Fig. 4: A NAD⁺ chronotherapy at ZT11 corrects abnormal gene and protein expression from crucial molecular effectors of liver disease and triggers a specific transcriptional signature.

a, b Overlap (a) and heatmap (b) of DE genes when comparing CD-HF and HF-HFN groups either at day (ZT6) or ant night (ZT18) (n = 3). c, d Functional annotation of CD-HF and HF-HFN shared genes at daytime (c) or nighttime (d). e Homer de novo motif discovery analyses from promoters of genes DE exclusively in the HF group. f, g Circadian protein expression of AKT, p-AKT(S473), AMPK, and p-AMPK(T172) (F) or the mTOR pathway (g) in whole cell extracts from CD, HF, and HFN livers determined by western blot. Tubulin or p84 were used as a loading control. Images represent 3–4 independent experiments. Uncropped blots in Source Data. h, i Overlap (h) and heatmaps (i) of DE genes when comparing CD-HFN and HF-HFN groups either at day (ZT6) or at night (ZT18) (n = 3). j Functional annotation of shared genes DE in analyses CD-HFN and HF-HFN at nighttime. k Homer de novo motif discovery analyses from promoters of genes whose expression is altered exclusively in the HFN group. Adj. P value corresponds to the FDR q-value. This is the false discovery rate analog of hypergeometric p value after correction for multiple hypothesis testing according to Benjamini and Hochberg. CD control diet fed mice, HF high-fat diet fed mice, HFN High-fat diet fed, NAD⁺ treated mice at ZT11. See also Supplementary Figs. S3, S4 and Supplementary Data 4.