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Distinguishing active versus passive DNA demethylation using Illumina MethylationEPIC BeadChip microarrays

ABSTRACT: The 5-carbon position on cytosine nucleotides preceding guanines in genomic DNA (CpG) are common targets for DNA methylation (5mC). Genomic locations enriched for 5mC contribute to the regulation of chromatin structure and gene expression. While largely stable, massive waves of DNA demethylation are observed in key developmental windows in mammals. Additionally, DNA methylation is therapeutically targeted in cancer via DNA methyltransferase inhibition, in part to reverse abnormal silencing of tumor suppressor genes. DNA methylation removal can occur through both active and passive mechanisms. Ten-eleven translocation enzymes (TETs) oxidize 5mC in a stepwise manner to 5-hydroxymethylcytosine (5hmC), 5-formylcytosine (5fC), and 5-carboxylcytosine (5caC). These oxidized cytosine forms have reported function in gene regulation and also serve as intermediates in an active 5mC removal process that involves the base excision repair pathway. 5mC can also be removed passively through sequential cell divisions in the absence of DNA methylation maintenance. Distinguishing active versus passive 5mC removal on a genome-wide scale remains a challenge. In this chapter, we describe approaches that couple TET-assisted bisulfite (TAB) and oxidative bisulfite (OxBS) conversion to the Illumina MethylationEPIC BeadChIP (EPIC array) and show how these technologies can be used to distinguish active versus passive DNA demethylation. We also describe integrative bioinformatics pipelines to facilitate this analysis.

ORGANISM(S): Homo sapiens

PROVIDER: GSE145113 | GEO | 2021/01/31

REPOSITORIES: GEO

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