Extended Data Fig. 3: NADH inhibition of SIRT1 activity.

a, Lineweaver-Burk transformation of data from SAMDI-MS. b, Relative NADH quantified by HPLC following supplementation with pyruvate or lactate compared to controls (n = 3). c, Densitometric quantification of western blots from liver of 4–6 mo old male mice in indicated genotypes at ZT4 for p53-(K379)Ac (left) and FOXO1-(K242, K245, K262)Ac (right) relative to control TRF-Reg (from Fig. 3d) (n = 3). d, FOXO1 ChIP-seq from liver collected during the daytime comparing effect of FOXO1 binding in TRF-CR (x-axis) with the effect of hepatic SIRT1 ablation (L-Sirt1^−/−) (y-axis). Each point indicates a FOXO1 peak in control liver. Peaks that have absolute log₂(fold change) >0.5 for both comparisons are colored blue or black and counted by quadrant (n = 2). e, Densitometric quantification of western blots as in (c) relative to null-transduced TRF-Reg mice (from Fig. 3e) (Ac-p53: n = 4 for TRF-CR, LbNOX. n = 6 for all other conditions; Ac-FOXO1: n = 2 for TRF-CR, LbNOX. n = 3 for all other conditions). f, FOXO1 ChIP-seq from liver during the daytime comparing effect of TRF-CR (x-axis) (n = 3) with effect of (f) LbNOX in TRF-CR mice (y-axis) (n = 2) with coloring and counting as in panel d. (n = 2–3). g, Western blotting for Ac-FOXO1 during the nighttime (ZT16) in null- and LbNOX-transduced mice on TRF-Reg and TRF-CR (n.s. non-specific) and densitometric quantification (n = 3). h, BMAL1 ChIP-seq from liver during the daytime comparing the effect of TRF-CR and L-Sirt1^−/− as in panel d (n = 3). Uncropped Western blot scans labelled with molecular weight markers are presented in the Source Data Files. i, RNA-seq reads mapping to the exon of Sirt1 that is flanked by LoxP sites (Ex4) relative to exon 9 (Ex9) of Sirt1, and RNA-seq reads per 10 million sequenced reads (RPM) that align to iCre and LbNOX in null- or LbNOX-transduced TRF-CR mice co-transduced with iCre (n = 6). Null-expressing mice on TRF-CR (blue) (n = 5) are shown as reference. j, Quadrant plot comparing transcriptional responses to TRF-CR in null-transduced mice (x-axis) (n = 6) and LbNOX-expression in TRF-CR, L-Sirt1^−/− mice (y-axis) (n = 6). Each point indicates a gene that is DE by TRF-CR in null-transduced mice (930 genes). Percentages of genes within each quadrant are shown. k, Average 48-hour fasting body temperature in 4–6 mo old female liver-specific Sirt1^−/− mice (n = 5). Data are presented as mean values ± SEM. Statistics were performed with unpaired, two-tailed student’s t test except as otherwise noted in the figure. *p < 0.05.

Source data