Fig. 7. Metabolic rewiring of glucose metabolism into the PPP and de novo lipogenesis participates to ChREBP pro-proliferative effects.

a–c Parental and ChREBP overexpressing SNU449 cells were treated with 6-AN (6-aminonicotinamide, 40 μM) for 24 h. Cells were then incubated for 30 min with 11 mM of ¹³C₆-glucose. a Enrichment in (m+6) 6-phosphogluconate from ¹³C₆-glucose (n = 4 independent experiments). b Enrichment in (m+5) ribose 5-phopshate from ¹³C₆-glucose in response to ChREBP overexpression (n = 4 independent experiments). c Enrichment in (m+3) ribose 5-phopshate from ¹³C₆-glucose in response to ChREBP overexpression (n = 4 independent experiments). d De novo nucleotide synthesis from parental and ChREBP overexpressing SNU449 cells incubated 6 h with 11 mM of ¹⁴C₆-labeled glucose (n = 4 independent experiments). e Effect of 6-AN treatment (40 μM, 24 h) on ChREBP-mediated increase in hepatocyte proliferation was studied in SNU449 cells. Representative clonogenic assays shown (n = 7 independent experiments). f Effect of 6-AN treatment (40 μM, 24 h) and dNTPs rescue (100 μM each) on ChREBP-mediated increase in cell proliferation. Cell proliferation index was determined by measuring the % of BrdU positive cells (n = 4 independent experiments). g Representative Western blot analysis of proteins involved in PPP pathway (n = 10 biologically independent mice per group). h De novo nucleotide synthesis from ¹⁴C₆-labeled glucose in ChREBP tumors (n = 3 biologically independent mice per group). i ChIP experiments measuring ChREBP occupancy at the G6PDH, PGD, TKT and RPIA promoters in ChREBP tumors relative to IgG controls (n = 4 biologically independent mice per group). j Relative NADPH/NADP ratio in ChREBP overexpressing tumors (n = 6 biologically independent mice per group). k De novo lipid synthesis from ¹⁴C₆-labeled glucose in ChREBP tumors (n = 3 biologically independent mice per group). l Representative Western blot analysis of proteins involved in de novo lipogenic pathways (n = 10 biologically independent mice per group). m ChIP experiments measuring ChREBP occupancy at the ACC, FAS and SCD1 promoters in ChREBP tumors relative to IgG controls (n = 4 biologically independent mice per group). n Measurement of SCD1 activity in ChREBP tumors (n = 3 biologically independent mice per group). o–r FAS expression was knockdown by Crispr/Cas9 in ChREBP overexpressing SNU449 cells (FASi). o Representative Western blot showing FAS deletion in ChREBP overexpressing SNU449 (n = 3 independent experiments). p De novo lipid synthesis from ¹⁴C₆-labeled glucose. SNU449 cells were incubated 6 h with 11 mM of ¹⁴C₆-labeled glucose (n = 3 independent experiments). q Representative clonogenic assays shown after FAS silencing (n = 3 independent experiments). r Effect of FAS silencing and oleate supplementation (50 μM) on ChREBP-mediated increase in cell proliferation. Cell proliferation index was determined by measuring the % of BrdU positive cells (n = 3 independent experiments). All error bars represent mean ± SEM. a–d, f, h, k, n, p, r Statistical analyses were made using two-way ANOVA and Tukey’s multiple-comparisons test. i, j and m Statistical analyses were determined by unpaired two-sided Student’s t test. Source data are provided as a Source Data file.