Discovery of epigenetically silenced genes by methylated DNA lmmunoprecipitation in colon cancer cells
Discovery of epigenetically silenced genes by methylated DNA lmmunoprecipitation in colon cancer cells
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DOI：
10.1158/0008-5472.can-07-2687
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发表时间：
2007-12-15
期刊：
影响因子：
11.2
通讯作者：
Esteller, Manel
中科院分区：
文献类型：
--
作者：
Jacinto, Filipe V.;Ballestar, Esteban;Esteller, Manel
肿瘤抑制基因的CpG岛启动子超甲基化是人类癌症的共同标志，新的大规模表观基因组技术可能有助于我们尝试定义肿瘤细胞的完整DNA超甲基化。在这里，我们报告了一个功能搜索高甲基化的CpG岛使用结肠直肠癌细胞系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）。使用甲基化DNA免疫沉淀方法结合启动子微阵列分析，我们发现，DKO细胞经历了一个显着的高甲基化CpG岛的损失。对这些候选序列的进一步表征显示CpG岛启动子高甲基化和在肿瘤发生中具有潜在重要作用的基因的沉默，例如Ras鸟嘌呤核苷酸释放因子（RASGRF 2）、乳腺癌相关碱性螺旋环转录因子（BHLHB 9）和同源框基因（HOXD 1）。这些基因的高甲基化发生在癌前病变中，并在肿瘤发生过程中积累。因此，我们的研究结果表明，DNMT遗传破坏策略结合甲基化DNA免疫沉淀在寻找人类癌症中未知的高甲基化候选基因中的有用性。可能有助于我们理解疾病的生物学，并具有潜在的翻译用途。
CpG island promoter hypermethylation of tumor suppressor genes is a common hallmark of human cancer, and new large-scale epigenomic technologies might be useful in our attempts to define the complete DNA hypermethylome of tumor cells. 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 methylated DNA immunoprecipitation methodology in conjunction with promoter microarray analyses, we found that DKO cells experience a significant loss of hypermethylated CpG islands. Further characterization of these candidate sequences shows CpG island promoter hypermethylation and silencing of genes with potentially important roles in tumorigenesis, such as the Ras guanine nucleotide-releasing factor (RASGRF2), the apoptosis-associated basic helix-loop transcription factor (BHLHB9), and the homeobox gene (HOXD1). Hypermethylation of these genes occurs in premalignant lesions and accumulates during tumorigenesis. Thus, our results show the usefulness of DNMT genetic disruption strategies combined with methylated DNA immunoprecipitation in searching for unknown hypermethylated candidate genes in human cancer that. might aid our understanding of the biology of the disease and be of potential translational use.