The Extreme Anterior Domain and Face Formation

 描述(申请人提供)：这项提案将定义与极前区(EAD)和脸部形成相关的信号机制，以非洲爪蛙和热带爪蛙为模型。EAD是一个保守的胚胎区，外胚层和内胚层并列在一起，发育到嘴、垂体前叶和鼻孔。在之前的资助期间，我们有了几个新的发现。(1)通过激肽释放酶-激肽释放酶途径和一氧化氮(NO)，EAD是发育脑神经嵴所必需的组织中心；(2)通过Wnt/PCP途径，传入的CnD诱导EAD经历汇聚延伸，这是口腔形成的新步骤；(3)Wnt拮抗剂frzb1+新月体，定位于EAD ACT全球其他面部区域。我们的数据让我们深入了解 面部发育的最早阶段，并提出了两个具有很大影响的假设，将在本提案中解决。来自EAD的WNT拮抗剂调节额鼻隆起(FNP)和第一个ARCH NC的发展。EAD通过缓激肽和一氧化氮以浓度依赖的方式调节NC的迁移。有两个目标。第一个目标将描述来源于EAD的Frzb1+新月体在神经脊发育中的作用。将在局部功能丧失(LOF)后评估EAD来源的Frzb1+新月体在FNP和First ARCH NC确定、迁移、增殖和生存中的作用(S)。X.热带突变株将补充和扩展分析。将使用表达Frzb1的异种细胞或浸泡在Frzb-Ig G蛋白中的珠子来检测Frzb1+新月体是否足以指导神经脊的发育。FRZB1+新月体抑制β-连环素介导的WNT报告活性的能力将在第一个ARCH NC中被评估。受frzb1+新月体调控的信号通路和候选靶基因将被确定。第二个目标将确定EAD和激肽释放酶途径在调节一氧化氮(NO)信号转导和NC迁移中的作用。使用NO传感器DAF2和NO传感器纳米管，可以识别发育中面部区域产生NO的细胞。面部NO产生中对EAD和EAD激肽释放酶因子的需求将通过摘除和局部LOF进行评估。将检测ccnc细胞在胚胎或培养中是否直接对BDK多肽做出反应的能力。NO对靶细胞的下游影响将集中在信号通路和细胞骨架的变化上。脸部是人类个体的定义特征。面部畸形很常见(约1/700的新生儿)，导致身体和心理障碍。面部的大部分可能会受到EAD组织者活动减少的影响。本文获得的数据将用于产前诊断和矫正，在颅面领域具有重要意义。

 DESCRIPTION (provided by applicant): This proposal will define signaling mechanisms associated with the extreme anterior domain (EAD) and face formation using the frogs Xenopus laevis and X. tropicalis as models. The EAD is a conserved embryonic region where ectoderm and endoderm are juxtaposed, that develops into the mouth, anterior pituitary and nostrils. In the previous funding period, we made several novel findings. (1) The EAD is an organizing center necessary for cranial neural crest (CNC) development, using the Kinin-Kallikrein pathway and nitric oxide (NO); (2) Incoming CNC induces the EAD to undergo convergent extension, a novel step in mouth formation, using the Wnt/PCP pathway; (3) Wnt antagonists frzb1+crescent that are localized in the EAD act globally, in other facial regions. Our data give insight into the earliest stages of facial development and suggest two hypotheses with high impact that will be addressed in this proposal. Wnt antagonists from the EAD regulate frontonasal prominence (FNP) and first arch CNC development. The EAD signals through Bradykinin and nitric oxide to regulate CNC migration, in a concentration-dependent manner. There are two Aims. The first Aim will delineate the role of Frzb1+Crescent derived from the EAD in neural crest development. The role(s) of EAD-derived Frzb1+Crescent in FNP and first arch CNC determination, migration, proliferation and survival will be assessed after local loss of function (LOF). X. tropicalis mutants will complement and extend analyses. Frzb1+Crescent will be assayed for sufficiency to direct neural crest development, using heterologous cells expressing Frzb1 or beads soaked in Frzb-IgG protein. Ability of Frzb1+Crescent to inhibit β-catenin-mediated Wnt reporter activity in the FNP and first arch CNC will be assessed. Signaling pathways and candidate target genes modulated by frzb1+crescent will be determined. The second Aim will define the role of the EAD and the Kinin-Kallikrein pathway in modulating nitric oxide (NO) signaling and CNC migration. Cells in the developing facial region which produce NO will be identified, using the NO sensor DAF2 and NO-sensor nanotubes. A requirement for the EAD and EAD Kinin-Kallikrein factors in facial NO production will be assessed by extirpation and local LOF. cCNC cells will be assayed for ability to respond directly to Bdk peptides or NO in the embryo or in culture. Downstream effects of NO on target cells will be examined focusing on changes in signaling pathways and the cytoskeleton. The face is the defining feature of the individual human. Facial abnormalities are frequent (~1/700 births), resulting in physical and psychological disturbances. Much of the face may be impacted by reduced activity of the EAD organizer. The data obtained here will inform pre-natal diagnosis and correction, and are highly significant in the craniofacial field.