Kohno T, Kawanishi M, Inazawa J, Yokota J
Biology Division, National Cancer Center Research Institute, Tokyo, Japan.
Hum Genet. 1998 Mar;102(3):258-64. doi: 10.1007/s004390050689.
DNA hypermethylation is believed to be involved in human carcinogenesis, since it suppresses the transcription of defined genes and is associated with chromosomal instability. In this study, we identified CpG islands that are hypermethylated in human lung cancer by a modified arbitrarily primed-polymerase chain reaction method using genomic DNAs digested with a methylation-sensitive restriction enzyme, HpaII, as templates. When we analyzed genomic DNAs from normal lung tissues and non-small cell lung carcinoma cell lines using three arbitrary primers, three DNA fragments were amplified from lung cancer DNAs but not from normal lung DNAs. Restriction mapping and Southern blot analysis revealed that all of these bands were amplified from CpG islands that were hypermethylated in the lung cancer cell lines. These islands were mapped to chromosomes 4q34, 10q26 and 17p13.1-p13.2, respectively, and these chromosomal regions were also hypermethylated in a subset of primary lung tumors in vivo. Thus, diverse chromosomal regions are hypermethylated in lung cancer cells. The results also indicate that this method is simple and effective for screening of CpG islands that are hypermethylated in cancer cells.
DNA高甲基化被认为与人类致癌作用有关，因为它会抑制特定基因的转录并与染色体不稳定相关。在本研究中，我们使用经甲基化敏感限制酶HpaII消化的基因组DNA作为模板，通过改良的任意引物聚合酶链反应方法，鉴定了在人类肺癌中发生高甲基化的CpG岛。当我们使用三种任意引物分析来自正常肺组织和非小细胞肺癌细胞系的基因组DNA时，从肺癌DNA中扩增出了三个DNA片段，而正常肺DNA中未扩增出。限制性图谱分析和Southern印迹分析表明，所有这些条带均从在肺癌细胞系中发生高甲基化的CpG岛扩增而来。这些岛分别定位于染色体4q34、10q26和17p13.1 - p13.2，并且在体内的一部分原发性肺肿瘤中这些染色体区域也发生了高甲基化。因此，肺癌细胞中不同的染色体区域发生了高甲基化。结果还表明，该方法对于筛选癌细胞中发生高甲基化的CpG岛简单有效。
