Iws1参与小鼠胚胎干细胞中胚层分化发育

Systematic Elucidation of the Mechanism of the Role of Iws1 in the Early Heart Development

目的揭示Iws1在小鼠胚胎干细胞分化发育中的作用。方法利用CRISPR-Cas9基因编辑建立稳定敲除Iws1的小鼠胚胎干细胞细胞系后,利用悬滴法诱导培养拟胚体(embryoid body,EB),在体外探究该基因是否会影响中胚层的发育。同时分离分别出生1天、7天、21天、56天的小鼠心肌细胞进行该基因的定量检测。并使用GeneMANIA数据库确定该基因的潜在靶标。此外,使用WebGestalt进行基因功能(gene ontology,GO)与信号通路(Kyoto Encyclopedia of Genes and Genomes,KEGG)富集分析。结果在小鼠胚胎干细胞诱导分化体系中,Iws1敲除抑制EB中胚层的发育。该基因在出生后21天的小鼠心肌细胞表现出高表达。Iws1具有19种潜在靶向基因。GO和KEGG分析表明,这些潜在靶基因与氧化还原反应,C型瘦蛋白受体信号通路和其他生理过程有关。结论Iws1参与小鼠胚胎干细胞中胚层分化,可能潜在调控中胚层源性终末分化器官的发育。

Objective To uncover the role of Iws1 in mouse embryonic stem cell differentiation.Methods We applied CRISPR-Cas9 to generate a Iws1 knock out mouse embryonic stem cell(mESC)line,and evaluated the role of Iws1 in mESC differentiation using embryoid body(EB)formation approach.Then,we determined Iws1 expression in cardiomyocytes isolated from hearts at postnatal day 1(P1),7(P7),21(P21)and 56(P56),respectively.Afterwards,we predicted potential target genes of Iws1 using GeneMANIA,and assessed their functional enrichment using gene ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)analyses.Results That the knockout of Iws1 repressed mesoderm specification in EB formation.In addition,Iws1 highly expressed in cardiomyocytes isolated from P21 hearts,compared to that from P1,P7 and P56 hearts.GeneMANIA analysis predicted 19 target genes of Iws1 which were enriched in biological functions related to redox reactions,C-type leptin receptor signaling pathways,and other physiological processes.Conclusion Iws1 is required for mesoderm specification in EB formation which may potentially affect mesoderm-derived organ development.