Extended Data Fig. 5: Bmal1-regulating nucleotide pool is required for cell proliferation and survival.
a, Measurement of the nucleoside/nucleotide contents by LC-MS/MS in WT mice with or without shG6pdx via the AAV8 system. b, Examples of oscillating nucleosides/nucleotides from the metabolome of WT and Bmal1-null mouse livers. The processing data were included in Supplementary Table 3. Box plots extend from the 25th and 75th percentile, with a horizontal line representing the median value, and the whiskers indicate the min and max values. c, Measurement of the contents of nucleotides in the livers from WT and LBKO mice before and after PH by LC-MS/MS. d, Representative images of hepatic organoids cultured in Matrigel assessed by bright field microscopy, with quantification of the numbers of hep-organoids formed from 104 cells. Primary hepatocytes from LBKO mice were treated with or without a combination of NAC and NTPs during the 14-day organoid culture. n = 3 independent experiments. e, Immunoblotting showing the protein levels of Bmal1 and G6pdx in primary hepatocytes, which were isolated from Bmal1f/f mouse livers and then treated with AAV-Cre. f-h, Graphs showing representative images (f) of hepatic organoids cultured in Matrigel and quantification of the numbers of hep-organoids formed from 104 cells (g) and the mean diameter (h). Primary hepatocytes, as pretreated in e, were subjected to 3D culture with or without a combination of NAC and dNTPs (N + N) for 14 days. n = 3 independent experiments. Scale bar, 50 μm. For a,c, n = 3 or 4 mice per group. Data are mean ± s.e.m. P-values were calculated using multiple unpaired two-tailed Student’s t-test (a), one-way ANOVA with Dunnett’s test (c,g,h) and two-tailed unpaired t-test (d).
