Involvement of p300 in TGF-β/Smad-Pathway-Mediated α2(I) Collagen Expression in Mouse Mesangial Cells

Abstract

Background: Transforming growth factor beta 1 (TGF-β1) induces α2(I) collagen gene (COL1A2) expression in mesangial cells through physical and functional cooperation of Smad proteins and Sp1. A transcriptional coactivator, p300, is also suggested to play an important role in TGF-β1/Smad signal transduction. However, the role of p300 in TGF-β1/Smad-pathway-mediated transcriptional activation of the COL1A2 gene in mesangial cells is still obscure. Methods: Endogenous p300 expression and its modulation by TGF-β1 were evaluated by Western blotting and immunofluorescence. The physical interaction of p300 with Smad2/3 was examined by immunoprecipitation followed by Western blotting. The functional role of p300 in TGF-β1/Smad-pathway-mediated COL1A2 transcription was investigated in cotransfection experiments using a COL1A2 promoter-luciferase reporter gene construct and p300 expression plasmids. Results: TGF-β1 induced COL1A2 gene expression in cultured mouse mesangial cells which was blocked by overexpression of inhibitory Smad7. In addition, TGF-β1-induced nuclear export of endogenous Smad7 was observed in mouse mesangial cells. Endogenous p300 was expressed in the nucleus of the cells. TGF-β1 induced interaction of endogenous p300 with Smad2/3, and a dominant negative construct of p300 inhibited the TGF-β1-induced COL1A2 expression in cultured mouse mesangial cells. Conclusions: p300 may be involved in TGF-β1/Smad-pathway-mediated type I collagen gene transcription in mouse mesangial cells. Our findings would reveal a molecular basis of TGF-β1-induced type I collagen gene transcription in mouse mesangial cells.