Figure 3
PTENWT cells have reduced metabolism in response to WNT/β-catenin signaling. (a) Bioenergetic metrics were calculated from the XFe96 extracellular flux bioanalyzer OCR and ECAR traces (see Supplementary Figure 3 for example traces). Upper panel PTENWT A375 cells, Lower panel PTENMut A2058 cells. Basal respiration was calculated from the OCR data at 26 min, subtracting the non-mitochondrial respiration. ATP production was calculated by the change in OCR before and after injection of oligomycin. Maximal respiratory capacity was calculated after injection of FCCP, as the maximum OCR reading subtracting non-mitochondrial respiration. Non-mitochondrial respiration was calculated from the OCR data following injection of complex I and III inhibitors; antimycin A and rotenone, respectively. Basal ECAR was calculated from the ECAR data at 26 min, by subtracting the non-glycolytic acidification. Mean shown±s.d. *P<0.05 or **P<0.01 and ***P<0.001. (b) The shift in cellular energy phenotypes was calculated from basal OCR and basal ECAR readings for control and WNT3A overexpressing PTENWT A375, and PTENMut A2058 cells. (c) Fold-change in CS activity normalized to total protein concentration. In parental lines, readings were taken following 48 h of stimulation with carrier or 50 ng/ml rWNT3A. (d) Fold-change in lactate secretion normalized to total protein concentration, using the same experimental conditions as described for c. For panels c and d, mean shown±s.d. *P<0.05 or **P<0.01 and ***P<0.001.
