Extended Data Fig. 8: Metabolic changes downstream of Pik3caH1047R/wt expression.
From: Organismal metabolism regulates the expansion of oncogenic PIK3CA mutant clones in normal esophagus
a-b, Proportion of WT-RYFP cells in mixed cultures with Pik3caH1047R/wt cells, after 15 days in minimal media normalized to baseline value. +/–Gluc indicates culture with/without glucose and 2-DG indicates culture with 5 mM 2-deoxyglucose. Each dot is a culture from a different animal, lines are mean values. n = 4 cultures per condition. c-e, In vitro cell competition assay. c, Protocol: Confluent mixed cultures of WT-RYFP and induced Pik3caH1047R/wt cells with/without DCA 25 mM for 28 days in minimal medium. d, Representative confocal images of basal cell layer of cultures from c, WT-RYFP cells, yellow, DAPI, blue. Scale bar, 20 μm. e, Flow cytometric analysis from mixed cultures in c. Each dot is a culture from a different mouse and lines connect mean values (n = 7-15 cultures). Two-tailed paired t-test. f, Protocol: Cre-RYFP control and Cre-Pik3caH1047R-YFP/wt mice were induced and treated with DCA. Clones with >1 basal cell sizes were analyzed at 28 days. Mice and clone numbers are shown. g, Heatmaps showing the differences between each treatment versus control in Cre-RYFP (upper panels) or Cre-Pik3caH1047R-YFP/wt (lower panels) mice from f. Two-tailed 2D Kolmogorov-Smirnov test. h-i, Average basal clone sizes h and proportion of first suprabasal layer cells i for each strain and treatment from f. Data includes clones with at least one basal cell. Bars are SD. Two-tailed unpaired t-test. j, Wild-type and Pik3caH1047R mutant basal cell clone size distributions from untreated or DCA-treated Cre-RYFP and Cre-Pik3caH1047R-YFP/wt mice, respectively, 28 days post- induction. n = 311-799 clones from 5-8 animals per condition. Dots indicate mean and lines standard deviation. Two-tailed Kolmogorov-Smirnov test and Contrast ART-C Post-hoc test of differences of differences between distributions. k. Proportion of WT-RYFP cells in mixed cultures with Pik3caH1047R/wt cells, at 15 days normalized to baseline value. Cells were culture in minimal media (CTL) or with Betulin (6 µg/ml), CB839 (10 µM), TOFA (30 µM) or TVB-2640 (0.1 µM). Each dot represents a culture from a different animal, lines correspond to mean values. n = 4 cultures from individual animals per condition. Two tailed paired t-test.
