Molecular mechanisms and epigenetic signatures that specify thymus fate

描述(由申请人提供)：尽管胸腺在T细胞发育中起着关键作用，但在胎儿发育过程中确定胸腺初始命运所需的机制尚不清楚。了解这些机制将有助于制定策略，在培养中产生功能强大的胸腺器官，并使老化、退化的胸腺恢复活力。在小鼠胚胎发育期间，胸腺的命运决定于E9.5和E11之间的腹侧第三咽囊(Pp)内胚层；该袋的背侧区域形成甲状旁腺。由于第三个PP必须被模式化成两个不同的细胞命运，因此必须有适当的机制来确保两个器官都得到适当的指定。我们实验室和其他实验室之前的工作表明，BMP2/4和FGF8/10信号与确定胸腺的命运有关，而SHH信号是甲状旁腺命运所必需的。这两条信号通路似乎是相互对立的；然而，它们相互作用和调节器官特异性转录网络的机制仍有待建立。Richie和Manley实验室最近的合作实验表明，转录因子TBX1是决定胸腺和甲状旁腺细胞命运的关键调节因子。我们已经证明，TBX1在第三内胚层的甲状旁腺区域表达，但不在腹侧胸腺命运域表达，并且异位的TBX1足以抑制Foxn1的表达。基于我们的初步数据，我们建立了一个胸腺命运规范的可测试模型，该模型表明FGF8和BMP4是正调节因子，而SHH和TBX1拮抗胸腺发育。这个项目使用一种新的方法来破译分子网络，该分子网络将胚胎学、遗传学、分子生物学、生物信息学和表观遗传学结合在一起，指定内胚层祖细胞与胸腺的命运有关。这一结果将为规划早期祖细胞以生成支持胸腺生成的功能性胸腺上皮细胞网络提供理论基础。我们将在以下具体目标中测试我们的模型。目的1.验证一种假说，即多种机制抑制了第三PP内胚层腹侧区域中TBX1的表达，并且这种抑制是建立胸腺命运所必需的。目的2.检测Tbx1抑制胸腺分化的能力是否受到时间限制，并确定异位Tbx1和/或SHH信号阻断胸腺命运的机制。目的3.验证器官特异性表观遗传特征的逐步建立决定胸腺命运的假设，并依赖于BMP和成纤维细胞生长因子信号通路之间的合作

DESCRIPTION (provided by applicant): Despite the critical role of the thymus in T cell development, the mechanisms required to specify initial thymus fate during fetal development are not known. Understanding these mechanisms will facilitate the development of strategies to generate functional thymus organs in culture, and to rejuvenate the aged, involuted thymus. Thymus fate is specified in the ventral 3rd pharyngeal pouch (pp) endoderm between E9.5 and E11 during mouse fetal development; the dorsal domain of this pouch gives rise to the parathyroid glands. As the 3rd pp must be patterned into two distinct cell fates, mechanisms must be in place to ensure that both organs are properly specified. Previous work from our labs and others have implicated BMP2/4 and FGF8/10 signals in establishing thymus fate, while SHH signaling is required for parathyroid fate. These two signaling pathways appear to act in opposition; however, the mechanisms by which they interact with each other and regulate organ-specific transcriptional networks remain to be established. Recent collaborative experiments from the Richie and Manley labs suggest that the transcription factor TBX1 is a key regulator of the decision between thymus and parathyroid cell fate. We have shown that Tbx1 is expressed in the parathyroid domain of the 3rd pp endoderm, but not in the ventral thymus fated domain, and that ectopic TBX1 is sufficient to suppress Foxn1 expression. Based on our preliminary data we generated a testable model of thymus fate specification in which suggest that FGF8 and BMP4 are positive regulators, whereas SHH and Tbx1 antagonize thymus development. This project uses a novel approach to decipher the molecular network that specifies endodermal progenitors to a thymus fate, combining embryology, genetics, molecular biology, bioinformatics, and epigenetics. The results will provide a rationale basis for programming early progenitors to generate a functional thymic epithelial cell network that supports thymopoiesis. We will test our model in the following Specific Aims. Aim 1. Test the hypothesis that multiple mechanisms suppress Tbx1 expression in the ventral domain of 3rd pp endoderm, and that this suppression is required to establish thymus fate. Aim 2. Test whether the ability of TBX1 to inhibit thymus differentiation is temporally restricted, and identify the mechanism by which ectopic TBX1 and/or SHH signaling blocks thymus fate. Aim 3. Test the hypothesis that progressive establishment of organ-specific epigenetic signatures determines thymus fate, and depends on cooperation between the BMP and FGF signaling pathways