Fig. 2

BRD7 formed a negative feedback loop with c-Myc in miR-141 transcription. a IF detected the subcellular distribution pattern of BRD7 and c-Myc protein, as well as the colocalization of BRD7 with endogenous c-Myc in 5-8F and HNE1 cells transfected with pEGFP-C2/BRD7. c-Myc: endogenous c-Myc protein, BRD7: EGFP-tagged BRD7 protein. b Co-IP assays and western blotting confirmed the interaction between BRD7 (GFP-tagged) and c-Myc (Flag-tagged) in HEK293 cells. c The dual-luciferase reporter assays assessed the miR-141 activity promoter when overexpressing BRD7 in c-Myc-overexpressing 5-8F and HEK293 cells (c-Myc) and control cells (Vector). d qRT-PCR assays evaluated the expression of miR-141 in c-Myc-overexpressing 5-8F and HEK293 cells when overexpressing BRD7. e A schematic diagram showing the locations of BRD7-binding peaks in chromosome 12 identified by ChIP-sequencing. f Western blotting and g qRT-PCR assays evaluated the expression of BRD7 protein and mRNA, respectively, in c-Myc knockdown 5-8F cells (shc-Myc) and control cells (shCtrl). h The dual-luciferase reporter assays determined the BRD7 promoter activity in c-Myc knockdown 5-8F cells and control cells. i Western blotting and j qRT-PCR assays analyzed the expression of BRD7 protein and mRNA in c-Myc-overexpressing HEK293 cells and controls. k The dual-luciferase reporter assays determined the BRD7 promoter activity in c-Myc-overexpressing HEK293 cells. l ChIP-PCR assays using antibodies specific for Flag tags validated the c-Myc-binding sites in the BRD7 promoter. miR-141 was used as a positive control in this experiment. b, f and i β-actin served as an internal control. D), G) and J) U6 or GAPDH served as an internal control. c, d, g, h, j and k The error bars represent the mean ± S.E.M. ***P < 0.001