Identifying new genes involved in thymic involution

项目摘要/摘要 胸腺是负责产生T细胞的主要免疫器官，其功能随着 年龄(退化)被认为是与衰老相关的免疫衰老的重要因素。有很强的 有证据表明，与衰老相关的胸腺退化是在青年时期通过胸腺中的机制启动的 上皮细胞(TECs)导致其耗尽和间隔室组织的丧失。尽管它扮演着核心角色 T细胞获得性免疫的形成和维持及胸腺的多效性负性影响 衰老过程中的退化，令人惊讶的是胸腺器官功能的分子调节器很少，动态平衡和 已经确定了卷土重来的问题。一种关键的转录因子FOXN1已知是这两种基因的主要调节者 胸腺发育和出生后的维持。然而，它的调控机制和途径 它通过什么影响TEC的发育、功能和衰老相关的退化在很大程度上仍不清楚。TECS 是胸腺细胞总数的一小部分，通过实验分离是出了名的困难。因此，识别 通过基于基因表达或生化分析的调节器和效应器是困难的，而且无论如何都会是困难的 专门针对TEC子集或特定年龄。鉴于胸腺退化的渐进性，设计 这种识别胸腺功能和随年龄增长而维持的关键调节因素的实验将是加倍的 很有挑战性。正向基因筛查是根据基因功能识别基因的有力工具，因此 允许公正地发现新的途径和机制，但通常不适用于与衰老相关的 表型。在最近由R21资助的一个项目中，我的实验室使用ENU执行了一个主要的修饰符筛选 突变，以确定新的突变，当杂合子时，要么增强(增加严重性)，要么 抑制(挽救/减轻严重程度)Foxn1Z/Z胸腺退化表型。我们确定了18个主要的 修改器行，包括增强器和抑制器。因为它们影响Foxn1的表达和/或 在胸腺早退的Foxn1Z/Z模型中的作用，我们的假设是这些修饰物代表 以前未被识别的基因，通常作用于促进或抑制与衰老相关的胸腺退化。在 目前的提案，我们将跟进我们成功的初始项目，以确定在我们的试验中发现的修改 筛选，并测试它们是否影响正常老化期间的退化轨迹。这一新颖的方法可以 研究胸腺退化的分子基础是一种潜在的变革性方法 识别控制胸腺功能和衰老相关退化的新基因、途径和机制。

Project Summary/Abstract The thymus is the primary immune organ responsible for the generation of T cells, and its functional decline with age (involution) is considered a significant contributor to aging-associated immunosenescence. There is strong evidence that aging-related thymic involution is initiated during youth by mechanisms that operate in thymic epithelial cells (TECs) to result in their depletion and loss of compartmental organization. Despite its central role in the formation and maintenance of T cell adaptive immunity and the pleiotropic negative impacts of thymus involution during aging, surprisingly few molecular regulators of thymus organ function, homeostasis, and involution have been identified. A key transcription factor, FOXN1, is known to be a primary regulator of both thymus development and postnatal maintenance. However, the mechanisms of its regulation and the pathways through which it affects TEC development, function, and aging-related involution remain largely unknown. TECs are a small subset of total thymus cells and are notoriously difficult to isolate experimentally. Thus, identifying regulators and effectors by gene expression-based or biochemical analysis is difficult, and in any case would be specifically targeted to a TEC subset or specific age. Given the gradual nature of thymic involution, designing such experiments to identify key regulators of thymus function and maintenance with aging would be doubly challenging. Forward genetic screens are powerful tools for identifying genes based on their function, thus allowing unbiased discovery of novel pathways and mechanisms, but are not generally practical for aging-related phenotypes. In a recent R21-funded project, my lab performed a dominant modifier screen using ENU mutagenesis to identify novel mutations that, when heterozygous, either enhance (increase the severity) or suppress (rescue/reduce the severity) the Foxn1Z/Z thymic involution phenotype. We identified 18 dominant modifier lines, including both enhancers and suppressors. Because they impact Foxn1 expression and/or function in the Foxn1Z/Z model of premature thymic involution, our hypothesis is that these modifiers represent previously unrecognized genes that normally act to promote or restrain aging-related thymic involution. In the current proposal, we will follow up on our successful initial project to identify the modifiers found in our pilot screen, and test whether they affect the trajectory of involution during normal aging. This novel approach to investigating the molecular basis of thymus involution represents a potentially transformative approach to identifying new genes, pathways, and mechanisms that control thymus function and aging-related involution.