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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

Knockdown of TAZ suppresses Twist-induced cell migration and invasion of T47D cells.(A) Effect of TAZ siRNA on cell migrating activity in Twist-overexpressing T47D cells and T47D cells expressing control vector using a wound healing assay. A scratch (“wound”) was inflicted to a cell layer produced 48 hours post-plating, and culture continued for an additional 24 hrs. Wound closures were photographed at 0 and 24 hr. Presented data are the mean ± SD from three independent experiments, with *and #indicating significant difference of p < 0.05 from control values. (B) Effect of TAZ siRNA on cell invasiveness in Twist-overexpressing T47D cells and T47D cells expressing control vector using a modified Boyden Chamber invasion assay as described in the Materials and Methods. Presented data are a graphic representation of the mean ± SD of percentage of invasive cells obtained from three separate experiments, with *and #indicating significant difference of p < 0.05 from control values. (C) Expression of Twist results in increased expression of PAR1, which promotes invasion, migration, and induces CSC-like properties in breast cancer cells by upregulating the expression of TAZ.
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f6: Knockdown of TAZ suppresses Twist-induced cell migration and invasion of T47D cells.(A) Effect of TAZ siRNA on cell migrating activity in Twist-overexpressing T47D cells and T47D cells expressing control vector using a wound healing assay. A scratch (“wound”) was inflicted to a cell layer produced 48 hours post-plating, and culture continued for an additional 24 hrs. Wound closures were photographed at 0 and 24 hr. Presented data are the mean ± SD from three independent experiments, with *and #indicating significant difference of p < 0.05 from control values. (B) Effect of TAZ siRNA on cell invasiveness in Twist-overexpressing T47D cells and T47D cells expressing control vector using a modified Boyden Chamber invasion assay as described in the Materials and Methods. Presented data are a graphic representation of the mean ± SD of percentage of invasive cells obtained from three separate experiments, with *and #indicating significant difference of p < 0.05 from control values. (C) Expression of Twist results in increased expression of PAR1, which promotes invasion, migration, and induces CSC-like properties in breast cancer cells by upregulating the expression of TAZ.

Mentions: If TAZ activation is crucial for the Twist-mediated EMT, depletion of TAZ should reverse the changes induced by Twist. To test this possibility, we employed a luciferase reporter assay to determine if TAZ is required for the Twist-mediated activation of CTGF promoter. We found that expression of Twist increased the promoter activity of CTGF, however, this effect was blocked by knocking down the expression of TAZ (Fig. 5C). Taken together, these results indicate that Twist expression activates TAZ, which leads to an increase in CTGF promoter transactivation as measured by luciferase activity. To further confirm the effect of TAZ, we knocked down the expression of TAZ by siTAZ in T47D-Twist cells and control vector cells (Fig. 5D), and performed wound healing and invasion analyses. We found that the migration induced by Twist in T47D cells was reduced from 3-fold to 1.3-fold (Fig. 6A), and that the invasion capability was reduced from 14-fold to 5-fold (Fig. 6B); these data indicate that Hippo pathway inhibition is required for the increased migratory and invasiveness in Twist-mediated EMT.


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)

Knockdown of TAZ suppresses Twist-induced cell migration and invasion of T47D cells.(A) Effect of TAZ siRNA on cell migrating activity in Twist-overexpressing T47D cells and T47D cells expressing control vector using a wound healing assay. A scratch (“wound”) was inflicted to a cell layer produced 48 hours post-plating, and culture continued for an additional 24 hrs. Wound closures were photographed at 0 and 24 hr. Presented data are the mean ± SD from three independent experiments, with *and #indicating significant difference of p < 0.05 from control values. (B) Effect of TAZ siRNA on cell invasiveness in Twist-overexpressing T47D cells and T47D cells expressing control vector using a modified Boyden Chamber invasion assay as described in the Materials and Methods. Presented data are a graphic representation of the mean ± SD of percentage of invasive cells obtained from three separate experiments, with *and #indicating significant difference of p < 0.05 from control values. (C) Expression of Twist results in increased expression of PAR1, which promotes invasion, migration, and induces CSC-like properties in breast cancer cells by upregulating the expression of TAZ.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4837350&req=5

f6: Knockdown of TAZ suppresses Twist-induced cell migration and invasion of T47D cells.(A) Effect of TAZ siRNA on cell migrating activity in Twist-overexpressing T47D cells and T47D cells expressing control vector using a wound healing assay. A scratch (“wound”) was inflicted to a cell layer produced 48 hours post-plating, and culture continued for an additional 24 hrs. Wound closures were photographed at 0 and 24 hr. Presented data are the mean ± SD from three independent experiments, with *and #indicating significant difference of p < 0.05 from control values. (B) Effect of TAZ siRNA on cell invasiveness in Twist-overexpressing T47D cells and T47D cells expressing control vector using a modified Boyden Chamber invasion assay as described in the Materials and Methods. Presented data are a graphic representation of the mean ± SD of percentage of invasive cells obtained from three separate experiments, with *and #indicating significant difference of p < 0.05 from control values. (C) Expression of Twist results in increased expression of PAR1, which promotes invasion, migration, and induces CSC-like properties in breast cancer cells by upregulating the expression of TAZ.
Mentions: If TAZ activation is crucial for the Twist-mediated EMT, depletion of TAZ should reverse the changes induced by Twist. To test this possibility, we employed a luciferase reporter assay to determine if TAZ is required for the Twist-mediated activation of CTGF promoter. We found that expression of Twist increased the promoter activity of CTGF, however, this effect was blocked by knocking down the expression of TAZ (Fig. 5C). Taken together, these results indicate that Twist expression activates TAZ, which leads to an increase in CTGF promoter transactivation as measured by luciferase activity. To further confirm the effect of TAZ, we knocked down the expression of TAZ by siTAZ in T47D-Twist cells and control vector cells (Fig. 5D), and performed wound healing and invasion analyses. We found that the migration induced by Twist in T47D cells was reduced from 3-fold to 1.3-fold (Fig. 6A), and that the invasion capability was reduced from 14-fold to 5-fold (Fig. 6B); these data indicate that Hippo pathway inhibition is required for the increased migratory and invasiveness in Twist-mediated EMT.

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