Fig. 3: BH3-only proteins mediate FASN inhibition-induced apoptosis.

a FASN inhibition activates pro-apoptotic BH3-only proteins. Left. Representative immunoblot analysis of NOXA, BIM, PUMA, BAK, BAX, BCL-2, MCL-1, and FASN in cell lysates from BT-474 cells treated with graded concentrations of C75 for 48 h. Right. Q-PCR analyses of the expression of Bim, Noxa, and Puma. Before RNA extraction, cells were treated with C75 in the presence of the pan caspase inhibitor Q-VD-OPh (5 µmol/L) for 48 h, collected by trypsinization, sedimented at 250×g for 10 min and frozen on dry ice. Data are presented as mean ± SD (n = 3), p < 0.05 and p < 0.005 (* and **, respectively). b. Depletion of BH3-only proteins prevents FASNi-induced apoptotic cell death. Annexin V/propidium iodide staining-based flow cytometric assessment of apoptotic cell death in C75-treated BT-474 cells previously transfected with either control, BIM, NOXA, or PUMA single siRNAs (top panels) or their combinations (bottom panels). Annexin V⁺ cells data are presented as mean ± SD (n = 3), p < 0.05 and p < 0.005 (* and **, respectively). Also shown are representative immunoblots (n = 3) of the BIM, NOXA, and PUMA knockdown efficiencies achieved by the corresponding siRNAs.