Figure 3.

Characterization of c‐Myc as a transcription factor regulating survivin expression. (A) Progressive deletion mutation analysis of survivin promoter activity and its response to imatinib. K562 cells were transfected with pGL3‐S1764 and its progressive deletion mutants and cultured for 8 hrs, after which cells were incubated for 16 hrs in the absence or presence of 2.5 uM imatinib. The promoter activity was measured by normalizing firefly to renilla luciferase activity. Mean values ± S.D. from three independent experiments in triplicate are shown.*P≤ 0.001,#no significance. (B) A schematic diagram of the proximal survivin promoter indicating potential transcription factor binding sites. (C) Site‐directed mutation analysis of survivin promoter activity and its response to imatinib. Constructs of Sp1, STAT/TCF and c‐Myc binding sites mutation described in the illustration above were generated and transfected into K562 cells. The promoter activity was measured by normalizing firefly to renilla luciferase activity. Mean values ± S.D. from three independent experiments in triplicate are shown. *P < 0.01, #no significance. (D) ChIP assay analysed in vivo binding activity between c‐Myc and survivin promoter. Treated K562 cells were subjected to the ChIP protocol. The immunoprecipitations were performed with antibody against c‐Myc, and an isotype IgG as negative control. A parallel analysis were performed using an antibody against RNA‐polymerase as systematic control.