Identification of downstream signals for Pitx2a

描述(由申请人提供)： Pitx2是一种双类同源结构域转录因子，与人类Rieger综合征有关，在小鼠、鸡、青蛙和斑马鱼的左右发育过程中发挥着重要作用。在器官发生过程中，细胞的增殖、运动以及对细胞命运的指示信号的变化被认为是引导左右不对称的重要因素。作为一种转录因子，Pitx2应该激活和/或抑制其靶基因的转录，以执行特定的细胞功能。然而，人们对Pitx2的下游靶点和信号通路知之甚少。在这项研究中，我建议寻找受Pitx2a(Pitx2亚型之一)调控的靶基因和相关的信号通路。我将使用一种异位表达Pitx2a的可诱导HeLa细胞系，它已经在实验室中获得。这个可诱导的细胞系将使我能够根据细胞表型追踪Pitx2a的下游信号通路，并通过比较诱导和未诱导条件下从细胞中分离的RNA来确定Pitx2a的靶基因。转基因小鼠模型也将被用来评估可能的下游靶基因在介导小鼠体内Pitx2a功能方面的重要性。利用这些细胞培养和小鼠模型系统识别这些下游靶基因和信号通路，应该能让我在理解Pitx2a功能的潜在机制以及胚胎发育过程中Pitx2a表达与其在形态发生中的作用之间缺失的联系方面取得进展。

DESCRIPTION (provided by applicant): Pitx2, a bicoid-type homeodomain transcription factor, has been implicated in Rieger's syndrome in humans and plays an important role in mediating left-right development in mice, chickens, frogs and zebrafish. It is believed that cell proliferation, cell motility as well as changes in instructive signals for cell fate in local areas are important for directing left-right asymmetry during organogenesis. As a transcription factor, Pitx2 should activate and/or repress the transcription of its target genes to execute specific cellular functions. However, little is known about the downstream targets and signaling pathways for Pitx2. I propose in this study to search for the target genes regulated by Pitx2a (one of the Pitx2 isoforms) and the relevant signaling pathways. I will use an inducible HeLa cell line ectopically expressing Pitx2a, which is already available in the laboratory. This inducible cell line will allow me to trace the downstream signaling pathways for Pitx2a based on cell phenotype and to identify the target genes for Pitx2a by comparing the RNA isolated from cells under the induced and uninduced conditions. The transgenic mouse model will also be used to assess the importance of the putative downstream target genes in mediating the function of Pitx2a in mice. Identification of these downstream target genes and signaling pathways using these cell culture and mouse model systems should allow me to make progress towards understanding of the mechanism underlying the functions of Pitx2a as well as the missing links between Pitx2a expression and its roles in morphogenesis during embryonic development.