Dynamic and differential regulation of stem cell factor FoxD3 in the neural crest is Encrypted in the genome.
Dynamic and differential regulation of stem cell factor FoxD3 in the neural crest is Encrypted in the genome.
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DOI：
10.1371/journal.pgen.1003142
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发表时间：
2012
期刊：
影响因子：
4.5
通讯作者：
Bronner ME
中科院分区：
文献类型：
--
作者：
Simões-Costa MS;McKeown SJ;Tan-Cabugao J;Sauka-Spengler T;Bronner ME
关键的干细胞转录因子 FoxD3 由迁移前和迁移神经嵴表达，这是一种形成多种衍生物的胚胎干细胞群。尽管 FoxD3 在发育和干细胞生物学中发挥着重要作用，但人们对这些细胞中介导 FoxD3 活性的因素知之甚少。我们发现了两个 FoxD3 增强子 NC1 和 NC2，它们以空间和时间上不同的方式驱动报告基因表达。 NC1 活动重现了颅神经嵴中的初始 FoxD3 表达，而 NC2 活动重现了迷走神经/躯干水平上的初始 FoxD3 表达，而仅稍后出现在迁移性颅嵴中。详细的突变分析、体内染色质免疫沉淀和吗啉代敲低表明，转录因子 Pax7 和 Msx1/2 与神经嵴特异性基因 Ets1 配合，与颅骨 NC1 调节元件结合。然而，在迷走神经/躯干水平，它们与神经板边缘基因 Zic1 一起发挥作用，Zic1 直接与 NC2 增强子结合。这些结果揭示了 FoxD3 在不同神经嵴亚群中的动态和差异调节，表明异质性在调节水平上是加密的。神经嵴增强子的分离不仅可以建立神经嵴形成的直接调节连接，而且还为组织特异性操作和羊膜动物神经嵴细胞身份的研究提供了有价值的工具。 FoxD3是一种重要的干细胞因子，在包括神经嵴细胞在内的多种胚胎细胞中表达。在胚胎中，神经嵴细胞是一种干细胞，可形成多种衍生物，包括神经细胞、色素细胞和面部结构。为了更好地了解神经嵴发育和分化，我们探索了这些细胞中 FoxD3 表达的调节机制。通过检查非编码 DNA，我们已经确定了以重现 FoxD3 表达模式介导绿色荧光蛋白 (GFP) 表达的不同基因组区域。有趣的是，我们发现了两个基因组“开关”或增强子，称为 NC1 和 NC2，它们以一种模式驱动 GFP 表达，该模式重现了神经嵴发育过程中不同时间和地点的 FoxD3 表达。我们发现 Pax 和 Msx 蛋白通过直接结合 NC1 和 NC2 增强子来打开它们。此外，由 NC1 驱动的颅表达需要一种名为 Ets1 的蛋白质，而 NC2 的躯干表达则需要另一种名为 Zic1 的蛋白质。结果表明，FoxD3 在不同的神经嵴细胞群中以基因组中特定编码的方式进行差异调节。这些增强子为了解鸟类和哺乳动物的神经嵴发育提供了有价值的工具。
The critical stem cell transcription factor FoxD3 is expressed by the premigratory and migrating neural crest, an embryonic stem cell population that forms diverse derivatives. Despite its important role in development and stem cell biology, little is known about what mediates FoxD3 activity in these cells. We have uncovered two FoxD3 enhancers, NC1 and NC2, that drive reporter expression in spatially and temporally distinct manners. Whereas NC1 activity recapitulates initial FoxD3 expression in the cranial neural crest, NC2 activity recapitulates initial FoxD3 expression at vagal/trunk levels while appearing only later in migrating cranial crest. Detailed mutational analysis, in vivo chromatin immunoprecipitation, and morpholino knock-downs reveal that transcription factors Pax7 and Msx1/2 cooperate with the neural crest specifier gene, Ets1, to bind to the cranial NC1 regulatory element. However, at vagal/trunk levels, they function together with the neural plate border gene, Zic1, which directly binds to the NC2 enhancer. These results reveal dynamic and differential regulation of FoxD3 in distinct neural crest subpopulations, suggesting that heterogeneity is encrypted at the regulatory level. Isolation of neural crest enhancers not only allows establishment of direct regulatory connections underlying neural crest formation, but also provides valuable tools for tissue specific manipulation and investigation of neural crest cell identity in amniotes. FoxD3 is an important stem cell factor expressed in many types of embryonic cells including neural crest cells. In the embryo, neural crest cells are a type of stem cell that forms diverse derivatives, including nerve cells, pigment cells, and facial structures. To better understand neural crest development and differentiation, we have explored how FoxD3 expression is regulated in these cells. By examining non-coding DNA, we have identified distinct genomic regions that mediate expression of green fluorescent protein (GFP) in a pattern that recapitulates FoxD3 expression. Interestingly, we find two genomic “on–off” switches or enhancers, called NC1 and NC2, that drive GFP expression in a pattern that recapitulates FoxD3 expression at different times and places during neural crest development. We find that Pax and Msx proteins turn on both NC1 and NC2 enhancers by directly binding to them. In addition, cranial expression driven by NC1 requires a protein called Ets1, whereas trunk expression of NC2 requires a different protein called Zic1. The results show that FoxD3 in differentially regulated in distinct neural crest cell populations in a manner that is specifically encoded in the genome. These enhancers provide valuable tools for understanding neural crest development in birds and mammals.
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