Interaction of smad3 with a proximal smad-binding element of the human alpha2(I) procollagen gene promoter required for transcriptional activation by TGF-beta.

Transcription of the alpha2(I) collagen gene (COL1A2) in fibroblasts is potently induced by transforming growth factor-beta (TGF-beta). Smad family proteins function as intracellular signal transducers for TGF-beta that convey information from the cell membrane to the nucleus. In the present study, we establish the functional requirement for endogenous Smad3 and Smad4 in TGF-beta-stimulated COL1A2 transcription in human skin fibroblasts in vitro. Furthermore, using transfections with a series of 5' deletions of the human COL1A2 promoter, we identify a proximal region between -353 and -148 bp, which is required for full stimulation of transcription by a constitutively active TGF-beta type I receptor. This region of the COL1A2 promoter contains a CAGA motif also found in the promoter of the plasminogen activator inhibitor-1. Substitutions disrupting this sequence decreased the binding of nuclear extracts or recombinant Smad3 to the CAGACA oligonucleotide, and markedly reduced the transcriptional response to TGF-beta or overexpressed Smad3 in transient transfection assays. The insertion of tandem repeats of CAGACA conferred TGF-beta stimulation to a heterologous minimal promoter-reporter construct. Inhibition of endogenous Smad expression in fibroblasts by antisense oligonucleotides or cDNA against Smad3 or Smad4, and transfection of COL1A2 promoter constructs into Smad4-deficient breast adenocarcinoma cells, indicated the critical role of Smads for the full TGF-beta response. The importance of Smad binding to the CAGACA box of COL1A2 was further established by transcriptional decoy oligonucleotide competition. Taken together, the results identify a functional Smad-binding element of the COL1A2 promoter harboring a CAGACA consensus sequence that is both necessary and sufficient for stimulation by TGF-beta, and demonstrate that interaction of this Smad-binding element with endogenous Smads is required for the full TGF-beta response in fibroblasts.

Chen SJ ，Yuan W ，Lo S ，Trojanowska M ，Varga J ... -  《JOURNAL OF CELLULAR PHYSIOLOGY》

Smad-dependent stimulation of type I collagen gene expression in human skin fibroblasts by TGF-beta involves functional cooperation with p300/CBP transcriptional coactivators.

Ghosh AK ，Yuan W ，Mori Y ，Varga J ... -  《ONCOGENE》

Stimulation of type I collagen transcription in human skin fibroblasts by TGF-beta: involvement of Smad 3.

Chen SJ ，Yuan W ，Mori Y ，Levenson A ，Trojanowska M ，Varga J ... -  《JOURNAL OF INVESTIGATIVE DERMATOLOGY》

Tenascin-C upregulation by transforming growth factor-beta in human dermal fibroblasts involves Smad3, Sp1, and Ets1.

Jinnin M ，Ihn H ，Asano Y ，Yamane K ，Trojanowska M ，Tamaki K ... -  《ONCOGENE》

Constitutively phosphorylated Smad3 interacts with Sp1 and p300 in scleroderma fibroblasts.

Ihn H ，Yamane K ，Asano Y ，Jinnin M ，Tamaki K ... -  《RHEUMATOLOGY》