N6-methyladenine DNA modification in Drosophila
Authors: Dapeng Zhang
Ruichuan Yin
Ning Zhang
Ying Cheng
Di Liu
Hua Huang
Guoqiang Zhang
Dahua Chen
Chaoyi Li
Wenxin Zhang
Shunmin He
Chuan He
Yuanxiang Zhu
Peng Zhang
Baodong Liu
Yuewan Luo
Hailin Wang
Xiaoling Liu
Jianzhao Liu
Journal: Cell (2015)
Discipline: MOLECULAR BIOLOGY & GENETICS
Keywords: DNA modification; N(6)-methyladenine; microbial genomes; biological processes; bacteria
Affiliations: State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, China., State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, Beijing 100085, China., Key Laboratory of the Zoological Systematics and Evolution, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, China., Department of Chemistry and Institute for Biophysical Dynamics, Howard Hughes Medical Institute, The University of Chicago, Chicago, IL 60637, USA., State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-Environmental Sciences, Chinese Academy of Sciences, Beijing 100085, China. Electronic address: ., State Key Laboratory of Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, China. Electronic address: .

Abstract:  DNA N(6)-methyladenine (6mA) modification is commonly found in microbial genomes and plays important functions in regulating numerous biological processes in bacteria. However, whether 6mA occurs and what its potential roles are in higher-eukaryote cells remain unknown. Here, we show that 6mA is present in Drosophila genome and that the 6mA modification is dynamic and is regulated by the Drosophila Tet homolog, DNA 6mA demethylase (DMAD), during embryogenesis. Importantly, our biochemical assays demonstrate that DMAD directly catalyzes 6mA demethylation in vitro. Further genetic and sequencing analyses reveal that DMAD is essential for development and that DMAD removes 6mA primarily from transposon regions, which correlates with transposon suppression in Drosophila ovary. Collectively, we uncover a DNA modification in Drosophila and describe a potential role of the DMAD-6mA regulatory axis in controlling development in higher eukaryotes. 

Retracted: No

PubPeer comments:
 No evidence for 6mA DNA modification in eukaryotic genomes https: bmcgenomics.biomedcentral.com articles 10.1186 s12864-019-5754-6 , 
 #1 Now this paper was just published , suggesting that all commonly used techniques to profile DNA N6-methyladenine (6mdA) in mammals and other higher eukaryotes have inherent flaws, resulting in false detection. These errors can explain the purported discovery of 6mdA in higher organisms, including flies. , 
 Readers should become aware of a recent paper entitled Critical assessment of DNA adenine methylation in eukaryotes using quantitative deconvolution , which reanalyzed the 6mA DIP-seq data from this study and found that: reads that mapped to multiple bacterial genomes. , indicating that the values obtained at Zhang et al., 2015 are overestimated. From the text: A previous study quantified 6mA in D. melanogaster using UHPLC-MS MS and reported that 6mA A reaches the peak level of ~700 ppm (parts per million) in ~0.75-hour embryos and falls to ~10 ppm at later stages such as adult tissues (8). We first collected the fly embryo sample at ~0.75 hours and got 674,650 SMRT CCS reads for single-molecule 6mA analysis (table S2). Despite strict measures to avoid contamination (31), we found that 96.12% of the CCS reads mapped to the D. melanogaster genome reference, whereas 3.88% of the CCS reads mapped to a few microbes (Fig. 4A). Specifically, the contamination reads came from S. cerevisiae (1.65%), the major food source of Drosophila (33), and two genera of bacteria, Acetobacter (0.86%) and Lactobacillus (0.23%), the main gut commensal bacteria of D. melanogaster (34). We separately quantified 6mA A levels in the D. melanogaster genome and in each contamination source and found that the level of 6mA A in total gDNA was 100 ppm (CI, 50 to 200 ppm, consistent with the ~121 ppm UHPLC-MS MS estimate), 2 ppm in D. melanogaster (CI, 1 to 10 ppm), 2 ppm in Saccharomyces (CI, 1 to 10 ppm), 5495 ppm in Acetobacter (CI, 3162 to 10,000 ppm), 977 ppm in Lactobacillus (CI, 501 to 1995 ppm), and 7413 ppm in Others (including additional bacterial genera and unannotated sequences; CI, 3981 to 12,589 ppm) (Fig. 4B and fig. S9) (31). Despite their relatively low abundance (3.88%), bacteria contributed to most of the 6mA events in the total gDNA (Fig. 4C). In Acetobacter, we observed a high-confidence bacterial 6mA motif (GANTC) (Fig. 4B), consistent with the REBASE database (35). The 6mA A level of 2 ppm (CI, 1 to 10 ppm) estimated for D. melanogaster, in contrast to the ~700 ppm previously reported, only explains 1.44% of the total 6mA events in the gDNA sample (consideringtaxonomy abundances; Fig. 4C) ,