拟南芥miRNA172b-5p、miRNA172e-5p和miRNA472-3p靶向MSH6基因参与Cd应激响应

生态学杂志

拟南芥miRNA172b-5p、miRNA172e-5p和miRNA472-3p靶向MSH6基因参与Cd应激响应

成智博^1,5,王鹤潼²,赵强³,张延召⁴,贾春云¹,何蕾^1,5,崔伟娜^1,5,台培东¹,刘宛^1*

(¹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；²沈阳大学，沈阳 110044；³沈阳农业大学，沈阳 110866；⁴洛阳师范学院，河南洛阳 471022； ⁵中国科学院大学，北京 100049)

出版日期:2019-12-10 发布日期:2019-12-10

MiRNA172b-5p, miRNA172e-5p and miRNA472-3p responded to Cd stress by targeting MSH6 gene in Arabidopsis thaliana.

CHENG Zhi-bo^1,5, WANG He-tong², ZHAO Qiang³, ZHANG Yan-zhao⁴, JIA Chun-yun¹, HE Lei^1,5, CUI Wei-na^1,5, TAI Pei-dong¹, LIU Wan^1*

(¹Key Laboratory of Pollution Ecology and Environmental Engineering, Institute of Applied Ecology, Chinese Academy of Sciences, Shenyang 110016, China; ²Shenyang University, Shenyang 110044, China; ³Shenyang Agricultural University, Shenyang 110866, China; ⁴Luoyang Normal University, Luoyang 471022, Henan, China; ⁵University of Chinese Academy of Sciences, Beijing 100049, China).

Online:2019-12-10 Published:2019-12-10

摘要/Abstract

摘要： MSH6在激活细胞周期阻滞和修复镉诱导的DNA损伤等方面具有重要功能。本研究以拟南芥为实验材料，通过生物信息学分析发现，MSH6具有miRNA172b-5p、miRNA172e-5p、miRNA472-3p等microRNAs (miRNAs) 的作用靶点。利用烟草双荧光素酶报告系统体外验证检测发现，与阴性对照组相比，miRNA172b-5p、miRNA172e-5p和miRNA472-3p均可显著抑制荧光素酶的相对活性，其比值分别下降了25.5%、47.2%和49.5%。采用拟南芥植株瞬时侵染体内验证结果显示，miRNA172b-5p、miRNA172e-5p和miRNA4723p分别上升了178.9%、123.6%和37.6%；同时，qRT-PCR（real time quantitative reverse transcript polymerase chain reaction）结果表明，拟南芥幼苗在1.25、2.5、4.0 mg·L^-1 Cd胁迫7 d后，与对照组相比，MSH6表达显著降低，分别下降了20.3%、22.7%和38.2%。而miRNA172b-5p、miRNA172e-5p和miRNA472-3p表达显著增加，二者呈显著的负相关关系(r=-0.997，P<0.01；r=-0.997，P<0.01；r=-0.952，P<0.05)，表明miRNA172b-5p、miRNA172e-5p和miRNA472-3p是拟南芥Cd胁迫响应的miRNAs，并且靶向MSH6基因。本研究为DNA mismatch repair (MMR)对Cd毒理调控机制的研究提供miRNA理论基础。

关键词: 火烧迹地, 土壤有机碳, 土壤微生物生物量碳, 土壤可溶性有机碳

Abstract: MSH6 plays an important role in the activation of checkpoints in cell cycles and in repairing of DNA damage under Cd stress. In this study, we found that MSH6 gene had targeting sequence of microRNAs (miRNAs), including miRNA172b-5p, miRNA172e-5p and miRNA472-3p in Arabidopsis thaliana by bioinformatics analysis. The results of dual luciferase reporter assay showed that miRNA172b-5p, miRNA172e-5p, and miRNA472-3p significantly reduced the related luciferase activity of reporter construct, which was decreased by 25.5%, 47.2% and 49.5%, respectively, compared with the negative control group in an Agrobacterium tumefaciensmediated in vitro assay (P<0.01). Results from Arabidopsis thaliana leaves injected transient expression in vivo assay showed that the over-expression level of miRNA172b-5p, miRNA172e-5p and miRNA472-3p was 178.9%, 123.6% and 37.6%, respectively. Results of qRT-PCR(real time quantitative reverse transcript polymerase chain reaction) showed that the expression of MSH6 was decreased by 20.3%, 22.7% and 38.2% in Arabidopsis thaliana leaves when exposed to 1.25, 2.5, 4.0 mg·L^-1 Cd stress for seven days, but the expression levels of miRNA172b-5p, miRNA172e-5p and miRNA472-3p were enhanced. There were negative relationships between these three miRNAs and MSH6gene (r=-0.995, P<0.01; r=-0.995, P<0.01; r=-0.984, P<0.05), indicating that miRNA172b-5p, miRNA172e-5p and miRNA472-3p were Cd responsive miRNAs and could directly target toMSH6 gene in Arabidopsis thaliana. Ourresults provide theoretical basis of miRNA for the study of Cd toxicological mechanism regulated by DNA mismatch repair (MMR) system.

Key words: burned area, soil dissolved organic carbon., soil organic carbon, soil microbial biomass carbon

成智博,王鹤潼,赵强,张延召,贾春云,何蕾,崔伟娜,台培东,刘宛. 拟南芥miRNA172b-5p、miRNA172e-5p和miRNA472-3p靶向MSH6基因参与Cd应激响应[J]. 生态学杂志.

CHENG Zhi-bo, WANG He-tong, ZHAO Qiang, ZHANG Yan-zhao, JIA Chun-yun, HE Lei, CUI Wei-na, TAI Pei-dong, LIU Wan. MiRNA172b-5p, miRNA172e-5p and miRNA472-3p responded to Cd stress by targeting MSH6 gene in Arabidopsis thaliana.[J]. cje.

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