Activation of β-Catenin and Yap1 in Human Hepatoblastoma and Induction of Hepatocarcinogenesis in Mice

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• 作者：Junyan Tao¹ ; ^&lowast ; ; Diego F. Calvisi² ; ^&lowast ; ; Sarangarajan Ranganathan³ ; ^&lowast ; ; Antonio Cigliano² ; Lili Zhou³ ; ⁴ ; Sucha Singh³ ; Lijie Jiang¹ ; Biao Fan¹ ; Luigi Terracciano⁵ ; Sorin Armeanu&ndash ; Ebinger⁶ ; Silvia Ribback² ; Frank Dombrowski² ; Matthias Evert² ; Xin Chen¹ ; ⁷ ; ^{xin.chen@ucsf.edu" class="auth_mail" title="E-mail the corresponding author} ; Satdarshan P.S. Monga³ ; ^{smonga@pitt.edu" class="auth_mail" title="E-mail the corresponding author}
• 关键词：Liver Cancer ; Gene Regulation ; Oncogene ; Tumor Formation
• 刊名：Gastroenterology
• 出版年：2014
• 出版时间：September 2014
• 年：2014
• 卷：147
• 期：3
• 页码：690-701
• 全文大小：6611 K

文摘

Aberrant activation of β-catenin and Yes-associated protein 1 (Yap1) signaling pathways have been associated with the development of multiple tumor types. Yap functions as a transcriptional coactivator by interacting with TEA domain DNA binding proteins. We investigated the interactions among these pathways during hepatic tumorigenesis.

Methods

ra0015">We used immunohistochemical analysis to determine expression of β-catenin and Yap1 in liver cancer specimens collected from patients in Europe and the United States, consisting of 104 hepatocellular carcinoma, 62 intrahepatic cholangiocarcinoma, and 94 hepatoblastoma samples. We assessed β-catenin and Yap1 signaling and interactions in hepatoblastoma cell lines ((HuH6, HepG2, HepT1, HC-AFW1, HepG2, and HC-AFW1); proteins were knocked down with small interfering RNAs, and effects on proliferation and cell death were measured. Sleeping beauty–mediated hydrodynamic transfection was used to overexpress constitutively active forms of β-catenin (ΔN90/β-catenin) and Yap1 (YapS127A) in livers of mice; tissues were collected, and histological and immunohistochemical analyses were performed.

Results

ra0020">We observed nuclear localization of β-catenin and Yap1 in 79% of hepatoblastoma samples but not in most hepatocellular carcinoma or intrahepatic cholangiocarcinoma samples. Yap1 and β-catenin coprecipitated in hepatoblastoma but not hepatocellular carcinoma cells. Small interfering RNA–mediated knockdown of Yap1 or β-catenin in hepatoblastoma cells reduced proliferation in an additive manner. Knockdown of Yap1 reduced its ability to coactivate transcription with β-catenin; β-catenin inhibitors inactivated Yap1. Overexpression of constitutively active forms of Yap1 and β-catenin in mouse liver led to rapid tumorigenesis, with 100% mortality by 11 weeks. Tumor cells expressed both proteins, and human hepatoblastoma cells expressed common targets of their 2 signaling pathways. Yap1 binding of TEA domain factors was required for tumorigenesis in mice.

Conclusions

ra0025">β-catenin and the transcriptional regulator Yap1 interact physically and are activated in most human hepatoblastoma tissues; overexpression of activated forms of these proteins in livers of mice leads to rapid tumor development. Further analysis of these mice will allow further studies of these pathways in hepatoblastoma pathogenesis and could lead to the identification of new therapeutic targets.