Global analysis of DNA methylation in hepatocellular carcinoma by a liquid hybridization capture-based bisulfite sequencing approach.

Fei Gao,Tong Wang,Hanlin Lu,Yu Yao,Arian Laurence,Zhimei Yuan,Junwen Wang,Meng Xia,Qian Chen,Huifang Liang,Ming Zhou,Xiaoping Chen,Xiaolong Tan,Jingjing Yu,Wei Chen,Hua Xu,Wei Xiao,Xiuqing Zhang

doi:10.1186/s13148-015-0121-1

Abstract

BackgroundEpigenetic alterations, such as aberrant DNA methylation of promoter and enhancer regions, which lead to atypical gene expression, have been associated with carcinogenesis. In hepatocellular carcinoma (HCC), genome-wide analysis of methylation has only recently been used. For a better understanding of hepatocarcinogenesis, we applied an even higher resolution analysis of the promoter methylome to identify previously unknown regions and genes differentially methylated in HCC.ResultsOptimized liquid hybridization capture-based bisulfite sequencing (LHC-BS) was developed to quantitatively analyze 1.86 million CpG sites in individual samples from eight pairs of HCC and adjacent tissues. By linking the differentially methylated regions (DMRs) in promoters to the differentially expressed genes (DEGs), we identified 12 DMR-associated genes. We further utilized Illumina MiSeq combining the bisulfite sequencing PCR approach to validate the 12 candidate genes. Analysis of an additional 78 HCC pairs on the Illumina MiSeq platform confirmed that 7 genes showed either promoter hyper-methylation (SMAD6, IFITM1, LRRC4, CHST4, and TBX15) or hypo-methylation (CCL20 and NQO1) in HCC.ConclusionsNovel methylome profiling provides a cost-efficient approach to identifying candidate genes in human HCC that may contribute to hepatocarcinogenesis. Our work provides further information critical for understanding the epigenetic processes underlying tumorigenesis and development of HCC.Electronic supplementary materialThe online version of this article (doi:10.1186/s13148-015-0121-1) contains supplementary material, which is available to authorized users.

Highlights

Epigenetic alterations, such as aberrant DNA methylation of promoter and enhancer regions, which lead to atypical gene expression, have been associated with carcinogenesis
The promoter methylome differentiates tumor tissue from adjacent non-tumor tissue in hepatocellular carcinoma (HCC) The clinicopathologic features of the 8 patients with HCC in this promoter-wide methylation study are described in Additional file 1: Table S1
Promoters were denoted as regions from −2200 bp to +500 bp of the transcriptional start sites (TSS) [16]

Summary

Introduction

Epigenetic alterations, such as aberrant DNA methylation of promoter and enhancer regions, which lead to atypical gene expression, have been associated with carcinogenesis. In hepatocellular carcinoma (HCC), genome-wide analysis of methylation has only recently been used. For a better understanding of hepatocarcinogenesis, we applied an even higher resolution analysis of the promoter methylome to identify previously unknown regions and genes differentially methylated in HCC. The development of HCC is a multistep process characterized by the accumulation of genetic mutations and epigenetic aberrations. Epigenetic alterations such as aberrant methylation and histone modification occur far more frequently than genetic mutations in cancers and can significantly affect the efficacy of messenger RNA (mRNA) synthesis without changing the primary DNA sequence [2].

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Discussion

Conclusion