Determining the role of REV-ERBs in the SCN and liver circadian hierarchy

确定 REV-ERB 在 SCN 和肝脏昼夜节律层次中的作用

• 批准号: 10314798
• 负责人: Lauren Nicole Woodie
• 金额: $ 6.6万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-01-01 至 2022-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10314798
• 关键词: Determining; role; REV; ERBs; SCN

PROJECT SUMMARY/ABSTRACT The maintenance and synchrony of circadian rhythms throughout the body is essential for metabolic health and homeostasis. In mammals, the hypothalamic suprachiasmatic nucleus (SCN) acts as the master circadian regulator, but organs such as the liver possess cell-autonomous clocks that regulate circadian gene expression in a tissue-specific manner. The REV-ERBα and β nuclear receptors are important components of the mammalian molecular clock that synchronize metabolic and circadian rhythms. Published data from the Lazar lab using a REV-ERBα/β hepatocyte double knock-out (HepDKO) mouse suggests that there are distinct sets of genes in the liver with varying degrees of cell-autonomy. Indeed, hundreds of normally oscillating genes lose rhythmicity under HepDKO conditions, but a nearly equal number of circadian oscillating genes retained rhythmicity in HepDKO livers. This implies that these genes have non-cell-autonomous rhythms dictated by signals from the SCN. There are several proposed SCN-derived outputs that can dictate hepatic rhythmicity, but innervation via the common hepatic branch (CHB) of the vagus nerve an understudied option. The vagus nerve is an established mode of communication between the brain and periphery that can integrate circadian rhythms of fed/fasting state. However, the role of the CHB in relaying circadian signals between the SCN and liver has yet to be examined. Therefore, I hypothesize that REV-ERB-mediated signals from the SCN clock are necessary for regulation of genes retained in HepDKO livers and that the CHB acts as an integral communicator of these regulatory signals. Specific Aim 1 will characterize hepatic physiology and rhythmicity in a mouse model lacking REV-ERBs in the SCN and liver by knocking out REV-ERBs in the SCN of HepDKO animals and performing circadian and metabolic phenotyping experiments. I will also collect tissues from these animals every 3 hours for 24 hours to compare molecular and gene regulatory phenotypes resulting from REV-ERBs DKO using next generation sequencing (NGS)-based -omics techniques. Specific Aim 2 will determine the role of the CHB in relaying REV-ERB-mediated circadian signals by severing the CHB through vagotomy (CHBx) in HepDKO mice. I will perform circadian and metabolic phenotyping experiments and collect tissues every 3 hours for 24 hours to compare molecular and gene regulatory phenotypes resulting from CHBx using NGS-based -omics. I predict that the rhythm of gene expression and regulation as well as behavioral and metabolic rhythmicity in HepDKO mice will be significantly disrupted without SCN REV-ERBs and in response to CHBx. The experiments outlined herein not only provide significant advances in the understanding of circadian and metabolic rhythm synchronization but provide substantial training opportunities for me by utilizing the Lazar lab's unique REV-ERB DKO mouse models and their expertise in state-of-the-art NGS tools to probe the role of REV-ERB-meditated rhythmicity.

项目总结/摘要 全身昼夜节律的维持和同步对于代谢健康和 体内平衡在哺乳动物中，下丘脑视交叉上核（SCN）作为主昼夜节律 调节器，但器官，如肝脏拥有细胞自主时钟，调节昼夜基因表达 以组织特异性的方式。REV-ERBα和β核受体是细胞核的重要组成部分。 哺乳动物的分子钟，同步新陈代谢和昼夜节律。来自Lazar的数据 使用REV-ERBα/β肝细胞双敲除（HepDKO）小鼠的实验室表明， 肝脏中的基因具有不同程度的细胞自主性。事实上，数百个正常振荡的基因 HepDKO条件下的节律性，但保留了几乎相同数量的昼夜节律振荡基因 HepDKO肝脏中的节律性。这意味着这些基因具有非细胞自主的节律， 来自SCN的信号。有几种提出的SCN衍生的输出可以指示肝节律性， 但通过迷走神经的肝总分支（CHB）的神经支配是一个未充分研究的选择。迷走 神经是大脑和外周之间建立的通信模式， 进食/禁食状态的节律。然而，CHB在SCN和CNS之间传递昼夜节律信号的作用， 肝脏还有待检查。因此，我假设来自SCN时钟的REV-ERB介导的信号是 对于HepDKO肝脏中保留的基因的调节是必要的，并且CHB作为一个整体 这些监管信号的传播者。具体目标1将表征肝脏生理学， 通过敲除SCN和肝脏中的REV-ERB， 图10示出了HepDKO动物的SCN和进行昼夜节律和代谢表型分析实验。我也会收集 每3小时采集这些动物的组织，持续24小时，以比较分子和基因调节表型 使用基于下一代测序（NGS）的组学技术从REV-ERBs DKO产生。具体 目的2将确定CHB在传递REV-ERB介导的昼夜节律信号中的作用， HepDKO小鼠通过迷走神经切断术（CHBX）的CHB。我会进行昼夜节律和代谢表型分析 每3小时进行实验并收集组织，持续24小时，以比较分子和基因调控 使用基于NGS的组学从CHBx产生的表型。我预测基因表达的节奏和 HepDKO小鼠中的调节以及行为和代谢节律将被显著破坏 无SCN REV-ERB和对CHBX有反应。本文概述的实验不仅提供了显著的 对昼夜节律和代谢节律同步的理解的进展， 通过利用Lazar实验室独特的REV-ERB DKO小鼠模型及其 拥有最先进的NGS工具的专业知识，以探索REV-ERB冥想节律的作用。