BTG3 tumor suppressor gene promoter demethylation, histone modification and cell cycle arrest by genistein in renal cancer
BTG3 tumor suppressor gene promoter demethylation, histone modification and cell cycle arrest by genistein in renal cancer
复制标题
DOI：
10.1093/carcin/bgp042
复制
发表时间：
2009-04-01
期刊：
影响因子：
4.7
通讯作者：
Dahiya, Rajvir
中科院分区：
文献类型：
--
作者：
Majid, Shahana;Dar, Altaf A.;Dahiya, Rajvir
BTG 3/ANA/APRO 4基因在某些恶性肿瘤中是一个抑癌基因。它构成Src介导的信号传导的重要负调节机制，是细胞周期的负调节剂，并抑制转录因子E2 F1。我们报告说，BTG 3在肾癌中下调，失活的机制是通过启动子超甲基化。实时定量PCR结果显示，BTG 3在肿瘤组织和细胞中表达下调。Genistein和5-Aza-C诱导A498、ACHN和HEK-293肾细胞癌（RCC）细胞系中BTG 3信使RNA（mRNA）的表达。亚硫酸氢盐修饰的PCR和DNA测序结果显示，在肿瘤样品和癌细胞系中，BTG 3启动子完全甲基化。Genistein和5Aza-C处理显著降低启动子甲基化，重新激活BTG 3表达。染色质免疫沉淀分析显示，染料木黄酮和5氮杂-C增加乙酰化组蛋白3，4，2 H3 K4，3 H3 K4和RNA聚合酶II的水平在BTG 3启动子指示活性组蛋白修饰。酶活性测定结果表明，染料木黄酮和5-氮杂-C降低DNA甲基转移酶和甲基化CpG结合结构域2的活性，增加HAT的活性。细胞周期和3-（4，5-二甲基噻唑-2-基）-2，5-联苯四氮唑溴化物细胞增殖实验表明，染料木黄酮通过诱导细胞周期阻滞对癌细胞生长具有抗增殖作用。这是第一个报告表明，BTG 3是表观遗传沉默在肾细胞癌，并可以重新激活染料木素诱导的启动子去甲基化和活性组蛋白修饰。染料木黄酮具有类似于5-氮杂-C的作用，5-氮杂-C是一种强有力的脱甲基剂，具有高毒性和不稳定性。染料木黄酮是一种天然、无毒的膳食补充剂，可有效抑制肾癌细胞的生长，使其成为肾癌表观遗传学治疗的一个有希望的候选药物。
BTG3/ANA/APRO4 has been reported to be a tumor suppressor gene in some malignancies. It constitutes important negative regulatory mechanism for Src-mediated signaling, a negative regulator of the cell cycle and inhibits transcription factor E2F1. We report that BTG3 is downregulated in renal cancer and that the mechanism of inactivation is through promoter hypermethylation. Quantitative real-time polymerase chain reaction (PCR) showed that BTG3 was downregulated in cancer tissues and cells. Genistein and 5-aza-2'-deoxycytidine (5Aza-C) induced BTG3 messenger RNA (mRNA) expression in A498, ACHN and HEK-293 renal cell carcinoma (RCC) cell lines. Bisulfite-modified PCR and DNA sequencing results showed complete methylation of BTG3 promoter in tumor samples and cancer cell lines. Genistein and 5Aza-C treatment significantly decreased promoter methylation, reactivating BTG3 expression. Chromatin immunoprecipitation assay revealed that genistein and 5Aza-C increased levels of acetylated histones 3, 4, 2H3K4, 3H3K4 and RNA polymerase II at the BTG3 promoter indicative of active histone modifications. Enzymatic assays showed genistein and 5Aza-C decreased DNA Methyltransferase, methyl-CpG-binding domain 2 activity and increased HAT activity. Cell cycle and 3-(4,5-dimethylthiazole-2-yl)-2,5-biphenyl tetrazolium bromide cell proliferation assays showed that genistein has antiproliferative effect on cancer cell growth through induction of cell cycle arrest. This is the first report to show that BTG3 is epigenetically silenced in RCC and can be reactivated by genistein-induced promoter demethylation and active histone modification. Genistein had similar effects to that of 5Aza-C, which is a potent demethylating agent with high toxicity and instability. Genistein being a natural, non-toxic, dietary isoflavone is effective in retarding the growth of RCC cells, making it a promising candidate for epigenetic therapy in renal carcinoma.