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The Hippo/YAP pathway interacts with EGFR signaling and HPV oncoproteins to regulate cervical cancer progression.

He C, Mao D, Hua G, Lv X, Chen X, Angeletti PC, Dong J, Remmenga SW, Rodabaugh KJ, Zhou J, Lambert PF, Yang P, Davis JS, Wang C - EMBO Mol Med (2015)

Bottom Line: The Hippo signaling pathway controls organ size and tumorigenesis through a kinase cascade that inactivates Yes-associated protein (YAP).Our results suggest that YAP expression is associated with a poor prognosis for cervical cancer.Our study indicates that combined targeting of the Hippo and the ERBB signaling pathways represents a novel therapeutic strategy for prevention and treatment of cervical cancer.

View Article: PubMed Central - PubMed

Affiliation: Olson Center for Women's Health, Department of Obstetrics & Gynecology, University of Nebraska Medical Center, Omaha, NE, USA College of Animal Science and Technology, Huazhong Agricultural University, Wuhan, China.

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YAP regulated EGFR, AREG, and TGF-α expression in cervical cancer cellRT–PCR (left) and Western blot (right) were used to determine the expression of EGFR, AREG, and TGF-α in ME180-MXIV, ME180-YAP, and ME180-YAPS127A cells. GAPDH and actin were used as controls.The concentration of AREG in cell culture medium from ME180-MXIV, ME180-YAP, and ME180-YAPS127A cells. The concentration of AREG in the medium was determined by AREG ELISA kit. Each bar represents mean ± SEM (n = 5). Bars with different letters are significantly different from each other (ME-MX vs. ME-YAP, P < 0.0001; ME-MX vs. ME-YAPS127A, P < 0.0001).The concentrations of AREG in ME180 cells with or without siYAP treatment. ME180-CTRL and ME180-siRNA cells were incubated in the FBS 1% medium for 48 h. Each bar represents mean ± SEM (n = 5). Bars with different letters are significantly different from each other (P = 0.0006).Data information: Quantitative data in (B) were analyzed for significance using one-way ANOVA in GraphPad Prism 5 with Tukey’s post hoc tests. Data in (C) were analyzed for significance with unpaired t-test in GraphPad Prism 5 with Welch’s correction.
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fig14ev: YAP regulated EGFR, AREG, and TGF-α expression in cervical cancer cellRT–PCR (left) and Western blot (right) were used to determine the expression of EGFR, AREG, and TGF-α in ME180-MXIV, ME180-YAP, and ME180-YAPS127A cells. GAPDH and actin were used as controls.The concentration of AREG in cell culture medium from ME180-MXIV, ME180-YAP, and ME180-YAPS127A cells. The concentration of AREG in the medium was determined by AREG ELISA kit. Each bar represents mean ± SEM (n = 5). Bars with different letters are significantly different from each other (ME-MX vs. ME-YAP, P < 0.0001; ME-MX vs. ME-YAPS127A, P < 0.0001).The concentrations of AREG in ME180 cells with or without siYAP treatment. ME180-CTRL and ME180-siRNA cells were incubated in the FBS 1% medium for 48 h. Each bar represents mean ± SEM (n = 5). Bars with different letters are significantly different from each other (P = 0.0006).Data information: Quantitative data in (B) were analyzed for significance using one-way ANOVA in GraphPad Prism 5 with Tukey’s post hoc tests. Data in (C) were analyzed for significance with unpaired t-test in GraphPad Prism 5 with Welch’s correction.

Mentions: Amphiregulin (AREG), a known YAP target gene, was up-regulated by YAP in cervical cancer ME180 cells (Figs5A and EV2A). Most importantly, overexpression or constitutive activation of YAP induced significant increase in the secretion of AREG in the culture medium (FigEV2B). Knockdown of YAP significantly reduced AREG concentrations in the culture medium (FigEV2C). Interestingly, overexpression of wild-type YAP or constitutively active YAP also dramatically increased expression of TGF-α and EGFR mRNA (Figs5A and EV2A). This observation is supported by the RNA sequencing data extracted from TCGA datasets, in which we found that YAP expression is significantly correlated with TGF-α and EGFR expression in cervical cancer (P = 0.0009 and P = 0.0122, respectively, FigEV3). Consistent with our observations in ovarian granulosa cell tumors (Wang et al, 2012a), TGF-α significantly enhanced proliferation of ME180 cells (Fig5B) and promoted cell cycle progression (Fig5C). Treatment of ME180 cells with TGF-α for 24 h resulted in elongation of ME180 cells (Fig5D). In growth medium, control ME180 cells formed a monolayer upon reaching confluence and had a marked reduction in growth rate. TGF-α-treated ME180 cells, however, continued to grow even after the cells reached confluence, leading to the formation of multilayer cell plaques (Fig5D). Wound-healing assays showed that TGF-α (10 h) treatment dramatically induced wound closure in ME180 cells, indicating that TGF-α also induced cervical cancer cell migration (Fig5E).


