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Twist-mediated Epithelial-mesenchymal Transition Promotes Breast Tumor Cell Invasion via Inhibition of Hippo Pathway.

Wang Y, Liu J, Ying X, Lin PC, Zhou BP - Sci Rep (2016)

Bottom Line: However, the mechanisms that facilitate the functions of Twist remain unclear.Here we report that Twist overexpression increased expression of PAR1, an upstream regulator of the Hippo pathway; PAR1 promotes invasion, migration, and CSC-like properties in breast cancer by activating the transcriptional co-activator TAZ.Our study indicates that Hippo pathway inhibition is required for the increased migratory and invasiveness ability of breast cancer cells in Twist-mediated EMT.

View Article: PubMed Central - PubMed

Affiliation: Cancer Institute of Integrative Medicine, Zhejiang Academy of Chinese Medicine, Hangzhou, Zhejiang, 310007, China.

ABSTRACT
Twist is a key transcription factor for Epithelial-mesenchymal transition (EMT), which is a cellular de-differentiation program that promotes invasion and metastasis, confers tumor cells with cancer stem cell (CSC)-like characteristics, and increases therapeutic resistance. However, the mechanisms that facilitate the functions of Twist remain unclear. Here we report that Twist overexpression increased expression of PAR1, an upstream regulator of the Hippo pathway; PAR1 promotes invasion, migration, and CSC-like properties in breast cancer by activating the transcriptional co-activator TAZ. Our study indicates that Hippo pathway inhibition is required for the increased migratory and invasiveness ability of breast cancer cells in Twist-mediated EMT.

No MeSH data available.


Related in: MedlinePlus

Overexpression of Snail or Twist induces EMT in T47D and MCF7 cells.(A) Representative images show expression of E-cadherin and ERα in Snail- or Twist-expressing T47D and MCF7 cells analyzed by immunofluorescent staining. Nuclei were visualized with DAPI staining (blue). The morphologic changes associated with EMT are shown in representative phase contrast images. Scale bars, 50 μm. (B) Expression of E-cadherin, N-cadherin and ERα in these cells was assessed by western blot analysis; actin served as a loading control. (C) Quantification of the relative mRNA levels of E-cadherin and ERα in Twist expressing T47D cells compared with vector-control cells using real-time PCR. Presented data are the mean ± SD from three separate experiments, with *and **indicate p < 0.01 in comparison with that of control.
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f2: Overexpression of Snail or Twist induces EMT in T47D and MCF7 cells.(A) Representative images show expression of E-cadherin and ERα in Snail- or Twist-expressing T47D and MCF7 cells analyzed by immunofluorescent staining. Nuclei were visualized with DAPI staining (blue). The morphologic changes associated with EMT are shown in representative phase contrast images. Scale bars, 50 μm. (B) Expression of E-cadherin, N-cadherin and ERα in these cells was assessed by western blot analysis; actin served as a loading control. (C) Quantification of the relative mRNA levels of E-cadherin and ERα in Twist expressing T47D cells compared with vector-control cells using real-time PCR. Presented data are the mean ± SD from three separate experiments, with *and **indicate p < 0.01 in comparison with that of control.

Mentions: To determine the role of Snail and Twist in EMT, we expressed Snail or Twist in immortalized human mammary epithelial cells (HMLE). Expression of Snail or Twist induced morphologic changes in HMLE cells, from a cobble-stone-like epithelial appearance to a spindle-shaped fibroblastic-like phenotype; these cells became elongated in shape and disassociated from their neighboring cells (Fig. 1A). Immunofluorescence staining showed downregulation of the epithelial marker E-cadherin, and upregulation of the mesenchymal marker Vimentin. Western blot analysis confirmed these results (Fig. 1B). We also expressed Snail or Twist in two luminal breast cancer cell lines, T47D and MCF7, that contain little endogenous Snail and Twist. Expression of Snail or Twist induced EMT in these cells, and converted the morphology of luminal cells to a basal-like phenotype (Fig. 2A). In addition, we found downregulation of the luminal epithelial markers E-cadherin and ERα, and the upregulation of the mesenchymal marker N-cadherin by immunofluorescence and western blot analysis (Fig. 2A,B). Long term (over 10 days) expression of Snail in T47D and MCF7 cells led to apoptosis in both cell lines (Table 1), and expression of Twist in MCF7 cells also led to apoptosis in this cell line. Interestingly, overexpression of Twist in T47D cells did not result in apoptosis, but led to the formation of a stable cell line with morphologic changes typical of EMT (Table 1). The mRNA levels of E-cadherin and ERα were dramatically decreased in this transformed cell line (Fig. 2C).


