Transforming growth factor beta-inducible independent binding of SMAD to the Smad7 promoter

SMAD proteins can mediate transforming growth factor beta (TGF-beta)-induci ble transcriptional responses. Whereas SMAD can recognize specific DNA sequ ences, it is usually recruited to a promoter through interaction with a DNA -binding partner. In an effort to search for TGF-beta-inducible genes, we u sed a subtractive screening method and identified human Smad7. which can an tagonize TGF-beta signaling and is rapidly up-regulated by TCF-beta. In thi s report, we show that TGF-beta can stabilize Smad7 mRNA and activate Smad7 transcription. The Smad7 promoter is the first TGF-beta responsive promote r identified in vertebrates that contains the 8-bp palindromic SMAD-binding element (SBE), an optimal binding site previously identified by a PCR-base d selection from random oligonucleotides by using recombinant Smad3 and sma d4. We demonstrate that on TGF-beta treatment, endogenous SMAD complex can bind to a Smad7 promoter DNA as short as 14 or 16 bp that contains the 8-bp see in gel mobility shift and supershift assays. Our studies provide stron g evidence that SMAD proteins can bind to a natural TGF-beta responsive pro moter independent of other sequence-specific transcription factors. We furt her show that, whereas recombinant Smad3 binds to the SEE. endogenous or ev en transfected Smad3 cannot bind to the SEE in the absence of Smad4. These findings have important implications in the identification of target genes of the TGF-beta/SMAD signaling pathways.