The Hippo/YAP pathway interacts with EGFR signaling and HPV oncoproteins to regulate cervical cancer progression.

He C, Mao D, Hua G, Lv X, Chen X, Angeletti PC, Dong J, Remmenga SW, Rodabaugh KJ, Zhou J, Lambert PF, Yang P, Davis JS, Wang C - EMBO Mol Med (2015)

YAP regulated EGFR, AREG, and TGF-α expression in cervical cancer cellRT–PCR (left) and Western blot (right) were used to determine the expression of EGFR, AREG, and TGF-α in ME180-MXIV, ME180-YAP, and ME180-YAPS127A cells. GAPDH and actin were used as controls.The concentration of AREG in cell culture medium from ME180-MXIV, ME180-YAP, and ME180-YAPS127A cells. The concentration of AREG in the medium was determined by AREG ELISA kit. Each bar represents mean ± SEM (n = 5). Bars with different letters are significantly different from each other (ME-MX vs. ME-YAP, P < 0.0001; ME-MX vs. ME-YAPS127A, P < 0.0001).The concentrations of AREG in ME180 cells with or without siYAP treatment. ME180-CTRL and ME180-siRNA cells were incubated in the FBS 1% medium for 48 h. Each bar represents mean ± SEM (n = 5). Bars with different letters are significantly different from each other (P = 0.0006).Data information: Quantitative data in (B) were analyzed for significance using one-way ANOVA in GraphPad Prism 5 with Tukey’s post hoc tests. Data in (C) were analyzed for significance with unpaired t-test in GraphPad Prism 5 with Welch’s correction.
© Copyright Policy - open-access
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4644376&req=5

fig14ev: YAP regulated EGFR, AREG, and TGF-α expression in cervical cancer cellRT–PCR (left) and Western blot (right) were used to determine the expression of EGFR, AREG, and TGF-α in ME180-MXIV, ME180-YAP, and ME180-YAPS127A cells. GAPDH and actin were used as controls.The concentration of AREG in cell culture medium from ME180-MXIV, ME180-YAP, and ME180-YAPS127A cells. The concentration of AREG in the medium was determined by AREG ELISA kit. Each bar represents mean ± SEM (n = 5). Bars with different letters are significantly different from each other (ME-MX vs. ME-YAP, P < 0.0001; ME-MX vs. ME-YAPS127A, P < 0.0001).The concentrations of AREG in ME180 cells with or without siYAP treatment. ME180-CTRL and ME180-siRNA cells were incubated in the FBS 1% medium for 48 h. Each bar represents mean ± SEM (n = 5). Bars with different letters are significantly different from each other (P = 0.0006).Data information: Quantitative data in (B) were analyzed for significance using one-way ANOVA in GraphPad Prism 5 with Tukey’s post hoc tests. Data in (C) were analyzed for significance with unpaired t-test in GraphPad Prism 5 with Welch’s correction.
Mentions: Amphiregulin (AREG), a known YAP target gene, was up-regulated by YAP in cervical cancer ME180 cells (Figs5A and EV2A). Most importantly, overexpression or constitutive activation of YAP induced significant increase in the secretion of AREG in the culture medium (FigEV2B). Knockdown of YAP significantly reduced AREG concentrations in the culture medium (FigEV2C). Interestingly, overexpression of wild-type YAP or constitutively active YAP also dramatically increased expression of TGF-α and EGFR mRNA (Figs5A and EV2A). This observation is supported by the RNA sequencing data extracted from TCGA datasets, in which we found that YAP expression is significantly correlated with TGF-α and EGFR expression in cervical cancer (P = 0.0009 and P = 0.0122, respectively, FigEV3). Consistent with our observations in ovarian granulosa cell tumors (Wang et al, 2012a), TGF-α significantly enhanced proliferation of ME180 cells (Fig5B) and promoted cell cycle progression (Fig5C). Treatment of ME180 cells with TGF-α for 24 h resulted in elongation of ME180 cells (Fig5D). In growth medium, control ME180 cells formed a monolayer upon reaching confluence and had a marked reduction in growth rate. TGF-α-treated ME180 cells, however, continued to grow even after the cells reached confluence, leading to the formation of multilayer cell plaques (Fig5D). Wound-healing assays showed that TGF-α (10 h) treatment dramatically induced wound closure in ME180 cells, indicating that TGF-α also induced cervical cancer cell migration (Fig5E).

Bottom Line: The Hippo signaling pathway controls organ size and tumorigenesis through a kinase cascade that inactivates Yes-associated protein (YAP).Our results suggest that YAP expression is associated with a poor prognosis for cervical cancer.Our study indicates that combined targeting of the Hippo and the ERBB signaling pathways represents a novel therapeutic strategy for prevention and treatment of cervical cancer.

View Article: PubMed Central - PubMed

Affiliation: Olson Center for Women's Health, Department of Obstetrics & Gynecology, University of Nebraska Medical Center, Omaha, NE, USA College of Animal Science and Technology, Huazhong Agricultural University, Wuhan, China.

Show MeSH
Related in: MedlinePlus