Identification and Characterization of Genes from del22q

说明(申请人提供)：人、小鼠和斑马鱼的心血管发育需要TBX1。自从最初认识到该基因在发育中的作用以来，在突变分析方面取得了相当大的进展，导致了新的假说。Tbx1是一种T-box转录因子，在胚胎咽部的分割、主动脉弓的构型和心脏流出道的形态发生中都是必需的。然而，如果我们不解决两个基本问题：什么是调控TBX1基因的关键角色，什么是TBX1蛋白的靶标，那么关于该基因的进一步知识获取将是有限的。TBX1基因具有微妙的动态调节，在这里，我们表明这与胚胎发育过程中的特定时间作用有关。例如，心血管发育需要早期表达，而胸腺器官发生和腭部发生需要晚期表达。此外，我们还展示了在心血管发育过程中对TBX1功能的组织特异性要求。这项建议建立在这些新数据的基础上，以寻找在关键时间和组织中重要的TBX1的调节因子和靶标。 总体目标是在TBX1发挥作用的遗传途径中建立硬链接。我们相信，这项工作将导致导致或修改先天性心脏病的基因。在第一个具体目标中，我们将使用新的方法来确定咽内胚层中调节TBX1所需的遗传元件，咽内胚层是TBX1功能的关键组织，并将确定该组织中的TBX1表达是否足以挽救突变体中出现的心血管异常。在第二个特定目标中，我们将使用转录组分析和染色质免疫沉淀分析相结合的方法来识别转录靶标。初步数据显示了这种方法的威力。此外，我们还建议对候选目标基因进行体外和体内验证。 该项目直接阐述了TBX1在心血管发育中的作用，该功能由相关组织中基因和蛋白质转录靶标的上游调控所定义。数据应该提供DiGeorge和Velcore-Facial综合征以及可能影响心血管发育的其他遗传性疾病中破坏的遗传网络的解剖。

DESCRIPTION (provided by applicant): Tbx1 is required for cardiovascular development in human, mouse and zebra fish. Since the initial recognition of the role of this gene in development, there has been considerable progress in the analysis of mutants that led to novel hypotheses. Tbx1 is a T-box transcription factor required for segmentation of the embryonic pharynx, patterning of the aortic arch, and morphogenesis of the outflow tract of the heart. However, further knowledge acquisition about this gene would be limited if we do not address two fundamental questions: what are the critical players in the regulation of the Tbx1 gene, and what are the targets of the Tbx1 protein. The Tbx1 gene has an exquisitely dynamic regulation, and here we show that this is associated with time-specific roles during embryogenesis. For example, early expression is required for cardiovascular development while late expression is required for thymic organogenesis and palatogenesis. In addition, we show tissue specific requirements for Tbx1 function during cardiovascular development. This proposal builds upon these new data to search for regulators and targets of Tbx1 that are important at the critical times and tissues. The overall aim is to establish hard links in the genetic pathway within which Tbx1 functions. We believe that this work will lead to genes causing or modifying congenital heart disease. In the first specific aim we will use novel approaches to identify genetic elements necessary for Tbx1 regulation in the pharyngeal endoderm, a key tissue for Tbx1 function, and will also determine if Tbx1 expression in this tissue is sufficient to rescue the cardiovascular abnormalities seen in mutants. In the second specific aim, we will use a combination of transcriptome analysis and chromatin immunoprecipitation assay to identify transcriptional targets. Preliminary data show the power of this approach. In addition, we propose in vitro and in vivo validation of candidate target genes. This project addresses directly the function of Tbx1 in cardiovascular development as defined by upstream regulation of the gene and transcriptional targets of the protein in relevant tissues. Data should provide the dissection of a genetic network disrupted in DiGeorge and Velocardiofacial syndromes, and possibly in other genetic disorders affecting cardiovascular development.