Coupling between circadian rhythms and redox signaling in stem cell differentiation and adult neurogenesis

干细胞分化和成体神经发生中昼夜节律与氧化还原信号之间的耦合

基本信息

批准号：
10299608
负责人：
Daniel Maxim Iascone
金额：
$ 7.22万
依托单位：
UNIVERSITY OF PENNSYLVANIA
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-12-01 至 2023-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10299608
关键词：
ARNTL gene Acute Adult Alzheimer&apos s Disease Behavioral Biological Models Biology Bromodeoxyuridine CRISPR/Cas technology Cell Differentiation process Cell division Cells Circadian Dysregulation Circadian Rhythms Circadian gene expression Coupling Cre driver Development Disease Down-Regulation Embryonic Development Exhibits Fibroblasts Gene Expression Genes Genetic Genetic Recombination Genetic Transcription Glutamates Glycolysis Goals Health Hippocampus (Brain)Histology Homeostasis Hour Human Hydrogen Peroxide Image Knock-out Knockout Mice Label Light Link Mammals Maps Measures Mediating Metabolic Metabolism Modeling Molecular Mus NADP Neurodegenerative Disorders Neuronal Differentiation Neurons Oxidation-Reduction Oxidative Phosphorylation Oxidative Stress Parkinson Disease Pathway interactions Pentosephosphate Pathway Phase Physiological Processes Post-Translational Protein Processing Process Production Reactive Oxygen Species Regulation Reporter Research Research Training Role Signal Transduction Stains Stem cell pluripotency Synapses Tamoxifen Testing Thymidine Time Training Transgenes Transgenic Organisms Venus adult neurogenesis adult stem cell age related analog base biological adaptation to stress cell type circadian circadian pacemaker cofactor directed differentiation embryonic stem cell excitatory neuron genome editing granule cell human embryonic stem cell in vitro Model in vivo in vivo Model induced pluripotent stem cell nerve stem cell neurogenesis neuronal circuitry novel pluripotency self-renewal sensor stem cell differentiation stem cell division stem cell homeostasis stem cell model stem cells tool transcription factor

项目摘要

Project Summary Circadian rhythms are necessary to coordinate the timing of key behavioral and physiological processes in mammals [1-3]. However, while our understanding of the function of circadian clock genes in embryonic development is rapidly advancing [4-6], the molecular mechanisms through which these rhythms emerge during stem cell differentiation remain elusive [7]. Recently, signaling by reactive oxygen species (redox signaling) has emerged as an essential link between cellular metabolism and circadian rhythms in adult function [8, 9]. Signaling from the pentose phosphate pathway through production of redox cofactor NADPH is an important regulator of transcriptional oscillations, influencing the expression of core circadian clock genes through the redox-sensitive transcription factor NRF2 [10]. NRF2 is a crucial regulator of embryonic stem cell pluripotency and self-renewal, but whether redox signaling contributes to the development of circadian rhythms in differentiating stem cells remains completely unexplored [11]. Using fluorescent reporters of the hydrogen peroxide and Per2 expression, we propose to simultaneously visualize reactive oxygen species and circadian rhythms in single cells for the first time. By combining this novel model system with CRISPR/Cas9-mediated genome editing approaches, we will causally test the role of redox signaling in the development of circadian rhythms in human induced pluripotent stem cells undergoing directed differentiation to glutamatergic neurons. Using adult hippocampal neurogenesis as an in vivo model system for neuronal differentiation, we will further explore the function of redox-circadian coupling in coordinating the sequential development and circuit integration of adult-born granule cells. The long-term objective of this proposal is to create a novel model system to explore the mechanisms through which reciprocal interaction between redox and circadian transcription factor networks direct the proper sequential timing of development. While the current proposal seeks to investigate how redox-circadian coupling drives the differentiation of pluripotent and adult stem cells, we aim to describe a general paradigm for the coordination of metabolism, cell division, and stem cell homeostasis in health and disease. Hypothesis: We hypothesize that redox signaling drives the development of circadian rhythms in stem cells following the loss of pluripotency, and that reciprocal regulation between redox signaling and circadian rhythms drives the cellular maturation. We predict that disrupting redox-circadian coupling in adult neural stem cells through acute Nrf2 knockout will induce cell division and differentiation, but hinder the development of circadian rhythms and normal maturation of adult-born granule cells. We will test this hypothesis in the following aims: Aim 1: Causally link redox signaling to circadian rhythm development in human induced pluripotent stem cells Aim 2: Determine impact of Nrf2 KO-mediated disruption of redox-circadian coupling on adult neurogenesis

项目概要昼夜节律对于协调关键行为和生理过程的时间是必要的。哺乳动物 [1-3]。然而，虽然我们对胚胎中生物钟基因的功能的了解发育正在迅速推进[4-6]，这些节律出现的分子机制干细胞分化过程中的变化仍然难以捉摸[7]。最近，活性氧（氧化还原信号）已成为成人细胞代谢和昼夜节律之间的重要联系函数 [8, 9]。戊糖磷酸途径通过产生氧化还原辅因子 NADPH 发出的信号是转录振荡的重要调节因子，影响核心生物钟基因的表达通过氧化还原敏感转录因子 NRF2 [10]。 NRF2是胚胎干细胞的重要调节因子多能性和自我更新，但氧化还原信号是否有助于昼夜节律的发展分化干细胞的作用尚未完全被探索[11]。使用过氧化氢和 Per2 表达的荧光报告基因，我们建议同时首次将单细胞中的活性氧和昼夜节律可视化。通过结合这个采用 CRISPR/Cas9 介导的基因组编辑方法的新型模型系统，我们将因果测试其作用氧化还原信号在人类诱导多能干细胞昼夜节律发育中的作用定向分化为谷氨酸能神经元。使用成人海马神经发生作为体内模型神经元分化系统中，我们将进一步探讨氧化还原-昼夜节律耦合在神经元分化中的功能协调成年颗粒细胞的顺序发育和电路整合。该提案的长期目标是创建一个新颖的模型系统，通过以下方式探索机制：氧化还原和昼夜节律转录因子网络之间的相互作用指导正确的发展的顺序时间。虽然当前的提案旨在研究氧化还原昼夜节律如何耦合驱动多能干细胞和成体干细胞的分化，我们的目标是描述一个通用范例健康和疾病中新陈代谢、细胞分裂和干细胞稳态的协调。假设：我们假设氧化还原信号驱动干细胞昼夜节律的发展多能性丧失后，以及氧化还原信号和昼夜节律之间的相互调节促进细胞成熟。我们预测，破坏成体神经干细胞中的氧化还原-昼夜节律耦合通过急性Nrf2敲除会诱导细胞分裂和分化，但会阻碍细胞的发育昼夜节律和成年颗粒细胞的正常成熟。我们将在以下目标：目标 1：将氧化还原信号传导与人类诱导多能干细胞的昼夜节律发育联系起来目标 2：确定 Nrf2 KO 介导的氧化还原-昼夜节律耦合破坏对成人神经发生的影响