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Autocrine transforming growth factor β signaling regulates extracellular signal-regulated kinase 1/2 phosphorylation via modulation of protein phosphatase 2A expression in scleroderma fibroblasts.

Samuel GH, Bujor AM, Nakerakanti SS, Hant FN, Trojanowska M - Fibrogenesis Tissue Repair (2010)

Bottom Line: We show for the first time that PP2A mRNA and protein expression are significantly reduced in SSc fibroblasts and correlate with an increase in ERK1/2 phosphorylation and collagen expression.Accordingly, blockade of autocrine TGFβ signaling in SSc fibroblasts using soluble recombinant TGFβ receptor II (SRII) restored PP2A levels and decreased ERK1/2 phosphorylation and collagen expression.Taken together, these studies suggest that decreased PP2A levels in SSc is a result of constitutively activated autocrine TGFβ signaling and could contribute to enhanced phosphorylation of ERK1/2 and matrix production in SSc fibroblasts.

View Article: PubMed Central - HTML - PubMed

Affiliation: Arthritis Center, Division of Rheumatology, Boston University Medical Campus, Boston, MA, USA. trojanme@bu.edu.

ABSTRACT

Background: During scleroderma (SSc) pathogenesis, fibroblasts acquire an activated phenotype characterized by enhanced production of extracellular matrix (ECM) and constitutive activation of several major signaling pathways including extracellular signal-related kinase (ERK1/2). Several studies have addressed the role of ERK1/2 in SSc fibrosis however the mechanism of its prolonged activation in SSc fibroblasts is still unknown. Protein phosphatase 2A (PP2A) is a key serine threonine phosphatase responsible for dephosphorylation of a wide array of signaling molecules. Recently published microarray data from cultured SSc fibroblasts suggests that the catalytic subunit (C-subunit) of PP2A is downregulated in SSc. In this study we examined the role and regulation of PP2A in SSc fibroblasts in the context of ERK1/2 phosphorylation and matrix production.

Results: We show for the first time that PP2A mRNA and protein expression are significantly reduced in SSc fibroblasts and correlate with an increase in ERK1/2 phosphorylation and collagen expression. Furthermore, transforming growth factor β (TGFβ), a major profibrotic cytokine implicated in SSc fibrosis, downregulates PP2A expression in healthy fibroblasts. PP2A-specific small interfering RNA (siRNA) was utilized to confirm the role of PP2A in ERK1/2 dephosphorylation in dermal fibroblasts. Accordingly, blockade of autocrine TGFβ signaling in SSc fibroblasts using soluble recombinant TGFβ receptor II (SRII) restored PP2A levels and decreased ERK1/2 phosphorylation and collagen expression. In addition, we observed that inhibition of ERK1/2 in SSc fibroblasts increased PP2A expression suggesting that ERK1/2 phosphorylation also contributes to maintaining low levels of PP2A, leading to an even further amplification of ERK1/2 phosphorylation.

Conclusions: Taken together, these studies suggest that decreased PP2A levels in SSc is a result of constitutively activated autocrine TGFβ signaling and could contribute to enhanced phosphorylation of ERK1/2 and matrix production in SSc fibroblasts.

No MeSH data available.


Related in: MedlinePlus

Extracellular signal-regulated kinase (ERK)1/2 phosphorylation is a negative regulator of protein phosphatase 2A (PP2A) expression in scleroderma (SSc) fibroblasts. (a) After treatment with ERK1/2 inhibitor U0126 (10 nm) for 24 h, PP2A expression was determined using western blot analysis in SSc fibroblasts; P < 0.05. (b) Bar graph showing quantification of PP2A expression from (a).
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Figure 5: Extracellular signal-regulated kinase (ERK)1/2 phosphorylation is a negative regulator of protein phosphatase 2A (PP2A) expression in scleroderma (SSc) fibroblasts. (a) After treatment with ERK1/2 inhibitor U0126 (10 nm) for 24 h, PP2A expression was determined using western blot analysis in SSc fibroblasts; P < 0.05. (b) Bar graph showing quantification of PP2A expression from (a).

Mentions: The activity of kinases and phosphatases is tightly regulated in the cell and often involve feedback mechanisms, which help maintain the levels of cellular phosphorylation. A study performed in human lung fibroblasts suggests that silencing of ERK1/2 is associated with a decrease in PP2A activity [25]. In order to further explore the relationship between PP2A and ERK1/2 phosphorylation, we examined the possibility that ERK1/2 activation could play a role in regulating the PP2A levels in SSc fibroblasts. SSc and normal dermal fibroblasts were treated with the pharmacological inhibitor U0126 to block ERK1/2 phosphorylation. Interestingly, only SSc fibroblasts showed increased PP2A expression upon treatment with U0126, suggesting that ERK1/2 activation contributes to maintaining decreased PP2A levels in SSc (Figure 5a,b). No significant change in PP2A levels in normal fibroblasts was observed (Data not shown). Taken together, these results suggest that autocrine TGFβ signaling in SSc fibroblasts leads to activation of ERK1/2 which in turn downregulates PP2A levels, thereby leading to even more prolonged phosphorylation of ERK1/2.


