The Linchpin that Joins the Circadian Oscillator to Clock Output

描述(由申请人提供)：生物体利用可预测的环境光/暗周期，系统地随着昼夜同步地改变它们的新陈代谢、生理和行为。这些由分子时钟产生的昼夜节律，如果被破坏，可能会对健康和疾病产生深远的后果。然而，这些生物钟的机制在任何生物体中都只被部分了解。由于对这些机制的严格理解对于解决与昼夜节律相关的疾病是必不可少的，理旺研究小组的长期目标是阐明时钟和时钟控制模型系统中细胞过程的机制。文献中的报道和本实验室的数据强烈表明，作为计时机制的一部分，时钟蛋白重新安排了它们的全球构象，从而改变了它们的功能特性。例如，步速设定蛋白，在动物中的PER和在真菌中的FRQ，随着时间的推移，在全局紧凑和开放状态之间经历构象变化。同样，我们最近发现蓝藻时钟蛋白Kaib也执行全球构象重排，Kaib？Kaib*，称为“折叠切换”。令人惊讶的是，我们还发现，Kaib折叠切换不仅在昼夜节律的产生中起着至关重要的作用，而且还调节着这些节律的下游传递。因此，这里的目标是阐明蛋白质大规模构象变化在时钟机制中的作用。这一目标的实现预计将对原核生物和真核生物的时间生物学领域产生巨大影响。这一提议的中心假设是凯布？KAIB*折叠开关是连接振荡器功能和时钟输出的关键。为了检验中心假说，我们将追求三个具体目标：1)确立凯布的角色？Kaib*折叠开关在振荡器功能中；2)如何确定Kaib？Kaib*折叠开关调节SASA输出途径；以及3)如何决定Kaib？Kaib*折叠开关调节Cika的输出途径。使用综合方法获得的初步数据有力地支持了中心假说：结构？变种人的？体外实验？计算建模？活体实验。一个在每个领域都有专门知识的强大的合作者团队加强了拟议工作的可行性。这一提议是创新的，因为文献中缺乏报道表明，这一提议的核心概念被忽视了：蛋白质结构的巨大变化支撑着生物钟的机制。这项提议意义重大，因为这里的发现有望为真核生物钟以及之前可能没有认识到蛋白质折叠切换的其他过程打开新的、可操作的洞察力。归根结底，这些知识有可能创造出应对昼夜节律相关疾病的策略。

DESCRIPTION (provided by applicant): Organisms exploit predictable environmental light/dark cycles by systematically varying their metabolism, physiology, and behavior in synchrony with day and night. These circadian rhythms, which are produced by molecular clocks, can have profound consequences to health and disease if disrupted. However, the mechanisms of these circadian clocks are only partially understood in any organism. Because a rigorous understanding of these mechanisms will be indispensable for tackling circadian elated diseases, the long-term goal of the LiWang research group is to elucidate the mechanisms of clocks and clock control over cellular processes in model systems. Reports in the literature and data from this laboratory strongly suggest that clock proteins rearrange their global conformations, and thus their functional properties, as part of the timekeeping mechanism. For example, pace-setter proteins, PER in animals and FRQ in fungi, undergo conformational changes between globally compact and open states as they keep time. Similarly, we recently discovered that the cyanobacteria clock protein, KaiB, also executes global conformational rearrangements, KaiB ? KaiB*, called "fold switching". Surprisingly, we also found that KaiB fold switching not only plays an essential role in the generation of circadian rhythms, but regulates the transmission of those rhythms downstream. Thus, the objective here is to elucidate the roles of large-scale conformational changes by proteins in clock mechanisms. Attaining this goal is predicted to have an enormous influence on the field of both prokaryotic and eukaryotic chronobiology. The central hypothesis of this proposal is that KaiB ? KaiB* fold switching is the linchpin that joins oscillator function to clock output. To test the central hypothesis we will pursue three specific aims: 1) Establish the role of KaiB ? KaiB* fold switching in oscillator functions; 2) Determine how KaiB ? KaiB* fold switching regulates the SasA output pathway; and 3) Determine how KaiB ? KaiB* fold switching regulates the CikA output pathway. The central hypothesis is strongly supported by preliminary data obtained by using an integrative approach: structures ? mutants' ? in vitro experiments ? computational modeling ? in vivo experiments. A strong team of collaborators with expertise in each area enhances the feasibility of the work proposed. This proposal is innovative, because a lack of reports in the literature reveals that the central concept of this proposal has been overlooked: Large changes in protein structure underpin the mechanisms of circadian clocks. The proposal is significant, because the findings here are expected to open new and actionable insights into eukaryotic clocks, and to other processes in which protein fold switching may not have been previously recognized. Ultimately, such knowledge has the potential to create strategies with which to tackle circadian-related diseases.