Biochemical reconstitution of TET1-TDG-BER-dependent active DNA demethylation reveals a highly coordinated mechanism.
Biochemical reconstitution of TET1-TDG-BER-dependent active DNA demethylation reveals a highly coordinated mechanism.
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TET1-TDG-BER依赖性的活性DNA脱甲基化的生化重构揭示了高度协调的机制。
DOI：
10.1038/ncomms10806
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发表时间：
2016-03-02
影响因子：
16.6
通讯作者：
Schär P
中科院分区：
文献类型：
--
作者：
Weber AR;Krawczyk C;Robertson AB;Kuśnierczyk A;Vågbø CB;Schuermann D;Klungland A;Schär P
CpG二核苷酸中的胞嘧啶甲基化是由DNA甲基转移酶和去甲基化酶动态建立和维持的表观遗传DNA修饰。随着5-甲基胞嘧啶（5mC）导向羟化酶和10 - 11易位（TET）蛋白和胸腺嘧啶DNA糖基化酶（TDG）的碱基切除活性的发现，活性DNA去甲基化的分子机制最近才开始出现。这暗示了一个通过TET蛋白氧化5mC的途径，该途径产生tgd依赖性碱基切除修复（BER）的底物，然后用c取代5mC。然而，TET与BER有效偶联的直接证据从未被提出。在这里，我们发现TET1和TDG物理相互作用氧化和去除5mC，并通过生化重构证明TET-TDG-BER系统能够产生DNA去甲基化。我们发现该机制确保了对称甲基化CpGs的顺序去甲基化，从而避免了DNA双链断裂的形成，但导致了甲基化CpGs的易变性。胞嘧啶甲基化是一种动态的DNA修饰，涉及被怀疑参与去甲基化的碱基切除修复途径。在这里，作者表明TET1和TDG相互作用以靶向修饰的碱基并协调误码以避免双链断裂。
Cytosine methylation in CpG dinucleotides is an epigenetic DNA modification dynamically established and maintained by DNA methyltransferases and demethylases. Molecular mechanisms of active DNA demethylation began to surface only recently with the discovery of the 5-methylcytosine (5mC)-directed hydroxylase and base excision activities of ten–eleven translocation (TET) proteins and thymine DNA glycosylase (TDG). This implicated a pathway operating through oxidation of 5mC by TET proteins, which generates substrates for TDG-dependent base excision repair (BER) that then replaces 5mC with C. Yet, direct evidence for a productive coupling of TET with BER has never been presented. Here we show that TET1 and TDG physically interact to oxidize and excise 5mC, and proof by biochemical reconstitution that the TET–TDG–BER system is capable of productive DNA demethylation. We show that the mechanism assures a sequential demethylation of symmetrically methylated CpGs, thereby avoiding DNA double-strand break formation but contributing to the mutability of methylated CpGs. Cytosine methylation is a dynamic DNA modification with the involvement of the base excision repair pathway suspected to be involved in demethylation. Here the authors show that TET1 and TDG interact to target modified bases and coordinate BER to avoid double strand breaks.