Fig. 3

Complex I inhibition using BAY 87-2243 reduces ATP levels and induces an energy crisis in melanoma cells. a Melanoma cells were treated with different concentrations of BAY 87-2243 and ATP was measured after 8 h. Oligomycin (1 μM) served as positive control (n = 3). b, c Melanoma cells were treated with BAY 87-2243 (10 nM). Cell lysates were collected from treated cells (16 h) and probed with antibodies recognizing phospho-AMPK (Thr172), total AMPK, c phospho-RAPTOR (Ser792), d anti-phospho-ERK1/2 (Thr202/Tyr204), and anti-ERK1/2. Actin was used as a loading control. e Melanoma cells were treated with various concentrations of BAY 87-2243. Cell viability was measured in high (25 mM) and low (5 mM) glucose medium after 72 h (n = 3). IC50 values were calculated using GraphPad Prism. f A-375 cells were treated with BAY 87-2243 (10 nM) under low- (5 mM) and high (25 mM) glucose conditions. Cell lysates were collected from treated cells (16 h) and probed with antibodies recognizing phospho-AMPK (Thr172) and total AMPK. Actin was used as a loading control. g Melanoma cells were treated with BAY 87-2243 (10 nM) under low- (5 mM) and high (25 mM) glucose conditions and ATP was measured after 16 h (n = 3). h A-375 cells were treated with BAY 87-2243 (10 nM) under low- (5 mM) and high (25 mM) glucose conditions. Cytosolic ROS levels were measured using the ROS-reactive dye CM-H₂DCFDA after 16 h (n = 3). Data are represented as the mean ± SD. *p < 0.05