Smad3 Mediates Transforming Growth Factor-β-induced Collagenase-3 (Matrix Metalloproteinase-13) Expression in Human Gingival Fibroblasts: EVIDENCE FOR CROSS-TALK BETWEEN Smad3 AND p38 SIGNALING PATHWAYS

Suvi-Katri Leivonen,Andrew Chantry,Lari Häkkinen,Jiahuai Han,Veli-Matti Kähäri

doi:10.1074/jbc.m206535200

Abstract

Transforming growth factor-beta (TGF-beta) is a potent inducer of collagenase-3 (MMP-13) gene expression in human gingival fibroblasts, and this requires activation of the p38 mitogen-activated protein kinase pathway. Here, we have constructed recombinant adenoviruses harboring genes for hemagglutinin-tagged Smad2, Smad3, and Smad4 and used these in dissecting the role of Smads, the signaling mediators of TGF-beta, in regulation of endogenous MMP-13 gene expression in human gingival fibroblasts. Adenoviral expression of Smad3, but not Smad2, augmented the TGF-beta-elicited induction of MMP-13 expression. In addition, adenoviral gene delivery of dominant negative Smad3 blocked the TGF-beta-induced MMP-13 expression in gingival fibroblasts. Co-expression of Smad3 with constitutively active MKK3b and MKK6b, the upstream activators of p38, resulted in nuclear translocation of Smad3 in the absence of TGF-beta and in induction of MMP-13 expression. The induction of MMP-13 expression by Smad3 and constitutively active mutants of MKK3b or MKK6b was blocked by specific p38 inhibitor SB203580 and by the dominant negative form of p38alpha. These results show that TGF-beta-induced expression of human MMP-13 gene in gingival fibroblasts is dependent on the activation of two distinct signaling pathways (i.e. Smad3 and p38alpha). In addition, these findings provide evidence for a novel type of cross-talk between Smad and p38 mitogen-activated protein kinase signaling cascades, which involves activation of Smad3 by p38alpha.

Highlights

Transforming growth factor-␤ (TGF-␤) is a potent inducer of collagenase-3 (MMP-13) gene expression in human gingival fibroblasts, and this requires activation of the p38 mitogen-activated protein kinase pathway
These results show that TGF-␤induced expression of human Matrix metalloproteinases (MMPs)-13 gene in gingival fibroblasts is dependent on the activation of two distinct signaling pathways (i.e. Smad3 and p38␣)
Activation of p38␣ induces the nuclear translocation of Smad3, indicating that Smad3 is activated by p38␣ either directly or indirectly via endogenous mediator. Together these results indicate that Smad3 is involved in TGF␤-elicited induction of the expression of MMP-13 in human gingival fibroblasts and that this involves cross-talk between Smad3 and the p38 mitogen-activated protein kinase (MAPK) pathway

Summary

Introduction

Transforming growth factor-␤ (TGF-␤) is a potent inducer of collagenase-3 (MMP-13) gene expression in human gingival fibroblasts, and this requires activation of the p38 mitogen-activated protein kinase pathway. Total cellular RNA was analyzed with Northern blot hybridization for expression of MMP-13, pro␣1(I) collagen, PAI-1, and GAPDH mRNAs. B and C, to examine the effect of Smad3 together with Smad4, human gingival fibroblasts were infected with adenoviruses for Smad3 (RAdSmad3) and Smad4 (RAdSmad4) and with empty control virus RAd66 and treated with TGF-␤1 (5 ng/ml) for 24 h.

Results

Conclusion