N6-methyladenine is incorporated into mammalian genome by DNA polymerase

Abstract

Dear Editor, DNA N 6-methyladenine (6mA), one of the most prevalent epigenetic base modifications in prokaryotes, 1 is recently found in multicellular eukaryotes. 2-8 This nucleobase may have epigenetic roles in regulation of retrotransposons, chromatin organization, and so on. 2-8 However, both our group 9 and Greer's group 10 noticed that eukaryotic DNA is easily contaminated with a minute of bacterial DNA, which carries overwhelmingly abundant 6mA (~2% 6mA/dA). 1 This brings great challenges for accurate detection of DNA 6mA in eukaryotes in terms of both sample pretreatments and analytical technologies. 9,10 For example, inconsistent with the report of Wu et al., 8 Schiffers et al. failed to detect 6mA above background levels in mouse embryonic stem (mES) cells using sensitive ultra-high-performance liquid chromatography-quadruple mass spectrometry (UHPLC-MS/MS) analysis. 11 To date, it is of intensive interest to seek conclusive evidence to support the prevalence of this post-replicative adenine modification in mammals.

ALKBH1 deficiency leads to loss of homeostasis in human diploid somatic cells

“…Despite that ALKBH1 has recently been reported as a 6mA demethylase (Xiao et al, 2018;Xie et al, 2018), which is by far the only one identified in human, we did not observe discernible difference in the 6mA levels in hESCs, hMSCs or hVSMCs caused by ALKBH1 deficiency, consistent with the recent report that ALKBH1 fails to eliminate 6mA in mESCs or HEK293T cells (Liu et al, 2020). Given that two studies have shown that ALKBH1 preferably demethylates 6mA on unpaired DNAs (Tian et al, 2020;Zhang et al, 2020a), our results imply that there might be few unpaired DNAs in the normally cultured hESCs, hMSCs and hVSMCs.…”

“…Another study reported 6mA as merely a false-positive signal likely caused by mycoplasma contamination and/or nonspecific antibodies (Douvlataniotis et al, 2020). While we and others performed experiments to exclude the possibilities of 6mA contaminations from cell culture or from the reagents used in LC-MS/ MS assay (Liu et al, 2020), proving the existence and primary origin of human genomic DNA 6mA is still a challenge in this field. With full consideration of these caveats, our study indicates that ALKBH1 is a key homeostatic regulator of human diploid adult stem cells and terminally differentiated cells.…”

“…6mA-independent roles of ALKBH1 in human diploid somatic cells LETTER demonstrated that 6mA is incorporated into the mammalian genomic DNA in the process of DNA replication by DNA polymerase in a 6mA methylase-independent manner (Liu et al, 2020;Musheev et al, 2020). Another study reported 6mA as merely a false-positive signal likely caused by mycoplasma contamination and/or nonspecific antibodies (Douvlataniotis et al, 2020).…”

Biochemical and structural basis for YTH domain of human YTHDC1 binding to methylated adenine in DNA

“…Other studies have failed to detect N6mA in several mouse tissues and mitochondrial DNA from human placenta ( 14 ), DNA isolated from mouse embryonic stem cells or brain and liver tissue ( 15 ) or cultured human cells ( 16 ). Nevertheless, more recent reports present evidence for low levels of N6mA in DNA from cultured human and mouse cells; however, that modified adenosine is incorporated (at least in part) by DNA polymerase using deoxy-N6mA converted from RNA-derived ribo-N6mA, via the nucleotide salvage pathway ( 17 , 18 ).…”

DNA6mA-MINT: DNA-6mA Modification Identification Neural Tool

“…Therefore, this makes it necessary to identify a 6mA position in the genome sites. Mammalian 6mA largely originates from the genomic incorporation mediated by DNA polymerase, while the methylase-generated 6mA in mice remains elusive [ 8 ].…”