Regulation of CD8 T Cell Immunity by CDK Inhibitor p27Kip1

描述(由申请人提供)：CD8 T细胞在抵御艾滋病毒、乙肝和丙型肝炎、爱泼斯坦-巴尔病毒、巨细胞病毒和阿拉伯病毒等病毒感染方面发挥着至关重要的作用。因此，一个主要的驱动力是开发新的疫苗接种策略，可以诱导强大的CD8 T细胞记忆，并产生针对这些病毒病原体的有效保护性免疫。最近的研究表明，记忆性CD8 T细胞前体作为效应细胞的子集存在于初级T细胞反应的高峰期。然而，与有功能的记忆细胞不同，这些记忆前体细胞处于细胞周期停滞状态；它们缺乏通过自身平衡增殖进行自我更新的能力，或者在抗原重新调用时进行增殖，并提供较差的保护性免疫。这些记忆前体开始进行分化程序，在抗原清除后持续几周的过程中成为有能力的记忆细胞。尽管人们认识到记忆分化的效应是CD8 T细胞依赖的保护性免疫发展的限速步骤，但记忆前体细胞增殖阻断的诱导和随后逆转的机制尚不清楚。我们在小鼠淋巴细胞性脉络膜脑膜炎病毒(LCMV)模型上的研究表明，细胞周期蛋白依赖性激酶抑制剂p27Kip1的遗传缺陷不仅加速了效应CD8T细胞向记忆细胞的分化，而且显著增加了CD8T细胞的记忆量。基于这些发现，我们的中心假设是：p27Kip1通过在有丝分裂信号的反应中反对细胞周期重新进入来阻碍记忆CD8 T细胞与主效应子CD8 T细胞的分化。我们进一步假设，记忆性CD8 T细胞分化包括p27Kip1的表达受上游FOXO转录因子的调控。本研究通过以下几个具体目的来阐明FOXO/p27Kip1通路在调节记忆CD8 T细胞分化中的关键作用：1)检验p27Kip1调节记忆细胞分化的作用是CD8 T细胞自主的假说；2)检验FOXO转录因子调控记忆CD8 T细胞分化的假说。AIMS 1和2将利用互补的方法，包括TCR转基因(TG)小鼠、骨髓嵌合体、T细胞中p27Kip1或FOXO条件缺陷的小鼠，以及可诱导的p27Kip1/FOXO TG小鼠，以确定p27Kip1和FOXO在体内调节CD8T细胞从效应者向功能记忆细胞分化过程中发生的表型和功能改变的动态作用。这些研究将为控制记忆细胞分化的机制提供重要的洞察力，并有可能确定p27Kip1和FOXO作为开发新的免疫疗法的分子靶点，以加速和增强疫苗诱导的CD8 T细胞依赖的保护性免疫对病毒感染的诱导。公共卫生报道：CD8 T细胞在抵御人类多种病毒感染方面发挥着关键作用。在这项建议中，我们试图确定关键的分子调节因子(p27Kip1和FOXO)，它们可以通过免疫疗法调节CD8T细胞，以增强疫苗诱导的保护性抗病毒免疫。这些研究应该有助于开发有效的疫苗，以保护人类免受病毒引起的疾病的侵袭。

DESCRIPTION (provided by applicant): CD8 T cells play a crucial role in defense against viral infections like HIV, hepatitis B and C, Epstein Barr virus, cytomegalovirus, and arenaviruses. A major drive therefore has been to develop novel vaccination strategies that can elicit potent CD8 T cell memory and engender effective protective immunity against these viral pathogens. Recent studies have indicated that memory CD8 T cell precursors are present as a subset of effector cells at the peak of the primary T cell response. However, unlike competent memory cells, these memory precursors are in a state of cell cycle arrest; they lack the ability to self renew by homeostatic proliferation or proliferate in response to antigen re-call, and provide poor protective immunity. These memory precursors embark on a program of differentiation to become competent memory cells by a process lasting up to several weeks after antigen clearance. Despite the recognition that effector to memory differentiation is a rate-limiting step in the development of CD8 T cell-dependent protective immunity, the mechanisms underlying the induction and subsequent reversal of proliferative block in memory precursors are poorly defined. Our studies using the mouse model of lymphocytic choriomeningitis virus (LCMV) have shown that genetic deficiency of cyclin dependent kinase inhibitor p27Kip1 not only accelerated the differentiation of memory cells from effector CD8 T cells, but also significantly increased the magnitude of CD8 T cell memory. Based on these findings, our central hypothesis is: p27Kip1 impedes differentiation of memory CD8 T cells from primary effector CD8 T cells by opposing cell cycle re-entry in response to mitogenic cues. We further hypothesize that memory CD8 T cell differentiation including p27Kip1 expression is regulated by the upstream FOXO transcription factors. This proposal focuses on elucidating the key roles of FOXO/p27Kip1 pathway in regulating the differentiation of memory CD8 T cells through these specific aims: 1) To test the hypothesis that effects of p27Kip1 in regulating memory cell differentiation are CD8 T-cell autonomous 2) To test the hypothesis that FOXO transcription factors regulate differentiation of memory CD8 T cells. Aims 1 and 2 will utilize complementary approaches including TCR transgenic (tg) mice, bone marrow chimeras, mice that are conditionally deficient for p27Kip1 or FOXOs in T cells, and inducible p27Kip1/FOXO tg mice to determine the in vivo role of p27Kip1 and FOXOs in regulating the dynamics of phenotypic and functional modifications that occur in CD8 T cells during differentiation from effectors to competent memory cells. These studies would provide critical insight into the mechanisms governing memory cell differentiation, and also have the potential to identify p27Kip1 and FOXO as molecular targets for development of novel immunotherapies to accelerate and enhance the induction of vaccine-induced CD8 T cell-dependent protective immunity against viral infections. PUBLIC HEALTH RELEVENCE: CD8 T cells play a critical role in defense against several viral infections of humans. In this proposal, we seek to identify key molecular regulators (p27Kip1 and FOXO) that can be modulated in CD8 T cells by immunotherapies to enhance vaccine-induced protective anti-viral immunity. These studies should aid in the development of effective vaccines to protect humans against virally induced diseases.