Abstract 63: Methylation status of histone H3 Lysine 4 in normal melanocytes is a major determinant of DNA hypermethylation in melanoma

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Abstract DNA hypermethylation in gene promoter CpG islands plays a crucial role in tumorigenesis. The determinants of the switch from unmethylated in normal to methylated in cancer remain poorly understood. It has been proposed that a bivalent chromatin domain (histone H3 Lysine 4 trimethylation (H3K4me3) and H3K27me3) in ES cells predisposes to aberrant DNA methylation, but this only accounts for a small fraction of the actual DNA hypermethylation. Here, we asked whether H3K4me3 alone affects DNA methylation changes in the tumorigenic process. To answer this question, we directly compared the epigenomes of melanoma cell lines LOXIMVI and SK-MEL-28 to normal epidermal melanocytes isolated from adults (HEMa) and neonates (HEMn). Briefly, we applied Digital Restriction Enzyme Analysis of Methylation (DREAM) to quantitatively map DNA methylation on a genome-wide scale, and ChIP-seq analysis to map the H3K4me3 pattern. Among the 5000∼5500 genes analyzed, promoter CpG island methylation (>50%) was seen in 1% in both HEMa and HEMn, compared to 17% and 6% in LOXIMVI and SK-MEL-28, respectively. H3K4me3 peaks were present in 67% and 75% of the genes in HEMa and HEMn, compared to 58% and 41% in LOXIMVI and SK-MEL-28. Defining a DNA methylation switch of a gene as going from below 20% methylation in normal to over 50% methylation in cancer cells, we found that the presence of H3K4me3 in normal cells was highly protective against such a switch when we compare the melanoma cell lines to normal cells. Taking the changes from HEMa to LOXIMVI as an example, 30% of the 1892 H3K4me3 negative genes in HEMa gained methylation in LOXIMVI, compared to only 8% of the 3603 H3K4me3-enriched genes (P<0.01). Strikingly, among these H3K4me3-enriched genes that gain DNA methylation, 87% lost H3K4me3 marks. Conversely, H3K4me3-enriched genes who lost the mark in cancer had higher chance of gaining DNA methylation than those who maintained it (72% vs. 0.4%). Thus, genes without H3K4me3 in normal cells are more likely to gain methylation in melanoma than genes with H3K4me3. To characterize further the relationship between normal and neoplastic DNA methylation, we took advantage of the highly quantitative nature of DREAM and divided genes into four groups with different DNA methylation levels in HEMa: 0-1%, 1-5%, 5-10% and 10-20%. The DNA methylation switch (to >50%) was seen in 13%, 43%, 40% and 50% of each of these groups. Consistent with the previous findings, in the normal cells 70% of the 0-1% methylated genes were H3K4me3 enriched, compared to 41%, 31%, and 40% of the other three groups, respectively. Taken together, we conclude that H3K4me3-enriched genes are relatively protected from DNA methylation. The presence of H3K4me3 is associated with lower levels of DNA methylation in normal cells and less chance of gaining methylation in cancer. Thus, the chromatin status in normal cells is a major determinant of aberrant DNA methylation in cancer. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 63. doi:10.1158/1538-7445.AM2011-63