Genome-wide characterization of 5-hydoxymethylcytosine in melanoma reveals major differences with nevus

Melanoma demonstrates altered patterns of DNA methylation that are associated with genetic instability and transcriptional repression of numerous genes. Active DNA demethylation is mediated by TET enzymes that catalyze conversion of 5-methylcytosine (mC) to 5-hydroxymethylcytosine (hmC). Loss of hmC occurs in melanoma and correlates with disease progression. Here we analyzed the genomic distribution of hmC along with mC in nevus and melanoma using oxidative bisulfite chemistry combined with high-density arrays. HmC was enriched relative to mC at enhancers, 5'UTR regions and CpG shores in nevus and melanoma samples, pointing to specific TET enzyme activity. The proportion of interrogated CpG sites with high hmC levels was lower in melanoma (0.54%) than in nevus (2.0%). Depletion of hmC in melanoma was evident across all chromosomes and intragenic regions, being more pronounced in metastatic than in non-metastatic tumors. The patterns of hmC distribution in melanoma samples differed significantly from those in nevus samples, exceeding differences in mC patterns. We identified specific CpG sites and regions with significantly lower hmC levels in melanoma than in nevus that might serve as diagnostic markers. Differentially hydroxymethylated regions localized to cancer-related genes, including the PTEN gene promoter, suggesting that deregulated DNA hydroxymethylation may contribute to melanoma pathogenesis.