The role of the transcriptional repressor ZEB2 in human neural crest cell formation and craniofacial pathology

项目摘要 这项建议的目的是描述转录抑制因子ZEB2在人类神经中的作用。 脊细胞形成、颅面分化和莫瓦特-威尔逊综合征。神经脊是一种多能的 脊椎动物特有的胚胎细胞群，可广泛迁移并分化为各种 衍生品，包括大多数头面部骨骼、软骨和牙齿器官组织。无数的人类 病理与异常的神经脊细胞发育有关，包括最近描述的 神经瘤性莫瓦特-威尔逊综合征(MWS)。巨噬细胞综合征是一种由新生杂合子引起的罕见综合征。 ZEB2基因突变导致不同的面部表型、牙齿异常、智力残疾和其他 异常包括先天性巨结肠症，这表明神经脊细胞在多个轴位的缺陷。 然而，在MWS患者中观察到了显著的基因和表型变异，并且 导致这种疾病的潜在机制尚不清楚。此外，虽然ZEB2已经 被认为是神经系统和神经脊细胞的主要发育调节器 上皮到间充质的转变和迁移：人类神经脊细胞模型中的最新证据 提示ZEB2在神经板边缘和神经脊的早期形成中起重要作用。已知ZEB2 负向调节TGFb超家族信号，其调节对于正常的神经峰是必不可少的 形成和分化。因此，假设ZEB2反复发挥作用，以建立 抑制表观遗传景观贯穿神经脊细胞形成和颅面发育 调节适当的TGFb超家族信号，这一调控的失败突显了MWS 病理学。为了验证这一假设，本文提出了三个具体目标。目标1将研究模块化功能 ZEB2对野生型人颅神经脊细胞中TGFb信号的调节作用 队形。特别是，ZEB2和TGFb受体之间的顺式调节相互作用激活了Smad 导致染色质可及性改变的蛋白质和共抑制物的功能要求将是 勾勒出来的。在目标2中，将通过建立诱导多能性来解决MWS的病理 来自不同基因型别的MWS患者的干细胞。使用这些工具，精确的转录和表观遗传学 在基于MWS的神经脊细胞形成期间的错误调节将被询问，提供急需的 对这一未被研究的病理的分子洞察力。在本提案的目标3中，ZEB2在野生型和 MWS人类牙齿的形成将通过建立以人类神经脊细胞为基础的 牙齿器官模型。这些结果将首次揭示这种转录抑制因子的分子作用。 贯穿人类神经脊细胞形成、分化和病理的多个阶段。

Project Summary The objective of this proposal is to delineate the role of the transcriptional repressor ZEB2 during human neural crest cell formation, craniofacial differentiation, and Mowat-Wilson Syndrome. The neural crest is a multipotent embryonic cell population unique to vertebrates which migrates extensively and differentiates into a variety of derivatives, including most of the craniofacial bone, cartilage, and dental organ tissues. Numerous human pathologies are associated with abnormal neural crest cell development, including the recently delineated neurocristopathy Mowat-Wilson Syndrome (MWS). MWS is a rare syndrome caused by de novo heterozygous mutations in ZEB2 leading to a distinct facial phenotype, dental abnormalities, intellectual disability, and other anomalies including Hirschprung disease which are indicative of neural crest cell defects at multiple axial levels. However, significant genotypic and phenotypic variability has been observed among MWS patients, and the underlying mechanisms that contribute to this disease are not well understood. Furthermore, while ZEB2 has been implicated as a major developmental regulator of the nervous system as well as in neural crest cell epithelial-to-mesenchymal transition and migration, recent evidence in our human neural crest cell model indicates a crucial role for ZEB2 in the early formation of the neural plate border and neural crest. ZEB2 is known to negatively regulate TGFb superfamily signaling, whose modulation is essential for proper neural crest formation and differentiation. Therefore, it is hypothesized that ZEB2 functions reiteratively to establish the repressive epigenetic landscape throughout neural crest cell formation and craniofacial development by modulating proper TGFb superfamily signaling, and that the failure of this regulation underscores the MWS pathology. To test this hypothesis, three specific aims are proposed. Aim 1 will examine the modular functionality of ZEB2 in regulating the proper levels of TGFb signaling throughout wild-type human cranial neural crest cell formation. In particular, the cis-regulatory interactions between ZEB2 and TGFb receptor activated Smad proteins and the functional requirement of co-repressors leading to changes in chromatin accessibility will be delineated. In Aim 2, the MWS pathology will be addressed through the establishment of induced pluripotent stem cells from MWS patients of diverse genotypes. Using these tools, the precise transcriptomic and epigenetic misregulation during MWS-based neural crest cell formation will be interrogated, providing much needed molecular insight into this understudied pathology. In Aim 3 of this proposal, the role of ZEB2 in wild-type and MWS human tooth formation will be interrogated through the establishment of a human neural crest cell-based tooth organoid model. These results will reveal for the first time the molecular role of this transcriptional repressor throughout multiple stages of human neural crest cell formation, differentiation, and pathology.