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. 2018 Jan 25;9:366. doi: 10.1038/s41467-017-02740-5

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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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Fig. 6 — Lsd1 represses expression of the pro-adipogenic factor Glis1. a ChIP-qPCR analyses using antibodies directed against Lsd1, REST corepressor 1 (Rcor1), histone deacetylase 1 (Hdac1), dimethylated histone 3 lysine 4 (H3K4me2), or rabbit immunoglobulin G (rIgG) in control (Ctrl) and LSD1(OE) C2C12 cells (left panel), or Lsd1 inhibitor-treated [Lsd1(i)] C2C12 cells (right panel) differentiated for 1 day in adipogenic medium. Immunoprecipitated chromatin was quantified by qPCR using primers flanking Lsd1-binding sites at the promoter of Glis1 gene. b qRT-PCR analyses showing relative transcript levels of Glis1 in Ctrl, LSD1(OE), Lsd1 knockdown [Lsd1(KD)], and Lsd1(i) C2C12 cells (left panel), or Ctrl, LSD1-GFP, Lsd1^iKO, and Lsd1(i) satellite cells (right panel) differentiated for 1 day in adipogenic medium. Significance was calculated by two-tailed Student’s t-test. c Western blot analysis showing protein levels of Glis1 in Ctrl, Lsd1^iKO, and Lsd1(i) satellite cells differentiated for 1 day in adipogenic medium. d–f Analyses of Ctrl and Lsd1^iKO EDL fiber culture cells transfected with control (siCtrl) or siRNA directed against Glis1 (siGlis1) and differentiated for 7 days in adipogenic medium. d qRT-PCR analyses showing relative transcript levels of Glis1 and Lsd1, e ORO staining, and f qRT-PCR analyses showing relative transcript levels of indicated genes. (a, d, f: n = 3, b: n = 6; mean + SEM, *p < 0.05, **p < 0.01, ***p < 0.001; scale bars: e: 50 µm)