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The dermatan sulfate proteoglycan decorin modulates ?2?1 integrin and the vimentin intermediate filament system during collagen synthesis.

ABSTRACT: Decorin, a small leucine-rich proteoglycan harboring a dermatan sulfate chain at its N-terminus, is involved in regulating matrix organization and cell signaling. Loss of the dermatan sulfate of decorin leads to an Ehlers-Danlos syndrome characterized by delayed wound healing. Decorin-null (Dcn(-/-)) mice display a phenotype similar to that of EDS patients. The fibrillar collagen phenotype of Dcn(-/-) mice could be rescued in vitro by decorin but not with decorin lacking the glycosaminoglycan chain. We utilized a 3D cell culture model to investigate the impact of the altered extracellular matrix on Dcn(-/-) fibroblasts. Using 2D gel electrophoresis followed by mass spectrometry, we identified vimentin as one of the proteins that was differentially upregulated by the presence of decorin. We discovered that a decorin-deficient matrix leads to abnormal nuclear morphology in the Dcn(-/-) fibroblasts. This phenotype could be rescued by the decorin proteoglycan but less efficiently by the decorin protein core. Decorin treatment led to a significant reduction of the ?2?1 integrin at day 6 in Dcn(-/-) fibroblasts, whereas the protein core had no effect on ?1. Interestingly, only the decorin core induced mRNA synthesis, phosphorylation and de novo synthesis of vimentin indicating that the proteoglycan decorin in the extracellular matrix stabilizes the vimentin intermediate filament system. We could support these results in vivo, because the dermis of wild-type mice have more vimentin and less ?1 integrin compared to Dcn(-/-). Furthermore, the ?2?1 null fibroblasts also showed a reduced amount of vimentin compared to wild-type. These data show for the first time that decorin has an impact on the biology of ?2?1 integrin and the vimentin intermediate filament system. Moreover, our findings provide a mechanistic explanation for the reported defects in wound healing associated with the Dcn(-/-) phenotype.

SUBMITTER: Jungmann O

PROVIDER: S-EPMC3513320 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature

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The dermatan sulfate proteoglycan decorin modulates α2β1 integrin and the vimentin intermediate filament system during collagen synthesis.

Jungmann Oliver O Nikolovska Katerina K Stock Christian C Schulz Jan-Niklas JN Eckes Beate B Riethmüller Christoph C Owens Rick T RT Iozzo Renato V RV Seidler Daniela G DG

PloS one 20121203 12

Decorin, a small leucine-rich proteoglycan harboring a dermatan sulfate chain at its N-terminus, is involved in regulating matrix organization and cell signaling. Loss of the dermatan sulfate of decorin leads to an Ehlers-Danlos syndrome characterized by delayed wound healing. Decorin-null (Dcn(-/-)) mice display a phenotype similar to that of EDS patients. The fibrillar collagen phenotype of Dcn(-/-) mice could be rescued in vitro by decorin but not with decorin lacking the glycosaminoglycan ch ...[more]

PMID: 23226541

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Project description:Decorin is a small leucine-rich proteoglycan harboring a single glycosaminoglycan chain, which, in skin, is mainly composed of dermatan sulfate (DS). Mutant mice with targeted disruption of the decorin gene (Dcn(-/-)) exhibit an abnormal collagen architecture in the dermis and reduced tensile strength, collectively leading to a skin fragility phenotype. Notably, Ehlers-Danlos patients with mutations in enzymes involved in the biosynthesis of DS display a similar phenotype, and recent studies indicate that DS is involved in growth factor binding and signaling. To determine the impact of the loss of DS-decorin in the dermis, we analyzed the glycosaminoglycan content of Dcn(-/-) and wild-type mouse skin. The total amount of chondroitin/dermatan sulfate (CS/DS) was increased in the Dcn(-/-) skin, but was overall less sulfated with a significant reduction in bisulfated ΔDiS2,X (X=4 or 6) disaccharide units, due to the reduced expression of uronyl 2-O sulfotransferase (Ust). With increasing age, sulfation declined; however, Dcn(-/-) CS/DS was constantly undersulfated vis-à-vis wild-type. Functionally, we found altered fibroblast growth factor (Fgf)-7 and -2 binding due to changes in the micro-heterogeneity of skin Dcn(-/-) CS/DS. To better delineate the role of decorin, we used a 3D Dcn(-/-) fibroblast cell culture model. We found that the CS/DS extracts of wild-type and Dcn(-/-) fibroblasts were similar to the skin sugars, and this correlated with the lack of uronyl 2-O sulfotransferase in the Dcn(-/-) fibroblasts. Moreover, Ffg7 binding to total CS/DS was attenuated in the Dcn(-/-) samples. Surprisingly, wild-type CS/DS significantly reduced the binding of Fgf7 to keratinocytes in a concentration dependent manner unlike the Dcn(-/-) CS/DS that only affected the binding at higher concentrations. Although binding to cell-surfaces was quite similar at higher concentrations, keratinocyte proliferation was differentially affected. Higher concentration of Dcn(-/-) CS/DS induced proliferation in contrast to wild-type CS/DS. 3D co-cultures of fibroblasts and keratinocytes showed that, unlike Dcn(-/-) CS/DS, wild-type CS/DS promoted differentiation of keratinocytes. Collectively, our results provide novel mechanistic explanations for the reported defects in wound healing in Dcn(-/-) mice and possibly Ehlers-Danlos patients. Moreover, the lack of decorin-derived DS and an altered CS/DS composition differentially influence keratinocyte behavior.

Project description:BACKGROUND: Human neuroblastoma (NB) cell lines may present with either one of the so-called S-and N-subtypes. We have previously reported a strong correlation between protein expression levels of vimentin, an S-subtype marker, and the p21Waf1 cyclin-dependent kinase inhibitor. We here investigated whether this correlation extend to the mRNA level in NB cell lines as well as in patients' tumors. We also further explored the relationship between expression of vimentin and p21, by asking whether vimentin could regulate p21 expression. METHODS: Vimentin and p21 mRNA levels in NB cell lines as well as in patients' tumors (n = 77) were quantified using Q-PCR. Q-PCR data obtained from tumors of high risk NB patients (n = 40) were analyzed in relation with the overall survival using the Log-rank Kaplan-Meier estimation. siRNA-mediated depletion or overexpression of vimentin in highly or low expressing vimentin cell lines, respectively, followed by protein expression and promoter activation assays were used to assess the role of vimentin in modulating p21 expression. RESULTS: We extend the significant correlation between vimentin and p21 expression to the mRNA level in NB cell lines as well as in patients' tumors. Overall survival analysis from Q-PCR data obtained from tumors of high risk patients suggests that lower levels of p21 expression could be associated with a poorer outcome. Our data additionally indicate that the correlation observed between p21 and vimentin expression levels results from p21 transcriptional activity being regulated by vimentin. Indeed, downregulating vimentin resulted in a significant decrease in p21 mRNA and protein expression as well as in p21 promoter activity. Conversely, overexpressing vimentin triggered an increase in p21 promoter activity in cells with a nuclear expression of vimentin. CONCLUSION: Our results suggest that p21 mRNA tumor expression level could represent a refined prognostic factor for high risk NB patients. Our data also show that vimentin regulates p21 transcription; this is the first demonstration of a gene regulating function for this type III-intermediate filament.

Dataset Information

The dermatan sulfate proteoglycan decorin modulates ?2?1 integrin and the vimentin intermediate filament system during collagen synthesis.

Publications

The dermatan sulfate proteoglycan decorin modulates α2β1 integrin and the vimentin intermediate filament system during collagen synthesis.

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