Kim Myungjin, Trinh Binh N, Long Tiffany I, Oghamian Shirley, Laird Peter W
Department of Surgery, University of Southern California, Keck School of Medicine, Norris Comprehensive Cancer Center, Los Angeles, CA 90089-9176, USA.
Nucleic Acids Res. 2004 Oct 27;32(19):5742-9. doi: 10.1093/nar/gkh912. Print 2004.
DNA hypomethylation is frequently seen in cancer and imparts genomic instability in mouse models and some tissue culture systems. However, the effects of genomic DNA hypomethylation on mutation rates are still elusive. We have developed a model system to analyze the effects of DNA methyltransferase 1 (Dnmt1) deficiency on DNA mismatch repair (MMR) in mouse embryonic stem (ES) cells. We generated sibling ES cell clones with and without functional Dnmt1 expression, containing a stable reporter gene that allowed us to measure the slippage rate at a mononucleotide repeat. We found that Dnmt1 deficiency led to a 7-fold increase in the microsatellite slippage rate. Interestingly, the region flanking the mononucleotide repeat was unmethylated regardless of Dnmt1 status, suggesting that it is not the local levels of DNA methylation that direct the increase in microsatellite instability (MSI). The enhanced MSI was associated with higher levels of histone H3 acetylation and lower MeCP2 binding at regions near the assayed microsatellite, suggesting that Dnmt1 loss may decrease MMR efficiency by modifying chromatin structure.
DNA低甲基化在癌症中经常出现，并在小鼠模型和一些组织培养系统中导致基因组不稳定。然而，基因组DNA低甲基化对突变率的影响仍不清楚。我们开发了一个模型系统来分析DNA甲基转移酶1（Dnmt1）缺陷对小鼠胚胎干细胞（ES细胞）中DNA错配修复（MMR）的影响。我们生成了具有和不具有功能性Dnmt1表达的同胞ES细胞克隆，其中含有一个稳定的报告基因，使我们能够测量单核苷酸重复序列处的滑动率。我们发现Dnmt1缺陷导致微卫星滑动率增加7倍。有趣的是，无论Dnmt1状态如何，单核苷酸重复序列侧翼区域均未甲基化，这表明不是DNA甲基化的局部水平导致微卫星不稳定性（MSI）增加。增强的MSI与所检测微卫星附近区域更高水平的组蛋白H3乙酰化和更低水平的MeCP2结合相关，这表明Dnmt1缺失可能通过改变染色质结构降低MMR效率。
