Transcriptional regulatory landscapes underlying FEZ Formation

总结： 脊椎动物组织和器官的形态发生是通过精心组织的相互作用发生的。期间 上颌骨的发育，前脑，面部外胚层和 介入的神经嵴细胞调节上颌的发育。我们的研究表明， 和神经嵴细胞共同作用，诱导Sonic hedgehog（SHH）在 额鼻外胚层区（FEZ），位于额鼻外胚层表面的信号中心 引导上颌原基图案化生长的过程（FNP）。我们的初步数据还表明， 前B细胞白血病同源盒（PBX）转录因子参与诱导和维持这种模式 通过相互作用的遗传网络在FEZ中表达SHH。本申请中提出的研究是 旨在揭示基因调控机制，这些机制是由来自前脑和大脑皮层的信号激活的。 神经嵴，以及研究PBX转录因子在调节SHH表达中的作用， 菲斯.具体来说，我们假设1）来自前脑的SHH信号诱导了表面的“能力”， 2）随后，在到达时，神经嵴细胞诱导SHH在外胚层细胞中的转录。 感受态细胞的FEZ，和3）PBX转录因子参与调节SHH的表达， 菲斯.我们将使用禽类和鼠类胚胎在三个特定目标中检验这一假设。在每个目标中，我们将 使用鸡胚，因为它们允许我们操纵大脑和大脑之间的信号相互作用。 FEZ（Aim 1）、神经嵴细胞和外胚层（Aim 2）或PBX 1和PBX 3的表达（Aim 3）。我们 将使用各种基因组方法（ATAC-seq、ChIP-seq、GRO-seq）来评估染色质的变化 和激活SHH基因座的转录调控元件（TREs）。在目标3中，我们还将评估 改变PBX基因表达的形态学、细胞学和分子学结果，我们将使用小鼠 胚胎利用Selleri实验室已经产生的数据。的工作成果 本申请中提出的方法将阐明面部发育的分子方面， 了解该地区发生的疾病过程。SHH或SHH通路中的等位基因变体 成员，与人类颅面复合体的畸形有关， 前脑无裂和唇裂伴或不伴腭裂。最近，PBX基因已被确定为风险 颅面畸形患者的变异。因此，我们的创新方法将研究与 染色质分析在发育生物学中的坚实基础将产生直接 适用于理解人类畸形学。

Summary: Morphogenesis of vertebrate tissues and organs occurs through well-orchestrated tissue interactions. During development of the upper jaw, signaling interactions among the forebrain, the facial ectoderm, and the intervening neural crest cells regulate development of the upper jaw. Our research has revealed that signals from the brain and the neural crest cells act together to induce expression of Sonic hedgehog (SHH) in the Frontonasal Ectodermal Zone (FEZ), a signaling center located in the surface ectoderm of the Frontonasal Process (FNP) that directs patterned growth of the upper jaw anlagen. Our preliminary data also suggest that Pre-B-cell leukemia homeobox (PBX) transcription factors are involved in inducing and maintaining the pattern of SHH expression in the FEZ by an interacting genetic network. The research proposed in this application is designed to uncover the gene regulatory mechanisms that are activated by signals from the forebrain and the neural crest, as well as, to investigate the role of PBX transcription factors in regulating SHH expression in the FEZ. Specifically, we hypothesize that 1) SHH signaling from the forebrain induces “competence” in surface ectoderm cells to express SHH; 2) subsequently, upon arrival, neural crest cells induce transcription of SHH in competent cells of the FEZ, and 3) PBX transcription factors participate in regulating expression of SHH in the FEZ. We will test this hypothesis in three specific aims using avian and murine embryos. In each Aim, we will use chick embryos because they allow us to manipulate the signaling interactions between the brain and the FEZ (Aim 1), the neural crest cells and the ectoderm (Aim 2), or expression of PBX1 and PBX3 (Aim 3). We will use a variety of genomic approaches (ATAC-seq, ChIP-seq, GRO-seq) to evaluate changes in chromatin and activated transcriptional regulatory elements (TREs) at the SHH locus. In Aim 3, we will also assess morphological, cellular, and molecular outcomes of altering PBX gene expression, and we will use mouse embryos to take advantage of data already generated in the Selleri laboratory. The results of the work proposed in this application will shed light on the molecular aspects of facial development, and will aid understanding of disease processes that occur in this region. Allelic variants in SHH, or SHH pathway members, are associated with human dysmorphologies of the craniofacial complex in diseases such as Holoprosencephaly and cleft lip with our without cleft palate. Recently, PBX genes have been identified as risk variants in patients with craniofacial dysmorphology. Hence, our innovative approach combining studies with a solid foundation in developmental biology with chromatin analyses will yield information that is directly applicable to understanding human dysmorphology.