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Sonic hedgehog-Gli1 signaling pathway regulates the epithelial mesenchymal transition (EMT) by mediating a new target gene, S100A4, in pancreatic cancer cells.

Xu X, Su B, Xie C, Wei S, Zhou Y, Liu H, Dai W, Cheng P, Wang F, Xu X, Guo C - PLoS ONE (2014)

Bottom Line: Five members of the S100 genes family, S100A2, S100A4, S100A6, S100A11, and S100A14 were found to be downregulated significantly upon Gli1 knockdown.Indeed, the data indicate S100A4 and vimentin genes were upregulated significantly by Shh/Gli1-expression increasing and E-cadherin was significantly reduced at the same time.Migration of PC cells was increased significantly in a dose-dependent manner of Gli1 expression (P<0.05) and siS100A4 significantly reversed the response of PC cells induced by L-Shh transduction (P<0.01).

View Article: PubMed Central - PubMed

Affiliation: Department of Gastroenterology, the Tenth People's Hospital of Shanghai, Tongji University, Shanghai, China.

ABSTRACT

Aims: The hedgehog signaling pathway plays an important role in EMT of pancreatic cancer cells, but the precise mechanisms remain elusive. Because S100A4 as a key EMT moleculer marker was found to be upregulated upon Gli1 in pancreatic cancer cells, we focused on the relationship between Shh-Gli1 signals and S100 genes family.

Methods: On the base of cDNA microarray data, we investigated regulating mechanism of Gli1 to some members of S100A genes family in pancreatic cancer cell lines firstly. Then, the regulation of Gli1 to S100A4 gene was studied by molecular biology assays and the pro-metastasis effection of Gli1-dependent S100A4 was investigated in vitro. Finally, the expressions of Shh, Gli1, S100A4 and E-cadherin in pancreatic cancer tissues were studied by using immunohistochemistry assays.

Results: Five members of the S100 genes family, S100A2, S100A4, S100A6, S100A11, and S100A14 were found to be downregulated significantly upon Gli1 knockdown. Gli1 enhancer prediction combining with in vitro data demonstrated that Gli1 primarily regulates S100A family members via cis-acting elements. Indeed, the data indicate S100A4 and vimentin genes were upregulated significantly by Shh/Gli1-expression increasing and E-cadherin was significantly reduced at the same time. Migration of PC cells was increased significantly in a dose-dependent manner of Gli1 expression (P<0.05) and siS100A4 significantly reversed the response of PC cells induced by L-Shh transduction (P<0.01).

Conclusion: Our data establish a novel connection between Shh-Gli1 signaling and S100A4 regulation, which imply that S100A4 might be one of the key factors in EMT mediated by Shh-Gli1 signaling in pancreatic cancer.

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Hh signaling pathway promoting invasion/migration of pancreatic cancer cells through mediating S100A4.A: The relative transcription levels of Shh, Gli1, S100A4, E-cadherin and vimentin were regulated by L-Gli1i/L-Shh transduction. B: The relative transcription levels of S100A4, E-cadherin and vimentin in L-Shh transfected cells were reversed by siS100A4 transduction. C: The expression levels of Gli1, S100A4, E-cadherin and vimentin proteins regulated by L-Gli1i/L-Shh transduction. D: The expression levels of S100A4, E-cadherin and vimentin proteins were reversed by siS100A4 transduction. E: The invasion/migration of pancreatic cancer cells regulated by L-Gli1i/L-Shh transduction were analyzed by transwell assays. F: The invasion/migration of L-Shh transfected cells were reversed by siS100A4 transduction. * P<0.05, ** P<0.01.
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pone-0096441-g004: Hh signaling pathway promoting invasion/migration of pancreatic cancer cells through mediating S100A4.A: The relative transcription levels of Shh, Gli1, S100A4, E-cadherin and vimentin were regulated by L-Gli1i/L-Shh transduction. B: The relative transcription levels of S100A4, E-cadherin and vimentin in L-Shh transfected cells were reversed by siS100A4 transduction. C: The expression levels of Gli1, S100A4, E-cadherin and vimentin proteins regulated by L-Gli1i/L-Shh transduction. D: The expression levels of S100A4, E-cadherin and vimentin proteins were reversed by siS100A4 transduction. E: The invasion/migration of pancreatic cancer cells regulated by L-Gli1i/L-Shh transduction were analyzed by transwell assays. F: The invasion/migration of L-Shh transfected cells were reversed by siS100A4 transduction. * P<0.05, ** P<0.01.

