Integrative epigenomic analysis of functional enhancers in liver carcinogenesis
Refereed conference paper presented and published in conference proceedings

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AbstractHepatocellular carcinoma (HCC) is a leading cause of cancer-related mortality worldwide, driven in part by late stage of diagnosis and therapy resistance. Accumulating evidence suggests that epigenetics plays a crucial role in converting the hostile immune and metabolic microenvironments into aberrant transcriptional activities, underscoring the fundamental roles of epigenetic regulation in HCC pathogenesis. Enhancer is a critical cis-regulatory element controlling cell-specific transcriptional programmes. Recent studies demonstrate that gene enhancers are frequently altered epigenetically in cancers, and deregulation of gene expression is much more associated with DNA methylation changes at enhancers than promoters. However, there are few comprehensive studies to date which focus on the deregulation of enhancer methylation. In this study, we focus on decoding the HCC methylome to identify enhancer epigenetic alterations that confer HCC malignant phenotypes. By applying whole genome bisulfite sequencing of human HCC samples, we have depicted the overall methylation profile of HCC. We further performed integrative analysis using enhancer database (FANTOM5) and identified 894 aberrantly methylated enhancers (854 are hypomethylated and 40 are hypermethylated) with target gene deregulation. Based on tissue-specificity and gene ontology data, we focused on a novel enhancer which targets a liver-specific transcription factor CCAAT/enhancer-binding protein beta (CEBPB). Quantitative pyrosequencing analysis confirmed that CEBPB enhancer was significantly hypomethylated in HCCs compared to adjacent liver tissues, which was associated with CEBPB over-expression. Importantly, patients with enhancer hypomethylation were associated with poor survival rates. Accumulating evidence suggests that enhancer RNA (eRNA), which belong to non-coding RNA family, emerges as potent regulators for chromatin modification and transcription regulation. We detected a 3 kb CEBPB eRNA, whose expression level was inversely correlated with enhancer methylation in 8 liver cell lines. Pharmacological demethylation using 5-aza-2-deoxycytidine significantly reduced CEBPB enhancer methylation and restored both CEBPB eRNA and target gene expression. Using CRISPR/Cas9 knockout of CEBPB enhancer or siRNA knockdown targeting CEBPB eRNA, we found both treatments resulted in the reduction in eRNA and mRNA expression, confirming its role on gene regulation. Functional analysis also confirmed that knockout of CEBPB enhancer led to a dramatic reduction of cell proliferative and invasive abilities, and nearly abolished the in vivo tumorigenicity. Luciferase assay showed that CEBPB was critical to its own enhancer transcription activity and mutation of CEBPB binding motif impaired such activity, suggesting a positive feedback regulatory circuit. Our study highlights the critical role of a novel aberrantly methylated enhancer in HCC, providing insights of eRNA in mediating the enhancer methylation and transcriptional enhancement during HCC carcinogenesis.
All Author(s) ListLei XIONG, Wei KANG, Qiong WU, Feng WU, Raymond W. LUNG, Ken H. YU, Sau-Dan LEE, Joseph LIU, Guangcun HUANG, Chiou-Miin WANG, Jian-Liang ZHOU, Michael W. CHAN, Nathalie WONG, Tim H. HUANG, Kevin Y. YIP, Alfred S. CHENG, Ka F. TO
Name of ConferenceGordon Research Conference on Cancer Genetics and Epigenetics
Start Date of Conference23/04/2017
End Date of Conference28/04/2017
Place of ConferenceLucca (Barga)
Country/Region of ConferenceItaly
LanguagesEnglish-United States

Last updated on 2018-17-05 at 12:36