苯并芘诱导细胞转化过程中PP2AB56ε表达与DNA损伤修复功能的研究

摘要

目的：探讨蛋白磷酸酶2A-B56ε亚基(PP2A B56ε,由PPP2R5E基因编码)的表达在苯并芘诱导细胞转化过程中对DNA损伤修复功能的影响。方法实时荧光定量-PCR(QRT-PCR)检测BEAS-2B细胞(人正常肺上皮细胞)、和BEAS-2BT细胞(苯并芘诱导恶变的人肺上皮细胞)中PPP2R5E的mRNA转录水平；免疫印迹法(Western Blot)检测细胞中 B56ε编码蛋白的水平；中性单细胞凝胶电泳实验(彗星实验,SCGE)检测喜树碱( camptothecin,CPT)处理的细胞DNA断裂损伤及修复情况；免疫印迹法检测 CPT 处理诱导γH2AX(磷酸化组蛋白 H2AX)水平。结果与 BEAS-2B 细胞相比较,BEAS-2BT 细胞中PPP2R5E其mRNA水平及B56ε蛋白水平均显著高表达(P<0.05)。与BEAS-2B细胞相比,经CPT处理恢复后,BEAS-2BT细胞尾距显著降低(P<0.05)。表明BEAS-2BT细胞中H2AX的磷酸化水平明显降低。结论在苯并芘诱导细胞转化过程中PP2A B56ε通过介导γH2AX去磷酸化影响DNA损伤修复功能。%Objective To investigate protein phosphatase 2A-B56εsubunit(PP2A B56ε,encoded by the gene PPP2R5E) expression to DNA damage and repair functions during benzopyrene induced cell transformation. Methods Real-time quantitative-PCR ( QRT-PCR) was used to detect the BEAS-2B cells (normal human lung epithelial cells) and BEAS-2BT cells (benzo(α)pyrene induced malignant human lung epithelial cells) mRNA transcript levels in PPP2R5E. SCGE assay was adopted to evaluate their genotoxicity. Western blot was used to detect theγH2AX(phosphorylated histone H2AX ) level. Results Compared with the BEAS-2B cells,the mRNA and pro-tein encoded by PPP2R5E and its expression levels were significantly higher(P<0. 05). SCGE analysis showed that,compared with BEAS-2B cells,BEAS-2BT cells tail was significantly reduced (P<0. 05). Compared with BEAS-2B cells,γH2AX expression levels of BEAS-2BT cells were significantly higher(P<0. 05). Conclusions In the process of benzo(α)pyrene induced cell transformation, PP2A B56εaffects DNA damage and repair by mediating throughγH2AX dephosphorylation.