Fig. 4

Nrf2-dependent activation of the PPP is evolutionarily conserved and required to stimulate regeneration (A) Volcano plot of differentially expressed genes and Nrf2 target genes (green) identified by comparing WT and Nrf2KO animals treated with APAP at 4 dpf for 24 h. Genes within red area of volcano plots are significantly upregulated (log2FC > 0.6, p < 0.05), and genes within blue area of volcano plots are significantly downregulated (log2FC < 0.6, p < 0.05). (B) Heatmap depicting the differentially expressed genes in the pentose phosphate pathway following 24 h of DMSO or APAP treatment in WT and Nrf2KO zebrafish. (C) ChIP-qPCR analysis of NRF2 binding to the PRDX1, G6PD, and chromosome 2 (chr 2) gene desert loci in untreated HepG2 cells and HepG2 cells treated with 10 mM APAP for 6 or 24 h, n = 3. p values were calculated using an unpaired two-tailed Student?s t test comparing each sample to the respective untreated control. Data are shown as mean ± SEM. (D) Survival of WT zebrafish exposed to DMSO, APAP, G6PDi-1, or APAP in the presence of G6PDi-1 for 72 h from 4 dpf. p value was calculated using a Mantel-Cox test comparing the survival of APAP+G6PDi-1 to APAP alone. Data are shown as mean ± SEM. (E) Representative multiphoton images of hepatocytes in G1 phase (magenta) and S/G2-M phase (cyan) in livers of 4 dpf lf:FastFUCCI animals treated with DMSO, APAP, G6PDi-1, or APAP in the presence of G6PDi-1 for 24 h. (F) Quantification of mitotic hepatocytes (% S/G2-M) derived from lf:fastFUCCI analysis. p value was calculated using an unpaired two-tailed Student?s t test comparing each sample to its respective control. Data are shown as median and interquartile range, n = 15. (G) Representative images of BrdU incorporation (yellow) in hepatocytes (magenta) derived from lf:NLS-mCherry zebrafish exposed to DMSO, APAP, G6PDi-1, or APAP in the presence of G6PDi-1 for 24 h. (H) Quantification of BrdU incorporation (% BrdU+ hepatocytes) in livers of WT zebrafish following 24 h exposure to DMSO, APAP, G6PDi-1, or APAP in the presence of G6PDi-1. p value was calculated using an unpaired two-tailed Student's t test. Data are shown as median and interquartile range, n = 15. See also Figure S4.