Mentions: To further study the pro-metastasis function of Gli1-derived S100A4 in pancreatic cancer cells, five groups of PC cells, L-Gli1i, L-C, L-Shh, L-Shh+siS100A4 MIS and L-Shh + siS100A4, were used to analize the regulation of Shh-Gli1 signals to S100A4, E-cadherin and VIM genes by real time RT-PCR and western-blotting assays. And then the same five group cells were used to evaluate invasion/migration of PC cells in vitro by transwell assays. The results of qRT-PCR and western-blotting showed that expression levels of S100A4 and VIM genes were increased significantly by Shh/Gli1-expression increasing. In contrast, the expression levels of E-cadherin were significantly reduced at the same time. (Figure 4A, C). Moreover, the S100A4 knocked-down signifiantly reversed downregulated E-cadherin and upregulated VIM induced by L-Shh transduction. (Figure 4B, D). The results of transwell assays showed the migration of PC cells were significantly decreased in L-Gli1i group and increased in L-Shh group respectively compared with L-C group (P<0.05). (Figure 4E). Moreover, siS100A4 significantly reversed the response of PC cells induced by L-Shh transduction (P<0.01). (Figure 4F). Thus, it seems that S100A4 mediated by abnomal activated Shh-Gli1 signaling pathway could be one of key pro-migration factors in pancreatic cancer cells.


Sonic hedgehog-Gli1 signaling pathway regulates the epithelial mesenchymal transition (EMT) by mediating a new target gene, S100A4, in pancreatic cancer cells.

Xu X, Su B, Xie C, Wei S, Zhou Y, Liu H, Dai W, Cheng P, Wang F, Xu X, Guo C - PLoS ONE (2014)

Hh signaling pathway promoting invasion/migration of pancreatic cancer cells through mediating S100A4.A: The relative transcription levels of Shh, Gli1, S100A4, E-cadherin and vimentin were regulated by L-Gli1i/L-Shh transduction. B: The relative transcription levels of S100A4, E-cadherin and vimentin in L-Shh transfected cells were reversed by siS100A4 transduction. C: The expression levels of Gli1, S100A4, E-cadherin and vimentin proteins regulated by L-Gli1i/L-Shh transduction. D: The expression levels of S100A4, E-cadherin and vimentin proteins were reversed by siS100A4 transduction. E: The invasion/migration of pancreatic cancer cells regulated by L-Gli1i/L-Shh transduction were analyzed by transwell assays. F: The invasion/migration of L-Shh transfected cells were reversed by siS100A4 transduction. * P<0.05, ** P<0.01.
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pone-0096441-g004: Hh signaling pathway promoting invasion/migration of pancreatic cancer cells through mediating S100A4.A: The relative transcription levels of Shh, Gli1, S100A4, E-cadherin and vimentin were regulated by L-Gli1i/L-Shh transduction. B: The relative transcription levels of S100A4, E-cadherin and vimentin in L-Shh transfected cells were reversed by siS100A4 transduction. C: The expression levels of Gli1, S100A4, E-cadherin and vimentin proteins regulated by L-Gli1i/L-Shh transduction. D: The expression levels of S100A4, E-cadherin and vimentin proteins were reversed by siS100A4 transduction. E: The invasion/migration of pancreatic cancer cells regulated by L-Gli1i/L-Shh transduction were analyzed by transwell assays. F: The invasion/migration of L-Shh transfected cells were reversed by siS100A4 transduction. * P<0.05, ** P<0.01.
Mentions: To further study the pro-metastasis function of Gli1-derived S100A4 in pancreatic cancer cells, five groups of PC cells, L-Gli1i, L-C, L-Shh, L-Shh+siS100A4 MIS and L-Shh + siS100A4, were used to analize the regulation of Shh-Gli1 signals to S100A4, E-cadherin and VIM genes by real time RT-PCR and western-blotting assays. And then the same five group cells were used to evaluate invasion/migration of PC cells in vitro by transwell assays. The results of qRT-PCR and western-blotting showed that expression levels of S100A4 and VIM genes were increased significantly by Shh/Gli1-expression increasing. In contrast, the expression levels of E-cadherin were significantly reduced at the same time. (Figure 4A, C). Moreover, the S100A4 knocked-down signifiantly reversed downregulated E-cadherin and upregulated VIM induced by L-Shh transduction. (Figure 4B, D). The results of transwell assays showed the migration of PC cells were significantly decreased in L-Gli1i group and increased in L-Shh group respectively compared with L-C group (P<0.05). (Figure 4E). Moreover, siS100A4 significantly reversed the response of PC cells induced by L-Shh transduction (P<0.01). (Figure 4F). Thus, it seems that S100A4 mediated by abnomal activated Shh-Gli1 signaling pathway could be one of key pro-migration factors in pancreatic cancer cells.

