Ab133664

The protein concentration was measured by Bradford assay (BioRad). Approximately 20–50 μg of protein was separated on SDS-PAGE and subsequently transferred to a nitrocellulose membrane (GE Healthcare). The membranes were blocked with 5% BSA in TBS-T for 1 h at room temperature and then incubated with respective primary antibodies at 4 °C overnight. The primary antibodies used were: NAA40 (1:1000, ab106408, Abcam), GCN5 (1:1000, sc-365321, Santa-Cruz), ACSS2 (1:1000, ab133664, Abcam), phospho-Akt (Ser473) (1:1000, cat. 9271, Cell signalling), Akt (pan) (1:000, cat. C67E7, Cell Signaling) and β-actin (1:1000; sc-1616-R, Santa-Cruz). The membranes were incubated with the secondary antibody, Horseradish peroxide (HRP)-conjugated goat anti-rabbit IgG (1:30,000, Scientific) or Horseradish peroxide (HRP)-conjugated goat anti-mouse IgG (1:1000, p0447, Dako Denmark). Bands were visualised by the enhanced chemiluminescence system (BioRad) and analysed using the densitometry tool in ImageJ analysis software (NJH).

Human HCC sections were de-paraffinized in xylene and hydrated in graded ethanol, followed by deionized water. Endogenous peroxidase activity was inactivated with 3% hydrogen peroxidase in methanol for 30 min. Antigen retrieval was performed by boiling the sections in 0.01 M citrate buffer (pH 6.0) for 25 min in microwave oven. The sections were incubated with normal goat serum for 20 min to block non-specific staining and then incubated overnight at 4 °C with primary antibodies. After incubation for 45 min with a HRP-conjugated anti-rabbit secondary antibody, the sections were detected using diaminobenzidine (DAB) kit according to the manufacturer's instructions, followed by counterstaining with hematoxylin. The primary antibodies were used as follows: anti-H3K9 acetyl (ab32129, Abcam, 1:150 dilution), anti-H3K27 acetyl (ab45173, Abcam, 1:80 dilution), anti-H3K56 acetyl (ab76307, Abcam, 1:60 dilution), anti-ACSS2 (ab133664, Abcam, 1:50 dilution), anti-ACSS1 antibody (HPA041014, Sigma, 1:50 dilution), and anti-FASN (C-20140, Santa Cruz, 1:40 dilution). The negative controls were performed by omitting the primary antibodies.

Anti-histone H3 (9715S, CST), anti-H3K9 acetyl (1328-1, Epitomics), anti-H3K27 acetyl (EP865Y, Epitomics), anti-H3K56 acetyl (EPR996Y, Epitomics), anti-ACSS2 (ab133664, Abcam), anti-ACSS1 antibody (17138-1-AP, Proteintech), anti-FASN (C-20140, Santa Cruz), anti-ACLY antibody (1699-1, Epitomics) and anti-β-actin (a00702, Genescript) antibodies were commercially obtained. ChIP grade antibodies for H3K9 acetyl (ab4441, Abcam), H3K27 acetyl (ab4729, Abcam), H3K56 acetyl (17-10259, Milipore) were commercially obtained.
[U-¹³C₂]-Acetate (CLM-440-1, Cambridge Isotope Laboratories), lipoprotein depleted fetal bovine serum (LPDS) (880100, Kalen Biomedical), sodium palmitate (P9767, Sigma), BSA with low fatty acid (0219989925, MP) and TSA (9950, CST) were commercially obtained. Myristic acid (D27, 98%) as internal standard was obtained from Cambridge Isotope Laboratories (Tewksbury, MA, USA). Certified ACS grade sodium hydroxide was purchased from Standard Chemical and hydrochloric acid was from VWR International LLC (Pennsylvania, USA). Formic acid and BSTFA (with 1% TMCS) were purchased from Sigma-Aldrich. Methanol and chloroform were obtained from Tedia (Fairfield, USA) and hexane obtained from Duksan Pure Chemicals Co., Ltd (South Korea).