Autocrine transforming growth factor β signaling regulates extracellular signal-regulated kinase 1/2 phosphorylation via modulation of protein phosphatase 2A expression in scleroderma fibroblasts.

Samuel GH, Bujor AM, Nakerakanti SS, Hant FN, Trojanowska M - Fibrogenesis Tissue Repair (2010)

Extracellular signal-regulated kinase (ERK)1/2 phosphorylation is a negative regulator of protein phosphatase 2A (PP2A) expression in scleroderma (SSc) fibroblasts. (a) After treatment with ERK1/2 inhibitor U0126 (10 nm) for 24 h, PP2A expression was determined using western blot analysis in SSc fibroblasts; P < 0.05. (b) Bar graph showing quantification of PP2A expression from (a).
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3008687&req=5

Figure 5: Extracellular signal-regulated kinase (ERK)1/2 phosphorylation is a negative regulator of protein phosphatase 2A (PP2A) expression in scleroderma (SSc) fibroblasts. (a) After treatment with ERK1/2 inhibitor U0126 (10 nm) for 24 h, PP2A expression was determined using western blot analysis in SSc fibroblasts; P < 0.05. (b) Bar graph showing quantification of PP2A expression from (a).
Mentions: The activity of kinases and phosphatases is tightly regulated in the cell and often involve feedback mechanisms, which help maintain the levels of cellular phosphorylation. A study performed in human lung fibroblasts suggests that silencing of ERK1/2 is associated with a decrease in PP2A activity [25]. In order to further explore the relationship between PP2A and ERK1/2 phosphorylation, we examined the possibility that ERK1/2 activation could play a role in regulating the PP2A levels in SSc fibroblasts. SSc and normal dermal fibroblasts were treated with the pharmacological inhibitor U0126 to block ERK1/2 phosphorylation. Interestingly, only SSc fibroblasts showed increased PP2A expression upon treatment with U0126, suggesting that ERK1/2 activation contributes to maintaining decreased PP2A levels in SSc (Figure 5a,b). No significant change in PP2A levels in normal fibroblasts was observed (Data not shown). Taken together, these results suggest that autocrine TGFβ signaling in SSc fibroblasts leads to activation of ERK1/2 which in turn downregulates PP2A levels, thereby leading to even more prolonged phosphorylation of ERK1/2.

Bottom Line: We show for the first time that PP2A mRNA and protein expression are significantly reduced in SSc fibroblasts and correlate with an increase in ERK1/2 phosphorylation and collagen expression.Accordingly, blockade of autocrine TGFβ signaling in SSc fibroblasts using soluble recombinant TGFβ receptor II (SRII) restored PP2A levels and decreased ERK1/2 phosphorylation and collagen expression.Taken together, these studies suggest that decreased PP2A levels in SSc is a result of constitutively activated autocrine TGFβ signaling and could contribute to enhanced phosphorylation of ERK1/2 and matrix production in SSc fibroblasts.

View Article: PubMed Central - HTML - PubMed

Affiliation: Arthritis Center, Division of Rheumatology, Boston University Medical Campus, Boston, MA, USA. trojanme@bu.edu.

ABSTRACT

Background: During scleroderma (SSc) pathogenesis, fibroblasts acquire an activated phenotype characterized by enhanced production of extracellular matrix (ECM) and constitutive activation of several major signaling pathways including extracellular signal-related kinase (ERK1/2). Several studies have addressed the role of ERK1/2 in SSc fibrosis however the mechanism of its prolonged activation in SSc fibroblasts is still unknown. Protein phosphatase 2A (PP2A) is a key serine threonine phosphatase responsible for dephosphorylation of a wide array of signaling molecules. Recently published microarray data from cultured SSc fibroblasts suggests that the catalytic subunit (C-subunit) of PP2A is downregulated in SSc. In this study we examined the role and regulation of PP2A in SSc fibroblasts in the context of ERK1/2 phosphorylation and matrix production.

Results: We show for the first time that PP2A mRNA and protein expression are significantly reduced in SSc fibroblasts and correlate with an increase in ERK1/2 phosphorylation and collagen expression. Furthermore, transforming growth factor β (TGFβ), a major profibrotic cytokine implicated in SSc fibrosis, downregulates PP2A expression in healthy fibroblasts. PP2A-specific small interfering RNA (siRNA) was utilized to confirm the role of PP2A in ERK1/2 dephosphorylation in dermal fibroblasts. Accordingly, blockade of autocrine TGFβ signaling in SSc fibroblasts using soluble recombinant TGFβ receptor II (SRII) restored PP2A levels and decreased ERK1/2 phosphorylation and collagen expression. In addition, we observed that inhibition of ERK1/2 in SSc fibroblasts increased PP2A expression suggesting that ERK1/2 phosphorylation also contributes to maintaining low levels of PP2A, leading to an even further amplification of ERK1/2 phosphorylation.

Conclusions: Taken together, these studies suggest that decreased PP2A levels in SSc is a result of constitutively activated autocrine TGFβ signaling and could contribute to enhanced phosphorylation of ERK1/2 and matrix production in SSc fibroblasts.

No MeSH data available.


Related in: MedlinePlus