| DNA甲基转移酶3B4过表达对293A细胞增殖影响及其机制的探讨 |
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| 引用本文： | 邹明新,姜树原,张姝,闫少春,邵国,刘晓光. DNA甲基转移酶3B4过表达对293A细胞增殖影响及其机制的探讨[J]. 中国癌症杂志, 2014, 24(2): 99-105. DOI: 10.3969/j.issn.1007-3969.2014.02.004 |
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| 作者姓名： | 邹明新 姜树原 张姝 闫少春 邵国 刘晓光 |
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| 作者单位： | 1.包头师范学院生命科学与技术学院，内蒙古 包头 014030；2.包头医学院生物医学研究中心及基础医学院，内蒙古 包头 014060 |
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| 基金项目： | 国家自然科学基金(No: 81060212，81160244，81360316)；中国博士后基金(No: 20080430851)；内蒙古自然科学基金(No: 2010BS1104)；内蒙古自治区高等学校科研项目(No: NJZY12221，NJZY12225)；内蒙古自治区高等学校青年科技英才支持计划资助(No: NJYT-13-A10)；教育部留学回国人员科研启动基金（No:46批） |
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| 摘 要： | 背景与目的：DNA甲基转移酶(DNA methyltransferase，DNMT)3B有近40种异构体，它们的表达有组织和疾病的特异性，细胞中这些异构体的功能尚不清楚。本研究旨在探讨外源性DNMT3B4基因过表达对人胚肾细胞株293A增殖的影响及其机制。方法：将携带DNMT3B4基因的质粒pCMV-DNMT3B4及空质粒pCMV-2B转染293A细胞(293A-vector)并形成稳定表达细胞系，培养300 d后用MTT法检测细胞的增殖；流式细胞仪检测细胞的周期分布；real-time PCR蛋白质印迹法(Western blot)检测细胞中p21表达情况；甲基化特异PCR(MS-PCR)检测p21基因启动子区甲基化状态。结果：在接种于96孔板的第4天，2株过表达DNMT3B4的293A细胞(293A-DNMT3B4-1和293A-DNMT3B4-2)吸光度(A)值分别是293A-vector细胞的(58.92±3.47)%和(68.82±5.64)%，过表达DNMT3B4抑制细胞增殖，差异有统计学意义(P<0.05)；293A-DNMT3B4-1和293A-DNMT3B4-2的S期细胞比例分别为(35.88±2.00)%和(37.00±1.79)%，293A-vector细胞比例为(40.44±0.91)%，过表达DNMT3B4降低S期细胞比例，差异有统计学意义(P<0.05)。过表达DNMT3B4增加p21的表达，但不改变p21基因启动子区甲基化状态。结论：DNMT3B4基因过表达可抑制293A细胞增殖并增加p21表达。
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| 关 键 词： | DNA甲基转移酶 3B 293A细胞 细胞增殖 |
Effect of overexpression of DNA methyltransferase 3B4 gene on proliferation of 293A cells |
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| ZOU Ming-xin,JIANG Shu-yuan,ZHANG Shu,YAN Shao-chun,SHAO Guo,LIU Xiao-guang. Effect of overexpression of DNA methyltransferase 3B4 gene on proliferation of 293A cells[J]. China Oncology, 2014, 24(2): 99-105. DOI: 10.3969/j.issn.1007-3969.2014.02.004 |
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| Authors: | ZOU Ming-xin JIANG Shu-yuan ZHANG Shu YAN Shao-chun SHAO Guo LIU Xiao-guang |
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| Affiliation: | 1.Biology Department, BaoTou Teachers College, Baotou Inner Mongolia 014030, China;2. Biomedicine Research Center and Basic Medical College, BaoTou Medical College, Baotou Inner Mongolia 014060, China |
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| Abstract: | Background and purpose: DNMT3B has nearly 40 known splice variants expressed in a tissueand disease-specific manner, but the roles of these splice variants in the cell are still unclear. The aim of this study was to investigate the effects of overexpression of DNA methyltransferase 3B4 (DNMT3B4) gene on proliferation of human embryo kidney 293A cells. Methods: 293A cells were transfected with plasmid pCMV-DNMT3B4 or pCMV-2B and then treated with G418 to get the stable cell line. The stable cell lines were determined for proliferation level by MTT method, and for cell cycle distribution by flow cytometry. The expression of p21 was detected by real-time PCR and Western blot. The methylation status of p21 gene promoter was detected by methylation-specific PCR (MS-PCR). Results: The absorbance value in DNMT3B4-1 and DNMT3B4-2 clone were (58.92±3.47)% and (68.82±5.64)% as compared to 293A-vector cells using MTT method. DNMT3B4 overexpression significantly decreased cell proliferation (P<0.05). S phase fraction of 293A-vector cells was (40.44±0.91)%. While in DNMT3B4-1 and DNMT3B4-2 clone cells, the S phase fraction was (35.88±2.00)% and (37.00±1.79)% respectively. Overexpression of DNMT3B4 could significantly decrease S phase fraction (P<0.05). The expression of p21 in DNMT3B4 overexpressed cells was increased, but the methylation status of p21 gene promoter was unchanged.Conclusion: Overexpression of DNMT3B4 can inhibit 293A cell proliferation and can facilitate p21 expression. |
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| Keywords: | DNA methyltransferase 3B 293A cell Cell proliferation |
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