Project 2 - The role of Foxn1 in controlling the transition from thymus expansion to homeostasis

摘要 越来越多的证据表明，胸腺上皮细胞（TEC）是负责协调 胎儿发育和产后功能。已知一个转录因子FOXN 1是一个关键的转录因子。 TEC增殖的多个方面的调节剂和在整个生命周期中的功能。TEC隔间 从胎儿/新生儿扩张计划切换到青少年体内平衡计划，具有关键的过渡点 小鼠出生后约7天（P0.07），人类出生后约4个月。小鼠实验数据显示， 这种开关由视网膜母细胞瘤（RB）通路控制，至少部分通过抑制Foxn 1基因起作用 通过E2 F转录因子表达。缺乏多个RB家族成员的小鼠不能进行这种转换 并继续扩张。其中CyclinD 1在TEC中特异性过表达的K5.D1转基因系， 通过激活抑制RB功能的细胞周期蛋白依赖性激酶来模拟这种RB功能丧失表型。无论是 RB突变体和K5.D1转基因Foxn 1基因表达升高，抑制Foxn 1表达 使用出生后特异性亚型等位基因（Foxn 1 Z/Z）恢复胎儿到幼年的转换，使胸腺正常化 尺寸我们还表明，Foxn 1对TEC增殖的调节主要发生在MHCIIlo TEC中，这与Foxn 1对TEC增殖的调节一致。 根据Richie实验室的初步数据，Sca 1-MHCIIlo TEC子集可能包含一个关键的增殖因子， 在从胎儿扩张到幼年稳态的转换过程中，这些数据表明 在围产期到幼年期的过渡期，RB蛋白通过E2 F转录因子调节Foxn 1的转录 在特定的TEC亚群中差异化，以“阻止”TEC增殖，从而调节器官大小。那里 越来越多的证据表明，新生儿和成人胸腺具有阶段特异性功能， 细胞群由于FOXN 1也被认为是TEC分化的关键调节因子，因此FOXN 1如何改变TEC的分化， 调控表达以控制TEC增殖也应该对TEC在细胞中的功能有显著影响。 新生儿和成人胸腺。我们提出了三个具体的目标来检验假设，即RB依赖 新生儿时期Foxn 1基因表达水平的变化对建立器官至关重要 体内平衡也是产生特定微环境所必需的， 器官功能。这个项目的目标是确定：如何在Foxn 1表达的变化， 转换影响小鼠中的基质组成变化，并与人胸腺中的基质变化进行比较; 如何Foxn 1的表达是由RB通路在这个过渡过程中调节;以及这些Foxn 1依赖性 过程影响免疫功能。

Abstract There is growing evidence that thymic epithelial cells (TEC) are the key cell type responsible for orchestrating the fetal development and postnatal function. A single transcription factor, FOXN1, is known to be a critical regulator of multiple aspects of TEC proliferation and function throughout the lifespan. The TEC compartment switches from a fetal/neonatal expansion program to a juvenile homeostasis program, with a key transition point occurring at about 7 days postnatal in mice (P07), and about 4 months of age in humans. Data in mice show that this switch is controlled by the retinoblastoma (RB) pathway, acting at least in part by suppression of Foxn1 gene expression by E2F transcription factors. Mice deficient for multiple RB family members fail to make this switch and continue to expand. The K5.D1 transgenic line, in which CyclinD1 is specifically overexpressed in TEC, mimics this RB loss of function phenotype by activating cyclin-dependent kinases that inhibit RB function. In both RB mutants and K5.D1 transgenics Foxn1 gene expression is elevated, and suppressing Foxn1 expression using a postnatal-specific hypomorphic allele (Foxn1Z/Z) restores the fetal to juvenile switch, normalizing thymus size. We have also shown that Foxn1 regulation of TEC proliferation occurs primarily in MHCIIlo TEC, consistent with preliminary data from the Richie lab that a Sca1-MHCIIlo TEC subset may contain a key proliferating progenitor population during the switch from fetal expansion to juvenile homeostasis. These data suggest that during the perinatal to juvenile transition, RB proteins modulate Foxn1 transcription via E2F transcription factors differentially in specific TEC subsets to ‘put the brakes on’ TEC proliferation, and thus regulate organ size. There is also growing evidence that the neonatal and adult thymi have stage-specific functions that generate distinct T cell populations. As FOXN1 is also known to be a key regulator of TEC differentiation, the changes in how Foxn1 expression is regulated to control TEC proliferation should also have significant impacts on TEC function in the neonatal and adult thymus. We propose three specific aims to test the hypothesis that the RB-dependent changes in Foxn1 gene expression levels during the neonatal period that are essential for establishing organ homeostasis are also required to generate specific microenvironments that are necessary for neonatal-specific organ functions. The goals of this project are to determine: how changes in Foxn1 expression across this transition impact stromal composition changes in mouse, and compare to stromal changes in human thymus; how Foxn1 expression is regulated by the RB pathway during this transition; and how these Foxn1-dependent processes impact immune function.