Epigenetic dysfunction is a known contributor in carcinogenesis, and is emerging as a mechanism involved in toxicant-induced malignant transformation for environmental carcinogens such as arsenicals. In addition to aberrant DNA methylation of single genes, another manifestation of epigenetic dysfunction in cancer is agglomerative DNA methylation, which can participate in long-range epigenetic silencing that targets many neighboring genes and has been shown to occur in several types of clinical cancers. Using in vitro model systems of toxicant-induced malignant transformation, we found hundreds of aberrant DNA methylation events that emerge during malignant transformation, some of which occur in an agglomerative fashion. In an arsenitetransformed prostate epithelial cell line, the protocadherin (PCDH), HOXC and HOXD gene family clusters are targeted for agglomerative DNA methylation. Aberrant DNA methylation in general occurred more often within H3K27me3 stem cell domains. We found a striking association between enrichment of H3K9me3 stem cell domains and toxicant-induced agglomerative DNA methylation. Global gene expression profiling of the arsenite-transformed prostate epithelial cells showed that gene expression changes and DNA methylation changes were negatively correlated, but less than 10% of the hypermethylated genes were down-regulated. These studies confirm that a majority of the DNA hypermethylation events occur at transcriptionally repressed, H3K27me3 marked genes. In contrast to aberrant DNA methylation targeting H3K27me3 premarked silent genes, we found that actively expressed ZNF genes marked with H3K9me3 on their 3’ ends, are preferred targets of DNA methylation linked gene silencing.
