PCNA clamp facilitates action of DNA cytosine methyltransferase 1 on hemimethylated DNA
PCNA clamp facilitates action of DNA cytosine methyltransferase 1 on hemimethylated DNA
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DOI：
10.1046/j.1365-2443.2002.00584.x
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发表时间：
2002-10-01
期刊：
影响因子：
2.1
通讯作者：
Tsurimoto, T
中科院分区：
文献类型：
--
作者：
Iida, T;Suetake, I;Tsurimoto, T
背景：增殖细胞核抗原（PCNA）是一种环状蛋白，是DNA聚合酶δ的合成因子。除了这种作用，PCNA与许多其他蛋白质相互作用，以增加它们在复制DNA位点的局部浓度。DNA胞嘧啶甲基转移酶1（Dnmt 1）通过在DNA复制后完成半甲基化DNA的甲基化来保存表观遗传信号，已被先前的工作指示为这些PCNA结合蛋白之一。结果：Dnmt 1可通过PCNA亲和层析从细胞核提取物中分离得到，但其表达的分子机制和功能意义尚不清楚。这两种蛋白之间的相互作用的研究表明，N-末端区域的Dnmt 1，其中包含一个典型的PCNA结合基序，具有核心的PCNA结合活性，和蛋白质的其余部分施加的Dnmt 1与PCNA的相互作用的负面影响。Dnmt 1与PCNA结合的DNA的亲和力比游离DNA高得多。此外，DNA甲基化检测与半甲基化的DNA作为底物显示，PCNA钳结合的DNA甲基化更有效地由Dnmt 1比游离DNA.Conclusion：这些结果提供了第一个生化证据表明，PCNA和Dnmt 1之间的物理相互作用促进新nepplicated DNA的甲基化，PCNA仍然作为一个功能性的钳。
Background: Proliferating cell nuclear antigen (PCNA) is a ring-shaped protein known as a processivity factor of DNA polymerase delta. In addition to this role, PCNA interacts with a number of other proteins to increase their local concentration at replicated DNA sites. DNA cytosine methyltransferase 1 (Dnmt1), which preserves epigenetic signals by completing the methylation of hemimethylated DNA after DNA replication, has been indicated as one of these PCNA binding proteins by a previous work. However, the molecular mechanisms and functional significance of their association have not yet been studied.Results: Dnmt1 can be readily isolated from nuclear extracts by PCNA affinity chromatography. Studies of the interactions between the two proteins demonstrate that the N-terminal region of Dnmt1, which contains a typical PCNA binding motif, has core PCNA binding activity, and that the remaining portion of the protein exerts a negative influence on the interaction of Dnmt1 with PCNA. The affinity of Dnmt1 for DNA is much higher for DNA bound by PCNA than for free DNA. Furthermore, DNA methylation assays with hemimethylated DNA as a substrate revealed that PCNA clamp-bound DNA is methylated more efficiently by Dnmt1 than is free DNA.Conclusion: These results provide the first biochemical evidence that physical interactions between PCNA and Dnmt1 facilitate the methylation of newly neplicated DNA, on which PCNA remains associated as a functional clamp.