Figure 2.

EGF activates multiple signaling pathways in A172 cells. A) A172 cells were stimulated with EGF for various times, and activation of the indicated signaling molecules was analyzed by Western blotting using specific antibodies. Total ERK is included as a loading control. B) Nuclear extracts were prepared from control and EGF-treated A172 cells as indicated. DNA binding activity of AP-1, STAT, and NF-κB was then analyzed by EMSA using the ³²P-labeled oligonucleotide probes. C–D) A172 cells were transiently transfected with the indicated reporter plasmids and a β-galactosidase expression vector. One day after transfection, cells were stimulated with EGF, cultured for an additional 24 h, and harvested. CAT activities were normalized to β-galactosidase activities to account for transfection efficiency. Results are expressed as fold induction.