MOLECULAR MECHANISMS OF VERTEBRATE TOOTH INITIATION

脊椎动物牙齿萌生的分子机制

• 批准号: 6728124
• 负责人: Yiping Chen
• 金额: $ 37.13万
• 依托单位: TULANE UNIVERSITY OF LOUISIANA
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-02-01 至 2009-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6728124
• 关键词: RNA interference; biological signal transduction; bone morphogenetic proteins; cell cell interaction; dental development; developmental genetics; electroporation; epithelium; gene expression; genetically modified animals; laboratory mouse; mesenchyme; organ culture

DESCRIPTION (provided by applicant): Multiple signaling molecules, including BMPs, FGFs, Shh, and Wnt proteins, have been implicated in mediating tissue interactions that regulate tooth initiation, morphogenesis and differentiation. In mice, the odontogenic potential, defined as the capability of an odontogenic tissue to elicit tooth developmental program in non-dental tissues, resides initially in dental epithelium, and then shifts to dental mesenchyme. Our long-term goal is to delineate the molecular basis of the odontogenic potential. Based on our previous studies, we hypothesize that during tooth initiation, BMP4 functions as an essential component of the odontogenic potential in the early dental epithelium, to induce the formation of the basal layer epithelium as well as to stimulate local cell proliferation to form a tooth bud. We further hypothesize that Wnt signaling pathway constitutes a component of the odontogenic potential in dental mesenchyme. Two specific aims are proposed to test these hypotheses. In Aim 1, the role of BMP4 in the early dental epithelium in tooth initiation will be defined by conditional knockout studies via the Cre/LoxP approach to achieve tissue-specific mutation of Bmp4 in the dental epithelium. This approach will also be used to delete the Bmpr-lA and Bmpr-IB genes in the dental epithelium to study if BMP4 acts intra-epithelially. Whether signaling of BMP4 to dental mesenchyme is essential for the acquisition of odontogenic potential in the mesenchyme will be tested by tissue recombination studies. Furthermore, whether Shh mediates Bmp4 activity in dental epithelium will be examined by conditional knockout and transgenic rescue experiments. In Aim 2, if Wnt signaling acts as a critical component of the odontogenic potential in dental mesenchyme will be studied by disrupting Writ signaling pathway in tissues used for tissue recombination. This will be achieved by overexpressing Wnt antagonist sFrp3 in dental mesenchyme, and Dkk1 and a dominant negative form of Tcf in responding non-dental epithelia via microelectroporation and virus-mediated gene delivery. The potential role of Wnt5a in the induction of tooth formation by dental mesenchyme will also be tested. Lastly, whether different sFrps confer the different inductive properties to incisor and molar mesenchyme will be determined by virus-mediated gene overexpression and RNAi-mediated gene knock-down studies, together with tissue recombination and organ culture studies.

描述（由申请人提供）：多种信号分子，包括 BMP、FGF、Shh 和 Wnt 蛋白，与介导调节牙齿萌生、形态发生和分化的组织相互作用有关。在小鼠中，成牙潜能定义为牙源组织在非牙组织中引发牙齿发育程序的能力，最初存在于牙上皮中，然后转移到牙间充质中。我们的长期目标是描绘成牙潜力的分子基础。基于我们之前的研究，我们假设在牙齿萌生过程中，BMP4作为早期牙上皮成牙潜能的重要组成部分，诱导基底层上皮的形成以及刺激局部细胞增殖形成牙齿芽。我们进一步假设 Wnt 信号通路构成牙间充质中成牙潜能的一个组成部分。提出了两个具体目标来检验这些假设。在目标 1 中，将通过 Cre/LoxP 方法的条件敲除研究来定义 BMP4 在牙齿萌生过程中早期牙上皮中的作用，以实现牙上皮中 Bmp4 的组织特异性突变。该方法还将用于删除牙上皮中的 Bmpr-1A 和 Bmpr-IB 基因，以研究 BMP4 是否在上皮内起作用。 BMP4 向牙齿间充质发出的信号是否对于获得间充质中的成牙潜力至关重要，将通过组织重组研究进行测试。此外，Shh是否介导牙上皮细胞中的Bmp4活性将通过条件敲除和转基因拯救实验进行检查。在目标 2 中，将通过破坏用于组织重组的组织中的 Writ 信号通路来研究 Wnt 信号是否作为牙齿间充质中成牙潜力的关键组成部分。这将通过微电穿孔和病毒介导的基因传递在牙齿间充质中过表达 Wnt 拮抗剂 sFrp3，以及在响应非牙齿上皮细胞中过表达 Dkk1 和 Tcf 的显性失活形式来实现。 Wnt5a 在牙齿间充质诱导牙齿形成中的潜在作用也将得到测试。最后，不同的 sFrps 是否赋予门牙和磨牙间充质不同的诱导特性，将通过病毒介导的基因过表达和 RNAi 介导的基因敲低研究，以及组织重组和器官培养研究来确定。