Fig. 5 | Nature Communications

Fig. 5

From: Spatial-fluxomics provides a subcellular-compartmentalized view of reductive glutamine metabolism in cancer cells

Fig. 5

Reversed CS flux supports cell survival and growth in SDH-deficient cells. a, b Isotopic labeling kinetics of cytosolic and mitochondrial citrate m + 5 and mitochondrial malate m + 3 when feeding SDH-WT (a) and SDH-KO cells (b) with [U-13C]-glutamine. c Mitochondrial to cytosolic metabolite pool size ratios (y-axis) in SDH-WT (green) and SDH-KO cells (orange); average fold change in whole-cell pool size upon SDH deficiency is indicated by color (x-axis). d Gibbs free energy of mitochondrial (IDH2/3) and cytosolic (IDH1) IDH isozymes (in the oxidative direction) in SDH-proficient (green) and deficient cells (orange). e Mitochondrial and cytosolic fluxes in SDH-KO cells, showing percentages from citrate synthase flux in the SDH-positive cells. Arrow represents the direction of net flux; number represents net flux in the direction of the arrow and number in parenthesis correspond to the backward flux. Confidence intervals for estimated fluxes shown in Supplementary Data 8. f Protein expression of mitochondrial and cytosolic markers in isolated mitochondria from SDH-WT and KO cells. g Fractional labeling of acetyl-CoA (m + 2) and malate (m + 4) in isolated mitochondria cultured with [U-13C]-citrate. h Fractional labeling of acetyl-CoA (m + 2) and malate (m + 4) in isolated SDH-KO mitochondria cultured with [U-13C]-citrate with or without ATP and ACLY inhibitor (BMS-303141). i Relative cell viability for SDH-WT (green) and SDH-KO cells (orange) following the addition of increasing concentration of dichloroacetate (DCA). XTT cell viability assay was performed 72 h after DCA treatment. j Relative metabolite pool sizes in SDH-WT (green) and SDH-KO cells (orange) with 10 mM DCA (24 h); compared to untreated control. k Metabolite fractional labeling from [U-13C]-glutamine in SDH-KO cells without (green) or with (orange) 10 mM DCA (24 h). *P < 0.05, **P < 0.01, and ***P < 0.001 by multiple t-test analysis with FDR correction (f) or by two-sample t-test (c, e, g, and h). n.d. not detected. Data are mean ± SD, n = 3 independent biological replicates

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