Replication-independent chromatin loading of Dnmt1 during G2 and M phases
Replication-independent chromatin loading of Dnmt1 during G2 and M phases
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DOI：
10.1038/sj.embor.7400295
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发表时间：
2004-12-01
期刊：
影响因子：
7.7
通讯作者：
Cardoso, MC
中科院分区：
文献类型：
--
作者：
Easwaran, HP;Schermelleh, L;Cardoso, MC
主要的DNA甲基转移酶DNMT1与S期的DNA复制位点相结合，维持了新合成链中的甲基化模式。鉴于DNMT1在体外的缓慢动力学与复制分叉的快速进展，我们已经测试了DNMT1是否与染色质在S期之后相关联。利用时间推移显微镜对表达绿色荧光蛋白标记的Dnmt1和DsRed标记的DNA连接酶I作为细胞周期推进标记的哺乳动物细胞，我们发现Dnmt1在G2和M期间与染色质结合，这种结合是由特定的靶向序列介导的，表现出对结构性而不是兼性异染色质的强烈偏好，并且不依赖于异染色质特异性的组蛋白H3-Lys9三甲基化、SUV39H和HP1。此外，光漂白分析表明，Dnmt1在整个G2和M期间持续被加载到染色质上，这表明Dnmt1具有复制无关的作用，这可能代表了维持DNA甲基化的一种新的单独途径。
The major DNA methyltransferase, Dnmt1, associates with DNA replication sites in S phase maintaining the methylation pattern in the newly synthesized strand. In view of the slow kinetics of Dnmt1 in vitro versus the fast progression of the replication fork, we have tested whether Dnmt1 associates with chromatin beyond S phase. Using time-lapse microscopy of mammalian cells expressing green-fluorescent- protein-tagged Dnmt1 and DsRed-tagged DNA Ligase I as a cell cycle progression marker, we have found that Dnmt1 associates with chromatin during G2 and M. This association is mediated by a specific targeting sequence, shows strong preference for constitutive but not facultative heterochromatin and is independent of heterochromatin-specific histone H3 Lys9 trimethylation, SUV39H and HP1. Moreover, photobleaching analyses showed that Dnmt1 is continuously loaded onto chromatin throughout G2 and M, indicating a replication-independent role of Dnmt1 that could represent a novel and separate pathway to maintain DNA methylation.