N6-Methyladenine DNA Modification in the Human Genome
Authors: Ying Chen
Jinbao Liu
Minghui He
Qian Zhang
Shang-Qian Xie
De Chen
Chuan-Le Xiao
Feng Luo
Guang-Rong Yan
Zhe Liang
De-Peng Wang
Song Zhu
Xiao-Feng Gu
Guoliang Yu
Xiao-Chen Bo
Kai Wang
Journal: Molecular Cell (2018)
Discipline: MOLECULAR BIOLOGY & GENETICS
Keywords: 6mA; cancer; demethylase; human genome; m6A
Affiliations: Biomedicine Research Center, the Third Affiliated Hospital of Guangzhou Medical University, Guangzhou 510150, China; State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou 510060, China; College of Plant Protection, Hunan Agricultural University, Changsha 410128, China., Biomedicine Research Center, the Third Affiliated Hospital of Guangzhou Medical University, Guangzhou 510150, China; Key Laboratory of Protein Modification and Degradation, School of Basic Medical Sciences, Guangzhou Medical University, Guangzhou 511436, China., CookGene Biosciences Center, Guangzhou 510320, China., Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, China., State Key Laboratory of Ophthalmology, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou 510060, China., Grandomics Biosciences, Beijing 102206, China., Key Laboratory of Protein Modification and Degradation, School of Basic Medical Sciences, Guangzhou Medical University, Guangzhou 511436, China., School of Computing, Clemson University, Clemson, SC 29634-0974, USA., Department of Biological Sciences, National University of Singapore, 117543 Singapore, Singapore., Dapartment of Biotechnology, Beijing Institute of Radiation Medicine, Beijing 100850, China. Electronic address: ., Biotechnology Research Institute, Chinese Academy of Agricultural Sciences, Beijing 100081, China. Electronic address: ., Raymond G. Perelman Center for Cellular and Molecular Therapeutics, Children's Hospital of Philadelphia, Philadelphia, PA 19104, USA. Electronic address: ., Biomedicine Research Center, the Third Affiliated Hospital of Guangzhou Medical University, Guangzhou 510150, China; Key Laboratory of Protein Modification and Degradation, School of Basic Medical Sciences, Guangzhou Medical University, Guangzhou 511436, China. Electronic address: .

Abstract:  DNA N6-methyladenine (6mA) modification is the most prevalent DNA modification in prokaryotes, but whether it exists in human cells and whether it plays a role in human diseases remain enigmatic. Here, we showed that 6mA is extensively present in the human genome, and we cataloged 881,240 6mA sites accounting for ∼0.051% of the total adenines. [G/C]AGG[C/T] was the most significantly associated motif with 6mA modification. 6mA sites were enriched in the coding regions and mark actively transcribed genes in human cells. DNA 6mA and N6-demethyladenine modification in the human genome were mediated by methyltransferase N6AMT1 and demethylase ALKBH1, respectively. The abundance of 6mA was significantly lower in cancers, accompanied by decreased N6AMT1 and increased ALKBH1 levels, and downregulation of 6mA modification levels promoted tumorigenesis. Collectively, our results demonstrate that DNA 6mA modification is extensively present in human cells and the decrease of genomic DNA 6mA promotes human tumorigenesis. 

