Figure 6

Chromatin-Immunoprecipitation (ChIP) indicates that Smad4 binds to all three promoters of LM-322. Chromatin prepared from Smad4-negative, Smad4-reexpressing and TGFβ-treated Smad4-reexpressing BxPC3 cells was shared to obtain fragments of 300–500 bp length, immunoprecipitated and used for PCR amplification of ~200 bp promoter fragments. Input chromatin after sharing is directly used for PCR amplification as a control. Binding of Smad4 to the LAMA3 promoter region which harbors the SBE could repeatedly be shown as well as to the regions which incorporate the functional AP1 sites in all three promoters. Attempts to show binding of Smad4 to other regions within the 4 kb LM-332 promoters failed.