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. 2018 Oct 15;8:15226. doi: 10.1038/s41598-018-33149-9

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© The Author(s) 2018

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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H₂S-Deficient mice negatively regulate global gene expression and histone acetylation. (a) Western blot analysis of the indicated proteins (HDAC3, H3K27ac and H3) derived from femur bone extract of CBS^+/− mice. (b) Global Acetyl Histone H3K27 Quantification Kit confirmed that overall histone acetylation (H3K27) was indeed increased in femur bone extracts of CBS^+/− mice. (c) Histone Deacetylase (HDAC) Activity was decreased in CBS^+/− mice as assessed by HDAC activity fluorometric assay kit. (d) Histone deacetylase (HDAC) activity of BMMSCs derived from WT mice, were transfected with CBS CRISPR/Cas9 Knockout (KO) plasmid. (e) Differential gene expression of representative inflammatory genes in femur bone extracts of CBS^+/− mice. (f) Diagrammatic representation of H3K27ac binding to target genes of inflammatory cytokines. (g,h) Chromatin immune precipitation (ChIP) assay was performed assessing H3K27ac binding into the promoter of IL-6 and TNF-α. The results are expressed as the percentage of input. (i) Differential gene expression of representative osteogenic genes in femur bone extracts of CBS^+/− mice. (j,k) ChIP assay was performed assessing H3K27ac binding into the promoter of OCN and RunX2. The results are expressed as the percentage of input. Experiments were repeated three times. Data are expressed as mean ± SEM. n = 5 mice per group. *p < 0.05 compared with the WT mice, ^#p < 0.05 compared with the CBS^+/− mice.