作者投稿/查稿 Open Access
高级检索
潘雪莉, 张爱华. 亚砷酸钠对人皮肤角质形成细胞MGMT基因组蛋白乙酰化及转录与表达的影响[J]. 环境与职业医学, 2011, 28(4): 210-214.
引用本文: 潘雪莉, 张爱华. 亚砷酸钠对人皮肤角质形成细胞MGMT基因组蛋白乙酰化及转录与表达的影响[J]. 环境与职业医学, 2011, 28(4): 210-214.
shu
PAN Xue-li , ZHANG Ai-hua . Effects of Sodium Arsenite on Histone Acetylation, Transcription and Expression of O6-Methylguanine-DNA Methyltransferase Gene in HaCaT Cells[J]. Journal of Environmental and Occupational Medicine, 2011, 28(4): 210-214.
Citation: PAN Xue-li , ZHANG Ai-hua . Effects of Sodium Arsenite on Histone Acetylation, Transcription and Expression of O6-Methylguanine-DNA Methyltransferase Gene in HaCaT Cells[J]. Journal of Environmental and Occupational Medicine, 2011, 28(4): 210-214.
shu

亚砷酸钠对人皮肤角质形成细胞MGMT基因组蛋白乙酰化及转录与表达的影响

Effects of Sodium Arsenite on Histone Acetylation, Transcription and Expression of O6-Methylguanine-DNA Methyltransferase Gene in HaCaT Cells

    摘要
  • 摘要: 目的 了解不同剂量亚砷酸钠(NaAsO2)对人皮肤角质形成细胞(HaCaT 细胞)中O6-甲基鸟嘌呤DNA甲基转移酶(MGMT) 基因组蛋白乙酰化调控、mRNA转录及蛋白表达的影响。

     方法 以25.00、12.50、6.25、3.13 μmol/LNaAsO2 重复间隔处理HaCaT 细胞72 h(NaAsO2 处理24 h,隔天再次用NaAsO2 进行相同的处理,重复处理3 次)。定量染色质免疫沉淀技术(Q-ChIP)检测MGMT 基因转录调控区(ChIP1、ChIP2 区域)及MGMT 基因编码区(ChIP3 区域,对照区域)组蛋白乙酰化修饰水平;实时荧光定量PCR(Q-PCR)和免疫印迹分析分别检测MGMT 基因的mRNA转录和蛋白表达。设不染毒的HaCaT 细胞为空白对照(0.00 μmol/L),人表皮鳞癌细胞株A431 为阳性对照。

     结果 0.00、3.13、6.25、12.50、25.00 μmol/L NaAsO2 处理细胞中,MGMT 基因转录调控区ChIP1 区域的组蛋白H3K9 乙酰化水平分别为176.68& #177;8.50、175.71& #177;18.14、161.26& #177;16.28、146.23& #177;24.00 和82.64& #177;33.87,差异有统计学意义(F=28.809,P<0.05);组蛋白H4 乙酰化水平分别为183.59& #177;11.98、180.84& #177;24.10、166.52& #177;5.48、156.87& #177;10.64 和103.42& #177;7.04,差异有统计学意义(F=36.493,P<0.05)。ChIP2 区域的组蛋白H3K9 乙酰化水平分别为171.11& #177;16.54、167.55& #177;8.97、156.51& #177;8.59、135.88& #177;16.55 和82.01& #177;3.96,差异有统计学意义(F=49.626,P<0.05);组蛋白H4 乙酰化水平分别为117.23& #177;16.21、143.29& #177;10.59、135.87& #177;7.44、105.48& #177;7.56 和78.79& #177;6.92,差异有统计学意义(F=25.438,P<0.05)。编码区ChIP3 区域的组蛋白H3K9 乙酰化水平分别为37.53& #177;6.23、35.57& #177;5.85、40.81& #177;4.45、42.18& #177;1.23 和41.87& #177;5.71,差异无统计学意义(F=2.341,P>0.05);组蛋白H4 乙酰化水平分别为40.78& #177;2.42、38.56& #177;4.66、39.47& #177;2.88、33.13& #177;3.48和31.48& #177;4.99,差异无统计学意义(F=3.027,P>0.05)。MGMT 基因mRNA转录相对表达量分别为1.198 29& #177;0.159 97、1.518 31& #177;0.180 54、1.425 22& #177;0.180 39、1.014 54& #177;0.096 79 和0.887 72& #177;0.020 00,差异有统计学意义(F=37.359,P <0.05);MGMT 蛋白相对表达量分别为1.066 19& #177;0.061 24、1.174 47& #177;0.064 75、0.848 83& #177;0.057 01、0.471 63& #177;0.023 34 和0.240 34& #177;0.014 43,差异有统计学意义(F=20.687,P<0.05)。

