2α) and total eIF2α. MEFs of the indicated genotypes were treated with or without jasplakinolide 1 µM for 1 hr. Protein lysates were analysed by SDS-PAGE and subjected to immunoblot.
Figure 6—figure supplement 1. Immunoblot for P-eIF2α, total eIF2α, and ATF4 (specific band marked with an asterisk) in lysates of wild type (WT) or eIF2αAA MEFs following treatment with thapsigargin 300 nM for 4 hr and/or jasplakinolide 1 µM for 4 hr.
