Yan H, Tang G, Wang H, Hao L, He T, Sun X, Ting A H, Deng A, Sun S
Department of Laboratory Medicine, Changhai Hospital, The Second Military Medical University, Shanghai, China.
Institute of Genetics, Second Military Medical University, Shanghai, China.
Oncogene. 2016 Jul 28;35(30):3995-4008. doi: 10.1038/onc.2015.423. Epub 2015 Nov 9.
Levels of γ-aminobutyric acid (GABA) and glutamic acid decarboxylase 1 (GAD1), the enzyme that synthesizes GABA, are significantly increased in neoplastic tissues. However, the mechanism underlying this increase remains elusive. Instead of silencing gene transcription, we showed that the GAD1 promoter was hypermethylated in both colon and liver cancer cells, leading to the production of high levels of GAD1. GAD1 is a target gene that is silenced by H3K27me3. The key locus responsible for GAD1 reactivation was mapped to a DNA methylation-sensitive CTCF-binding site (CTCF-BS3) within the third intron of GAD1. Chromosome configuration capture (3C) analysis indicated that an intrachromosomal loop was formed by CTCF self-dimerisation in normal cells (CTCF binds to both unmethylated CTCF-BS3 and CTCF-BS2). The CTCF dimer then interacted with suppressor of zeste 12 homologue (SUZ12), which is a domain of Polycomb repressive complex 2 (PRC2), promoting the methylation of H3K27 and the silencing of GAD1 expression. This silencing was shown to be inhibited by DNA methylation in cancer cells. These findings strongly suggest that GAD1 is reactivated by DNA methylation, which provided a model for DNA methylation and the active orchestration of oncogenic gene expression by CTCF in cancer cells.
γ-氨基丁酸（GABA）以及合成GABA的谷氨酸脱羧酶1（GAD1）在肿瘤组织中的水平显著升高。然而，这种升高背后的机制仍然难以捉摸。我们发现，在结肠癌细胞和肝癌细胞中，GAD1启动子并非发生基因转录沉默，而是发生了高甲基化，从而导致高水平的GAD1产生。GAD1是一个被H3K27me3沉默的靶基因。负责GAD1重新激活的关键位点被定位到GAD1第三内含子内一个对DNA甲基化敏感的CCCTC结合因子结合位点（CTCF-BS3）。染色体构象捕获（3C）分析表明，在正常细胞中，CTCF通过自身二聚化形成了一个染色体内环（CTCF与未甲基化的CTCF-BS3和CTCF-BS2均结合）。然后，CTCF二聚体与多梳抑制复合物2（PRC2）的一个结构域——zeste同源物12抑制因子（SUZ12）相互作用，促进H3K27甲基化以及GAD1表达的沉默。研究表明，癌细胞中的DNA甲基化可抑制这种沉默。这些发现有力地表明，GAD1通过DNA甲基化被重新激活，这为癌细胞中DNA甲基化以及CTCF对致癌基因表达的主动调控提供了一个模型。
