Wnt signaling regulates the lineage differentiation potential of mouse embryonic stem cells through Tcf3 down-regulation.
Wnt signaling regulates the lineage differentiation potential of mouse embryonic stem cells through Tcf3 down-regulation.
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DOI：
10.1371/journal.pgen.1003424
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发表时间：
2013-05
期刊：
影响因子：
4.5
通讯作者：
Fodde R
中科院分区：
文献类型：
--
作者：
Atlasi Y;Noori R;Gaspar C;Franken P;Sacchetti A;Rafati H;Mahmoudi T;Decraene C;Calin GA;Merrill BJ;Fodde R
经典Wnt信号在调节小鼠胚胎干细胞（ESCs）的自我更新和分化中起限速作用。我们以前已经表明，突变的APC（腺瘤性息肉病大肠杆菌）肿瘤抑制基因组成型激活Wnt信号在胚胎干细胞和抑制他们的能力，向外，中，和内胚层谱系分化。然而，Wnt调控小鼠胚胎干细胞谱系分化的潜在分子和细胞机制迄今仍不清楚。为此，我们推导并研究了编码不同水平Wnt信号激活的几种Apc突变ESC系的基因表达谱。我们发现，下调Tcf 3，Tcf/Lef家族的成员和自我更新和多能性的控制中的关键球员，代表了一个特定的和主要的Wnt激活ESCs的反应。因此，拯救Tcf 3表达部分恢复了在Apc突变型ESC中观察到的神经缺陷，表明Tcf 3下调是Wnt介导的神经分化抑制的必要步骤。我们发现，在组成性激活Wnt信号传导的背景下，Tcf 3下调不是由启动子DNA甲基化引起的，而是可能由RNA和蛋白质水平上的多种机制引起的，如观察到的激活组蛋白标记物的减少所示。（H3 K4 me 3和H3-乙酰化）和miR-211的上调，miR-211是一种新型Wnt调节的microRNA，靶向Tcf 3并减弱小鼠ESCs的早期神经分化。我们的数据首次表明，Wnt信号下调Tcf 3的表达，可能在转录和转录后水平，从而突出了一个新的机制，通过Wnt信号抑制神经外胚层谱系分化小鼠胚胎干细胞。再生医学未来的成功在很大程度上取决于我们对控制干细胞分化的细胞和分子机制的了解和操纵能力。越来越多的证据表明，在小鼠胚胎干细胞中，典型的Wnt/β-连环蛋白信号不仅增强自我更新，而且还将细胞命运决定导向非神经外胚层谱系。然而，很少有人知道的机制，潜在的分化缺陷所造成的组成性活性Wnt信号。使用一组具有不同Wnt信号水平的Apc突变型ESC，我们发现，除其他外，下调Tcf 3（多能性回路的关键成员）以及诱导一种新的Wnt调节的microRNA（miR-211）代表了两种重要的下游效应，Wnt信号通过这两种效应抑制小鼠ESC的神经分化。我们还提供了一个更详细的图片Wnt信号如何抵消Tcf 3在干细胞中的功能，显示Tcf 3的抑制，在活跃的Wnt信号的背景下，涉及组蛋白修饰Tcf 3启动子和miR-211的激活，转录后稳定Tcf 3下调。了解Wnt信号在ESC中的下游作用具有基础和翻译相关性，因为它可以用于操纵ESC向特定细胞谱系分化。
Canonical Wnt signaling plays a rate-limiting role in regulating self-renewal and differentiation in mouse embryonic stem cells (ESCs). We have previously shown that mutation in the Apc (adenomatous polyposis coli) tumor suppressor gene constitutively activates Wnt signaling in ESCs and inhibits their capacity to differentiate towards ecto-, meso-, and endodermal lineages. However, the underlying molecular and cellular mechanisms through which Wnt regulates lineage differentiation in mouse ESCs remain to date largely unknown. To this aim, we have derived and studied the gene expression profiles of several Apc-mutant ESC lines encoding for different levels of Wnt signaling activation. We found that down-regulation of Tcf3, a member of the Tcf/Lef family and a key player in the control of self-renewal and pluripotency, represents a specific and primary response to Wnt activation in ESCs. Accordingly, rescuing Tcf3 expression partially restored the neural defects observed in Apc-mutant ESCs, suggesting that Tcf3 down-regulation is a necessary step towards Wnt-mediated suppression of neural differentiation. We found that Tcf3 down-regulation in the context of constitutively active Wnt signaling does not result from promoter DNA methylation but is likely to be caused by a plethora of mechanisms at both the RNA and protein level as shown by the observed decrease in activating histone marks (H3K4me3 and H3-acetylation) and the upregulation of miR-211, a novel Wnt-regulated microRNA that targets Tcf3 and attenuates early neural differentiation in mouse ESCs. Our data show for the first time that Wnt signaling down-regulates Tcf3 expression, possibly at both the transcriptional and post-transcriptional levels, and thus highlight a novel mechanism through which Wnt signaling inhibits neuro-ectodermal lineage differentiation in mouse embryonic stem cells. The future successes of regenerative medicine largely rely on our knowledge of, and our capacity to manipulate, the cellular and molecular mechanisms governing stem cell differentiation. A growing body of evidence suggests that, in mouse embryonic stem cells, canonical Wnt/β-catenin signaling not only enhances self-renewal but also directs the cell fate decision towards non-neuroectodermal lineages. However, little is known about the mechanisms underlying the differentiation defects caused by constitutive active Wnt signaling. Using a set of Apc-mutant ESCs harbouring different levels of Wnt signaling, we found that, among others, down-regulation of Tcf3, a key member of the pluripotency circuit, as well as induction of a novel Wnt-regulated microRNA, miR-211, represent two important downstream effects through which Wnt signaling inhibits neural differentiation in mouse ESCs. We also provide a more detailed picture on how Wnt signaling counteracts Tcf3 function in stem cells by showing that Tcf3 repression, in the context of active Wnt signaling, involves histone modifications at the Tcf3 promoter and the activation of miR-211, which post-transcriptionally stabilizes Tcf3 downregulation. Understanding the downstream effects of Wnt signaling in ESCs is of both fundamental and translational relevance, as it may be exploited to manipulate ESC differentiation towards specific cell lineages.
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