CyclinD1 and the mechanisms of thymic involution

描述（由申请方提供）：胸腺对T细胞发育至关重要。当胸腺细胞分化成熟为幼稚T细胞并迁移到外周时，胸腺上皮细胞（TEC）提供必要的生长、分化和生存信号。胸腺退化减少了幼稚T细胞的输出，这限制了TCR的库，并损害了对新遇到的抗原的免疫反应。TECs的消耗是导致衰老期间或细胞清除治疗后胸腺退化的主要因素。我们发现，在角蛋白5（K5）表达TEC中强制细胞周期蛋白D1的表达会导致胸腺增生，但功能正常，在衰老过程中不能进行退化。这些数据表明，持续高水平的细胞周期蛋白D1增强TEC的增殖和分化，这反过来又调节胸腺的稳态和退化。该建议的主要目标是确定K5.Cyclin D1转基因防止胸腺退化的机制。虽然细胞周期蛋白D1是一个众所周知的细胞周期调控因子，它也作为一个转录调控因子。细胞周期蛋白D1b是一种选择性剪接的同种型，其中细胞周期蛋白盒被保留，但外显子5被删除。外显子5含有LxxLL基序，其是通过募集转录辅激活因子来增强转录的核受体相互作用结构域。我们发现K5.CyclinD1b转基因小鼠的胸腺在正常的时间范围内退化。在K5.Cyclin D1和K5.Cyclin D1b小鼠中不同的退化表型暗示转录调节是Cyclin D1的强制表达防止退化的关键机制。与这一观点一致，我们发现K5.Cyclin D1转基因增强了Foxn1的表达，Foxn1是TEC发育和维持所需的转录因子。我们将使用K5.CyclinD1和K5.CyclinD1b转基因系来鉴定调节胸腺退化和幼稚T细胞输出的内源性因子和途径。目的1和2将检验K5.CyclinD1通过控制细胞周期进程和转录调节影响胸腺稳态的假设。目的3探讨防止胸腺退化对外周血T细胞池组成的影响。由于我们发现在年轻的K5.Cyclin D1小鼠的脾脏中最近胸腺移民（RTE）的数量增加，我们将测试这一假设，即防止胸腺退化维持RTE输出，这反过来又保持了不同的池幼稚外周T细胞在老化过程中或细胞清除治疗后。本研究将为制定预防或逆转胸腺退化的治疗策略提供理论依据。

DESCRIPTION (provided by applicant): The thymus is essential for T cell development. Thymic epithelial cells (TECs) supply essential growth, differentiation and survival signals as thymocytes differentiate mature to na¿ve T cells and emigrate to the periphery. Thymus involution reduces na¿ve T cell output, which restricts the TCR repertoire and impairs immune responses to newly encountered antigens. Depletion of TECs is a major factor contributing to thymus involution during aging or after cytoablative therapy. We find that enforcing Cyclin D1 expression in keratin 5 (K5) expressing TECs results in a hyperplastic, yet functional thymus that fails to undergo involution during aging. These data suggest that sustained high levels of Cyclin D1 enhances TEC proliferation and differentiation, which, in turn, regulates thymus homeostasis and involution. A major goal of this proposal is to determine the mechanism(s) by which the K5.Cyclin D1 transgene prevents thymus involution. Although Cyclin D1 is a well-known cell cycle regulator, it also functions as a transcriptional regulator. Cyclin D1b is an alternatively spliced isoform in which the cyclin box is retained, but exon 5 is deleted. Exon 5 contains an LxxLL motif, which is a nuclear receptor interaction domain that enhances transcription by recruiting transcriptional coactivators. We find that the thymus involutes within normal time frame in K5.CyclinD1b transgenic mice. The distinct involution phenotypes in K5.Cyclin D1 and K5.Cyclin D1b mice implicate transcriptional regulation as a key mechanism by which enforced expression of Cyclin D1 prevents involution. Consistent with this notion, we find that the K5.Cyclin D1 transgene enhances expression of Foxn1, a transcription factor required for TEC development and maintenance. We will use the K5.CyclinD1 and K5.CyclinD1b transgenic lines to identify endogenous factors and pathways that regulate thymus involution and naive T cell output. Aims 1 and 2 will test the hypothesis that K5.CyclinD1 influences thymus homeostasis both by control of cell cycle progression and by transcriptional regulation. Aim 3 explores the consequences of preventing thymus involution on the composition of the peripheral T cell pool. Since we find an increased number of recent thymic emigrants (RTEs) in spleens of young K5.Cyclin D1 mice, we will test the hypothesis that preventing thymus involution sustains RTE output, which in turn maintains a diverse pool of naive peripheral T cells during aging or after cytoablative therapy. This investigation will provid a rational basis for developing therapeutic strategies to prevent or reverse thymus involution.