Ab176316

The cells were quickly harvested and homogenized on ice in lysis buffer for Western and IP (Beyotime, P0013) plus 1 mM PMSF (Beyotime, ST506) and 1% phosphatase inhibitor (Boster, AR1183). After protein A + G magnetic beads were washed, antibodies against PCAF (Abcam, ab176316), p65 (Abcam, ab218533), HA (Cell Signaling Technology, 3724 S) and Flag (Cell Signaling Technology, 14,793 S) were incubated with the beads. Then, IgG from the corresponding species was used to eliminate nonspecific binding. Thereafter, protein A + G magnetic beads conjugated with antibody or normal IgG were added to the cell extracts and incubated according to the manufacturer’s instructions. Then, the magnetic beads were collected, washed, and resuspended in 60 µl of SDS loading buffer. All samples were boiled for 5 min and analyzed via western blotting.

Samples and cells were collected for Western blotting as previously described. Western blot analysis was performed using the following antibodies: LHPP (NBP1-83273, Novus, 0.2 ug/ml), METTL14 (ab220030, Abcam, 1:1000), HIF-1α (ab243861, Abcam, 1:1000), β-ACTIN (ab8226, Abcam, 1:2000), Acetyl Lysine (ab190479, Abcam, 1:1000), P300 (ab10485, Abcam, 1:5000), TIP60 (ab151432, Abcam, 1:1000), GCN5 (ab282176, Abcam, 1:1000), PCAF (ab176316, Abcam, 1:1000), GLUT1 (ab115730, Abcam, 1:5000), c-Myc (ab32072, Abcam, 1:1000), PKM2 (ab137852, Abcam, 1:1000), ALDOLASE (ab252953, Abcam, 1:1000), ENOLASE1 (H00002023-M01, Novus, 1:500), GLS1 (H00002744-M01, Novus, 1:500), GSK-3β (phospho S9) (ab75814, Abcam, 1:5000), GSK-3β (ab32391, Abcam, 1:5000), β-CATENIN (phospho S37) (ab75777, Abcam, 1:500).