Figure 4. Genetic manipulations can alter hypoxic reduction in PDH activity and OCR.
(a) PDH activity assay performed on immuno-captured PDH from MIA PaCa-2 Hg, shHIF1α, shPDHK1, and PDP1 cells treated with 16 h normoxia or hypoxia (0.5% O2) showing a blunted response to hypoxia. (mean ± SD, two-tailed Student’s t-test, **p < 0.01) (b) Western blot of captured PDH from samples in (a) showing a commensurate reduction in phosphorylation of E1α. (c) Quantization of the phospho signal in panel (b) normalized to the signal for total E1α in (b) by ImageJ software. (d) OCR to ECAR ratios of modified MIA PaCa-2 cells incubated with or without 1 mM DMOG overnight showing modification of hypoxic OCR/ECAR. (mean ± SEM, two-tailed Student’s t-test, *p < 0.05, ***p < 0.001).