Synergistic cooperation between Sp1 and Smad3/Smad4 mediates transforming growth factor β1 stimulation of α2(I)-collagen (COL1A2) transcription

Wen Zhang, Jiongwen Ou, Yutaka Inagaki, Patricia Greenwel, Francesco Ramirez

Research output: Contribution to journalArticlepeer-review

215 Scopus citations

Abstract

Transforming growth factor-β1 (TGFβ) is a strong activator of extracellular matrix accumulation. TGFβ stimulates the gene coding for human α2(I)-collagen (COL1A2) by inducing binding of an Sp1-containing complex to an upstream promoter element (TGFβ responsive element or TbRE) that contains a CAGA box. Here we report that the CAGA box of the TbRE is the binding site of the Smad3/Smad4 complex, and that the binding of the complex is required for TGFβ-induced COL1A2 up-regulation. Recombinant Smad3 and Smad4 bind in vitro to the CAGA box of COL1A2; TGFβ treatment of cultured fibroblasts induces Smad3/Smad4 binding to the TbRE; transient overexpression of Smad3 and Smad4 in fibroblasts transactivates TbRE-driven transcription; and COLIA2 gene up-regulation by TGFβ is abolished in cells stably transfected with plasmids that express dominant negative forms of Smad3 or Smad4. In Sp1-deficient Drosophila Schneider cells, there was cooperative synergy between Smad3/Smad4 and Sp1 at the TbRE site. The analysis also emphasized the requirement of both Sp1 - and Smad-binding sites for optimal promoter transactivation. In cells stably transfected with a plasmid expressing a dominant negative form of Sp1, the synergy was shown to be promoter-specific and dependent on the binding of Sp1 to the TbRE. Interestingly, overexpression of dominant negative Sp1 was found to block the antagonistic signal of tumor necrosis factor-α on COL1A2 transcription, as well. These results provide the first linkage between the Smad3 and Smad4 proteins and TGFβ stimulation of type I collagen biosynthesis.

Original languageEnglish
Pages (from-to)39237-39245
Number of pages9
JournalJournal of Biological Chemistry
Volume275
Issue number50
DOIs
StatePublished - 15 Dec 2000
Externally publishedYes

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