TY - JOUR
T1 - Synergistic cooperation between Sp1 and Smad3/Smad4 mediates transforming growth factor β1 stimulation of α2(I)-collagen (COL1A2) transcription
AU - Zhang, Wen
AU - Ou, Jiongwen
AU - Inagaki, Yutaka
AU - Greenwel, Patricia
AU - Ramirez, Francesco
PY - 2000/12/15
Y1 - 2000/12/15
N2 - 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.
AB - 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.
UR - http://www.scopus.com/inward/record.url?scp=0034671506&partnerID=8YFLogxK
U2 - 10.1074/jbc.M003339200
DO - 10.1074/jbc.M003339200
M3 - Article
C2 - 11007770
AN - SCOPUS:0034671506
SN - 0021-9258
VL - 275
SP - 39237
EP - 39245
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 50
ER -