Laboratory of Developmental Signalling, Cancer Research UK London Research Institute, London, United Kingdom.
PLoS Biol. 2011 Feb 15;9(2):e1000593. doi: 10.1371/journal.pbio.1000593.
Bone morphogenetic protein (BMP) gradients provide positional information to direct cell fate specification, such as patterning of the vertebrate ectoderm into neural, neural crest, and epidermal tissues, with precise borders segregating these domains. However, little is known about how BMP activity is regulated spatially and temporally during vertebrate development to contribute to embryonic patterning, and more specifically to neural crest formation. Through a large-scale in vivo functional screen in Xenopus for neural crest fate, we identified an essential regulator of BMP activity, SNW1. SNW1 is a nuclear protein known to regulate gene expression. Using antisense morpholinos to deplete SNW1 protein in both Xenopus and zebrafish embryos, we demonstrate that dorsally expressed SNW1 is required for neural crest specification, and this is independent of mesoderm formation and gastrulation morphogenetic movements. By exploiting a combination of immunostaining for phosphorylated Smad1 in Xenopus embryos and a BMP-dependent reporter transgenic zebrafish line, we show that SNW1 regulates a specific domain of BMP activity in the dorsal ectoderm at the neural plate border at post-gastrula stages. We use double in situ hybridizations and immunofluorescence to show how this domain of BMP activity is spatially positioned relative to the neural crest domain and that of SNW1 expression. Further in vivo and in vitro assays using cell culture and tissue explants allow us to conclude that SNW1 acts upstream of the BMP receptors. Finally, we show that the requirement of SNW1 for neural crest specification is through its ability to regulate BMP activity, as we demonstrate that targeted overexpression of BMP to the neural plate border is sufficient to restore neural crest formation in Xenopus SNW1 morphants. We conclude that through its ability to regulate a specific domain of BMP activity in the vertebrate embryo, SNW1 is a critical regulator of neural plate border formation and thus neural crest specification.
骨形态发生蛋白(BMP)梯度为细胞命运特化提供位置信息,例如将脊椎动物外胚层模式化为神经、神经嵴和表皮组织,精确的边界将这些区域分隔开来。然而,人们对 BMP 活性如何在脊椎动物发育过程中进行空间和时间调节以促进胚胎模式形成,更具体地说是促进神经嵴形成,知之甚少。通过在 Xenopus 中进行的大规模体内功能筛选,我们鉴定出一种 BMP 活性的必需调节因子 SNW1。SNW1 是一种已知调节基因表达的核蛋白。通过使用反义 morpholino 耗尽 Xenopus 和斑马鱼胚胎中的 SNW1 蛋白,我们证明了背部表达的 SNW1 对于神经嵴特化是必需的,这与中胚层形成和原肠胚形态发生运动无关。通过利用 Xenopus 胚胎中磷酸化 Smad1 的免疫染色和 BMP 依赖性报告基因转基因斑马鱼系的结合,我们表明 SNW1 调节神经板边界处背侧外胚层中 BMP 活性的特定域。我们使用双重原位杂交和免疫荧光显示了这个 BMP 活性域相对于神经嵴域和 SNW1 表达域的空间定位。进一步的体内和体外测定,包括细胞培养和组织外植体,使我们能够得出结论,SNW1 作用于 BMP 受体的上游。最后,我们表明 SNW1 对于神经嵴特化的要求是通过其调节 BMP 活性的能力,因为我们证明了靶向过表达 BMP 到神经板边界足以恢复 Xenopus SNW1 形态发生缺陷体中的神经嵴形成。我们得出结论,通过其调节脊椎动物胚胎中 BMP 活性的特定域的能力,SNW1 是神经板边界形成的关键调节因子,从而是神经嵴特化的关键调节因子。
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