GRNs Guiding Cranial Versus Trunk Neural Crest Formation

神经脊是一个多能的胚胎细胞群，对不同的衍生品有贡献， 包括外周神经节、面部软骨和骨骼，以及黑素细胞。我们已经提议并 测试了由一系列不同的调控步骤组成的多步骤基因调控网络(GRN) 它们协同作用，向脑神经脊部灌输其定义特征。然而，有重要的 不同神经脊种群发育潜力和迁移途径的差异 不同的轴向水平。在这里，我们建议沿着神经轴探索GRN的差异，重点是 来自两个不同区域的迁移前神经脊细胞：颅骨和躯干。我们的初步转录组 分析发现许多转录因子和信号分子专属于颅脑，但不是躯干 神经峰或反之亦然。我们的目标是确定这些基因在颅骨和躯干中的位置。 GRN。该系统级策略将提供对NC GRN为什么产生特定 用于将这些单元预编程到不同状态的调节状态。目标是： 目的1：分析GRN在颅脑和躯干水平的复合微扰。与全基因组 迁徙前脑脊和主干神经脊的活性转录组的表示，我们将 进行功能丧失实验以干扰基因功能并量化随后的全局 用纳米链分析单个胚胎中可能的靶基因的转录变化。 目的2：神经脊增强剂的系统基因组和功能分析/解剖。我们将确定CisRegulative 调节GRN关键因子在颅神经与主干神经脊中表达的分子 人口。我们将执行多维建模，将转录组数据和 带有功能扰动的活性增强子得到了神经脊GRN的代表性模型。 目的3：重建躯干神经脊程序以测试成骨潜能。使用GRN 信息，我们将通过重新设计干线NC的调节电路并观察是否 关键的GRN亚电路的错误表达/缺失会影响它们的识别和对软骨的贡献。

The neural crest is a multipotent embryonic cell population that contributes to diverse derivatives, including peripheral ganglia, cartilage and bone of the face, and melanocytes. We have proposed and tested a multistep gene regulatory network (GRN), comprised of a logical series of distinct regulatory steps that act in concert to imbue the cranial neural crest its defining traits. However, there are significant differences in developmental potential and migratory pathways of different neural crest populations arising at different axial levels. Here, we propose to explore GRN differences along the neural axis, focusing on premigratory neural crest cells from two distinct regions: cranial versus trunk. Our preliminary transcriptome analysis reveals many transcription factors and signaling molecules specific to the cranial but not trunk neural crest or vice versa. Our goal is to determine the position of these genes in the cranial versus trunk GRNs. This systems level strategy will provide understanding of why NC GRNs produces a particular regulatory state for use in preprogramming these cells to a different state. The aims are: Aim 1: Multiplex perturbation analysis of GRN connections at cranial and trunk levels. With the genomewide representation ofthe active transcriptome of premigratory cranial and trunk neural crest in hand, we will perform loss-of-function experiments to perturb gene function and quantitate subsequent global transcriptional changes in putative target genes in single embryos using Nanostring analysis. Aim 2: Phylogenomic and funcfional analysis/dissection of neural crest enhancers. We will identify cisregulatory elements that mediate expression of key GRN factors in cranial versus trunk neural crest populations. We will perform multidimensional modeling that incorporates results of transcriptome data and active enhancers with functional perturbation results into representational models of neural crest GRNs. Aim 3: Reengineering ofthe trunk neural crest program to test skeletogenic potenfial. Using GRN informafion, we will challenge the fate of trunk NC by reengineering their regulatory circuits and observing if misexpression/deletion of key GRN subcircuits affects their identify and ability to contribute to cartilage.