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Epigenetic inactivation of transforming growth factor‐β1 target gene HEYL , a novel tumor suppressor, is involved in the P53 ‐induced apoptotic pathway in hepatocellular carcinoma
Author(s) -
Kuo KungKai,
Jian ShuFang,
Li YiJin,
Wan ShiWei,
Weng ChingChieh,
Fang KuanTe,
Wu DengChyang,
Cheng KuangHung
Publication year - 2015
Publication title -
hepatology research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.123
H-Index - 75
eISSN - 1872-034X
pISSN - 1386-6346
DOI - 10.1111/hepr.12414
Subject(s) - biology , microbiology and biotechnology , cancer research , carcinogenesis , gene , biochemistry
Aim Hairy/enhancer‐of‐split related with YRPW motif‐like ( HEYL ) protein was first identified as a transcriptional repressor. It is a downstream gene of the N otch and transforming growth factor‐β pathways. Little is known about its role in the pathogenesis of hepatocellular carcinoma ( HCC ). Methods Eighty surgically resected paired HCC and adjacent non‐cancerous tissues were analyzed for HEYL expression by reverse transcription quantitative polymerase chain reaction ( RT – qPCR ) and immunohistochemistry ( IHC ). HCC cells were transfected with pHEYL ‐ EGFP vector to overexpress the HEYL gene or infected with specific sh HEYL lentiviral vector to silence HEYL gene expression. HEYL expressional analysis and functional characterization were assessed by 3‐(4 5‐dimethylthiazol‐2‐yl)‐2 5‐diphenyltetrazolium bromide assays, flow cytometry, RT – qPCR , western blotting and methylation‐specific PCR . Results We determined that HEYL expression was inactivated in more than 75% of HCC . In addition, overexpression of HEYL in SK ‐Hep 1 cells caused apoptosis by the cleavage of caspase 3 and poly ( ADP ‐ribose) polymerase. We discovered that HEYL apoptosis was preceded by serine 15 phosphorylation and accumulation of P53 . Molecular analysis revealed that HEYL overexpression led to increased p16, p19, p21, p27 and Bad protein expression, and reduced c‐ Myc , Bcl ‐2 and C yclin B1 expression. Epigenetic silencing of HEYL expression by DNA hypermethylation in HCC directly correlated with loss of HEYL expression in HCC . Conclusion HEYL is frequently downregulated by promoter methylation in HCC . HEYL may be a tumor suppressor of liver carcinogenesis through upregulation of P53 gene expression and activation of P53 ‐mediated apoptosis.