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. 2021 Mar 12;12:1630. doi: 10.1038/s41467-021-21893-y

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© This is a U.S. government work and not under copyright protection in the U.S.; foreign copyright protection may apply 2021

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 — Mll3^‒/‒Mll4^f/f or Smarcb1^f/f brown preadipocytes were infected with adenoviruses expressing GFP or Cre. Sub-confluent cells were collected for ChIP-Seq and ATAC-Seq. BAF⁺ active enhancers (AEs) were defined as those bound by SMARCA4 and ARID1A. a, b Deletion of Mll4 reduces BAF binding on BAF⁺ AEs in preadipocytes. a AEs shown in the heat maps were ranked by the intensity of SMARCA4 at the center in control (GFP) cells. b Fold changes of intensities between control and Mll4 KO (Cre) cells on MLL4⁺ (n = 6304) and MLL4^- (n = 6484) AEs in preadipocytes are shown in box plots. Statistical significance levels are as follows (Wilcoxon signed rank test, one-sided): SMARCA4 (p = 1); ARID1A (p = 0); ATAC (p = 5.2E-38); H3K27ac (p = 1.6E-293). Exact p-values cannot be computed for SMARCA4 and ARID1A due to ties in the ranks. c, d SMARCB1 is required for maintaining MLL4 binding on BAF⁺ MLL4⁺ AEs in preadipocytes. c AEs shown in the heat maps were ranked by the intensity of SMARCA4 at the center in control cells. d Fold changes of intensities between control (GFP) and Smarcb1 KO (Cre) cells on AEs (n = 6,304) in preadipocytes are shown in box plots. Statistical significance levels are as follows (Wilcoxon signed rank test, one-sided): SMARCA4 (p = 1); ARID1A (p = 4.0E-59); ATAC (p = 1); MLL4 (p = 0); H3K4me1 (p = 0); H3K27ac (p = 1.1E-274). Exact p-values cannot be computed for SMARCA4, MLL4 and H3K4me1 due to ties in the ranks. e, f SMARCA4 is required for maintaining MLL4 binding on BAF⁺ MLL4⁺ AEs in preadipocytes. Smarca4^AID/AID preadipocytes were infected with retroviruses expressing Tir1 or vector (Vec) only, followed by auxin (0.5 mM) treatment for 4 h. Cells were then collected for ATAC-Seq and ChIP-Seq of MLL4. e AEs shown in the heat maps were ranked by the intensity of MLL4 at the center in control cells. f Fold changes of intensities between SMARCA4-depleted (Tir1) and control (Vec) cells on AEs (n = 6,304) in preadipocytes are shown in box plots. Statistical significance levels are as follows (Wilcoxon signed rank test, one-sided): ATAC (p = 0); MLL4 (p = 0). Exact p-values cannot be computed due to ties in the ranks. In b, d and f, box represents the first and third quartiles with the horizontal line showing the median, and whiskers indicating minimum and maximum in all box plots. Outliers were not included.

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