Dissecting and engineering reversible cell cycle states

7.项目总结/摘要 器官系统中的许多疾病以及衰老的有害影响都可归因于衰老。 有助于器官功能的终末分化细胞的丧失。在这些情况下，没有活跃的干细胞 能够产生新的功能细胞的群体。此外，任何功能性细胞都不能 替代丢失的细胞，因为前者作为其分化过程的一部分已经永久退出细胞周期。一 使终末分化的细胞重新进入细胞周期的方法将显著减轻和 有可能治愈这些疾病。原则上，细胞分化不需要细胞周期退出。 事实上，一些分化的细胞类型，如肝细胞和淋巴细胞，保留了重新进入细胞的能力。 细胞周期这些细胞类型被认为是静止的，而不是终末分化的。一些 细胞外信号驱动静止细胞回到细胞周期以及细胞周期的途径 进入，细胞周期蛋白D-Cdk 4，6途径，已被定义。然而，目前还不清楚是什么使静止细胞 接受这些信号并重新激活细胞周期蛋白D-Cdk 4，6通路，而终末分化细胞不能 帐号吧.因此，终末分化细胞和静止细胞之间的差异是功能性的。 缺乏分子解释的分类。尽管哺乳动物细胞培养模型提供了易于处理的 尽管研究静止期的系统，他们未能概括细胞周期调控的复杂性， 组织中因此，有必要在生物体的背景下研究静止，以提供一个 全面了解这种状态，并促进其转化为再生医学。这个项目 将建立小鼠肝脏作为一个易处理的生理系统，以了解静止的可逆性。 为了赋予终末分化细胞这种能力，并使它们能够在细胞中增殖， 疾病的设置。该方法将考虑两种不相互排斥的平衡细胞周期蛋白D的方法， Cdk 4，6通路的再激活：通路本身的直接调节或外在因子的参与， 这个瀑布。目的1将确定细胞周期蛋白D的活性是否存在功能相关的差异， 与组织中其他细胞周期状态相比，静止肝细胞中的cdk 4，6通路组分。目标2将 在肝脏中建立第一个全基因组的功能缺失筛查，以识别该基因以外的任何基因。 这是静止的可逆性所必需的途径。最后，aim 3将定义保守的分子 通过确定目标1和目标2中确定的哪些特征也是 这种状态在淋巴细胞中的可逆性。总之，这些实验将阐明 静止的标志，从而区分组织中的静止和终末分化。 然后将核心因子引入终末分化的细胞中，并评估它们赋予 增殖能力这些发现不仅填补了我们对哺乳动物细胞认识的一个关键空白， 循环，而且还开启了一种治疗目前缺乏有效治疗的多种疾病的新方法。

7. PROJECT SUMMARY/ABSTRACT Numerous diseases across organ systems as well as the deleterious effects of aging are attributable to the loss of terminally differentiated cells that facilitate organ function. In these cases, there is no active stem cell population capable of generating new functional cells. Moreover, any functional cells that remain cannot replace lost cells as the former have permanently exited the cell cycle as part of their differentiation process. A means of making terminally differentiated cells re-enter the cell cycle would dramatically alleviate and potentially cure these disease states. In principle, cellular differentiation does not necessitate cell cycle exit. Indeed some differentiated cell types, such as hepatocytes and lymphocytes, retain the ability to re-enter the cell cycle. These cell types are considered to be quiescent rather than terminally differentiated. Some of the extracellular signals that drive quiescent cells back into the cell cycle as well as the pathway for cell cycle entry, the Cyclin D-Cdk4,6 pathway, have been defined. However, it is unclear what allows a quiescent cell to receive these signals and reactivate the Cyclin D-Cdk4,6 pathway while a terminally differentiated cell cannot do so. As such, the difference between terminally differentiated cells and quiescent cells is a functional classification lacking a molecular explanation. Although mammalian cell culture models have provided tractable systems for investigating quiescence, they have failed to recapitulate the complexity of cell cycle regulation in tissues. There is therefore a need to study quiescence in an organismal context in order to provide a comprehensive understanding of this state and facilitate its translation to regenerative medicine. This project will establish the mouse liver as a tractable physiologic system to understand the reversibility of the quiescent state ultimately in order to confer this ability to terminally differentiated cells and enable their proliferation in the setting of disease. The approach will consider two non-mutually exclusive means of poising the Cyclin D- Cdk4,6 pathway for reactivation: direct modulation of the pathway itself or involvement of a factor extrinsic to this cascade. Aim 1 will determine whether there are functionally relevant differences in the activity of Cyclin D- Cdk4,6 pathway components in quiescent hepatocytes compared to other cell cycle states in tissues. Aim 2 will establish the first genome-wide, loss-of-function screen in the liver to identify any genes outside of this pathway that are required for the reversibility of quiescence. Finally, aim 3 will define conserved molecular features of quiescence by determining which features identified in aims 1 and 2 are also required for the reversibility of this state in lymphocytes. Together, these experiments will elucidate the core molecular signature of quiescence and thus the distinction between quiescence and terminal differentiation in tissues. Core factors would then be introduced into terminally differentiated cells and evaluated for their ability to confer proliferative capacity. These findings will not only fill a critical gap in our understanding of the mammalian cell cycle but also unlock a novel means of treating myriad diseases currently lacking effective therapy.