Semiconservative transmission of DNA N 6 -adenine methylation in a unicellular eukaryote

作者：Yalan Sheng, Yuanyuan Wang, Wentao Yang, Xue Qing David Wang, Jiuwei Lu, Bo Pan, Bei Nan, Yongqiang Liu, Fei Ye, Chun Li, Jikui Song, Yali Dou, Shan Gao, Yifan Liu · 发表于：Genome Research · 年份：2024 · DOI：10.1101/gr.277843.123 · 被引用次数：22 · 研究领域：Protist diversity and phylogeny、Epigenetics and DNA Methylation、Environmental DNA in Biodiversity Studies

Although DNA N 6 -adenine methylation (6mA) is best known in prokaryotes, its presence in eukaryotes has recently generated great interest. Biochemical and genetic evidence supports that AMT1, an MT-A70 family methyltransferase (MTase), is crucial for 6mA deposition in unicellular eukaryotes. Nonetheless, the 6mA transmission mechanism remains to be elucidated. Taking advantage of single-molecule real-time circular consensus sequencing (SMRT CCS), here we provide definitive evidence for semiconservative transmission of 6mA in Tetrahymena thermophila . In wild-type (WT) cells, 6mA occurs at the self-complementary ApT dinucleotide, mostly in full methylation (full-6mApT); after DNA replication, hemi-methylation (hemi-6mApT) is transiently present on the parental strand, opposite to the daughter strand readily labeled by 5-bromo-2′-deoxyuridine (BrdU). In Δ AMT1 cells, 6mA predominantly occurs as hemi-6mApT. Hemi-to-full conversion in WT cells is fast, robust, and processive, whereas de novo methylation in Δ AMT1 cells is slow and sporadic. In Tetrahymena , regularly spaced 6mA clusters coincide with the linker DNA of nucleosomes arrayed in the gene body. Importantly, in vitro methylation of human chromatin by the reconstituted AMT1 complex recapitulates preferential targeting of hemi-6mApT sites in linker DNA, supporting AMT1's intrinsic and autonomous role in maintenance methylation. We conclude that 6mA is transmitted by a semiconservative mechanism: full-6mApT is split by DN...