Defining the role of non-clock genes in circadian physiology and pathophysiology

项目摘要 昼夜节律失调是许多疾病的危险因素，如2型糖尿病、心血管疾病、 高血压和癌症。时间疗法的概念越来越受到人们对药物改良的关注。 当在一天中的最佳时间提供药物时，有效并减少药物毒性。潜在的 昼夜节律的分子机制包括核心的互锁正反馈环和负反馈环 时钟分子基因。在我之前的研究中，在NIDDK提供的F32博士后培训资助下，我们 观察到的昼夜节律增强子和转录组重构不依赖于核心的昼夜表达 饮食诱导肥胖(DIO)小鼠的时钟基因。在我们新的成年肝细胞特异性REV-ERB基因敲除小鼠中 模型和限时饲养的小鼠，我们也观察到了类似的核心时钟基因独立转录 改建。这些观察结果为K01应用奠定了基础，并使我们假设 核心时钟基因在昼夜节律增强子活性和基因表达调控中发挥重要作用 各种正常生理学以及代谢性疾病的病理生理学。这项提议的目标是 建立昼夜节律表观基因组重塑与环境挑战的联系，并识别和 描述可应用于时序药理学和时序营养学的非核心时钟昼夜节律调节器 与代谢性疾病相关的战略。为了实现这一目标，我计划在全基因组范围内利用无偏见的 转录和生物信息学方法绘制增强子图谱和识别调控转录 成人肝细胞特异性Rev-ERB非肝细胞昼夜节律重塑的因子(目标1)和 随意喂食和限时喂食小鼠的肝脏也是如此(目标2)。实际上，我们没有偏见的整个基因组- 广谱增强子定位和转录组分析揭示了DIO选择性昼夜节律转录因子， 雌激素相关受体γ(ERRγ)。此外，我们不偏不倚的转录组分析显示 ERRγ在129S1/SvImJ(12 9)小鼠肝脏中的表达明显升高。值得注意的是，129只小鼠 抵抗体重增加和在B6小鼠中产生DIO的饮食中出现代谢功能障碍。作为一名 概念验证研究，我们将确定肝脏ERRγ在菌株特异性反应中的作用 我们的昼夜节律研究到特定品系的背景下(目标3)。 在米奇·拉扎尔博士的指导下，我加强了对转录调控的培训 肝脏新陈代谢。在K01颁奖期间，在Lazar博士的主要指导下和我的 咨询委员会，我将把我关于新陈代谢转录调控的培训扩展到细胞-细胞串扰水平 (单核序列)，并在昼夜节律生理学中非核心时钟基因的特征方面获得经验 和病理生理学。这次培训将使我能够系统地、定量地和功能地研究分子 参与新陈代谢的连接。我在科学和职业发展方面的进步将受到监督 我的顾问委员会从博士后实习生转变为独立调查员。

Project Abstract Circadian misalignment is a risk factor for many diseases, such as type-2 diabetes, cardiovascular disease, hypertension, and cancer. The concept of chronotherapy is attracting more and more attention to improving drug efficacy and diminishing drug toxicity when drugs are provided at the optimized time of the day. The underlying molecular mechanism of circadian rhythm includes the interlocking positive and negative feedback loops of core clock molecular genes. In my previous study, supported by an F32 post-doctoral training grant from NIDDK, we observed circadian enhancer and transcriptome remodeling is independent of the circadian expression of core clock genes in diet-induced obesity (DIO) mice. In our new adult hepatocyte-specific REV-ERB knockout mice model and time-restricted feeding mouse, we also observed similar core clock gene-independent transcriptomic remodeling. These observations set a foundation of this K01 application and lead us to hypothesize that non- core clock genes play an essential role in the regulation of circadian enhancer activity and gene expression in various normal physiology as well as the pathophysiology of metabolic diseases. The goal of this proposal is to establish the connection of circadian epigenomic remodeling and environmental challenge and to identify and characterize non-core clock circadian regulators that can apply to chrono-pharmacological and chrono-nutritive strategies relating to metabolic diseases. To accomplish this goal, I plan to utilize unbiased genome-wide transcriptional and bioinformatics methods to map enhancer landscape and identify regulatory transcription factors (TFs) for circadian remodeling in non-hepatocytes from adult hepatocyte-specific REV-ERB (Aim 1) and also in livers of ad libitum feeding and time-restricted feeding mice (Aim 2). Actually, our unbiased whole genome- wide enhancer mapping and transcriptome analysis revealed a DIO-selective circadian transcription factor, Estrogen Related Receptor Gamma (ERRγ). Moreover, our unbiased transcriptome profiling revealed that the expression of ERRγ is markedly higher in the livers of 129S1/SvImJ (129) mice. Of note, 129 mice are highly resistant to gaining weight and developing metabolic dysfunction on diets that produce DIO in B6 mice. As a proof-of-concept study, we will determine the role of hepatic ERRγ in strain-specific response to DIO and extend our circadian rhythm study to a strain-specific context (Aim 3). Under the mentorship of Dr. Mitch Lazar, I have strengthened my training in the transcriptional regulation of hepatic metabolism. In this K01 award period, with primary mentorship from Dr. Lazar and the guidance of my advisory committee, I will extend my training on transcription regulation of metabolism to cell-cell crosstalk level (single-nuclei seq) and gain experiences about characterization of non-core clock genes in circadian physiology and pathophysiology. This training will allow me to systemically, quantitatively and functionally study molecular connections involved in metabolism. My progress in science and career development will be under supervision of my advisory committee for the transition from a postdoctoral trainee to an independent investigator.