Cell cycle-related kinase mediates viral-host signalling to promote hepatitis B virus-associated hepatocarcinogenesis.
BACKGROUND
Androgen receptor (AR) signalling contributes to male predominance in hepatocellular carcinoma (HCC), which is more pronounced in HBV-endemic areas. Cell cycle-related kinase (CCRK) is essential for AR-induced hepatocarcinogenesis but its molecular function in HBV-associated HCC remains obscure.


OBJECTIVE
To determine the molecular function of CCRK in HBV-associated HCC.


METHODS
Transcriptional regulation was assessed by chromatin immunoprecipitation, promoter mutation and luciferase reporter assays. Hepatocellular proliferation and tumourigenesis were examined by colony formation, soft agar assays and using HBV X protein (HBx) transgenic mice with low-dose exposure to diethylnitrosamine. Protein expressions were examined in clinical samples and correlated with patient survival by log-rank Mantel-Cox test.


RESULTS
Overexpression of CCRK, but not its kinase-defective mutant, activated β-catenin/T cell factor signalling through phosphorylation of glycogen synthase kinase-3β (GSK-3β) at Ser9, led to upregulation of AR transcriptional activity and, subsequently, expression of HBx. The viral transactivator in turn induced CCRK expression through enhanced AR signalling, thus forming a positive regulatory loop. RNA interference silencing of CCRK, which suppressed the CCRK/GSK-3β/β-catenin/AR regulatory loop, significantly suppressed HBx-induced hepatocellular proliferation (p=0.001) and transformation (p<0.001) and remarkably reduced >80% diethylnitrosamine-mediated hepatocarcinogenesis in HBx transgenic mice. Finally, patients with HBV-associated HCC with concordant overexpression of CCRK, GSK-3β phosphorylation at Ser9, active dephosphorylated β-catenin and AR phosphorylation at Ser81 had poorer overall (HR=31.26, p<0.0001) and disease-free (HR=3.60, p<0.01) survival rates.


CONCLUSIONS
Our findings highlight the critical role of CCRK in a self-reinforcing circuitry that regulates HBV-associated hepatocarcinogenesis. Further characterisation of this intricate viral-host signalling may provide new prognostic biomarkers and therapeutic targets for HCC treatment.
Institute of Digestive Disease, The Chinese University of Hong Kong, Hong Kong SAR, P. R. China Department of Liver Disease, Shuguang Hospital, Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai, P. R. China.
Zhuo
Z
Yu
Department of Liver Disease, Shuguang Hospital, Affiliated to Shanghai University of Traditional Chinese Medicine, Shanghai, P. R. China.
Yue-Qiu
YQ
Gao
School of Biomedical Sciences, The Chinese University of Hong Kong, Hong Kong SAR, P. R. China.
Hai
H
Feng
Department of Medicine and Therapeutics, The Chinese University of Hong Kong, Hong Kong SAR, P. R. China.
Ying-Ying
YY
Lee
School of Biomedical Sciences, The Chinese University of Hong Kong, Hong Kong SAR, P. R. China.
May S
MS
Li
Department of Medicine and Therapeutics, The Chinese University of Hong Kong, Hong Kong SAR, P. R. China.
Yuan
Y
Tian
Department of Medicine and Therapeutics, The Chinese University of Hong Kong, Hong Kong SAR, P. R. China.
Minnie Y Y
MY
Go
Disease Model Research Laboratory, Aging Research Center and World Class Institute, Korea Research Institute of Bioscience and Biotechnology, Daejeon, Republic of Korea.
Dae-Yeul
DY
Yu
Department of Surgery, The Chinese University of Hong Kong, Hong Kong SAR, P. R. China.
Yue-Sun
YS
Cheung
Institute of Digestive Disease, The Chinese University of Hong Kong, Hong Kong SAR, P. R. China Department of Surgery, The Chinese University of Hong Kong, Hong Kong SAR, P. R. China.
Paul B S
PB
Lai
Institute of Digestive Disease, The Chinese University of Hong Kong, Hong Kong SAR, P. R. China Department of Medicine and Therapeutics, The Chinese University of Hong Kong, Hong Kong SAR, P. R. China State Key Laboratory of Digestive Disease, The Chinese University of Hong Kong, Hong Kong SAR, P. R. China.
Jun
J
Yu
Institute of Digestive Disease, The Chinese University of Hong Kong, Hong Kong SAR, P. R. China Department of Medicine and Therapeutics, The Chinese University of Hong Kong, Hong Kong SAR, P. R. China State Key Laboratory of Digestive Disease, The Chinese University of Hong Kong, Hong Kong SAR, P. R. China.
Vincent W S
VW
Wong
Institute of Digestive Disease, The Chinese University of Hong Kong, Hong Kong SAR, P. R. China State Key Laboratory of Digestive Disease, The Chinese University of Hong Kong, Hong Kong SAR, P. R. China.
Joseph J Y
JJ
Sung
Institute of Digestive Disease, The Chinese University of Hong Kong, Hong Kong SAR, P. R. China Department of Medicine and Therapeutics, The Chinese University of Hong Kong, Hong Kong SAR, P. R. China State Key Laboratory of Digestive Disease, The Chinese University of Hong Kong, Hong Kong SAR, P. R. China.
Henry L Y
HL
Chan
School of Biomedical Sciences, The Chinese University of Hong Kong, Hong Kong SAR, P. R. China State Key Laboratory of Digestive Disease, The Chinese University of Hong Kong, Hong Kong SAR, P. R. China.
Alfred S L
AS
Cheng
D063646:Carcinogenesis; D006528:Carcinoma, Hepatocellular / Q000378:metabolism* / Q000821:virology; D002478:Cells, Cultured; D018844:Cyclin-Dependent Kinases / Q000096:biosynthesis*; D015972:Gene Expression Regulation, Neoplastic; D038362:Glycogen Synthase Kinase 3 / Q000378:metabolism; D000071679:Glycogen Synthase Kinase 3 beta; D006509:Hepatitis B / Q000150:complications*; D054884:Host-Pathogen Interactions; D006801:Humans; D008113:Liver Neoplasms / Q000378:metabolism* / Q000821:virology; D011379:Prognosis; D011944:Receptors, Androgen / Q000378:metabolism; D051820:TCF Transcription Factors / Q000378:metabolism; D015534:Trans-Activators / Q000378:metabolism; D015533:Transcriptional Activation; D015854:Up-Regulation; D054334:Viral Regulatory and Accessory Proteins; D051176:beta Catenin / Q000378:metabolism; D000095043:Cyclin-Dependent Kinase-Activating Kinase
D011944:Receptors, Androgen; D051820:TCF Transcription Factors; D015534:Trans-Activators; D054334:Viral Regulatory and Accessory Proteins; D051176:beta Catenin; C057628:hepatitis B virus X protein; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; C000605507:Gsk3b protein, mouse; D018844:Cyclin-Dependent Kinases; D038362:Glycogen Synthase Kinase 3; D000095043:Cyclin-Dependent Kinase-Activating Kinase; C000722413:CCRK protein, human
Cell Signalling; Gene Regulation; Hepatitis B; Hepatocellular Carcinoma; Molecular Carcinogenesis