Retracted: No

PubPeer comments:
 Serious concerns were previously raised about the validity of this work while it was a preprint on bioarchive: https: pubpeer.com publications 9C514D1F6B7BE4E25ACE08184B3BEC , 
 The veracity of this paper reporting the existence and regulation of 6mA in humans will need to be independently verified by other labs before it can really be trusted. These data are in opposition to a recent study finding no evidence that 6mA occurs in human or mouse DNA ( lt;0.00004% 6mA A), and several other studies finding that the maximum possible level of 6mA in the human or mouse genome is just a few 6mA sites per million deoxyadenosines, only if we can assume that no bacterial contamination can account for the very low signal. If these previous studies are indeed correct, the results of this paper, Xiao et al 2018 cannot possibly be accurate, unless cell lines grown in this lab have magically gained extensive and enzymatically regulated 6mA methylation. https: onlinelibrary.wiley.com doi abs 10.1002 anie.201700424 https: www.ncbi.nlm.nih.gov pubmed 16684535 https: pubs.acs.org doi abs 10.1021 acs.analchem.7b01152?src=recsys journalCode=ancham https: www.ncbi.nlm.nih.gov pubmed 27027282 , 
 Serious concerns were raised about the validity and reproducibility of this paper while it was a preprint on bioarchive: https: pubpeer.com publications 9C514D1F6B7BE4E25ACE08184B3BEC To summarize, several recent studies have found that 6mA is undetectable ( lt;0.00004%), or detected at extremely low levels (~0.0001%) in human cells. These studies were performed using highly sensitive quantitation methods and have therefore called into question the existence of 6mA in humans: https: onlinelibrary.wiley.com doi abs 10.1002 anie.201700424 https: www.ncbi.nlm.nih.gov pubmed 16684535 https: pubs.acs.org doi abs 10.1021 acs.analchem.7b01152?src=recsys journalCode=ancham https: www.ncbi.nlm.nih.gov pubmed 27027282 Surprisingly, in this paper Xiao et al. reports that 6mA occurs at ~0.05% 6mA A in human cells. This is ~1000X higher than the maximum possible level of 6mA reported previously! This means that either all of the other labs were wrong (which seems unlikely), or this paper is seriously flawed. Alternatively, maybe human cell lines grown in this lab in China have magically acquired extensive and enzymatically regulated 6mA methylation, never seen before in other countries or labs. It will be important to consider how these findings can be reconciled with previous work on 6mA in humans. It will be similarly important for independent groups of scientists in other countries to test the validity and reproducibility of the claims in this paper. It is sad that this paper is misleading the community into thinking that a probably nonexistent DNA modification exists in humans and plays any role in cancer progression. , 
 maybe they are detecting the signal because their human cells are mycoplasma infected ? https: pubs.acs.org doi pdf 10.1021 acs.analchem.7b01152 , 
 A recent paper found no evidence for 6mA in the genome of mouse or human cells or tissues https: onlinelibrary.wiley.com doi abs 10.1002 anie.201700424 The method used in their paper was capable of detecting 6mA abundance as low as 0.00004% (1000X higher than what is reported here). I think mycoplasma contamination and or other experimental artifacts are almost certainly at play here. , 
 https: bmcgenomics.biomedcentral.com articles 10.1186 s12864-019-5754-6 , 
 https: www.ncbi.nlm.nih.gov pubmed 31636962 N6AMT1 co-crystal structure shows that it cannot methylate DNA and indicate that it is a protein methyltransferase , 
 Yet another well executed study showing that previous reports of 6mA in mammals (including this one) are artifactual: No evidence for DNA N6-methyladenine in mammals https: advances.sciencemag.org content 6 12 eaay3335  Unfortunately, in the absence of a 6mdA detection technique robust to the multiple sources of error that we have outlined, we conclude that the evidence published to date is not sufficient to support the presence of 6mdA in mammals. , 
 #8 After my study was published, the related reports have also showed that 6mA is existed in mammals in Cell 2018 and in Mol Cell 2020 . And the related structural analyses also indicated that mammalian ALKBH1 serves as an N6-mA demethylase in Cell Res 2020 . , 
 Did you not read this paper carefully? https: advances.sciencemag.org content 6 12 eaay3335 It shows that all of the popular 6mA  detection methods (dot blot, IP, SMRTseq, UHPLC-ms ms) used in prior studies are so problematic for measurements of mammalian DNA that they invariably give rise to false positive signals and false conclusions regarding the existence and regulation of 6mA in mammals. Your Mol Cell paper and the papers you ve just cited are irrelevant to your point, as they all use the same flawed methods. , 
 Claims of N6AMT1 as a u2018DNA (adenine-N6 )-methyltransferaseu2019 could not be reproduced by recent article published in Nature Chemical Biology . Also, this result could not be reproduced by Xie et al., 2018 ,