Chow KD CR HFD. Fed Fast Refed
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1 Supplementry Figure 1 Control d/d Chow KD CR Fed Fst Refed Supplementry Figure 1: Liver expression in diet nd disese models. () expression in the livers of ontrol nd d/d mie. () expression in the livers of mie fed how, ketogeni diet (KD), on lori restrition (CR) nd high ft diet (). () expression in the livers of d liitum fed, fsted (24 h) nd refed (24h) C57BL6/J mie. Nture Mediine: doi:1.138/nm.3882
2 Supplementry Figure 2 BXH19/TyJ SM/J CBA/J C3H/HeJ LG/J AKR/J AXB19/PgnJ BXA7/PgnJ SEA/GnJ AXB15/PgnJ BXA16/PgnJ BXD19/TyJ BXH2/KJ DBA/2J BXD14/TyJ FVB/NJ AXB19/PgnJ BXH4/TyJ A/J NZW/LJ NOD/ShiLtJ NZB/BlNJ NON/ShiLtJ CE/J MA/MyJ BXD36/TyJ BXD6/TyJ BXA8/PgnJ AXB19/PgnJ BXD39/TyJ BXA12/PgnJ BXD32/TyJ BXD24/TyJ BXD16/TyJ AXB2/PgnJ BXD24/TyJ-Cep29/J BXH22/KJ AXB1/PgnJ BXD9/TyJ BXA24/PgnJ I/LnJ 129X1/SvJ BXA13/PgnJ BXD21/TyJ RIIIS/J BXH1/TyJ SWR/J BXH7/TyJ BXA1/PgnJ BXD42/TyJ BTBR T+ tf/j BXA2/PgnJ BXD12/TyJ BXH9/TyJ AXB8/PgnJ BALB/J PL/J BXD8/TyJ BXD2/TyJ BXD29-Tlr4/J BXD38/TyJ C57BL/6J BXD4/TyJ BUB/BnJ LP/J C57L/J AXB13/PgnJ AXB12/PgnJ BXD18/TyJ BXH8/TyJ AXB23/PgnJ CAST/EiJ BXH14/TyJ AXB6/PgnJ BXA14/PgnJ BXD13/TyJ BXD15/TyJ BXD31/TyJ BXH6/TyJ BXD1/TyJ BXA11/PgnJ AXB5/PgnJ AXB4/PgnJ BXD2/TyJ B6C3-1/KJ AXB1/PgnJ BXD5/TyJ BXA26/PgnJ BXH2/TyJ AXB24/PgnJ BXA25/PgnJ BXA4/PgnJ BXD28/TyJ BXD11/TyJ C58/J BXD33/TyJ BXD34/TyJ KK/HlJ BXD27/TyJ Liver expression (Log 2) Supplementry Figure 2: Liver expression vries widely mong mouse inred strins. Liver expression (log2) ross HMDP strins otined from the System Genetis Resoure ( Nture Mediine: doi:1.138/nm.3882
3 Supplementry Tle 1: Correltion mong liver expression nd seleted metoli prmeters. Corr. Coef. Phenotype Nme (iweight midorreltion) P-vlue HDL TC FFA E-5 TG Ft_mss BW Len_mss Gene Symol: Proeset ID: t Cohort: HMDP Tissue: Liver Nture Mediine: doi:1.138/nm.3882
4 Supplementry Tle 2: Anthropometri nd linil hrteristis. n = 51 Men ± Stndrd devition Age (yers) ± 8.6 BMI (kg/m 2 ) 45 ± 6.5 Fsting gluose (mg/dl) 11.4 ± 14.4 Totl Cholesterol (mg/dl) ± 34.1 HDL Cholesterol (mg/dl) 48.2 ± 11.1 LDL Cholesterol (mg/dl) ± Fsting triglyerides (mg/dl) ± Gluose infusion rte (mg/kg/min) 3.73 ± 2.3 Cortisol (µg/dl) ± 4.9 Nture Mediine: doi:1.138/nm.3882
5 Supplementry Figure 3 Primry heptoyte SAM/SAH Liver SAM/SAH Primry heptoyte NAD + (pmoles/μg of protein) Liver NAD + (pmoles/mg of wet weight) Primry heptoyte NAM (pmol/μg protein) Liver NAM (pmol/mg of wet weight) d Primry heptoyte (pmol/μg protein) SH.5 AdGFP Ad Primry heptoytes (pmol/μg protein) Liver (pmol/mg wet weight) Supplementry Figure 3: Effet of expression on intrellulr SAM/SAH, NAD, NAM nd. () Effet of knokdown on SAM/SAH rtio in primry heptoytes (left) nd liver (right). () Effet of knokdown on NAD+ ontent in primry heptoytes (left) nd liver (right). () Effet of knokdown on NAM ontent in primry heptoytes (left) nd liver (right). (d) Intrellulr ontent in primry heptoytes overexpressing (left), in primry heptoytes with knokdown (middle), nd livers with knokdown (right). Sttistil signifine ws tested y unpired Student s t-test, P <.5). Nture Mediine: doi:1.138/nm.3882
