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TAZ promotes epithelial to mesenchymal transition via the upregulation of connective tissue growth factor expression in neuroblastoma cells.

Wang Q, Xu Z, An Q, Jiang D, Wang L, Liang B, Li Z - Mol Med Rep (2014)

Bottom Line: The Hippo signaling pathway regulates cell proliferation and apoptosis, and its primary downstream effectors are TAZ and yes‑associated protein 1 (YAP).Repressed expression of TAZ in SK‑N‑BE(2) cells was shown to result in a reduction in aggressiveness of the cell line, by Transwell migration and invasion assay.Furthermore, TAZ was shown by luciferase assay to induce CTGF expression by modulating the activation of the TGF‑β/Smad3 signaling pathway.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatric Surgery, The First Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang 150001, P.R. China.

ABSTRACT
Neuroblastoma (NB) is a neuroendocrine cancer that occurs most commonly in infants and young children. The Hippo signaling pathway regulates cell proliferation and apoptosis, and its primary downstream effectors are TAZ and yes‑associated protein 1 (YAP). The effect of TAZ on the metastatic progression of neuroblastoma and the underlying mechanisms involved remain elusive. In the current study, it was determined by western blot analysis that the migratory and invasive properties of SK‑N‑BE(2) human neuroblastoma cells are associated with high expression levels of TAZ. Repressed expression of TAZ in SK‑N‑BE(2) cells was shown to result in a reduction in aggressiveness of the cell line, by Transwell migration and invasion assay. In contrast, overexpression of TAZ in SK‑N‑SH human neuroblastoma cells was shown by Transwell migration and invasion assays, and western blot analysis, to result in epithelial‑mesenchymal transition (EMT) and increased invasiveness. Mechanistically, the overexpression of TAZ was demonstrated to upregulate the expression levels of connective tissue growth factor (CTGF), by western blot analysis and chromatin immunoprecipitation assay, while the knockdown of TAZ downregulated it. Furthermore, TAZ was shown by luciferase assay to induce CTGF expression by modulating the activation of the TGF‑β/Smad3 signaling pathway. In conclusion, the present study is, to the best of our knowledge, the first to demonstrate that the overexpression of TAZ induces EMT, increasing the invasive abilities of neuroblastoma cells. This suggests that TAZ may serve as a potential target in the development of novel therapies for the treatment of neuroblastoma.

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Migratory and invasive properties of SK-N-BE(2) cells expressing high levels of TAZ. (A) The mRNA and protein expression levels of TAZ were higher in SK-N-BE(2) cells compared with those observed in SK-N-SH cells. Top panel: results of reverse transcription quantitative polymerase chain reaction presented as a western blot from a representative study; bottom panel: graph of the relative intensities of TAZ, which were quantified with densitometry and normalized to GAPDH. Values shown are the mean ± standard deviation (SD) from three independent experiments. P<0.05 for SK-N-BE(2) compared with SK-N-SH. (B) SK-N-BE(2) cells had a higher migratory ability than that of SK-N-SH cells, as determined by Transwell® assays. Graph shows the relative fold changes in cell migration compared with SK-N-SH cells. Values shown are the mean ± SD from three independent experiments. Twelve images were randomly captured in each independent experiment. (C) SK-N-BE(2) cells had a greater invasive ability compared with that of SK-N-SH cells, as determined by Transwell® assays. Experiments were performed as described for B. (D) Expression levels of Vimentin, E-cadherin and β-catenin in SK-N-SH and SK-N-BE(2) cells.
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f1-mmr-11-02-0982: Migratory and invasive properties of SK-N-BE(2) cells expressing high levels of TAZ. (A) The mRNA and protein expression levels of TAZ were higher in SK-N-BE(2) cells compared with those observed in SK-N-SH cells. Top panel: results of reverse transcription quantitative polymerase chain reaction presented as a western blot from a representative study; bottom panel: graph of the relative intensities of TAZ, which were quantified with densitometry and normalized to GAPDH. Values shown are the mean ± standard deviation (SD) from three independent experiments. P<0.05 for SK-N-BE(2) compared with SK-N-SH. (B) SK-N-BE(2) cells had a higher migratory ability than that of SK-N-SH cells, as determined by Transwell® assays. Graph shows the relative fold changes in cell migration compared with SK-N-SH cells. Values shown are the mean ± SD from three independent experiments. Twelve images were randomly captured in each independent experiment. (C) SK-N-BE(2) cells had a greater invasive ability compared with that of SK-N-SH cells, as determined by Transwell® assays. Experiments were performed as described for B. (D) Expression levels of Vimentin, E-cadherin and β-catenin in SK-N-SH and SK-N-BE(2) cells.

Mentions: The SK-N-BE(2) cell line is a tumorigenic, aggressive and MYCN gene amplified neuroblastoma cell line. The TAZ mRNA and protein expression levels in SK-N-BE(2) cells were significantly higher than those in SK-N-SH cells, which are less aggressive and MYCN gene non-amplified (Fig. 1A). The migration and invasion abilities of SK-N-BE(2) cells, analyzed via Transwell® migration and Matrigel™ invasion assays, were higher than those in the SK-N-SH cells (Fig. 1B and C). These results indicated that TAZ may have a role in neuroblastoma cell migration and invasion. Subsequently, the EMT protein expression levels in the two cell lines were evaluated using western blot analysis. The results revealed that the mesenchymal marker Vimentin was upregulated in the SK-N-BE(2) cells compared with the levels in the SK-N-SH cells; furthermore, the expression levels of the epithelial markers E-cadherin and β-catenin were downregulated in the SK-N-BE(2) cells compared with those of the SK-N-SH cells (Fig. 1D).


TAZ promotes epithelial to mesenchymal transition via the upregulation of connective tissue growth factor expression in neuroblastoma cells.

