拟南芥MLH1基因启动子甲基化作为镉胁迫生物标记物潜力评估  

Methylation of MLH1 promoter in Arabidopsis thaliana as a biomarker of cadmium stress.

作　　者：何蕾[1,2] 王鹤潼[2] 宋婕[1,2] 崔伟娜[2,3] 曹霞[2] 惠秀娟[1] 台培东[2] 杨悦锁[4] 刘宛[2] 

机构地区：[1]辽宁大学环境学院,沈阳110036 [2]中国科学院沈阳应用生态研究所污染生态与环境工程重点实验室,沈阳110016 [3]上海应用技术学院,上海201418 [4]沈阳大学区域污染环境生态修复教育部重点实验室,沈阳110044

出　　处：《生态学杂志》2016年第5期1386-1393,共8页Chinese Journal of Ecology

基　　金：国家自然科学基金项目(21347007;40930739;20977095);沈阳大学区域污染环境生态修复教育部重点实验室基金项目(41472237)资助

摘　　要：应用重亚硫酸盐测序技术,研究了镉(Cd)胁迫21 d后拟南芥幼苗错配修复基因Mut L-homologue1(MLH1)启动子甲基化的变化趋势。结果显示,拟南芥MLH1启动子区域为-346^+42(391 bp),含有71个胞嘧啶:16个Cp G、6个CHG(H为C、A或T)和49个CHH位点。对照植株Cp G、CHH和CHG位点的甲基化率分别为44.8%、40.5%和52.0%。随Cd胁迫浓度的增加,MLH1启动子区胞嘧啶位点发生超甲基化和去甲基化的位点数逐渐增加;这些位点的甲基化率均呈上升趋势,且均高于对照组(除CHG位点外)。MLH1启动子区域71个胞嘧啶中,Cp G6、Cp G9、CHH44和CHG4位点的甲基化多态性对Cd胁迫更敏感,且具有剂量-效应关系;其中0.25 mg·L-1Cd胁迫下,Cp G9位点为超甲基化,其甲基化变化率为20.0%,其他3个位点为去甲基化,其甲基化变化率分别为12.0%、20.0%和20.0%。相对于幼苗的其他生物学性状(叶片数、生物量及叶绿素含量),Cd胁迫下MLH1启动子甲基化变化更显著且敏感,上述4个胞嘧啶热点的甲基化多态性可作为检测Cd胁迫对植物遗传毒性效应潜在的、有效的生物标记物。Using bisulfite DNA sequencing technique, this study investigated the methylation pat- terns of 71 cytosine residues including 16 CpG sites, 6 CHG sites （H as C, A or T） and 49 CHH sites within the region 391 bp between nucleotides -346 to ＋42 in MLH1 promoter region in Arabidopsis plantlets exposed to 0-5.0 mg~ L-1 cadmium （Cd） for 21 days. The methylation per- centages of CpG, CHH and CHG sites were 44.8%, 40.5% and 52.0%, respectively, in the control. With the increasing Cd level, the hyper- and hypomethylated cytosine residues, and the methylation percentages of those residues for CpG, CHG and CHH sites were increased in the Cd-treated seedlings, and were higher at CpG and CHH sites than in the controls except for CHG sites. Among 71 cytosines in MLH1 promoter regions, methylation polymorphism of CpG6, CpG9, CHH44 and CHG4 sites was much more sensitive to Cd stress than other cytosine resi- dues, and the effect was dose-dependent. The percentage of hypermethylation change for CpG9 site was 20.0%, while the percentages of hypomethylation change for CpG6, CHH44 and CHG4 sites were 12.0%, 20.0% and 20.0%, respectively, in the Cd-expoesd seedlings at Cd level of 0.25 mg·L-1. The presence of significant methylation polymorphism of cytocine residues for the above four hot spots in MLH1 promoter of Cd-treated seedlings, prior to the onset of leaf number, fresh mass and chlorophyll content of shoots, suggests that methylation polymorphisms of these hot loci are the most sensitive biomarkers for early diagnosis and risk assessment of genotoxic effects of Cd pollution.

关 键 词：Cd MLH1基因启动子 生物标记物 拟南芥 甲基化 

分 类 号：Q945.78[生物学—植物学] Q943.2