Role of actin cytoskeleton regulators in craniofacial development and disease

描述（由申请人提供）：神经嵴是在脊椎动物胚胎发生期间广泛迁移的独特细胞群。特别令人感兴趣的是，颅神经嵴（CNC）细胞在整个面部迁移，形成大部分颅面骨骼。它们的增殖、迁移或存活缺陷导致许多疾病和出生缺陷，包括唇腭裂、Treacher-Collins综合征和DiGeorge综合征。尽管最近在了解神经嵴发育方面取得了进展，但神经嵴迁移的机制仍然不清楚。因此，研究它们的迁移行为和其中肌动蛋白细胞骨架的调节对于理解先天性颅面疾病的基础至关重要。从微阵列屏幕上，我已经确定了多个肌动蛋白细胞骨架调节（ACR）基因，主要在神经嵴细胞表达。我的初步研究表明，Myosin-X和Caldesmon都是通过调节细胞形态和膜突起的适当CNC细胞迁移的关键。基于这些发现，我假设神经嵴细胞通过特定的ACR实现其独特的迁移能力，以快速精确地控制 它们的细胞骨架动力学。在这个计划中，我将通过进一步分析这些ACRs的活动，并建立它们与肌动蛋白结构的物理和功能相互作用来研究神经嵴迁移的机制。具体来说，我将结合联合收割机经典胚胎学技术与现代细胞生物学，生物化学，和在体内显微镜研究的动态安排肌动蛋白细胞骨架CNC迁移过程中，并检查如何在神经嵴ACRs调节这一过程。实验将在青蛙和小鸡中进行，考虑到 这两种模型系统的优点：青蛙的基因和组织操作简单，鸡的高分辨率成像和后期电穿孔。本研究的具体目的是：1）通过真实的时间成像技术研究正常神经嵴迁移过程中肌动蛋白细胞骨架的动态变化。2)确定肌球蛋白X和钙调蛋白在CNC迁移过程中调节肌动蛋白动力学的作用。3)功能特征Cdc 42 ep 1和其他候选基因在微阵列中确定的屏幕上的作用，在神经嵴的发展。4)确定CNC迁移中ACR之间的功能和物理相互作用。 公共卫生相关性：从背神经管迁移的CNC细胞对面部骨骼做出了重大贡献。这项工作的目的是研究脊椎动物胚胎发生过程中CNC细胞迁移的细胞生物学基础。了解神经嵴移位的机制将有助于了解先天性颅面缺损的病因。

DESCRIPTION (provided by applicant): Neural crest is a unique population of cells that migrate extensively during vertebrate embryogenesis. Of particular interest, cranial neural crest (CNC) cells migrate throughout the face to form most of the craniofacial skeleton. Defects in their proliferation, migration, or survival lead to numerous diseases and birth defects, including cleft lip/palate, Treacher-Collins syndrome, and DiGeorge syndrome. Despite recent progress in understanding aspects of neural crest development, the mechanisms underlying neural crest migration are still unclear. Thus, studying their migratory behavior and the regulation of actin cytoskeleton therein is crucial for understanding the basis of congenital craniofacial disorders. From a microarray screen, I have identified multiple actin cytoskeleton regulator (ACR) genes that are predominantly expressed in neural crest cells. My preliminary studies reveal that Myosin-X and Caldesmon are both critical for proper CNC cell migration via regulating cell morphology and membrane protrusions. Based on these findings, I hypothesis that neural crest cells achieve their unique migratory ability through specific ACRs to rapidly and precisely control their cytoskeletal dynamics. In this proposal, I will investigate the mechanism of neural crest migration by further analyzing the activities of these ACRs, and establishing their physical and functional interactions with actin structures. Specifically, I will combine classical embryology techniques with modern cell biology, biochemistry, and in vivo microscopy to study the dynamic arrangements of actin cytoskeleton during CNC migration and examine how ACRs in the neural crest regulate this process. Experiments will be carried out in frog and chick, taking into account the advantages of both model systems: simple gene and tissue manipulation in frog, and high-resolution imaging and late stage electroporation in chick. The following specific aims will be performed: 1) Examine the dynamics of actin cytoskeleton during normal neural crest migration by real time imaging. 2) Determine the role of Myosin-X and Caldesmon in regulating actin dynamics during CNC migration. 3) Functionally characterize Cdc42ep1 and additional candidate genes identified in the microarray screen for their roles in neural crest development. 4) Determine functional and physical interactions between ACRs in CNC migration. PUBLIC HEALTH RELEVANCE: CNC cells that migrate from dorsal neural tube make a major contribution to the facial skeleton. The proposed work aims to study the cell biological basis underlying the migration of CNC cells during vertebrate embryogenesis. Understanding the mechanism of neural crest migration will shed light on the etiology of congenital craniofacial defects.