Genistein depletes telomerase activity through cross-talk between genetic and epigenetic mechanisms.

Genistein, a natural isoflavone found in soybean products, has been reported to down-regulate telomerase activity and that this prevents cancer and contributes to the apoptosis of cancer cells. However, the precise molecular mechanism by which genistein represses telomerase is not clear. Here, we show that genistein inhibits the transcription of hTERT (human telomerase reverse transcriptase), the catalytic subunit of the human telomerase enzyme, in breast MCF10AT benign cells and MCF-7 cancer cells in a time- and dose-dependent manner. Three major DNA methyltransferases (DNMT1, 3a and 3b) were also decreased in genistein-treated breast cancer cells suggesting that genistein may repress hTERT by impacting epigenetic pathways. Sequential depletion of the hTERT promoter revealed that the hTERT core promoter region is responsible for the genistein-induced repression of hTERT transcription. Using a newly developed technique of chromatin immunoprecipitation (ChIP)-related bifulfite sequencing analysis, we found an increased binding of E2F-1 to the hTERT promoter is due to the site-specific hypomethylation of the E2F-1 recognition site. In addition, we found that genistein can remodel chromatin structures of the hTERT promoter by increasing trimethyl-H3K9 but decreasing dimethyl-H3K4 in the hTERT promoter. The repression of hTERT was enhanced by combination with genistein and the DNMT inhibitor, 5-aza-2′-deoxycytidine (5-aza-dCyd). These findings collectively show that genistein is working, at least in part, through epigenetic mechanisms of telomerase inhibition in breast benign and cancer cells and may facilitate approaches to breast cancer prevention and treatment using an epigenetic modulator combined with genistein.

染料木黄酮是一种在大豆制品中发现的天然异黄酮，据报道它能下调端粒酶活性，从而预防癌症并促进癌细胞凋亡。然而，染料木黄酮抑制端粒酶的确切分子机制尚不清楚。在此，我们发现染料木黄酮以时间和剂量依赖的方式抑制人乳腺MCF10AT良性细胞和MCF - 7癌细胞中hTERT（人端粒酶逆转录酶，人端粒酶的催化亚基）的转录。在染料木黄酮处理的乳腺癌细胞中，三种主要的DNA甲基转移酶（DNMT1、3a和3b）也有所减少，这表明染料木黄酮可能通过影响表观遗传途径抑制hTERT。对hTERT启动子的连续缺失分析表明，hTERT核心启动子区域负责染料木黄酮诱导的hTERT转录抑制。利用一种新开发的与染色质免疫沉淀（ChIP）相关的亚硫酸氢盐测序分析技术，我们发现E2F - 1与hTERT启动子的结合增加是由于E2F - 1识别位点的位点特异性低甲基化。此外，我们发现染料木黄酮可通过增加hTERT启动子中的三甲基 - H3K9但降低二甲基 - H3K4来重塑hTERT启动子的染色质结构。染料木黄酮与DNMT抑制剂5 - 氮杂 - 2′ - 脱氧胞苷（5 - aza - dCyd）联合使用可增强对hTERT的抑制。这些发现共同表明，染料木黄酮至少在部分程度上通过抑制乳腺良性和癌细胞中端粒酶的表观遗传机制发挥作用，并且可能促进使用表观遗传调节剂与染料木黄酮联合预防和治疗乳腺癌的方法。