Identification of the factors underlying tooth field size and competency

识别牙域大小和能力的潜在因素

• 批准号: 10284140
• 负责人: Tyler Square
• 金额: $ 15.27万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-01 至 2023-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10284140
• 关键词: Address; Adult; Advisory Committees; Area; Automobile Driving; Basic Science; Bioinformatics; Candidate Disease Gene; Cells; Competence; Data; Dental; Development; Dyes; Elements; Epithelial; Event; Exhibits; Face; Fishes; Future; Gasterosteidae; Gene Expression; Gene Expression Profile; Gene Family; Genes; Genetic; Genetic Determinism; Genetic Transcription; Goals; Growth; Head; Image; Implant; Individual; Institution; Label; Location; Mammals; Mesenchymal; Mesenchyme; Microdissection; Modeling; Morphogenesis; Morphology; Motion; Mus; Mutant Strains Mice; Mutation; Organ; Pathway interactions; Pattern; Pattern Formation; Phase; Phenotype; Positioning Attribute; Principal Investigator; Process; Regulator Genes; Reporter; Reporter Genes; Reproducibility; Research; Research Project Grants; Signal Pathway; Signal Transduction; Skeleton; Specific qualifier value; Specimen; Stains; Taste Buds; Techniques; Testing; Tissues; Tooth Germ; Tooth structure; Transcript; Transgenic Organisms; Tumor Necrosis Factor Receptor; Ursidae Family; Vertebrates; Work; Zebrafish; base; career development; cell type; craniofacial; craniofacial development; deciduous tooth; ectodysplasin; experimental study; face bone structure; genetic signature; in vivo; in vivo monitoring; insight; mRNA sequencing; mutant; neuromast; overexpression; receptor; response; single-cell RNA sequencing; skeletal; tool

Project Summary/Abstract The overall dental morphology of an adult vertebrate is set in motion by the initiation of tooth fields at specific regions in the body plan during early development. Thereafter, primary teeth form sequentially, expanding each tooth field as new teeth are added at the tooth field margin. Basic research into the genes and signaling pathways underlying these processes will reveal which cell types and genetic signatures are associated with tooth field initiation and expansion, thus informing future attempts to create and successfully implant live teeth. By determining which cell types are capable of undergoing transformation to a dental fate, new avenues for the creation of lab-made tooth organs will be revealed. Additionally, identifying the genetic signatures associated with tooth field expansion and arrest will provide information about the greater context under which dental arcade expansion is facilitated. The present study will use a newly developed set of stable transgenic stickleback fish and zebrafish to identify the cell types, transcript profiles, and signaling events that underlie the processes of tooth field initiation, expansion, and arrest. These model fishes present a unique opportunity to understand tooth field morphogenesis, because tooth field position and size can be manipulated using genetic tools. In response to Eda overexpression, both species form ectopic tooth fields in highly consistent locations in or on the head, while also expanding endogenous tooth fields. By contrast, Dkk2 overexpression in stickleback reduces tooth field size by inhibiting tooth field expansion. Aim 1 seeks to understand the gene expression dynamics of the switch to a tooth organ fate by assessing the fine spatial expression patterns of genes encoding Tumor Necrosis Factor Receptors (TNFRs), candidate receptors that may confer the response to Eda, and performing single-cell RNA sequencing on tooth-competent regions microdissected from Eda overexpressing and WT sticklebacks. Aim 2 of this project will longitudinally observe the process of ectopic tooth formation by repeatedly imaging fish as they grow ectopic facial teeth, utilizing lineage tracing techniques, reporter gene expression, and vital dye labeling to understand which cells contribute to ectopic teeth. Aim 3 will elucidate which developmental pathways are associated with tooth field size and tooth row number by comparing the gene expression profiles of the dissected tooth field margins derived from expanded (Eda overexpression), arrested (Dkk2 overexpression), and normal stickleback tooth fields. Overall, these experiments will yield information on how teeth can be specified in development (Aims 1 and 2), and which gene expression responses are concomitant with favorable or unfavorable conditions for tooth field expansion (Aim 3). The first portion of this work will occur at UC-Berkeley (K99 phase), facilitated by Dr. Square’s advisory committee, institution, and current lab. The completion of this work and the associated career development activities will support Dr. Square in his goal to become a principal investigator studying the interface of tooth and craniofacial development.

项目总结/摘要 成年脊椎动物的整体牙齿形态是由特定位置的牙区的起始而启动的。 在早期发展过程中，身体计划中的区域。此后，初级齿依次形成， 在齿域边缘添加新齿时的齿域。基因和信号通路的基础研究 这些过程的基础将揭示哪些细胞类型和遗传特征与牙齿区域相关 启动和扩展，从而为将来创建和成功植入活齿的尝试提供信息。通过 确定哪些细胞类型能够经历向牙齿命运的转化， 实验室制造的牙齿器官将被揭示。此外，识别相关的遗传特征 与牙齿领域的扩大和逮捕将提供有关更大的背景下， 促进了扩张。本研究将使用一套新开发的稳定的转基因棘鱼 和斑马鱼，以确定细胞类型，转录谱，和信号事件的基础过程， 齿区的起始、扩展和停止。这些模型鱼提供了一个独特的机会来了解牙齿 区域形态发生，因为牙齿区域的位置和大小可以使用遗传工具来操纵。响应 对于Eda过表达，两个物种在头部内或头部上高度一致的位置形成异位牙区， 同时也扩大了内源性牙区。相比之下，Dkk 2在棘鱼中的过表达减少了牙齿的生长。 通过抑制齿域扩展来实现齿域尺寸。目的1旨在了解基因表达动态的基因表达的 通过评估编码肿瘤坏死的基因的精细空间表达模式切换到牙齿器官命运 因子受体（TNFRs），可能赋予对EDA的反应的候选受体，并执行单细胞 对从Eda过表达和WT棘鱼中显微切割的牙齿感受态区域进行RNA测序。 本课题的目的二是通过对鱼类的重复成像，纵向观察异位牙的形成过程 当他们长出异位面牙时，利用谱系追踪技术，报告基因表达， 标记以了解哪些细胞导致异位牙齿。目标3将阐明哪些发展途径 与牙区大小和牙列数相关， 来自扩展（Eda过表达）、停滞（Dkk 2过表达）和 正常的棘鱼齿区总的来说，这些实验将产生如何指定牙齿的信息 在发育中（目的1和2），以及哪些基因表达反应伴随着有利的或 齿域扩展的不利条件（目标3）。这项工作的第一部分将发生在加州大学伯克利分校 (K99阶段），由Square博士的咨询委员会，机构和当前实验室提供便利。完成这一 工作和相关的职业发展活动将支持Square博士成为校长的目标 研究牙齿和颅面发育界面的研究者。