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ISL1是第二生心区的分子标志,在心血管发育中发挥重要作用.在心脏发育过程中,Isl1的表达具有鲜明的时空特异性.本研究利用P19CL6畸胎瘤干细胞作为心肌分化模型,探讨了Isl1在心肌诱导分化过程中的时间特异性表达及经典Wnt信号通路对其的调控.研究发现,Isl1在心肌分化早期高表达,于诱导第4 d到达高峰,随后快速下调.其表达趋势与经典Wnt通路的激活模式具有时间上的同步性.通过加入Wnt3a蛋白及Li Cl激活经典Wnt通路,能够促进Isl1基因的表达,而Wnt通路抑制分子Frizzled-4/Fc和DKK1能够下调Isl1表达.β-catenin过表达及RNAi实验也获得相似的结果.染色质免疫共沉淀实验证实,Wnt通路效应分子LEF1,在细胞分化第4 d与其在Isl1基因启动子上游-2 300 bp处的结合增强,因而促进了Isl1基因的表达.本研究表明,经典Wnt信号能够通过LEF1/β-catenin与Isl1启动子特异结合,调控Isl1基因在心肌早期分化阶段的表达.
ISL1 is a molecular marker of second-heart area and plays an important role in cardiovascular development.In the process of cardiac development, Isl1 expression has a distinct spatio-temporal specificity.In this study, P19CL6 teratocarcinoma stem cells were used as a model of myocardial differentiation, The expression of Isl1 in myocardium induced differentiation and its regulation by canonical Wnt signaling pathway were studied.It was found that Isl1 was highly expressed in the early stage of myocardial differentiation and peaked on the 4th day of induction and then rapidly downregulated.The expression trend of Isl1 was similar to that of classic Wnt pathway activation pattern with time synchronization.Wnt3a protein and LiCl activation of the canonical Wnt pathway, can promote Isl1 gene expression, and Wnt pathway inhibitor Frizzled-4 / Fc and DKK1 downregulation of Isl1 expression.β- catenin overexpression and RNAi experiments also obtained similar results.Chromatin co-immunoprecipitation experiments confirmed that Wnt pathway effector molecule LEF1, on the 4th day of cell differentiation and its promoter in Isl1 upstream -2300 bp at the junction of enhanced, thus promoting The expression of Isl1 gene.This study shows that canonical Wnt signaling can specifically bind to Isl1 promoter through LEF1 / β-catenin and regulate the expression of Isl1 gene in early myocardial cell The stage of expression.