CyclinD1 and the mechanisms of thymic involution

描述(申请人提供)：胸腺是T细胞发育所必需的。胸腺上皮细胞(TECs)提供必要的生长、分化和生存信号，因为胸腺细胞分化成熟为NA细胞并迁移到外周。胸腺退化会减少NAVE T细胞的输出，从而限制TCR谱系，并削弱对新遇到的抗原的免疫反应。TECs的耗竭是导致胸腺老化或细胞消融治疗后胸腺退化的一个主要因素。我们发现，在表达TECs的角蛋白5(K5)中加强Cyclin D1的表达会导致胸腺增生，但功能正常，在衰老过程中无法进行退缩。这些数据表明，持续高水平的Cyclin D1促进了TEC的增殖和分化，这反过来又调节了胸腺的动态平衡和退化。这一建议的一个主要目标是确定K5细胞周期蛋白D1转基因阻止胸腺退缩的机制(S)。虽然Cyclin D1是一种众所周知的细胞周期调节因子，但它也具有转录调节作用。细胞周期蛋白D1b是一种选择性剪接异构体，其中细胞周期蛋白框被保留，但外显子5被删除。外显子5包含一个LxxLL基序，它是一个核受体相互作用结构域，通过招募转录辅助激活因子来增强转录。我们发现，在K5细胞周期蛋白D1b转基因小鼠中，胸腺在正常的时间范围内退化。K5Cyclin D1和K5Cyclin D1b小鼠具有不同的卷曲表型，转录调控是其增强表达防止卷曲的关键机制。与这一观点一致，我们发现K5Cyclin 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.CylinD1b转基因株系来鉴定调节胸腺退化和幼稚T细胞输出的内源性因素和途径。AIMS 1和2将验证K5的假设，Cyclin D1通过控制细胞周期进程和转录调节影响胸腺的动态平衡。目的3探讨防止胸腺退缩对外周T细胞池组成的影响。由于我们在年轻的K5小鼠的脾中发现了数量增加的近期胸腺移居细胞(RTE)，我们将检验这样一种假设，即防止胸腺退缩维持RTE输出，而RTE输出反过来在衰老期间或细胞去除治疗后维持不同的幼稚外周T细胞池。本研究将为制定预防或逆转胸腺退缩的治疗策略提供合理的依据。

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.