Bottom Line: Five members of the S100 genes family, S100A2, S100A4, S100A6, S100A11, and S100A14 were found to be downregulated significantly upon Gli1 knockdown.Indeed, the data indicate S100A4 and vimentin genes were upregulated significantly by Shh/Gli1-expression increasing and E-cadherin was significantly reduced at the same time.Migration of PC cells was increased significantly in a dose-dependent manner of Gli1 expression (P<0.05) and siS100A4 significantly reversed the response of PC cells induced by L-Shh transduction (P<0.01).

View Article: PubMed Central - PubMed

Affiliation: Department of Gastroenterology, the Tenth People's Hospital of Shanghai, Tongji University, Shanghai, China.

ABSTRACT

Aims: The hedgehog signaling pathway plays an important role in EMT of pancreatic cancer cells, but the precise mechanisms remain elusive. Because S100A4 as a key EMT moleculer marker was found to be upregulated upon Gli1 in pancreatic cancer cells, we focused on the relationship between Shh-Gli1 signals and S100 genes family.

Methods: On the base of cDNA microarray data, we investigated regulating mechanism of Gli1 to some members of S100A genes family in pancreatic cancer cell lines firstly. Then, the regulation of Gli1 to S100A4 gene was studied by molecular biology assays and the pro-metastasis effection of Gli1-dependent S100A4 was investigated in vitro. Finally, the expressions of Shh, Gli1, S100A4 and E-cadherin in pancreatic cancer tissues were studied by using immunohistochemistry assays.

Results: Five members of the S100 genes family, S100A2, S100A4, S100A6, S100A11, and S100A14 were found to be downregulated significantly upon Gli1 knockdown. Gli1 enhancer prediction combining with in vitro data demonstrated that Gli1 primarily regulates S100A family members via cis-acting elements. Indeed, the data indicate S100A4 and vimentin genes were upregulated significantly by Shh/Gli1-expression increasing and E-cadherin was significantly reduced at the same time. Migration of PC cells was increased significantly in a dose-dependent manner of Gli1 expression (P<0.05) and siS100A4 significantly reversed the response of PC cells induced by L-Shh transduction (P<0.01).

Conclusion: Our data establish a novel connection between Shh-Gli1 signaling and S100A4 regulation, which imply that S100A4 might be one of the key factors in EMT mediated by Shh-Gli1 signaling in pancreatic cancer.

Show MeSH
Related in: MedlinePlus