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TGF-β-elicited induction of tissue inhibitor of metalloproteinases (TIMP)-3 expression in fibroblasts involves complex interplay between Smad3, p38α, and ERK1/2.

Leivonen SK, Lazaridis K, Decock J, Chantry A, Edwards DR, Kähäri VM - PLoS ONE (2013)

Bottom Line: Inhibition of extracellular signal-regulated kinase 1/2 (ERK1/2) activation with PD98059 and p38 mitogen-activated protein kinase activity by SB203580 resulted in suppression of TGF-β-induced TIMP-3 expression, indicating that ERK1/2 and p38 MAPK mediate the effect of TGF-β on TIMP-3 expression.Specific activation of p38α and ERK1/2 by constitutively active mutants of MKK3b or MEK1, respectively, and simultaneous co-expression of Smad3 resulted in induction of TIMP-3 expression in the absence of TGF-β indicating that Smad3 co-operates with p38 and ERK1/2 in the induction of TIMP-3 expression.These results demonstrate the complex interplay between Smad3, p38α, and ERK1/2 signaling in the regulation of TIMP-3 gene expression in fibroblasts, which may play a role in inflammation, tissue repair, and fibrosis.

View Article: PubMed Central - PubMed

Affiliation: Department of Dermatology, University of Turku, and Turku University Hospital, Turku, Finland.

ABSTRACT
Transforming growth factor-β (TGF-β) promotes extracellular matrix deposition by down-regulating the expression of matrix degrading proteinases and upregulating their inhibitors. Tissue inhibitor of metalloproteinases (TIMP)-3 is an ECM-associated specific inhibitor of matrix degrading metalloproteinases. Here, we have characterized the signaling pathways mediating TGF-β-induced expression of TIMP-3. Basal and TGF-β-induced TIMP-3 mRNA expression was abolished in Smad4-deficient mouse embryonic fibroblasts and restoring Smad4 expression rescued the response. Inhibition of Smad signaling by expression of Smad7 and dominant negative Smad3 completely abolished TGF-β-elicited expression of TIMP-3 in human fibroblasts, whereas overexpression of Smad3 enhanced it. Inhibition of extracellular signal-regulated kinase 1/2 (ERK1/2) activation with PD98059 and p38 mitogen-activated protein kinase activity by SB203580 resulted in suppression of TGF-β-induced TIMP-3 expression, indicating that ERK1/2 and p38 MAPK mediate the effect of TGF-β on TIMP-3 expression. Specific activation of p38α and ERK1/2 by constitutively active mutants of MKK3b or MEK1, respectively, and simultaneous co-expression of Smad3 resulted in induction of TIMP-3 expression in the absence of TGF-β indicating that Smad3 co-operates with p38 and ERK1/2 in the induction of TIMP-3 expression. These results demonstrate the complex interplay between Smad3, p38α, and ERK1/2 signaling in the regulation of TIMP-3 gene expression in fibroblasts, which may play a role in inflammation, tissue repair, and fibrosis.

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A schematic representation of the complex interplay of TGF-β-signaling pathways regulating TIMP-3 expression in human fibroblasts.Stimulation of human gingival fibroblasts with TGF-β results in activation of Smad3, ERK1/2 and p38 MAPK pathways. Activation of all three pathways is required for induction of TIMP-3 expression by TGF-β Smad3 associates with Smad4 and mediates induction of TIMP-3 expression by TGF-β. ERK1/2 and p38 MAPK pathways both co-operate with Smad3 in mediating the induction of TIMP-3 expression by TGF-β.
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pone-0057474-g005: A schematic representation of the complex interplay of TGF-β-signaling pathways regulating TIMP-3 expression in human fibroblasts.Stimulation of human gingival fibroblasts with TGF-β results in activation of Smad3, ERK1/2 and p38 MAPK pathways. Activation of all three pathways is required for induction of TIMP-3 expression by TGF-β Smad3 associates with Smad4 and mediates induction of TIMP-3 expression by TGF-β. ERK1/2 and p38 MAPK pathways both co-operate with Smad3 in mediating the induction of TIMP-3 expression by TGF-β.

Mentions: Here, p38 and ERK1/2 MAPK pathways mediated the effects of TGF-β on TIMP-3 gene expression in human gingival fibroblasts, since blocking their activity with chemical inhibitors SB203580 and PD98059 resulted in a marked suppression of TGF-β-induced TIMP-3 mRNA levels. In addition, activation of p38 by MKK3bE and ERK1/2 by MEK1CA in combination resulted in the induction of TIMP-3 expression in the absence of TGF-β, and this effect was augmented by simultaneous co-expression of Smad3. This indicates that p38, ERK1/2, and Smad3 synergistically mediate the up-regulation of the expression of TIMP-3. We have previously demonstrated, that p38 and Smad3 co-operate in mediating TGF-β-induced expression of MMP-13 in human gingival fibroblasts [25]. Activated p38 induced activation and nuclear translocation of Smad3 in gingival fibroblasts, indicating that p38 MAPK is able to activate Smad3. In addition, ERK1/2 and Smad3 co-operatively mediated the TGF-β-induced CTGF gene expression [26]. It is conceivable that also here, p38α and ERK1/2 influenced Smad3 activation, and together these signaling mediators induced TIMP-3 gene expression. The results are summarized Figure 5 demonstrating the complex regulation of TIMP-3 gene expression by crosstalk between ERK1/2, p38, and Smad3 pathways.


