Genetic unmasking of epigenetically silenced tumor suppressor genes in colon cancer cells deficient in DNA methyltransferases
Genetic unmasking of epigenetically silenced tumor suppressor genes in colon cancer cells deficient in DNA methyltransferases
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DOI：
10.1093/hmg/ddg226
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发表时间：
2003-09-01
影响因子：
3.5
通讯作者：
Esteller, M
中科院分区：
文献类型：
--
作者：
Paz, MF;Wei, S;Esteller, M
肿瘤抑制基因CpG岛的超甲基化相关沉默是人类癌症的常见标志。在这里，我们报告了一个功能搜索高甲基化的CpG岛使用大肠癌细胞系HCT-116，其中两个主要的DNA甲基转移酶，DNMT 1和DNMT 3b，已被遗传破坏（DKO细胞）。使用差异甲基化位点的两种分子筛选[差异甲基化杂交（DMH）和甲基化位点间扩增（AIMS）]，我们发现DKO细胞（而不是单个DNMT 1或DNMT 3b敲除）具有大量高甲基化CpG岛的丢失，这会诱导相邻基因的重新激活。我们已经鉴定了大量在肿瘤发生中具有潜在重要作用的CpG岛相关基因，如钙粘蛋白成员FAT或同源框基因LMX-1和DUX-4。对于其在转化中的作用尚未被表征的其他基因，例如钙通道all或血栓烷A2受体，它们在DKO细胞中的重新引入抑制了集落形成。因此，我们的研究结果表明DNMT 1和DNMT 3b在CpG岛甲基化相关沉默中的作用以及遗传破坏策略在寻找新的高甲基化位点中的有用性。
Hypermethylation associated silencing of the CpG islands of tumor suppressor genes is a common hallmark of human cancer. Here we report a functional search for hypermethylated CpG islands using the colorectal cancer cell line HCT-116, in which two major DNA methyltransferases, DNMT1 and DNMT3b, have been genetically disrupted (DKO cells). Using two molecular screenings for differentially methylated loci [differential methylation hybridization (DMH) and amplification of inter-methylated sites (AIMS)], we found that DKO cells, but not the single DNMT1 or DNMT3b knockouts, have a massive loss of hypermethylated CpG islands that induces the re-activation of the contiguous genes. We have characterized a substantial number of these CpG island associated genes with potentially important roles in tumorigenesis, such as the cadherin member FAT, or the homeobox genes LMX-1 and DUX-4. For other genes whose role in transformation has not been characterized, such as the calcium channel all or the thromboxane A2 receptor, their re-introduction in DKO cells inhibited colony formation. Thus, our results demonstrate the role of DNMT1 and DNMT3b in CpG island methylation associated silencing and the usefulness of genetic disruption strategies in searching for new hypermethylated loci.