Identification of DNMT1 (DNA methyltransferase 1) hypomorphs in somatic knockouts suggests an essential role for DNMT1 in cell survival
Identification of DNMT1 (DNA methyltransferase 1) hypomorphs in somatic knockouts suggests an essential role for DNMT1 in cell survival
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DOI：
10.1073/pnas.0604602103
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发表时间：
2006-09-19
影响因子：
11.1
通讯作者：
Liang, Gangning
中科院分区：
文献类型：
--
作者：
Egger, Gerda;Jeong, Shinwu;Liang, Gangning
先前的研究表明，DNA甲基转移酶（Dnmt）1是维持大量DNA甲基化所必需的，对小鼠发育至关重要。然而，在人癌细胞系HCT 116中DNMT 1的体细胞破坏不是致命的，并且仅引起甲基化的轻微降低。在这里，我们报告了一个截短的DNMT 1蛋白，这是由在HCT 116细胞中的DNMT 1的破坏产生的鉴定。截短的蛋白质，其中有部分的监管N-末端结构域删除，但保留催化C-末端结构域，是目前在所有DNMT 1单敲除和DNMT 1/DNMT 3b双敲除细胞系测试，并保留半甲基化酶活性在不同的水平。DNMT 1 RNAi导致WT和敲除细胞中细胞活力降低，并导致DNMT 1敲除细胞中DNA甲基化进一步丧失。此外，我们观察到在复制后甲基化的延迟和表达截短蛋白的细胞中特定CpG位点的半甲基化的增加。药物诱导的低甲基化后的再甲基化研究表明，DNMT 1在亚端粒重复序列D4 Z4的从头甲基化中起着假定的作用，D4 Z4在缺乏全长DNMT 1的细胞中丢失。我们的数据表明，DNMT 1可能是维持DNA甲基化，增殖和癌细胞存活所必需的。
Previous studies have shown that DNA methyltransferase (Dnmt) 1 is required for maintenance of bulk DNA methylation and is essential for mouse development. However, somatic disruption of DNMT1 in the human cancer cell line HCT116 was not lethal and caused only minor decreases in methylation. Here, we report the identification of a truncated DNMT1 protein, which was generated by the disruption of DNMT1 in HCT116 cells. The truncated protein, which had parts of the regulatory N-terminal domain deleted but preserved the catalytic C-terminal domain, was present at different levels in all DNMT1 single-knockout and DNMT1/DNMT3b double-knockout cell lines tested and retained hemimethylase activity. DNMT1 RNAi resulted in decreased cell viability in WT and knockout cells and further loss of DNA methylation in DNMT1 knockout cells. Furthermore, we observed a delay in methylation after replication and an increase in hemimethylation of specific CpG sites in cells expressing the truncated protein. Remethylation studies after drug-induced hypomethylation suggest a putative role of DNMT1 in the de novo methylation of a subtelomeric repeat, D4Z4, which is lost in cells lacking full-length DNMT1. Our data suggest that DNMT1 might be essential for maintenance of DNA methylation, proliferation, and survival of cancer cells.