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Critical assessment of DNA adenine methylation in eukaryotes using quantitative deconvolution

The discovery of N6-methyldeoxyadenine (6mA) across eukaryotes led to a search for additional epigenetic mechanisms. However, some studies have highlighted confounding factors that challenge the prevalence of 6mA in eukaryotes. We developed a metagenomic method to quantitatively deconvolve 6mA events from a genomic DNA sample into species of interest, genomic regions, and sources of contamination. Applying this method, we observed high-resolution 6mA deposition in two protozoa. We found that commensal or soil bacteria explained the vast majority of 6mA in insect and plant samples. We found no evidence of high abundance of 6mA in Drosophila, Arabidopsis, or humans. Plasmids used for genetic manipulation, even those from Dam methyltransferase mutant Escherichia coli, could carry abundant 6mA, confounding the evaluation of candidate 6mA methyltransferases and demethylases. On the basis of this work, we advocate for a reassessment of 6mA in eukaryotes. Description Reassessment of DNA 6mA in eukaryotes Certain forms of chemical modifications to DNA play important roles across the kingdoms of life; some forms have been widely studied and others are relatively new. DNA N6-methyldeoxyadenosine (6mA), which was recently reported to be prevalent across eukaryotes, created excitement for a new dimension to study biology and diseases. However, some studies have highlighted confounding factors, and there is an active debate over 6mA in eukaryotes. Kong et al. describe a method for quantitative 6mA deconvolution and report that bacterial contamination explains the vast majority of 6mA in DNA samples from insects and plants; the method also found no evidence for high 6mA levels in humans (see the Perspective by Boulias and Greer). This work advocates for a reassessment of 6mA in eukaryotes and provides an actionable approach. —DJ A metagenomic method called 6mASCOPE is developed to help clarify how limited DNA adenine methylation may be in eukaryotes.