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. 2022 Apr 20;8:215. doi: 10.1038/s41420-022-01009-1

Fig. 4. ABT-199 activates ATF3&ATF4 that induce NOXA expression by activation of the integrated stress response pathway.

Fig. 4

A SW982/WT cells were incubated in the presence or absence of 15 µM ABT-199 and/or 5 nM BTZ ( + 10 µM Q-VD-OPh) for 8 h. Expression of ATF3/ATF3 (left panel), ATF4/ATF4 (middle panel) or PMAIP1/NOXA (right panel) was analyzed by qRT-PCR and Western blot (β-ACTIN as loading control). B–D SW982/WT cells were incubated in the presence or absence of 15 µM ABT-199 and/or 5 nM BTZ ( + 10 µM Q-VD-OPh) with/without 200 nM ISRIB for 8 h and mRNA expression of ATF3, ATF4 and PMAIP1 was analyzed by qRT-PCR. E SW982/WT cells were incubated with 15 µM ABT-199 and/or 5 nM BTZ ( + 10 µM Q-VD-OPh) in the presence or absence of 200 nM ISRIB for 8 h. Each 40 µg protein were analyzed by Western blot for the expression of ATF4, (P)-eIF2α, eIF2α, ATF3 and NOXA (β-ACTIN as loading control). F, G SW982/WT cells were incubated with 15 µM ABT-199 and/or 5 nM BTZ with/without 200 nM ISRIB for 24 h and apoptosis was assessed flow cytometrically after staining cells with Annexin V-APC and TMRM. Graphs show mean values and individual data points. Statistical significance was analyzed by an unpaired student´s t-test. # indicates statistical power vs. control. BTZ bortezomib, CTRL control, ISRIB integrated stress response inhibitor.