Cell biology and Stem Cells Unit, Center for Cooperative Research in Biosciences, CIC bioGUNE, 48160 Derio, Spain.
Stem Cells. 2011 Jan;29(1):141-53. doi: 10.1002/stem.562.
Retinoic acid and Wnt/β-catenin signals play important roles during neuronal differentiation but less is known about noncanonical Wnt signals in this context. We examined retinoic acid and Wnt signaling in two human embryonal carcinoma cell lines, NTERA-2 (clone D1), which undergoes neuronal differentiation in response to retinoic acid, and 2102Ep, which does not. Retinoic acid treatment inhibited β-catenin/Tcf activity in NTERA-2 cells but not in 2102Ep cells. Inhibition occurred downstream of β-catenin but did not involve competition between retinoic acid receptors and β-catenin for binding to p300 or Tcf-4. Ectopic expression of FZD1 partially restored inhibition in 2102Ep cells, suggesting the involvement of Wnt ligands. Retinoic acid treatment of NTERA-2 cells induced the expression of Wnt-4 and Wnt-11, both of which were able to inhibit β-catenin/Tcf activity. Wnt-4 and Wnt-11 were found at cell borders in islands of cells that expressed OCT4 and GFAP and were predominantly negative for Nestin, PAX6, and GATA6. Gene silencing of Wnt-4, but not Wnt-11, reduced retinoic acid downregulation of OCT4 and Nanog and upregulation of PAX6, ASCL1, HOXC5, and NEUROD1, suggesting that Wnt-4 promotes early neuronal differentiation. Gene expression analysis of NTERA-2 cells stably overexpressing Wnt-11 suggested that Wnt-11 potentiates retinoic acid induction of early neurogenesis. Consistent with this, overexpression of Wnt-11 maintained a population of proliferating progenitor cells in cultures treated with retinoic acid for several weeks. These observations highlight the distinct roles of two noncanonical Wnts during the early stages of retinoic acid-induced neuronal differentiation.
视黄酸和 Wnt/β-catenin 信号在神经元分化过程中发挥重要作用，但在这种情况下，关于非经典 Wnt 信号知之甚少。我们检查了两种人胚胎癌细胞系 NTERA-2（克隆 D1）和 2102Ep 中的视黄酸和 Wnt 信号，前者在视黄酸的作用下经历神经元分化，而后者则不然。视黄酸处理抑制了 NTERA-2 细胞中的β-catenin/Tcf 活性，但不抑制 2102Ep 细胞。抑制发生在β-catenin 的下游，但不涉及视黄酸受体与β-catenin 竞争与 p300 或 Tcf-4 结合。FZD1 的异位表达部分恢复了 2102Ep 细胞中的抑制作用，表明 Wnt 配体的参与。视黄酸处理 NTERA-2 细胞诱导 Wnt-4 和 Wnt-11 的表达，两者都能抑制β-catenin/Tcf 活性。Wnt-4 和 Wnt-11 被发现存在于表达 OCT4 和 GFAP 的细胞岛的细胞边缘，主要对 Nestin、PAX6 和 GATA6 呈阴性。Wnt-4 的基因沉默，但不是 Wnt-11，减少了视黄酸对 OCT4 和 Nanog 的下调以及对 PAX6、ASCL1、HOXC5 和 NEUROD1 的上调，表明 Wnt-4 促进早期神经元分化。NTERA-2 细胞中稳定过表达 Wnt-11 的基因表达分析表明，Wnt-11 增强了视黄酸诱导的早期神经发生。这与过表达 Wnt-11 在视黄酸处理数周的培养物中维持增殖祖细胞群体的结果一致。这些观察结果突出了两种非经典 Wnt 在视黄酸诱导的神经元分化早期的不同作用。
