Fig. 1: The glycolysis‒oxPPP switch depends on GAPDH oxidation.
From: The GAPDH redox switch safeguards reductive capacity and enables survival of stressed tumour cells
a, GAPDH activity as measured in the lysate of HAP1 cells expressing either WT or mutant (Y314F) GAPDH, and following treatment of intact cells with 100 µM H2O2 for 5 min. Activity values are normalized relative to untreated WT cells. Data are presented as mean ± standard deviation (s.d.), based on n = 3 biological replicates with n = 3 technical replicates each. NS, non-significant; ***P < 0.001; P = 0.2413 and 0.0005, based on a two-tailed unpaired t-test. b, Glutathionylation of GAPDH Cys-152 in WT and mutant cells, before and after treatment of intact cells with 50 µM H2O2 for 5 min, as determined by LC–MS/MS analysis. Bars represent the mean of n = 3 technical replicates. c, Hyperoxidation (sulfinylation and/or sulfonylation) of GAPDH Cys-152 as visualized by immunoblotting before and after treatment of intact cells with up to 2 mM H2O2 for 5 min. Representative of n = 3 independent experiments. d, Partitioning of [1,2-13C]glucose flux into glycolysis and PPP, as determined by the isotopic signature of F6P. WT and mutant cells were left untreated, or treated with either 10 µM (low) or 50 µM (high) H2O2 for 30 s. Data are presented as mean ± s.d., based on n = 3 for H2O2 treated samples. *P < 0.05, **P < 0.01; P = 0.0016 and 0.0347, based on a two-tailed unpaired t-test. e, Change in the extracellular acidification rate (ECAR) in response to treatment of WT and mutant cells with 100 µM H2O2. Data are presented as mean ± s.d. (n = 3 biological replicates with n = 3 technical replicates each). *P < 0.05; P = 0.0291, based on a two-tailed unpaired t-test. f, Scheme depicting the rerouting of glucose flux between glycolysis and PPP in response to GAPDH oxidation. Blue arrows: glycolytic flux; red arrows: PPP flux. Please note that F6P and G3P are intermediates of both glycolysis and the PPP. Hence, GAPDH oxidation allows for the cycling of intermediates between the PPP and upper glycolysis, as indicated by reversed arrows in upper glycolysis (right).
