mRNA成熟调控因子NUDT21对K562细胞转录本选择性剪接的影响

Effect of NUDT21 on Alternative Splicing of Transcripts in K562 Cells

目的:研究mRNA前体3′端剪接调控因子NUDT21对白血病K562细胞转录本选择性剪接的影响。方法:以白血病K562细胞为研究对象,通过慢病毒敲减Nudix水解酶21(Nudix hydrolase 21,NUDT21)后,应用转录组测序RNA-Seq方法明确干扰前、后转录本的表达水平。生物信息学方法分析差异表达基因及可变剪接的变化,并用qPCR验证。结果:NUDT21敲减后K562细胞基因差异表达表现:上调5196条,下调3917条;转录本差异表达表现:上调22200条,下调19728条。通过GO和KEGG分析,NUDT21敲减后表达显著差异的转录本主要与细胞粘附分化、造血细胞系及自身免疫等过程相关。差异显著的选择性剪接共有436个,主要涉及细胞增殖、代谢等多个生物学过程的调控。ERBB2、MAPK激酶MKNK2、G蛋白偶联受体GRK6、真核细胞翻译延伸因子EEF1B2、细胞周期蛋白CCNL2、有丝分裂检查点蛋白BUB3等出现m RNA前体3′端的剪接改变。其中ERBB2mRNA变体1的表达减少,变体4的表达增多。结论:NUDT21通过包括mRNA前体3′端可变剪接在内的多种调控方式从较高层面影响细胞的生物学功能。

Objective:To explore the effect of nudix hydrolase 21(NUDT 21)on alternative splicing of transcripts in leukemia K562 cells.Methods:The K562 cell line was used as the research objects.The NUDT 21 was knocked-down by lentivirus vector,then the expression of transcripts before and after interference was determined by transcriptome sequencing(RNA seq).The bioinformatics methods including GO analysis and KEGG pathway analysis were used to analyze the changes of differentially expressed genes and 3′alternative splicing,then these changes were confirmed by q PCR.Results:After the NUDT 21 in K562 cells was knoced-down,the differentially expressed genes showed that 5196 were up-regulated,3917 were down-regulated.GO analysis and KEGG pathway analysis showed that the very differentially expressed transcripts mainly related with cell adhesion and differentiation,hematopoietic cell lines and autoimmunity.There were 436 significant alternative splicing,which mainly involved in the regulation of some biological processes such as cell proliferation and metabolism.The ERBB2,MAPK kinase MKNK2,G protein-coupled receptor GRK6,eukaryotic translation elongation factor EEF1 B2,cyclin L2 CCNL2,mitotic checkpoint protein BUB3 were changed by 3′alternative splicing.Among them the expression of variant 1 of ERBB2 mRNA decreased and variant 4 increased.Conclusion:NUDT21 influences the cell biological function at a higher level by variously regulating ways,including 3′end APA.