Studies of the Roles of Twist1 and E12 in Tooth Morphogenesis

Twist1 和 E12 在牙齿形态发生中的作用研究

• 批准号: 8328601
• 负责人: Yongbo Lu
• 金额: $ 10.95万
• 依托单位: TEXAS A&M UNIVERSITY HEALTH SCIENCE CTR
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-05 至 2014-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8328601
• 关键词: Affect; Apoptosis; BHLH Protein; Binding; Biological Assay; Cell Line; Cell Proliferation; Cell Survival; Cell surface; Cells; Coupled; Couples; Coupling; DNA Binding; Data; Defect; Dental; Development; Developmental Process; Drosophila genus; Embryo; Embryonic Development; Epithelial; Epithelium; Extracellular Matrix; Family; Fibroblast Growth Factor; Fibroblast Growth Factor Receptor 2; Genes; Goals; Growth; Growth Factor; In Vitro; Interleukin-2; Joint structure of suture of skull; Laboratories; Mammals; Mediating; Mesenchymal; Mesenchyme; Mesoderm; Methods; Modality; Molecular; Molecular Genetics; Morphogenesis; Mutant Strains Mice; Natural regeneration; Nature; Odontoblasts; Organogenesis; Pathogenesis; Play; Promoter Regions; Research; Research Personnel; Research Project Grants; Role; Signal Transduction; Staging; Testing; Time; Tooth structure; Work; base; career; cell motility; chromatin immunoprecipitation; dosage; gastrulation; in vivo; innovation; insight; mutant; novel; paralogous gene; prevent; promoter; response; transcription factor; twist protein

DESCRIPTION (provided by applicant): The developing tooth is a valuable paradigm for understanding the genetic and molecular basis for the way in which morphogenesis and terminal differentiation are achieved during organogenesis. Tooth morphogenesis involves reciprocal signaling interactions between the dental epithelium and mesenchyme mediated by transcription factors and growth factors, as well as cell surface and extracellular matrix (ECM) molecules. As mitogenic factors, fibroblast growth factors (FGFs) play essential roles in coupling dental epithelial and mesenchymal growth, since epithelial FGFs mainly regulate dental mesenchymal growth and mesenchymal FGFs regulate epithelial growth. Twist1 and Twist2 belong to the evolutionarily conserved Twist family of basic helix-loop-helix (bHLH) transcription factors and are highly expressed in the dental mesenchyme during tooth morphogenesis. Their expression in the dental mesenchyme can be induced by epithelial FGFs, and studies in our laboratory have shown that Twist1, together with its heterodimeric binding partner, E12, regulates FGF receptor 2 (Fgfr2) and Fgf10 promoter activities. These observations suggest that epithelial FGFs and mesenchymal FGFs might be coupled through Twist1 and Twist2 in the dental mesenchyme. The long-term goal of this research project is to understand the molecular mechanisms governing tooth morphogenesis and terminal differentiation of odontoblasts. The proposed studies will test the hypothesis that Twist1 forms heterodimers with E12 in the dental mesenchyme, and modulates tooth morphogenesis by regulating the expression of Fgfr2 and Fgf10. To test this hypothesis, two specific aims are proposed. Aim 1 will determine whether Twist1, along with its heterodimeric binding partner E12, regulates the expression of Fgfr2 and Fgf10 at the transcriptional level using in vitro approaches including quantitative real-time PCR, chromatin immunoprecipitation (ChIP) assay, and deletion analyses of the Fgfr2 and Fgf10 promoters. Aim 2 will determine if alterations in the expression of Twist1 and Twist2 affect tooth morphogenesis and odontoblast differentiation in vivo. This aim will be achieved by generating and analyzing the Twist1 and Twist2 compound mutant embryos to examine whether there are Twist dosage-dependent changes in the expression of Fgfr2 and Fgf10 in the dental mesenchyme as well as phenotypic alterations in the mutant teeth. Successful completion of the proposed studies will show that Twist1 and Twist2 play key roles in mediating FGF signaling triggered by epithelial FGFs and in generating recursive mesenchymal FGFs in the dental mesenchyme during advancing tooth morphogenesis (bud to early bell stages).

描述（由申请人提供）：发育中的牙齿是理解器官发生过程中形态发生和终末分化实现方式的遗传和分子基础的一个有价值的范例。牙齿形态发生涉及由转录因子和生长因子以及细胞表面和细胞外基质（ECM）分子介导的牙上皮和间充质之间的相互信号相互作用。作为有丝分裂因子，成纤维细胞生长因子（FGF）在耦合牙齿上皮和间质生长中发挥着重要作用，因为上皮FGF主要调节牙齿间质生长，间质FGF调节上皮生长。 Twist1 和 Twist2 属于进化上保守的碱性螺旋-环-螺旋 (bHLH) 转录因子 Twist 家族，在牙齿形态发生过程中在牙齿间充质中高度表达。它们在牙齿间充质中的表达可以由上皮 FGF 诱导，我们实验室的研究表明，Twist1 与其异二聚体结合伴侣 E12 一起调节 FGF 受体 2 (Fgfr2) 和 Fgf10 启动子活性。这些观察结果表明，上皮 FGF 和间充质 FGF 可能通过牙齿间充质中的 Twist1 和 Twist2 偶联。该研究项目的长期目标是了解控制牙齿形态发生和成牙本质细胞终末分化的分子机制。拟议的研究将检验以下假设：Twist1 与牙齿间充质中的 E12 形成异二聚体，并通过调节 Fgfr2 和 Fgf10 的表达来调节牙齿形态发生。为了检验这一假设，提出了两个具体目标。目标 1 将使用体外方法（包括定量实时 PCR、染色质免疫沉淀 (ChIP) 测定和 Fgfr2 缺失分析）确定 Twist1 及其异二聚体结合伴侣 E12 是否在转录水平调节 Fgfr2 和 Fgf10 的表达和 Fgf10 启动子。目标 2 将确定 Twist1 和 Twist2 表达的改变是否影响体内牙齿形态发生和成牙本质细胞分化。这一目标将通过生成和分析 Twist1 和 Twist2 复合突变体胚胎来检查牙齿间充质中 Fgfr2 和 Fgf10 的表达是否存在 Twist 剂量依赖性变化以及突变体牙齿的表型改变来实现。拟议研究的成功完成将表明，Twist1 和 Twist2 在介导上皮 FGF 触发的 FGF 信号传导以及在牙齿形态发生过程中（芽期到早期钟声阶段）在牙齿间充质中产生递归间充质 FGF 方面发挥着关键作用。