Decoding regulatory nodes controlling growth and proportion of the skull

解码控制头骨生长和比例的调节节点

• 批准号: 10348718
• 负责人: Katherine Christine Woronowicz
• 金额: $ 7.25万
• 依托单位: BOSTON CHILDREN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-03-01 至 2023-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10348718
• 关键词: Address; Adult; Affect; Anterior; Articulation; Base Pairing; Binding; Brain; Branchial arch structure; CRISPR/Cas technology; Cartilage; Cell Differentiation process; Cephalic; Cleft Palate; Clinical; Code; Complex; Congenital Abnormality; Craniofacial Abnormalities; Data; Development; Disease; Elements; Embryo; Enhancers; Essential Genes; Event; Evolution; Face; Fertilization; First Pharyngeal Arch; Fishes; Fluorescent in Situ Hybridization; Gene Expression; Gene Expression Regulation; Genes; Genetic Enhancer Element; Genomic approach; Genotype; Growth; Head; Homeobox; Homeobox Genes; Hour; Human; Human Development; Introns; Jaw; Knowledge; Light; Limb Bud; Link; Mastication; Mediating; Midbrain structure; Modification; Morphogenesis; Morphology; Mus; Mutation; Neural Crest; Neural Crest Cell; Neural Fold; Nucleic Acid Regulatory Sequences; Oral cavity; Oropharyngeal; Outcome Measure; Pan Genus; Pattern; Phenotype; Play; Primates; Prognathism; Regulation; Regulatory Element; Reporter; Research; Rest; Role; Sense Organs; Signal Transduction; Site; Syndrome; Synteny; Testing; Tissues; Untranslated RNA; Variant; Vertebrates; Work; Zebrafish; base; bone; cell motility; comparative genomics; craniofacial; craniofacial complex; craniofacial development; craniofacial microsomia; craniofacial structure; cranium; experimental study; gene regulatory network; hindbrain; improved; in vivo; loss of function; mutant; neurodevelopment; novel; preservation; pressure; single molecule; spatiotemporal; trait

PROJECT SUMMARY The skull is comprised of bones of different embryonic origins that articulate to encase the brain, to protect the sense organs, and to enable mastication. Well-orchestrated signaling and morphological events generate the seamless morphology of the craniofacial complex. Of central importance are the cranial neural crest cells (CNCCs) that migrate from the anterior neural folds to populate the oropharyngeal arches. CNCCs acquire axial identity from midbrain and hindbrain segmentation. Modifications to gene expression of CNCCs, their precursors, their derivatives, or even interacting tissues may underlie both normal variation and common craniofacial malformations. Although the gene regulatory networks that govern early specification of CNCCs are well known, we still lack detailed knowledge of later developmental events involving CNCC derivatives and how this relates to fundamental mechanisms of disease. The experiments outlined in this proposal will tease apart the morphological consequences of genotype. We expect that loci modifying the relative proportions of the skull and face will have commonalities amongst jawed vertebrates, as the head is an ancestral trait. Our prior phylogenomic comparisons identified fixed loci correlated with differential developmental prognathism. One of the regions identified encompasses a locus containing a large, cis-regulatory region highly conserved in all jawed vertebrates. This locus rests in an intron of agap1, and has retained directional synteny with the nearest neighbor, homeobox gene gbx2, over all of vertebrate evolution. This 343 base pair (bp) region is defined as having over 90% identity among vertebrates. A core of over 190bp is retained with 100% identity among primates, suggesting deep conservation preserved by strong selective pressure and a potential role in human development. Preliminary analyses suggest the region may participate in a broader regulatory hub that modulates expression of gbx2. As gbx2 is essential for patterning CNCCs, and is expressed in the oropharyngeal arches, my hypothesis is that the conserved non-coding region we identified acts as a specific enhancer for gbx2, mediating patterning of the forming arches and leading to proportional changes in outgrowth of the jaws. I will test this hypothesis by determining the function of components of the regulatory hub and their contributions to proper growth and form of the jaws. I will analyze necessity and function of orthologous sequences from zebrafish, chimp, and human, as well as determine the role of gbx2 in craniofacial morphology. Outcome measures include long-term assessment of CNCC migration and differentiation in vivo, and evaluating changes to spatiotemporal expression of gbx2 and related homeobox dlx genes in CNCCs and their derivatives. Findings from these experiments will lead to improved clinical strategies addressing disorders with disruptions to growth and form of the jaws as well as shed light on the contribution of coding and non-coding elements to craniofacial development and malformations.

项目摘要 头骨由不同胚胎起源的骨头组成，这些骨头连接起来包裹大脑，以保护大脑。 感觉器官，并使咀嚼。精心策划的信号和形态事件产生了 颅面复合体的完美形态最重要的是脑神经嵴细胞 （CNCC）从前神经褶皱迁移到口咽弓。CNCC收购 中脑和后脑分割的轴向一致性。CNCC基因表达的修饰，其 前体，它们的衍生物，甚至相互作用的组织可能是正常变异和共同变异的基础。 颅面畸形尽管控制CNCC早期特化的基因调控网络 众所周知，我们仍然缺乏涉及CNCC衍生物的后期发展事件的详细知识， 这与疾病的基本机制有什么关系本提案中概述的实验将梳理 除了基因型的形态学后果。我们预期，基因座改变的相对比例 由于头部是祖先的特征，头骨和面部在有颌脊椎动物中具有共性。我们 先前的基因组比较鉴定了与差异发育的多颌畸形相关的固定基因座。 鉴定的区域之一包括含有在细胞中高度保守的大的顺式调节区域的基因座， 都是有颚的脊椎动物该基因座位于agap1的内含子中，并保留了与 最近的邻居，同源盒基因gbx2，在所有的脊椎动物进化。这个343个碱基对（bp）的区域是 定义为在脊椎动物中具有超过90%的同一性。保留了超过190bp的核心，具有100%同一性 在灵长类动物中，这表明强烈的选择压力和潜在的作用， 人类发展初步分析表明，该地区可能会参与一个更广泛的监管中心， 调节Gbx2的表达。由于gbx2对于CNCC的图案化是必需的，并且在CNCC中表达。 口咽弓，我的假设是，保守的非编码区，我们确定作为一个 gbx2的特异性增强子，介导形成拱形的图案化并导致成比例的 颌骨生长的变化。我将通过确定的组件的功能来测试这个假设， 调节枢纽和他们的贡献，适当的增长和形式的下巴。我将分析必要性， 来自斑马鱼、黑猩猩和人类的正向序列的功能，以及确定gbx 2在 颅面形态学结果指标包括CNCC迁移的长期评估， 体内分化，并评估gbx 2和相关同源框dlx的时空表达的变化 CNCC及其衍生物中的基因。这些实验的发现将导致改进的临床策略 解决颌骨生长和形态中断的疾病，并阐明 颅面发育和畸形的编码和非编码因素。