Fig 2: Quantification of the expression of selected HDAC proteins. HepG2 and Huh7 cells were treated for 24 h with either solvent (Ctrl), bb at IC50 (0.6 µM), bb at two-fold IC50 (1.2 µM) or vs. at two-fold IC50 (4.0 µM). Western blotting was used to measure protein expression, using tubulin as loading control. (A) HDAC1 expression in Hep2G and Huh7 cells. (B) HDAC2 expression in Hep2G and Huh7 cells. (C) HDAC4 expression in Hep2G and Huh7 cells. **P<0.01 vs. ctrl. (D) HDAC6 expression in Hep2G and Huh7 cells. Corresponding representative western blotting images are provided in Fig. S2. n=3 per group. One-way ANOVA with Tukey's post hoc test was used to test for significance. HDAC, histone deacetylase; Ctrl,. control; bb, broxbam; vs, vorinostat.

Fig 3: Assessment of gene knockout frequencies by knockdown of HDAC1, HDAC2 and HDAC3. (A) HDAC1, HDAC2 and HDAC3 mRNA and protein levels were determined by RT-qPCR and western blot after transfection with scrambled siRNAs or HDAC1, HDAC2 or HDAC3 siRNAs. (B) Changes of EGFP knockout mediated by CRISPR/Cas9 upon knockdown of HDAC1, HDAC2 or HDAC3. Data in bar graphs are represented as mean ± SD (n ≥ 3), two-tailed unpaired Student's t-test: *P-values < 0.05; **P-values < 0.01; ***P-values < 0.001.

Fig 4: METTL3 interacts with HDAC1/2 and regulates H3K9ac and H3K27ac levels in the promoters of Cd36 and Ccl2 genes.a, b HA-HDAC1/2 expression vector was co-transfected with or without Flag-METTL3 expression vector in HEK293T cells. Total cell lysates were first incubated with DNase1 (200 U/ml) at 37 °C for 30 min. These lysates were immunoprecipitated with Flag or HA beads and then immunoblotted with anti-HA or anti-Flag antibodies. The samples were derived from the same experiment and the blots were processed in parallel. c Liver extracts from control and Mettl3-HOE mice at 8 weeks old were first incubated with DNase1 (200 U/ml) at 37 °C for 30 min. These lysates were immunoprecipitated with Flag beads and then immunoblotted with anti-HDAC1 or anti-HDAC2 antibodies. The samples were derived from the same experiment and the blots were processed in parallel. d Mettl3-HOE mice were fed an NC or MCD for 2 weeks. Liver extracts were first incubated with DNase1 (200 U/ml) at 37 °C for 30 min. These lysates were immunoprecipitated with Flag beads and then immunoblotted with anti-HDAC1 or anti-HDAC2 antibodies. The HDAC1/2 protein levels associated with Flag-METTL3 were quantified by ImageJ and normalized to input HDAC1/2 (n = 3 for each group; For HDAC1, NC versus MCD, P = 0.0096; For HDAC2, NC VS MCD, P = 0.0462). The samples were derived from the same experiment and the blots were processed in parallel. e, f H3K9ac and H3K27ac levels in the promoters of Ccl2, Cd36 and Actb genes in Mettl3-HOE and control mice fed an MCD for 2 weeks (n = 6 for each group; For H3K9ac, Ccl2, P = 0.0334, Cd36, P = 0.03, Actb, P = 0.8045; For H3K27ac, Ccl2, P = 0.0401, Cd36, P = 0.02585, Actb, P = 0.6527). g, h Primary hepatocytes were infected with Ad-βGal and Ad-METTL3 adenovirus. Ad-METTL3-infected hepatocytes were treated with or without TSA (2 μM) overnight. The relative Cd36 and Ccl2 mRNA levels were measured by qPCR (For Cd36, n = 6 for each group, βGal versus METTL3, P = 0.0016, METTL3 versus METTL3+TSA, P = 0.0009; For Ccl2, n = 4 for each group, βGal versus METTL3, P = 0.0054, METTL3 versus METTL3+TSA, P = 0.0167). CD36 and CCL2 protein levels were measured by immunoblotting, quantified by ImageJ and normalized to Tubulin (n = 5 for each group; For CD36, βGal versus METTL3, P = 0.0004, METTL3 versus METTL3+TSA, P = 0.0112; For CCL2, βGal versus METTL3, P < 0.0001, METTL3 versus METTL3+TSA, P = 0.0167). The samples were derived from the same experiment and the blots were processed in parallel. i, j H3K9ac and H3K27ac levels in the promoters of Ccl2, Cd36 and Actb genes in the livers of Mettl3-HKO and Mettl3flox/flox mice at 8 weeks old (n = 7 for each group; for H3K9ac, Ccl2, P = 0.0037, Cd36, P = 0.0113, Actb, P = 0.4987; For H3K27ac, Ccl2, P = 0.0097, Cd36, P = 0.0126, Actb, P = 0.4736). n was the number of biologically independent mice or cell samples. The cell culture experiments were repeated three times independently with similar results. Data represent the mean ± SEM. Significance was determined by unpaired two-tailed Student’s t test analysis. *P < 0.05. **P < 0.01. Source data are provided as a Source Data file.

Fig 5: Experimental procedures. CUMS, chronic unpredicted mild stress; BDNF, brain-derived neurotrophic factor; acH3K9, acetylation levels in histone H3 lysine 9; HDAC2, histone deacetylase 2; RT-polymerase chain reaction (PCR), reverse transcription-polymerase chain reaction; MS-qPCR, methylation-specific quantitative polymerase chain reaction; BW, body weight; SPT, sucrose preference test; OFT, open field test.