Anti h3k27ac d5e4

The following primary antibodies were used for western blots: anti-TUBULIN, T6199 mouse (1/10,000, Sigma Aldrich, St. Louis, MO, USA); anti-Beta-ACTIN, 13854 (1/20,000 Sigma Aldrich); anti-SMARCA4 49360S (1:1000, Cell Signaling Technology); anti-EZH2 5246S (1:1000, Cell Signaling); anti-H3K27ac D5E4 (1:1000, Cell Signaling) anti-H3K27me3 07-449 (1:1000, Cell Signaling); anti-UTX (KDM6A) D3Q1I (1:1000, Cell Signaling); anti-KDM6B (JMJD3) #3457 (1:1000, Cell Signaling) for western blots or anti-KDM6B (JMJD3) ab38113 (Abcam) for immunostaining (see also Supplementary Table 1). For western blots, whole-cell lysates were collected in a buffer containing 2% SDS 50 mM Tris–HCl (pH 7.4), 10% glycerol and protease inhibitor cocktail (Roche Applied Science). Protein concentrations were determined using a Bio-Rad DC Protein Assay kit (Life Science Research). Equal amounts of lysates (20 µg) were separated by SDS–PAGE and transferred to a nitrocellulose membrane that was blocked with 5% nonfat dry milk. Membranes were incubated with the primary antibody overnight at 4 °C, then washed before incubation with species-appropriate IRDye 680CW (925-68022) or IRDye 800CW (925-32213) fluorescent secondary antibodies (1:10,000 LI-COR, NE, USA) for 1 h at room temperature. Membrane imaging was performed on the Odyssey Li-Cor CLx i software: Image Studio Lite v5.2.