6 Supplementry Figure 4 MG132 AdGFP Ad AdGFP Ad Sirt1 Reltive expression MG132 + AdGFP Ad IP: HA IB: Flg HA-Sirt1 + Flg-U Protesome tivity (%) Ctrl IP: HA IB: Sirt1 MG132 + Supplementry Figure 4: regultes Sirt1 uiquitintion. () Effet of the protesome inhiitor MG132 (1 μm) on Sirt1 protein expression in primry heptoytes overexpressing (triplite determintion). () Effet of overexpression on Sirt1 uiquitintion (representtive of two independent experiments). () Effet of overexpression on generl protesome tivity. 5 μm MG132 ws used s positive ontrol (triplite determintion). Sttistil signifine ws tested y unpired Student s t-test, P <.5. Nture Mediine: doi:1.138/nm.3882
7 Supplementry Figure 5 S213 Y24 GFP Flg-wt Flg-Y2W Flg-A198W NAM Y2 F F A198 L164 GFP: + Flg: wt Y2W A198W IP:Flg IB:Flg d Sirt1: Flg: wt wt Y2W A198W GFP: + Sirt1 Flg e IP: HA WB: Flg IP:HA WB: HA HA-Sirt1 + Flg-U (mm) f Protesome tivity (%) (μm) MG132 (μm) 5 g Sirt1 tivity (%) (μm) Supplementry Figure 5: tivity is required for inresed Sirt1 protein expression. () Shown re seleted mino ids surrounding the modeled niotinmide in the rystl struture of. Muttions of Y2 nd A198 into tryptophn re overlid on the originl mino ids to show the position of the side hins with respet to NAM. () HPLC hromtogrms of immunopreipittes overexpressing GFP, -wt, -Y2W or -A198W nd treted with 1 mm NAM nd 1 μm SAM. () Immunolotting of wt nd mutnt proteins from the previous experiment. (d) Immunolotting of Sirt1 protein in HEK293T ells trnsfeted with GFP, -wt, -Y2W or -A198W expressing vetors (representtive of 2 independent experiments). (e) Effet of on Sirt1 uiquitintion. (f) Effet of ( 3 mm) on generl protesome tivity (5 μm of MG132 ws used s positive ontrol) (triplite determintions). (g) Effet of ( 1 mm) on Sirt1 tivity in vitro. Nture Mediine: doi:1.138/nm.3882
8 Supplementry Figure 6 e Reltive mrna expression Reltive mrna expression (pmoles/mg wet weight) Sref1 Fs Sd1 1%.3% 1% Body weight (g) Reltive mrna expression Sref2 Hmgs1 Hmgr Ldlr Nr1h3 Sr1 A1 Cyp71 Ag5 Ag8 Nr2 A11 d % 1% Time (dys).3% 1%.3% 1% f Reltive mrna expression Tnf Il6 Il1 Vm1.3% 1% Supplementry Figure 6: Effet of supplementtion of on weight gin nd liver gene expression. () Liver ontent of mie fed nd 1%. () Body weight gin of mie fed (n = 7), (n = 8),.3% (n = 8) nd 1% (n = 8). () Expression of genes in ftty id synthesis in the livers of mie fed (n = 5), (n = 5),.3% (n = 5) nd 1% (n = 5). (d-f) Expression of genes in holesterol metolism (d), holesterol lerne (e) nd inflmmtion genes (f) in the livers of mie fed (n = 7), (n = 8),.3% (n = 8) nd 1% (n = 8). Dt re presented s men ± s.e.m. Sttistil signifine ws evluted y unpired Student s t-test (, ) nd y one-wy ANOVA followed y postho Dunnett s with ontrol set to (d-f), P <.5. Nture Mediine: doi:1.138/nm.3882
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