Atovaquone induces phosphorylation of eIF2α, upregulates ATF4-specific target genes, and suppresses OXPHOS in a dose-dependent fashion. (A) MOLM13 cells were treated with atovaquone (AQ; 25 μM) for the indicated times, then analyzed by western blotting. Data are representative of 2 independent experiments. (B) MV4-11 cells were treated with AQ (20 μM) for 6 hours, followed by quantitative RT-PCR (qRT-PCR) analysis. Data are means ± standard errors of the mean (SEMs) from 3 independent experiments. (C) AQ consistently upregulates the ATF4-specific target genes ATF3, CHAC1, CHOP, and REDD1. Kasumi-1, MV4-11, and K562 cells were treated with AQ (20 μM), thapsigargin (1 μM), or tunicamycin (5 μg/mL) for 6 hours, followed by qRT-PCR using the indicated primers. Data are means ± SEMs from 3 independent experiments. (D) AQ induced dose-dependent reductions in OCR of Kasumi-1 cells and the primary patient samples AML3 and AML5. Patient samples were rested in HS5 CM for 24 hours before a 3-hour incubation with the indicated doses of AQ. Data are means ± SEMs of 2 independent experiments. Ant/Rot, antimycin A/rotenone (complex 3 and 1 inhibitors, respectively); FCCP, carbonyl cyanide-4-(trifluromethoxy) phenylhydrazone (mitochondrial uncoupler); oligo, oligomycin (ATP synthase inhibitor).