TGF-β-elicited induction of tissue inhibitor of metalloproteinases (TIMP)-3 expression in fibroblasts involves complex interplay between Smad3, p38α, and ERK1/2.

Leivonen SK, Lazaridis K, Decock J, Chantry A, Edwards DR, Kähäri VM - PLoS ONE (2013)

A schematic representation of the complex interplay of TGF-β-signaling pathways regulating TIMP-3 expression in human fibroblasts.Stimulation of human gingival fibroblasts with TGF-β results in activation of Smad3, ERK1/2 and p38 MAPK pathways. Activation of all three pathways is required for induction of TIMP-3 expression by TGF-β Smad3 associates with Smad4 and mediates induction of TIMP-3 expression by TGF-β. ERK1/2 and p38 MAPK pathways both co-operate with Smad3 in mediating the induction of TIMP-3 expression by TGF-β.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3585359&req=5

pone-0057474-g005: A schematic representation of the complex interplay of TGF-β-signaling pathways regulating TIMP-3 expression in human fibroblasts.Stimulation of human gingival fibroblasts with TGF-β results in activation of Smad3, ERK1/2 and p38 MAPK pathways. Activation of all three pathways is required for induction of TIMP-3 expression by TGF-β Smad3 associates with Smad4 and mediates induction of TIMP-3 expression by TGF-β. ERK1/2 and p38 MAPK pathways both co-operate with Smad3 in mediating the induction of TIMP-3 expression by TGF-β.
Mentions: Here, p38 and ERK1/2 MAPK pathways mediated the effects of TGF-β on TIMP-3 gene expression in human gingival fibroblasts, since blocking their activity with chemical inhibitors SB203580 and PD98059 resulted in a marked suppression of TGF-β-induced TIMP-3 mRNA levels. In addition, activation of p38 by MKK3bE and ERK1/2 by MEK1CA in combination resulted in the induction of TIMP-3 expression in the absence of TGF-β, and this effect was augmented by simultaneous co-expression of Smad3. This indicates that p38, ERK1/2, and Smad3 synergistically mediate the up-regulation of the expression of TIMP-3. We have previously demonstrated, that p38 and Smad3 co-operate in mediating TGF-β-induced expression of MMP-13 in human gingival fibroblasts [25]. Activated p38 induced activation and nuclear translocation of Smad3 in gingival fibroblasts, indicating that p38 MAPK is able to activate Smad3. In addition, ERK1/2 and Smad3 co-operatively mediated the TGF-β-induced CTGF gene expression [26]. It is conceivable that also here, p38α and ERK1/2 influenced Smad3 activation, and together these signaling mediators induced TIMP-3 gene expression. The results are summarized Figure 5 demonstrating the complex regulation of TIMP-3 gene expression by crosstalk between ERK1/2, p38, and Smad3 pathways.

Bottom Line: Inhibition of extracellular signal-regulated kinase 1/2 (ERK1/2) activation with PD98059 and p38 mitogen-activated protein kinase activity by SB203580 resulted in suppression of TGF-β-induced TIMP-3 expression, indicating that ERK1/2 and p38 MAPK mediate the effect of TGF-β on TIMP-3 expression.Specific activation of p38α and ERK1/2 by constitutively active mutants of MKK3b or MEK1, respectively, and simultaneous co-expression of Smad3 resulted in induction of TIMP-3 expression in the absence of TGF-β indicating that Smad3 co-operates with p38 and ERK1/2 in the induction of TIMP-3 expression.These results demonstrate the complex interplay between Smad3, p38α, and ERK1/2 signaling in the regulation of TIMP-3 gene expression in fibroblasts, which may play a role in inflammation, tissue repair, and fibrosis.

View Article: PubMed Central - PubMed

Affiliation: Department of Dermatology, University of Turku, and Turku University Hospital, Turku, Finland.

ABSTRACT
Transforming growth factor-β (TGF-β) promotes extracellular matrix deposition by down-regulating the expression of matrix degrading proteinases and upregulating their inhibitors. Tissue inhibitor of metalloproteinases (TIMP)-3 is an ECM-associated specific inhibitor of matrix degrading metalloproteinases. Here, we have characterized the signaling pathways mediating TGF-β-induced expression of TIMP-3. Basal and TGF-β-induced TIMP-3 mRNA expression was abolished in Smad4-deficient mouse embryonic fibroblasts and restoring Smad4 expression rescued the response. Inhibition of Smad signaling by expression of Smad7 and dominant negative Smad3 completely abolished TGF-β-elicited expression of TIMP-3 in human fibroblasts, whereas overexpression of Smad3 enhanced it. Inhibition of extracellular signal-regulated kinase 1/2 (ERK1/2) activation with PD98059 and p38 mitogen-activated protein kinase activity by SB203580 resulted in suppression of TGF-β-induced TIMP-3 expression, indicating that ERK1/2 and p38 MAPK mediate the effect of TGF-β on TIMP-3 expression. Specific activation of p38α and ERK1/2 by constitutively active mutants of MKK3b or MEK1, respectively, and simultaneous co-expression of Smad3 resulted in induction of TIMP-3 expression in the absence of TGF-β indicating that Smad3 co-operates with p38 and ERK1/2 in the induction of TIMP-3 expression. These results demonstrate the complex interplay between Smad3, p38α, and ERK1/2 signaling in the regulation of TIMP-3 gene expression in fibroblasts, which may play a role in inflammation, tissue repair, and fibrosis.

Show MeSH
Related in: MedlinePlus