Circadian Clock Regulation in Skin

项目摘要/摘要 生物钟，一种古老的、进化保守的计时系统，需要器官的最佳功能 和生物寿命，在包括皮肤在内的周围组织中活跃。外周器官中的时钟是 由视交叉上核的中央时钟协调，但我们也知道限时摄食 影响周围组织的生物钟、基因表达和动态平衡。尽管新的见解是 尤其是从对肝脏等新陈代谢器官的研究中，摄食时间、时钟、 上皮细胞的组织健康状况尚不清楚。特别是，我们不知道限时喂食如何影响 表皮干细胞的再生功能与皮肤老化。在老鼠身上，生物钟协调进程 细胞周期和DNA切除修复与中间代谢，如反映在氧化还原状态 表皮干细胞。耐人寻味的是，白天的限制摄食改变了相位并降低了 皮肤生物钟，它改变新陈代谢相关转录组的表达，而不改变 DNA合成中昼夜振荡的阶段。因此，白天限制进食会扰乱协调。 表皮干细胞的代谢和细胞周期进展之间的关系。而表皮干细胞中的这些循环 已知细胞调节对UVB诱导的DNA损伤的敏感性，它们在表皮动态平衡中的作用 在其他方面，干细胞仍然是未知的。在这里，我们将研究时钟协调振荡的想法 代谢产生的ROS水平与细胞周期和DNA修复机制一起最大限度地提高健康 和表皮干细胞的功能。具体地说，我们假设这种调节将新陈代谢降至最低- 当大多数表皮干细胞正在经历DNA复制、细胞周期最长阶段时产生ROS 对氧化DNA损伤敏感。这一假说预测，白天的摄食会导致昼夜节律 表皮干细胞的错位导致氧化代谢和细胞周期之间的不同步， 导致ROS诱导的DNA突变增加，表皮干细胞功能障碍，以及皮肤老化。为了测试这一点 假设，我们提出了两个目标。首先，我们将定义时间背后的基因调控机制- 限制摄食对表皮干细胞的生物钟和新陈代谢的调节。第二，我们将 确定限时喂养如何调节表皮干细胞的功能和影响年龄比率- 表皮干细胞中的相关DNA突变。这项提议意义重大，因为它测试了一种新的 生物钟如何协调上皮干细胞中间代谢和细胞周期的时间 细胞，以最大限度地减少体细胞DNA突变的积累，以及限制时间喂养如何强制或 打乱了这种协调。这项提议是创新的，因为它追求一种关于饮食的作用的新想法 干预和生物钟在皮肤老化中的作用，它使用最先进的方法，包括双链DNA- 测序、荧光寿命成像和单细胞RNA测序--以前没有应用过的方法 导致皮肤老化。

PROJECT SUMMARY/ABSTRACT The circadian clock, an ancient, evolutionary conserved timing system required for optimal function of organs and organismal lifespan, is active in peripheral tissues, including the skin. Clocks in peripheral organs are coordinated by the central clock in the suprachiasmatic nucleus, but we also know that time-restricted feeding affects circadian clocks, gene expression, and homeostasis in peripheral tissues. Although new insights are emerging, especially from studies in metabolic organs like the liver, the interplay between feeding time, clocks, and tissue health in epithelia is unclear. In particular, we don't know how time-restricted feeding affects the regenerative function of epidermal stem cells and skin aging. In mice, the circadian clock coordinates progression of the cell cycle and DNA excision repair with intermediary metabolism, as reflected in the redox state of epidermal stem cells. Intriguingly, daytime-restricted feeding shifts the phase and decreases the amplitude of the skin circadian clock, and it shifts the expression of the metabolism-related transcriptome without altering the phase of the diurnal oscillations in DNA synthesis. Daytime-restricted feeding, then, disrupts the coordination between metabolism and cell cycle progression in epidermal stem cells. Whereas these cycles in epidermal stem cells are known to modulate the sensitivity to UVB-induced DNA damage, their role in homeostasis of epidermal stem cells remains otherwise unknown. Here, we will investigate the idea that the clock coordinates oscillations of metabolism-generated ROS levels with the cell cycle and the DNA repair machinery to maximize the health and function of epidermal stem cells. Specifically, we hypothesize that this regulation minimizes metabolism- generated ROS when most epidermal stem cells are undergoing DNA replication, the cell cycle stage most sensitive to oxidative DNA damage. This hypothesis predicts that daytime feeding-induced circadian misalignment in epidermal stem cells causes asynchrony between oxidative metabolism and the cell cycle, leading to increased ROS-induced DNA mutations, epidermal stem cell dysfunction, and skin aging. To test this hypothesis, we propose two aims. First, we will define the gene-regulatory mechanisms underlying time- restricted feeding modulation of the circadian clock and metabolism in epidermal stem cells. Second, we will determine how time-restricted feeding modulates epidermal stem cell function and affects the rate of age- associated DNA mutations in epidermal stem cells. The proposal is significant because it tests a new model of how the circadian clock coordinates the timing of intermediary metabolism and the cell cycle in epithelial stem cells to minimize the accumulation of somatic DNA mutations, and how time-restricted feeding can enforce or disrupt this coordination. The proposal is innovative because it pursues a new idea about the role of dietary intervention and the circadian clock in skin aging, and it uses state of the art approaches, including duplex DNA- sequencing, fluorescence lifetime imaging, and single cell RNA-sequencing--approaches not previously applied to skin aging.