     结论 砷通过导致MGMT 基因转录调控区组蛋白去乙酰化,抑制MGMT 基因mRNA转录及蛋白表达,是砷致皮肤损害的重要机制之一。

     

    Abstract: Objective To observe the influences of different doses of sodium arsenite on histone acetylation regulation, mRNA transcription and protein expression of O6-methylguanine-DNA methyltransferase gene(MGMT)in HaCaT cells.

    Methods HaCaT cells were treated with 25.00, 12.50, 6.25 and 3.13 μmol/L NaAsO2 for 72 h at intervals and repeatedly. Histone acetylation modifications in two transcription regulatory region(ChIP1, ChIP2 region)and in coding region(ChIP3 region, the control region)of MGMT gene were detected by chromatin immuno-precipitation combined with quantitative PCR, the mRNA transcription and the protein expression of MGMT were detected by real-time quantitative PCR and Western blot. HaCaT cells untreated with NaAsO2(0.00 μmol/L)were set as the blank control group, human epidermal squamous carcinoma cell line A431 cells were set as the positive control group.

    Results Among the groups of HaCaT cells treated with 0.00, 3.13, 6.25, 12.50 and 25.00 μmol/L NaAsO2, the levels of histone acetylation of H3K9 in ChIP1 transcription regulatory region of MGMT gene were 176.68& #177;8.50, 175.71& #177;18.14, 161.26& #177;16.28, 146.23& #177;24.00 and 82.64& #177;33.87 respectively, the differences were significant(F=28.809, P<0.05). The levels of histone acetylation of H4 in ChIP1 transcription regulatory region were 183.59& #177;11.98, 180.84& #177;24.10, 166.52& #177;5.48, 156.87& #177;10.64 and 103.42& #177;7.04, the differences were significant(F=36.493, P<0.05). The levels of histone acetylation of H3K9 in ChIP2 transcription regulatory region were 171.11& #177;16.54, 167.55& #177;8.97, 156.51& #177;8.59, 135.88& #177;16.55 and 82.01& #177;3.96, the differences were significant(F=49.626, P<0.05). The levels of histone acetylation of H4 in ChIP2 transcription regulatory region were 117.23& #177;16.21, 143.29& #177;10.59, 135.87& #177; 7.44, 105.48& #177;7.56 and 78.79& #177;6.92, the differences were significant(F=25.438, P<0.05). The levels of histone acetylation of H3K9 in ChIP3 coding region were 37.53& #177;6.23, 35.57& #177;5.85, 40.81& #177;4.45, 42.18& #177;1.23 and 41.87& #177;5.71, the differences were not significant(F=2.341, P>0.05). The levels of histone acetylation of H4 in ChIP3 coding region were 40.78& #177;2.42, 38.56& #177;4.66, 39.47& #177;2.88, 33.13& #177;3.48 and 31.48& #177;4.99, the differences were not significant(F=3.027, P>0.05).The levels of MGMT mRNA transcription were 1.198 29& #177;0.159 97, 1.518 31& #177;0.180 54, 1.425 22& #177;0.180 39, 1.014 54& #177;0.096 79 and 0.887 72& #177;0.020 00, the differences were significant(F=37.359, P<0.05). The levels of MGMT protein expression were 1.066 19& #177;0.061 24, 1.174 47& #177; 0.064 75, 0.848 83& #177;0.057 01, 0.471 63& #177;0.023 34 and 0.240 34& #177;0.014 43, the differences were significant(F=20.687, P<0.05).

    Conclusion Arsenic can cause histone deacetylation of transcription regulatory region of MGMT gene, which results in MGMT mRNA transcription and protein expression to be inhibited. It might be one of important mechanisms of arsenic-induced skin lesion.

     

    • HTML全文
    • 参考文献(0)
    • 相关文章
    • 施引文献
  • 补充资料
    • 审稿意见
    • 勘误/撤稿

/

返回文章
返回
  • wechat

    • WeChat