Fig. 5: SIRT6 controls H3K56ac deposition and enhancer activity of the Utrn gene. | Nature Communications

Fig. 5: SIRT6 controls H3K56ac deposition and enhancer activity of the Utrn gene.

From: Inactivation of Sirt6 ameliorates muscular dystrophy in mdx mice by releasing suppression of utrophin expression

Fig. 5

a Venn diagram of H3K56ac, H3K27ac and H3K4me3 ChIP-seq data in wild type fiMuSCs. b Boxplot of upregulated H3K56ac peaks in fiMuSCs of Sirt6mKO mice. Y-axis: log2-fold changes (Log2FC), GW: all Genome-Wide peaks, DE: significantly deregulated peaks (n = 2). c, d Distribution of H3K56ac ChIP-seq peaks across genomic regions (c) and relative to TSS (d). e Boxplot demonstrating that the increase of H3K56ac peaks in Sirt6mKO is higher across enhancer-associated H3K56ac peaks (positive for H3K27ac but negative for H3K4me3) compared to promoter H3K56ac peaks (positive for H3K4me3) (n = 2). f Venn diagram of overlapping upregulated ATAC-seq and H3K56ac ChIP-seq peaks Sirt6-deficient fiMuSCs. g RT-qPCR analysis of Utrn expression in muscles from control (n = 8), mdx (n = 9) and Sirt6mKO/mdx (n = 9) mice. Reference gene: m36b4. (One-way ANOVA with Benjamini multiple comparisons test: *p = 0.0304, **p = 0.003 ****p < 0.0001). h Immunofluorescence staining for UTRN on TA sections from control, mdx and Sirt6mKO/mdx mice. Scale bar: 20 µm. n = 3 per group. i Western blot analysis of utrophin expression in muscles from control, mdx and Sirt6mKO/mdx mice. Loading control: α-tubulin. Quantification is on the right. (One-way ANOVA with Bonferroni multiple comparisons test: **p = 0.070, ***p = 0.0008, ****p < 0.0001; n = 3 per group). j H3K56ac distribution in the proximal promoter region and DUE of the Utrn gene showing differential H3K56ac peaks in control and Sirt6mKO MuSCs. k ChIP-qPCR analysis of Sirt6 (IgG: n = 7; SIRT6: n = 7), p300 (IgG: n = 7; p300: n = 7) and cJun (IgG: n = 5; cJun: n = 4) at the DUE of the Utrn gene in C2C12 myotubes. (**p = 0.0074, **p = 0.0054, **p = 0.0055). l ChIP-qPCR analysis of SIRT6 at the Utrn DUE in HA-SIRT6 overexpressing MuSCs derived myotubes (**p = 0.0026, n = 7 per condition). m ChIP-qPCR analysis of H3K56ac and H3K27ac at the DUE of the Utrn gene in mdx (H3K56ac: n = 12; H3K27ac: n = 9) and Sirt6mKO/mdx (H3K56ac: n = 9; H3K27ac: n = 5) muscles (***p = 0.005, *p = 0.0126). Data are presented as mean ± SEM (g, i, k–m). Box plots in b, e were defined by default in R, 1.5 × IQR (interquartile range). The whiskers show minimum and maximum values. Wilcoxon rank sum test, two-sided p < 2.2 × 1016. For k–m unpaired two-tailed t-test. Twelve to 16 weeks old male mice were used. Source data are provided in the Source Data file.

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