Mehdipour P, Karami F, Javan Firouzeh, Mehrazin M
Department of Medical Genetics, Tehran University of Medical Sciences, School of Medicine, Keshavarz Boulevard, Pour Sina Street, Tehran, Iran,
Mol Neurobiol. 2015 Aug;52(1):293-302. doi: 10.1007/s12035-014-8864-9. Epub 2014 Aug 27.
Ataxia telangiectasia mutated (ATM) is a key gene in DNA double-strand break (DSB), and therefore, most of its disabling genetic alterations play an important initiative role in many types of cancer. However, the exact role of ATM gene and its epigenetic alterations, especially promoter methylation in different grades of brain tumors, remains elusive. The current study was conducted to query possible correlations among methylation statue of ATM gene, ATM/ retinoblastoma (RB) protein expression, D1853N ATM polymorphism, telomere length (TL), and clinicopathological characteristics of various types of brain tumors. Isolated DNA from 30 fresh tissues was extracted from different types of brain tumors and two brain tissues from deceased normal healthy individuals. DNAs were treated with bisulfate sodium using DNA modification kit (Qiagen). Methylation-specific polymerase chain reaction (MSP-PCR) was implicated to determine the methylation status of treated DNA templates confirmed by promoter sequencing. Besides, the ATM and RB protein levels were determined by immunofluorescence (IF) assay using monoclonal mouse antihuman against ATM, P53, and RB proteins. To achieve an interactive correlation, the methylation data were statistically analyzed by considering TL and D1853N ATM polymorphism. More than 73% of the brain tumors were methylated in ATM gene promoter. There was strong correlation between ATM promoter methylation and its protein expression (p < 0.001). As a triangle, meaningful correlation was also found between methylated ATM promoter and ATM protein expression with D1853N ATM polymorphism (p = 0.01). ATM protein expression was not in line with RB protein expression while it was found to be significantly correlated with ATM promoter methylation (p = 0.01). There was significant correlation between TL neither with ATM promoter methylation nor with ATM protein expression nor with D1853N polymorphism. However, TL has shown strong correlation with patient's age and tumor grade (p = 0.01). Given the important role of cell cycle checkpoint proteins as well as RB and ATM in TL and cancer evolution, further assessment is warranted to shed more light on the pathway linking the telomere instability to tumor progression. High ATM methylation rate in brain tumor patients could open a new avenue toward early screening and cancer therapy.
共济失调毛细血管扩张症突变基因（ATM）是DNA双链断裂（DSB）中的关键基因，因此，其大多数功能丧失性基因改变在多种癌症中发挥着重要的起始作用。然而，ATM基因的确切作用及其表观遗传改变，尤其是不同分级脑肿瘤中的启动子甲基化，仍不清楚。本研究旨在探究ATM基因甲基化状态、ATM/视网膜母细胞瘤（RB）蛋白表达、D1853N ATM多态性、端粒长度（TL）与各类脑肿瘤临床病理特征之间可能存在的相关性。从不同类型的脑肿瘤以及两名已故正常健康个体的脑组织中提取30份新鲜组织的DNA。使用DNA修饰试剂盒（Qiagen）用亚硫酸氢钠处理DNA。甲基化特异性聚合酶链反应（MSP-PCR）用于确定经处理的DNA模板的甲基化状态，并通过启动子测序进行确认。此外，使用抗ATM、P53和RB蛋白的单克隆小鼠抗人抗体，通过免疫荧光（IF）测定法确定ATM和RB蛋白水平。为实现交互相关性，通过考虑TL和D1853N ATM多态性对甲基化数据进行统计分析。超过73%的脑肿瘤在ATM基因启动子处发生甲基化。ATM启动子甲基化与其蛋白表达之间存在强相关性（p < 0.001）。作为一个三角关系，在甲基化的ATM启动子与ATM蛋白表达和D1853N ATM多态性之间也发现了有意义的相关性（p = 0.01）。ATM蛋白表达与RB蛋白表达不一致，但发现其与ATM启动子甲基化显著相关（p = 0.01）。TL与ATM启动子甲基化、ATM蛋白表达或D1853N多态性之间均无显著相关性。然而，TL与患者年龄和肿瘤分级显示出强相关性（p = 0.01）。鉴于细胞周期检查点蛋白以及RB和ATM在TL和癌症演变中的重要作用，有必要进一步评估，以更深入了解将端粒不稳定性与肿瘤进展联系起来的途径。脑肿瘤患者中高ATM甲基化率可能为早期筛查和癌症治疗开辟一条新途径。
