Structural and functional characterization of the transforming growth factor-beta -induced Smad3/c-Jun transcriptional cooperativity.
Smads are intracellular proteins that act as central effectors for transforming growth factor-beta (TGF-beta) and related proteins from the activated receptor into the nucleus, where they regulate ligand-induced gene expression. AP-1 binding sites have been functionally linked to the transcriptional activation of various genes in response to TGF-beta. Accordingly, we have previously shown that the heteromeric complex of Smad3 and Smad4 synergizes with c-Jun/c-Fos at the AP-1 binding site of the collagenase I promoter to induce transcriptional activation in response to TGF-beta. Using the collagenase I promoter as model system, we have now investigated the role of the c-Jun and Smad3 interactions with the promoter DNA and have further characterized the physical basis of the c-Jun/Smad3 interaction in the transcriptional response. Mutational analyses of the c-Jun protein and the AP-1 binding site in the promoter revealed that the interaction of c-Jun with DNA is necessary for transcriptional activation by TGF-beta and Smad3. Similar analyses of Smad3 and the Smad binding sites revealed that binding of Smad3 to DNA is also required, but that its DNA sequence-specific recognition is not essential. We also found that the basic leucine zipper domain of c-Jun and a short sequence close to the N terminus of Smad3 mediate their physical interaction, and that these regions are critical for their DNA-binding function. Our studies provide a basis for understanding the functional cooperativity of Smads with the diversity of transcription factors, which underlies the Smad-induced transcriptional activation in response to TGF-beta and related factors.
Qing
J
J
Departments of Growth and Development, and Anatomy, Programs in Cell Biology and Developmental Biology, University of California, San Francisco, California 94143-0640, USA.
Zhang
Y
Y
Derynck
R
R
D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002460:Cell Line; D017364:Collagenases / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D004847:Epithelial Cells; D005786:Gene Expression Regulation / Q000187:drug effects; D017930:Genes, Reporter; D016758:Genes, jun; D006801:Humans; D008156:Luciferases / Q000235:genetics; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D016755:Proto-Oncogene Proteins c-jun / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D051900:Smad3 Protein; D015534:Trans-Activators / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic / Q000187:drug effects / Q000502:physiology*; D014162:Transfection; D016212:Transforming Growth Factor beta / Q000494:pharmacology*
D004268:DNA-Binding Proteins; D016755:Proto-Oncogene Proteins c-jun; D011994:Recombinant Proteins; C497790:SMAD3 protein, human; D051900:Smad3 Protein; D015534:Trans-Activators; D016212:Transforming Growth Factor beta; D008156:Luciferases; D017364:Collagenases; C417249:collagenase 1