Objective While oesophageal squamous cell carcinoma remains infrequent in Western populations, the incidence of oesophageal adenocarcinoma (EAC) has increased sixfold to eightfold over the past four decades. We aimed to characterise oesophageal cancer-specific and subtypes-specific gene regulation patterns and their upstream transcription factors (TFs).
Design To identify regulatory elements, we profiled fresh-frozen oesophageal normal samples, tumours and cell lines with chromatin immunoprecipitation sequencing (ChIP-Seq). Mathematical modelling was performed to establish (super)-enhancers landscapes and interconnected transcriptional circuitry formed by master TFs. Coregulation and cooperation between master TFs were investigated by ChIP-Seq, circularised chromosome conformation capture sequencing and luciferase assay. Biological functions of candidate factors were evaluated both in vitro and in vivo.
Results We found widespread and pervasive alterations of the (super)-enhancer reservoir in both subtypes of oesophageal cancer, leading to transcriptional activation of a myriad of novel oncogenes and signalling pathways, some of which may be exploited pharmacologically (eg, leukemia inhibitory factor (LIF) pathway). Focusing on EAC, we bioinformatically reconstructed and functionally validated an interconnected circuitry formed by four master TFs—ELF3, KLF5, GATA6 and EHF—which promoted each other’s expression by interacting with each super-enhancer. Downstream, these master TFs occupied almost all EAC super-enhancers and cooperatively orchestrated EAC transcriptome. Each TF within the transcriptional circuitry was highly and specifically expressed in EAC and functionally promoted EAC cell proliferation and survival.
Conclusions By establishing cancer-specific and subtype-specific features of the EAC epigenome, our findings promise to transform understanding of the transcriptional dysregulation and addiction of EAC, while providing molecular clues to develop novel therapeutic modalities against this malignancy.
- transcription factor
- gene regulation
- signal transduction
- oesophageal cancer
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LC, MH, JP, JP and YYJ contributed equally.
Contributors D-CL conceived and devised the study. D-CL, LC and MH designed the experiments and analysis. LC, JaP, JiP, YYJ, SC, WDL, JWS and ND performed the experiments. MH, TCS, OA, QY, HY and BB performed the bioinformatics and statistical analysis. YC, WNMD, KW, RT, SG, SJK and SJM contributed reagents and materials. LC, MH, JaP, D-CL and HPK analysed the data. D-CL and HPK supervised the research and wrote the manuscript.
Funding This research is supported by the National Research Foundation Singapore under its Singapore Translational Research (STaR) Investigator Award (NMRC/STaR/0021/2014) and administered by the Singapore Ministry of Health’s National Medical Research Council (NMRC), the NMRC Centre Grant awarded to National University Cancer Institute of Singapore, the National Research Foundation Singapore and the Singapore Ministry of Education under its Research Centres of Excellence initiatives (to HPK). This research is additionally supported by the RNA Biology Center at the Cancer Science Institute of Singapore, NUS, as part of funding under the Singapore Ministry of Education’s Tier 3 grants (grant number MOE2014-T3-1-006). SJM is supported by the Emerson Research Foundation and NIH grants DK118250, CA190040 and CA211457; he is also the Harry and Betty Myerberg Professor and American Cancer Society Clinical Research Professor. D-CL is supported by the DeGregorio Family Foundation, the Price Family Foundation as well as Samuel Oschin Comprehensive Cancer Institute (SOCCI) at Cedars-Sinai Medical Center through the Translational Pipeline Discovery Fund; he is member of UCLA Jonsson Comprehensive Cancer Center, UCLA Molecular Biology Institute as well as UCLA Cure: Digestive Diseases Research Center. SJK is supported by the Howard H Hall fund for oesophageal cancer research.
Competing interests None declared.
Patient consent for publication Obtained.
Provenance and peer review Not commissioned; externally peer reviewed.
Data availability statement Data are available in a public, open access repository.
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