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  • 李培昕,张星燎,周杨,等.lncRNA-135调控miR-592维持小鼠胚胎干细胞多能性[J].同济大学学报(医学版),2020,41(5):552-560.    [点击复制]
  • LI Pei-xin,ZHANG Xing-liao,ZHOU Yang,et al.lncRNA-135 maintains the pluripotency of mouse embryonic stem cells by functioning as a miRNA sponge of miR-592[J].同济大学学报(医学版),2020,41(5):552-560.   [点击复制]
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lncRNA-135调控miR-592维持小鼠胚胎干细胞多能性
李培昕,张星燎,周杨,张宁彦,张敬,张军
0
(同济大学医学院,上海200092;同济大学生命科学与技术学院,上海200092)
摘要:
目的筛选并确定调控miR-592作用的关键lncRNA,探讨其对miR-592沉默靶基因cep135的调控作用。方法利用lncRNA基因芯片筛选小鼠胚胎干细胞(mouse embryonic stem cells, mESCs)及KO miR-592 mESCs中差异的lncRNA从而找出关键调控分子lncRNA-135。利用转染试剂干扰lncRNA-135的表达,使用Western印迹法、双荧光素酶报告系统及免疫细胞化学染色等技术检测lncRNA-135对miR-592及其靶基因cep135的表达影响,进而阐明lncRNA-135对mESCs多能性的调控作用。结果lncRNA基因芯片分离确定了527条差异表达的lncRNA,经过生物信息学分析及qRT-PCR验证,确定了评分最高的lncRNA-135为关键作用分子。双荧光素酶报告系统证实了lncRNA-135能够结合miR-592以干扰其沉默靶基因cep135。而Western印迹法和免疫细胞化学染色结果则证实lncRNA-135通过作用于miR-592,阻止其对cep135的抑制作用,进而促进mESCs的多能性。结论lncRNA-135可以通过充当miR-592的“分子海绵”从而调节与mESCs多能性维持有关的多个基因的表达,这将可能帮助建立调控性miRNA网络与mESCs多能性之间的联系。
关键词:  lncRNA-135  miR-592  cep135  胚胎干细胞  多能性
DOI:10.16118/j.1008-0392.2020.05.003
投稿时间:2020-02-22
基金项目:国家自然科学基金面上项目(81771417)
lncRNA-135 maintains the pluripotency of mouse embryonic stem cells by functioning as a miRNA sponge of miR-592
LI Pei-xin,ZHANG Xing-liao,ZHOU Yang,ZHANG Ning-yan,ZHANG Jing,ZHANG Jun
(Tongji University School of Medicine, Shanghai 200092, China;Tongji University School of Life Science and Technology, Shanghai 200092, China)
Abstract:
ObjectiveTo screen and identify key lncRNAs that regulate the function of miR-592 and its mechanisms. MethodsThe key regulatory molecules that regulate miR-595 were screened and identified with lncRNA chip in mouse embryonic stem cells(mESCs) of wild-type and miR-592 knockout(KO) mice. The expression of lncRNA-135 was knocked down by transfection of interference fragments. The effect of lncRNA-135 on the expression of miR-592 and its target gene cep135 was confirmed by Western blot, dual luciferase reporter system and immunocytochemistry. Furthermore, the regulation of mESCs pluripotency by lncRNA-135 was verified. ResultsThrough lncRNA gene chip screening, 527 differentially expressed lncRNAs were identified. After bioinformatics analysis and qRT-PCR verification, the lncRNA-135 with the highest score was identified as the key lncRNA. The dual luciferase reporter system confirmed that lncRNA-135 was able to bind miR-592 to interfere with its silencing target gene cep135. Western blot and immunocytochemical results confirmed that lncRNA-135 promoted the pluripotency of mESCs by inhibiting the miR-592-cep135 system. ConclusionThe results suggest that lncRNA-135 can act as a sponge for miR-592 to regulate the expression of multiple genes related to the maintenance of mESCs pluripotency.
Key words:  lncRNA-135  miR-592  cep135  embryonic stem cells  pluripotency

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