Fig. 7.

Identification of a critical regulatory element responsive to mutant K-ras. A: sequences between 3.36 and 3.17 kb in the Bmp4 promoter were divided into 5 fragments (probes 1–5) and EMSAs were performed. Nuclear extracts of SW480^sicontrol (c) and SW480^siKrasV12 (kd) were utilized. *Specific shift obtained in SW480^sicontrol cells by probe 3. B: 200-fold molar excess of unlabeled probe 3 was used as a specific competitor (sc) to confirm specificity. C: specific shift was also observed in SW480^sicontrol cells incubated with probe 3A, spanning the region 3,285–3258 bp of the Bmp4 promoter. D: treatment of SW480^sicontrol cells with 20 μM PD98059 disrupts the binding of the critical element to the Bmp4 promoter. E: 3 different probes were generated that contain mutations of the first 10 bp (mut1), second 10 bp (mut2), and third 10 bp (mut3) of probe 3A. Specific shift (*) was lost in SW480^sicontrol cells (c) incubated with probe mut3.