Extended Data Fig. 2: Expression of key metabolic enzymes in MYC liver tumours and the method of genetically manipulating their expression in vivo.

a, A diagram depicting some of the most relevant transcriptomic changes in central carbon metabolism in MYC-driven liver tumours when compared with normal livers based on microarray data shown in (b). b, A heat map of gene expression of relevant metabolic enzymes and metabolite transporters in normal livers and MYC-driven liver tumours (n = 4 mice per group). Statistical analysis was performed using a two-tailed Student’s t-test. *, P < 0.05. c, Generation of liver specific conditional knockouts of the genes of interest in hepatocytes. CreER^T2 is expressed under the hepatocyte-specific albumin gene promoter. Cre is activated upon administration of tamoxifen. Simultaneously, the Enhanced Yellow Fluorescent Protein (eYFP) reporter gene is activated, following Cre-mediated excision of a loxP-flanked transcriptional “stop” sequence, after the Rosa26 locus. d–g, To confirm a cell of origin of MYC-driven liver tumours, the dose of AAV8-Cre (adenoassociated virus serotype 8 expressing Cre recombinase) required to activate the eYFP reporter expression in all the hepatocytes was titrated. Dose is expressed in genome copies (GC): d, Macroscopic fluorescent image of the livers of the mice treated with different doses of AAV8-Cre; e, Confocal fluorescent image of the mouse livers shown in (d) demonstrating that the dose of 8 × 10¹² GC induces eYFP expression in all the hepatocytes; f, Immunostaining with the cholangiocyte marker Pan-cytokeratin (PanCK) demonstrates that cholangiocytes are not targeted by AAV8-Cre; g, Rosa26-eYFP mice treated with 8 × 10¹² GC AAV8-Cre, were hydrodynamically transfected with MYC and MCL1, and the resulting tumours expressed eYFP, demonstrating that hepatocytes are the cell of origin of the tumours (n = 3 mice per dose). h, MYC and MCL1 are among genes frequently upregulated in human liver cancers. Analysis performed in cBioPortal^68,69, combining all available liver cancer databases, expressed as percentage of positive tumour samples.