DNA methylation in liver tumorigenesis in fish from the environment

The link between environment, alteration in DNA methylation and cancer has been well established in humans; yet, it is under-studied in unsequenced non-model organisms. The occurrence of liver tumors in the flatfish dab collected at certain UK sampling sites exceeds 20%, yet the causative agents and the molecular mechanisms of tumor formation are not known, especially regarding the balance between epigenetic and genetic factors. Methylated DNA Immunoprecipitation (MeDIP) combined with de novo high-throughput DNA sequencing were used to investigate DNA methylation changes in dab hepatocellular adenoma tumors for the first time in an unsequenced species. Novel custom-made dab gene expression arrays were designed and used to determine the relationship between DNA methylation and gene expression. In addition, the confirmatory techniques of bisulfite sequencing PCR (BSP) and RT-PCR were applied. Genes involved in pathways related to cancer, including apoptosis, wnt/β-catenin signaling and genomic and non-genomic estrogen responses, were altered both in methylation and transcription. Global methylation was statistically significantly 1.8-fold reduced in hepatocellular adenoma and non-cancerous surrounding tissues compared with liver from non-cancer bearing dab. Based on the identified changes and chemical exposure data, our study supports the epigenetic model of cancer. We hypothesize that chronic exposure to a mixture of environmental contaminants contributes to a global hypomethylation followed by further epigenetic and genomic changes. The findings suggest a link between environment, epigenetics and cancer in fish tumors in the wild and show the utility of this methodology for studies in non-model organisms.