Activation of β-catenin and Yap1 in human hepatoblastoma and induction of hepatocarcinogenesis in mice.
OBJECTIVE
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
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
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
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.
Department of Bioengineering and Therapeutic Sciences, University of California, San Francisco, CA.
Junyan
J
Tao
Institute of Pathology, University of Greifswald, Greifswald, Germany.
Diego F
DF
Calvisi
Department of Pathology, University of Pittsburgh, School of Medicine, Pittsburgh, PA.
Sarangarajan
S
Ranganathan
Institute of Pathology, University of Greifswald, Greifswald, Germany.
Antonio
A
Cigliano
Department of Pathology, University of Pittsburgh, School of Medicine, Pittsburgh, PA.|The Second Affiliated Hospital, School of Medicine, Xi'an Jiaotong University, Xi'an, P. R. China.
Lili
L
Zhou
Department of Pathology, University of Pittsburgh, School of Medicine, Pittsburgh, PA.
Sucha
S
Singh
Department of Bioengineering and Therapeutic Sciences, University of California, San Francisco, CA.
Lijie
L
Jiang
Department of Bioengineering and Therapeutic Sciences, University of California, San Francisco, CA.
Biao
B
Fan
Institute of Pathology, University Hospital Basel, Basel, Switzerland.
Luigi
L
Terracciano
Department of Pediatric Surgery and Pediatric Urology, Children's University Hospital Tuebingen, Tuebingen, Germany.
Sorin
S
Armeanu-Ebinger
Institute of Pathology, University of Greifswald, Greifswald, Germany.
Silvia
S
Ribback
Institute of Pathology, University of Greifswald, Greifswald, Germany.
Frank
F
Dombrowski
Institute of Pathology, University of Greifswald, Greifswald, Germany.
Matthias
M
Evert
Department of Bioengineering and Therapeutic Sciences, University of California, San Francisco, CA.|Liver Center, University of California, San Francisco, CA.
Xin
X
Chen
Department of Pathology, University of Pittsburgh, School of Medicine, Pittsburgh, PA.
Satdarshan P S
SPS
Monga
D048868:Adaptor Proteins, Signal Transducing / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001650:Bile Duct Neoplasms / Q000378:metabolism / Q000473:pathology; D001653:Bile Ducts, Intrahepatic / Q000378:metabolism / Q000473:pathology; D006528:Carcinoma, Hepatocellular / Q000378:metabolism / Q000473:pathology; D016923:Cell Death; D049109:Cell Proliferation; D002471:Cell Transformation, Neoplastic / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D018281:Cholangiocarcinoma / Q000378:metabolism / Q000473:pathology; D005060:Europe; D015972:Gene Expression Regulation, Neoplastic; D056945:Hep G2 Cells; D018197:Hepatoblastoma / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D006801:Humans; D008113:Liver Neoplasms / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D051379:Mice; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D034622:RNA Interference; D015398:Signal Transduction; D013997:Time Factors; D014157:Transcription Factors; D014158:Transcription, Genetic; D014162:Transfection; D014481:United States; D000091102:YAP-Signaling Proteins; D051176:beta Catenin / Q000235:genetics / Q000378:metabolism*
D048868:Adaptor Proteins, Signal Transducing; C495270:CTNNB1 protein, human; D010750:Phosphoproteins; D014157:Transcription Factors; D000091102:YAP-Signaling Proteins; C088374:YAP1 protein, human; D051176:beta Catenin
Gene Regulation; Liver Cancer; Oncogene; Tumor Formation