Mammalian ALKBH1 serves as an N6-mA demethylase of unpairing DNA

• 来源期刊: 细胞研究（英文版）
• 年，卷（期）: 2020,（3）
• 页码范围: 197-210
• 页数: 14页
• 语种: 英文

摘要

N6-methyladenine (N6-mA) of DNA is an emerging epigenetic mark in mammalian genome.Levels of N6-mA undergo drastic fluctuation during early embryogenesis,indicative of active regulation.Here we show that the 2-oxoglutarate-dependent oxygenase ALKBH1 functions as a nuclear eraser of N6-mA in unpairing regions (e.g.,SIDD,S_tress-!nduced DNA Double Helix _Destabilization regions) of mammalian genomes.Enzymatic profiling studies revealed that ALKBH1 prefers bubbled or bulged DNAs as substrate,instead of single-stranded (ss-) or double-stranded (ds-) DNAs.Structural studies of ALKBH1 revealed an unexpected "stretch-out" conformation of its "Flip1" motif,a conserved element that usually bends over catalytic center to facilitate substrate base flipping in other DNA demethylases.Thus,lack of a bending "Flip1" explains the observed preference of ALKBH1 for unpairing substrates,in which the flipped N6-mA is primed for catalysis.Co-crystal structural studies of ALKBH1 bound to a 21-mer bulged DNA explained the need of both flanking duplexes and a flipped base for recognition and catalysis.Key elements (e.g.,an ALKBH1-specific α1 helix) as well as residues contributing to structural integrity and catalytic activity were validated by structurebased mutagenesis studies.Furthermore,ssDNA-seq and DIP-seq analyses revealed significant co-occurrence of base unpairing regions with N6-mA in mouse genome.Collectively,our biochemical,structural and genomic studies suggest that ALKBH1 is an important DNA demethylase that regulates genome N6-mA turnover of unpairing regions associated with dynamic chromosome regulation.