c-Myc inhibits promoter activity and endogenous mRNA level of BRD7 gene. (A) Luciferase assay of 0.5μg pGL3/-266,-212 in COS7 cells cotransfected with various indicated amounts of pCMV-HA/c-Myc. The SV40 β-galactosidase vector was used for normalizing transfection efficiency. Data are the means ± S.D. of three independent experiments. (B) Top: 0.5 μg modified reporter construct pGL3/-266,-212/GFP was cotransfected with 2.5 μg pCMV-HA/c-Myc or 2.5 μg pCMV-HA into COS7 cells by Lipofectamine 2000 Reagent. 38 h after transfection, the signal of EGFP fluorescence driven by promoter fragment -266/-212 was observed by using an AX-80 analytical microscope system (Olympus, Tokyo, Japan). A1, B1: COS7 cells cotransfected with pGL3/-404,+46/GFP and pCMV-HA; A2, B2: COS7 cells cotransfected with pGL3/-266,-212/GFP and pCMV-HA-c-Myc. Bottom: After examining the direct GFP fluorescence, the analyzed COS7 cells were subjected to immunoblot assay with anti-GFP as the primary antibody to detect the EGFP expression driven by promoter fragment -266/-212 with or without overexpression of pCMV-HA-c-Myc. pCMV-HA was used as control. (C) and (D) Examination of BRD7 expression in COS7 cells transfected with pCMV-HA/c-Myc by RT-PCR and Real-time PCR, respectively. pCMV-HA was used as a negative control.