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. 2024 Jan 10;80:101876. doi: 10.1016/j.molmet.2024.101876

Figure 3.

Figure 3

NF1 deficiency constrains oxidative ATP production. A) NF1 deficiency decreases basal and maximal OCR across models. Monensin is an ionophore that pushes cells to the maximal glycolytic range. 2DG is a competitive hexokinase inhibitor. Oligomycin (oligo) is a complex V inhibitor, fluoro-carbonyl cyanide phenylhydrazone (FCCP) is an uncoupling agent, and rotenone and antimycin A (Rot/AntiA) are complex I and complex III inhibitors, respectively. B) NF1 deficiency restricts joules of ATP (JATP) production in MCF7 cells and rat mammary fibroblasts. JATP values were calculated using CEAS (R Package). Dotted lines outline the basal bioenergetic scope area. The symbols represent the geometric mean, and the lines represent the 95 % confidence interval. MCF7: n = 2, 12 technical replicates per biological replicate. Fibroblast: n = 2, 24 technical replicates per biological replicate. Tumor cells: n = 2, 28 technical replicates per biological replicate. C) Nf1 deficiency restricts max bioenergetic scope (JATP from glycolysis x JATP from OXPHOS) in MCF7 cells and rat mammary fibroblasts. The bar represents the geometric mean, and the black, horizontal lines represent the 95 % confidence interval boundaries. JATP values were calculated using CEAS (R Package). D) NF1 deficiency has a divergent effect on glycolytic JATP production based on cell type. JATP values were calculated using CEAS (R Package). The bar represents the geometric mean, and the black, horizontal lines represent the 95 % confidence interval boundaries. MCF7: n = 2, 12 technical replicates per biological replicate. Fibroblast: n = 2, 24 technical replicates per biological replicate. Tumor cells: n = 2, 28 technical replicates per biological replicate. E) NF1 deficiency restricts OXPHOS JATP production. JATP values were calculated using CEAS (R Package). The bar represents the geometric mean, and the black, horizontal lines represent the 95 % confidence interval boundaries. MCF7: n = 2, 12 technical replicates per biological replicate. Fibroblast: n = 2, 24 technical replicates per biological replicate. Tumor cells: n = 2, 28 technical replicates per biological replicate. F) NF1 deficiency impacts OCR without altering total mitochondrial load as measured by Western blot analysis of voltage-dependent anion channel (VDAC) expression. Immunoblotting was carried out using antibodies against VDAC and β-tubulin.