Extended Data Fig. 3: Transient proliferation of zone-1 and -2 GLUL^neg hepatocytes following β-catenin and MYC activation.

a-e, AAV8.TBG.Cre (2 × 10¹¹ GC/ml) treated livers sampled 4 and 10 days post administration a, LW/BW, Biological replicates: Day 4—WT n = 10, R26^LSL-MYC (M) n = 11, Ctnnb1^ex3/WT (B) n = 14, Ctnnb1^ex3/WT;R26^LSL-MYC (BM) n = 9; Day 10—WT n = 10, M n = 11, B n = 10, BM n = 11. Bars are mean ± s.d. One-way ANOVA with Holm-Sidak’s multiple comparisons test. b, Confocal IF staining for BrdU (red), GLUL (yellow), and HNF4α (white) 10 days post induction. Nuclei counterstained with DAPI (blue), n = 6. Scale bars, 100μm. c, Quantification of GLUL^pos hepatocytes. Biological replicates: Day 4—WT n = 10, M n = 11, B n = 14, BM n = 7; Day 10—WT n = 10, M n = 10, B n = 11, BM n = 7. Bars are mean ± s.d. One-way ANOVA with Holm-Sidak’s multiple comparisons test. d, Whole liver RNA-Seq, top reactome pathways enriched in downregulated and upregulated genes between day-4 and -10 BM livers. Dots are coloured according to their adjusted P-values. The enrichPathway() function in R, using a two-sided hypergeometric model determined the probability of geneset overlap occurring by chance. Multiple testing correction was applied using the Benjamini-Hochberg method. Biological replicates: Day 4, n = 5; Day 10, n = 3. e, Quantification of BrdU^pos hepatocytes, stratified into GLUL-positive and -negative subsets. Two-sided Wilcoxon matched-pairs test. Biological replicates: n = 6 per condition. f, Quantification of BrdU^pos hepatocytes in Day-4 vehicle- and rapamycin-treated mice. Biological replicates: M vehicle n = 5, rapamycin n = 7; B vehicle n = 4, rapamycin n = 4. Bars are mean ± s.d. One tailed Mann–Whitney test. g, Quantification of BrdU^pos hepatocytes in Igfbp2 knockout (Igfbp2^–/–) mice. Biological replicates: M Igfbp2^+/+ n = 13, Igfbp2^–/– n = 9; B Igfbp2^+/+ n = 17, Igfbp2^–/– n = 8. Bars are mean ± s.d. One tailed t-test and Mann–Whitney test. Data from Fig. 1c included in Igfbp2^+/+ groups. h, Acute BM activation combined with viral-vector delivery of an Igfbp2 expressing transgene. i-k, LW/BW and quantification of BrdU^pos and GLUL^pos hepatocytes 8 days after AAV8-vector administration. Biological replicates: n = 10. Bars are mean ± s.d. two tailed Mann–Whitney test. l, Acute BM activation combined with viral-vector delivery of a Akt transgene containing a myristoylation sequence. m-o, LW/BW and quantification of BrdU^pos and GLUL^pos hepatocytes 8 days after AAV8-vector administration. Biological replicates: n = 5. Bars are mean ± s.d. two tailed Mann–Whitney test. p, Acute BM activation combined with PTEN loss. The illustrations of the mouse and adenovirus in panels h,l,p were adapted from Medical Art Servier (https://servier.com) under a CC BY 4.0 licence. q-s, LW/BW and quantification of BrdU^pos and GLUL^pos hepatocytes 8 days after AAV8-vector administration. Biological replicates: BM n = 7, BMP n = 6. Bars are mean ± s.d. two tailed Mann–Whitney test.

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