Regulation of Runx2 Function by Twist-1 in Tooth Development

在这一竞争性更新中，我们建议继续研究Runx 2在牙齿发育中的作用。数据从 我们在上一个奖项期间的工作表明，Runx 2在指导牙齿上皮细胞的命运中起着关键作用。 形态发生以及控制成牙本质细胞分化的开始。我们的研究表明， 了解Runx 2活性在牙齿形态发生和细胞分化过程中是如何精确调节的， 在这些过程中的作用通过与其他分子的相互作用来调节。核蛋白Twist-1是 作为Runx 2的调节蛋白伴侣特别感兴趣。我们研究Runx 2，一种细胞分化， 因子，与Twist-1，一种细胞存活因子相互作用，来自我们和其他实验室的研究，表明， Runx 2和Twist-1之间的这些相互作用发生在蛋白质水平。我们的实验将直接测试 假设Runx 2在成牙本质细胞分化中的关键功能是由Twist-1在蛋白水平调节的， 蛋白质相互作用在本质上是功能性拮抗的。Runx 2功能的选择性和短暂性阻断 通过Twist-1提供了抑制成牙本质细胞分化直到形态发生完成的手段。我们进一步 我认为Runx 2和Twist-1之间的相互作用不是相互拮抗的，因为Twist-1可以介导细胞凋亡， 通过FGF介导的上皮间充质信号传导在形态发生期间增殖。因此， 人CCD中的多生牙和Runx 2（-/-）小鼠中的副芽可能反映了Twist-1活性的增加 而不是Runx 2水平降低的直接影响。目标1将确定Runx 2和Twist-1的模式是否 (mRNA和蛋白质）表达与它们在牙齿发育期间提出的伙伴关系相容， 将这些模式与牙齿形态发生和成牙本质细胞的分子标记物的表达相关联 分化目标2将用小鼠遗传功能丧失和功能获得方法评估是否 Twist-1表达的改变影响牙齿形态发生和成牙本质细胞分化。目标3将研究 牙齿间充质中Runx 2- Twist-1蛋白相互作用的分子基础及其功能后果 Runx 2上的这种相互作用在成牙本质细胞分化中起作用，Aim 4将测试BHLH结构域是否在成牙本质细胞分化中起作用。 扭曲瘦介导的牙齿形态发生通过FGF信号，这是独立的与Runx 2的相互作用。这些 研究将增加我们对Runx 2如何通过其在牙齿发育中的选择性功能的理解。 与Twist-1合作重要的是，他们将解释多生牙是如何形成的，以及成牙本质细胞的分化是否与多生牙的形成有关。 通过抑制的释放来确定。这些数据也将为理解发病机制提供一个框架 锁骨颅骨发育不良和Saethre-Chotzen综合征，这两种人类遗传性疾病威胁到牙列。

In this competing renewal, we propose to continue our research on the role of Runx2 in tooth development. Data from our work in the previous award period indicated key roles for Runx2 in directing the fate of dental epithelium during morphogenesis as well as in controlling the onset of odontoblast differentiation. Our studies point to the critical need to learn how Runx2 activities are precisely regulated during tooth morphogenesis and cell differentiation and whether its role in these processes is modulated through interactions with other molecules. The nuclear protein Twist-1 is of particular interest as a regulatory protein partner for Runx2. Our rationale for studying if Runx2, a cell differentiation factor, interacts with Twist-1, a cell survival factor, is derived from studies in our and other laboratories that suggest that these interactions between Runx2 and Twist-1 occur at the protein level. Our experiments will directly test the hypothesis that Runx2's key functions in odontoblast differentiation are regulated by Twist-1 at the level of protein- protein interactions that are functionally antagonistic in nature. The selective and transient blocking of Runx2 function by Twist-1 provides a means to restrain odontoblast differentiation until morphogenesis is complete. We further propose that interactions between Runx2 and Twist-1 are not mutually antagonistic as Twist-1 can mediate cell proliferation during morphogenesis via FGF-mediated epithelial mesenchymal signaling. Hence, the presence of supernumerary teeth in human CCD and accessory buds in Runx2(-/-) mice likely reflect increased activity of Twist-1 rather than a direct effect of decreased levels of Runx2. Aim 1 will determine if the patterns of Runx2 and Twist-1 (mRNA and protein) expression are compatible with their proposed partnership during tooth development and will correlate these patterns with the expression of molecular markers of tooth morphogenesis and odontoblast differentiation. Aim 2 will assess with mouse genetic loss-of-function and gain-of-function approaches whether alterations in Twist-1 expression affects tooth morphogenesis and odontoblast differentiation. Aim 3 will study the molecular basis of Runx2 - Twist-1 protein interactions in dental mesenchyme and the functional consequences of this interaction on Runx2 functions in odontoblast differentiation, and Aim 4 will test whether the bHLH domain of Twist-lean mediate tooth morphogenesis via FGF-signaling that is independent of its interactions with Runx2. These studies will increase our understanding of how Runx2 achieves its selective functions in tooth development through its partnership with Twist-1. Importantly, they will explain how supernumerary teeth form and if odontoblast differentiation is determined by the release of an inhibition. Such data will also provide a framework for understanding the pathogenesis of Cleidocranial Dysplasia and Saethre-Chotzen Syndrome, two human genetic disorders that threaten dentition.