Twist-mediated Epithelial-mesenchymal Transition Promotes Breast Tumor Cell Invasion via Inhibition of Hippo Pathway.

Wang Y, Liu J, Ying X, Lin PC, Zhou BP - Sci Rep (2016)

Overexpression of Snail or Twist induces EMT in T47D and MCF7 cells.(A) Representative images show expression of E-cadherin and ERα in Snail- or Twist-expressing T47D and MCF7 cells analyzed by immunofluorescent staining. Nuclei were visualized with DAPI staining (blue). The morphologic changes associated with EMT are shown in representative phase contrast images. Scale bars, 50 μm. (B) Expression of E-cadherin, N-cadherin and ERα in these cells was assessed by western blot analysis; actin served as a loading control. (C) Quantification of the relative mRNA levels of E-cadherin and ERα in Twist expressing T47D cells compared with vector-control cells using real-time PCR. Presented data are the mean ± SD from three separate experiments, with *and **indicate p < 0.01 in comparison with that of control.
© Copyright Policy - open-access
Related In: Results  -  Collection

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f2: Overexpression of Snail or Twist induces EMT in T47D and MCF7 cells.(A) Representative images show expression of E-cadherin and ERα in Snail- or Twist-expressing T47D and MCF7 cells analyzed by immunofluorescent staining. Nuclei were visualized with DAPI staining (blue). The morphologic changes associated with EMT are shown in representative phase contrast images. Scale bars, 50 μm. (B) Expression of E-cadherin, N-cadherin and ERα in these cells was assessed by western blot analysis; actin served as a loading control. (C) Quantification of the relative mRNA levels of E-cadherin and ERα in Twist expressing T47D cells compared with vector-control cells using real-time PCR. Presented data are the mean ± SD from three separate experiments, with *and **indicate p < 0.01 in comparison with that of control.
Mentions: To determine the role of Snail and Twist in EMT, we expressed Snail or Twist in immortalized human mammary epithelial cells (HMLE). Expression of Snail or Twist induced morphologic changes in HMLE cells, from a cobble-stone-like epithelial appearance to a spindle-shaped fibroblastic-like phenotype; these cells became elongated in shape and disassociated from their neighboring cells (Fig. 1A). Immunofluorescence staining showed downregulation of the epithelial marker E-cadherin, and upregulation of the mesenchymal marker Vimentin. Western blot analysis confirmed these results (Fig. 1B). We also expressed Snail or Twist in two luminal breast cancer cell lines, T47D and MCF7, that contain little endogenous Snail and Twist. Expression of Snail or Twist induced EMT in these cells, and converted the morphology of luminal cells to a basal-like phenotype (Fig. 2A). In addition, we found downregulation of the luminal epithelial markers E-cadherin and ERα, and the upregulation of the mesenchymal marker N-cadherin by immunofluorescence and western blot analysis (Fig. 2A,B). Long term (over 10 days) expression of Snail in T47D and MCF7 cells led to apoptosis in both cell lines (Table 1), and expression of Twist in MCF7 cells also led to apoptosis in this cell line. Interestingly, overexpression of Twist in T47D cells did not result in apoptosis, but led to the formation of a stable cell line with morphologic changes typical of EMT (Table 1). The mRNA levels of E-cadherin and ERα were dramatically decreased in this transformed cell line (Fig. 2C).

Bottom Line: However, the mechanisms that facilitate the functions of Twist remain unclear.Here we report that Twist overexpression increased expression of PAR1, an upstream regulator of the Hippo pathway; PAR1 promotes invasion, migration, and CSC-like properties in breast cancer by activating the transcriptional co-activator TAZ.Our study indicates that Hippo pathway inhibition is required for the increased migratory and invasiveness ability of breast cancer cells in Twist-mediated EMT.

View Article: PubMed Central - PubMed

Affiliation: Cancer Institute of Integrative Medicine, Zhejiang Academy of Chinese Medicine, Hangzhou, Zhejiang, 310007, China.

ABSTRACT
Twist is a key transcription factor for Epithelial-mesenchymal transition (EMT), which is a cellular de-differentiation program that promotes invasion and metastasis, confers tumor cells with cancer stem cell (CSC)-like characteristics, and increases therapeutic resistance. However, the mechanisms that facilitate the functions of Twist remain unclear. Here we report that Twist overexpression increased expression of PAR1, an upstream regulator of the Hippo pathway; PAR1 promotes invasion, migration, and CSC-like properties in breast cancer by activating the transcriptional co-activator TAZ. Our study indicates that Hippo pathway inhibition is required for the increased migratory and invasiveness ability of breast cancer cells in Twist-mediated EMT.

No MeSH data available.


Related in: MedlinePlus