Wang Q, Xu Z, An Q, Jiang D, Wang L, Liang B, Li Z - Mol Med Rep (2014)

Migratory and invasive properties of SK-N-BE(2) cells expressing high levels of TAZ. (A) The mRNA and protein expression levels of TAZ were higher in SK-N-BE(2) cells compared with those observed in SK-N-SH cells. Top panel: results of reverse transcription quantitative polymerase chain reaction presented as a western blot from a representative study; bottom panel: graph of the relative intensities of TAZ, which were quantified with densitometry and normalized to GAPDH. Values shown are the mean ± standard deviation (SD) from three independent experiments. P<0.05 for SK-N-BE(2) compared with SK-N-SH. (B) SK-N-BE(2) cells had a higher migratory ability than that of SK-N-SH cells, as determined by Transwell® assays. Graph shows the relative fold changes in cell migration compared with SK-N-SH cells. Values shown are the mean ± SD from three independent experiments. Twelve images were randomly captured in each independent experiment. (C) SK-N-BE(2) cells had a greater invasive ability compared with that of SK-N-SH cells, as determined by Transwell® assays. Experiments were performed as described for B. (D) Expression levels of Vimentin, E-cadherin and β-catenin in SK-N-SH and SK-N-BE(2) cells.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4262480&req=5

f1-mmr-11-02-0982: Migratory and invasive properties of SK-N-BE(2) cells expressing high levels of TAZ. (A) The mRNA and protein expression levels of TAZ were higher in SK-N-BE(2) cells compared with those observed in SK-N-SH cells. Top panel: results of reverse transcription quantitative polymerase chain reaction presented as a western blot from a representative study; bottom panel: graph of the relative intensities of TAZ, which were quantified with densitometry and normalized to GAPDH. Values shown are the mean ± standard deviation (SD) from three independent experiments. P<0.05 for SK-N-BE(2) compared with SK-N-SH. (B) SK-N-BE(2) cells had a higher migratory ability than that of SK-N-SH cells, as determined by Transwell® assays. Graph shows the relative fold changes in cell migration compared with SK-N-SH cells. Values shown are the mean ± SD from three independent experiments. Twelve images were randomly captured in each independent experiment. (C) SK-N-BE(2) cells had a greater invasive ability compared with that of SK-N-SH cells, as determined by Transwell® assays. Experiments were performed as described for B. (D) Expression levels of Vimentin, E-cadherin and β-catenin in SK-N-SH and SK-N-BE(2) cells.
Mentions: The SK-N-BE(2) cell line is a tumorigenic, aggressive and MYCN gene amplified neuroblastoma cell line. The TAZ mRNA and protein expression levels in SK-N-BE(2) cells were significantly higher than those in SK-N-SH cells, which are less aggressive and MYCN gene non-amplified (Fig. 1A). The migration and invasion abilities of SK-N-BE(2) cells, analyzed via Transwell® migration and Matrigel™ invasion assays, were higher than those in the SK-N-SH cells (Fig. 1B and C). These results indicated that TAZ may have a role in neuroblastoma cell migration and invasion. Subsequently, the EMT protein expression levels in the two cell lines were evaluated using western blot analysis. The results revealed that the mesenchymal marker Vimentin was upregulated in the SK-N-BE(2) cells compared with the levels in the SK-N-SH cells; furthermore, the expression levels of the epithelial markers E-cadherin and β-catenin were downregulated in the SK-N-BE(2) cells compared with those of the SK-N-SH cells (Fig. 1D).

Bottom Line: The Hippo signaling pathway regulates cell proliferation and apoptosis, and its primary downstream effectors are TAZ and yes‑associated protein 1 (YAP).Repressed expression of TAZ in SK‑N‑BE(2) cells was shown to result in a reduction in aggressiveness of the cell line, by Transwell migration and invasion assay.Furthermore, TAZ was shown by luciferase assay to induce CTGF expression by modulating the activation of the TGF‑β/Smad3 signaling pathway.

View Article: PubMed Central - PubMed

Affiliation: Department of Pediatric Surgery, The First Affiliated Hospital of Harbin Medical University, Harbin, Heilongjiang 150001, P.R. China.

ABSTRACT
Neuroblastoma (NB) is a neuroendocrine cancer that occurs most commonly in infants and young children. The Hippo signaling pathway regulates cell proliferation and apoptosis, and its primary downstream effectors are TAZ and yes‑associated protein 1 (YAP). The effect of TAZ on the metastatic progression of neuroblastoma and the underlying mechanisms involved remain elusive. In the current study, it was determined by western blot analysis that the migratory and invasive properties of SK‑N‑BE(2) human neuroblastoma cells are associated with high expression levels of TAZ. Repressed expression of TAZ in SK‑N‑BE(2) cells was shown to result in a reduction in aggressiveness of the cell line, by Transwell migration and invasion assay. In contrast, overexpression of TAZ in SK‑N‑SH human neuroblastoma cells was shown by Transwell migration and invasion assays, and western blot analysis, to result in epithelial‑mesenchymal transition (EMT) and increased invasiveness. Mechanistically, the overexpression of TAZ was demonstrated to upregulate the expression levels of connective tissue growth factor (CTGF), by western blot analysis and chromatin immunoprecipitation assay, while the knockdown of TAZ downregulated it. Furthermore, TAZ was shown by luciferase assay to induce CTGF expression by modulating the activation of the TGF‑β/Smad3 signaling pathway. In conclusion, the present study is, to the best of our knowledge, the first to demonstrate that the overexpression of TAZ induces EMT, increasing the invasive abilities of neuroblastoma cells. This suggests that TAZ may serve as a potential target in the development of novel therapies for the treatment of neuroblastoma.

Show MeSH
Related in: MedlinePlus