Biology of nuclear receptor rev-erb alpha in circadian rhythm and metabolism

描述(由申请人提供)： 肥胖症和糖尿病在美国已经达到流行水平。糖尿病及其相关并发症正在成为该国发病率的主要原因之一。尽管一些假说将肥胖和胰岛素抵抗联系在一起，但昼夜节律在糖尿病发病机制中的作用还没有得到很好的解决。有趣的是，空腹血糖、胰岛素敏感性和糖耐量存在明显的昼夜变化，在肥胖和糖尿病的情况下，这种变化会严重减弱。 孤儿核受体rev-erb(作为一种核心的昼夜节律基因出现。通过抑制另一个时钟基因Bma1的基因表达，Rev-erb(作为昼夜节律的负调节因子)发挥作用。我们证明了依赖于GSK3的磷酸化是维持REV-ERB蛋白稳定性所必需的(并且在体外同步Bma1基因的昼夜振荡是至关重要的。为了扩大我们对GSK3(依赖于Rev-erb的调控)的认识，我们在这一应用的第一阶段(K99阶段)提出了两个特定的目标，包括：目的1.阐明GSK3(介导Rev-erb(在肝细胞中的降解)上游的信号通路；以及目的2.确定Rev-erb(被GSK3(在昼夜节律和代谢中)磷酸化)在体内的作用。为了解决这些问题，将创建一个表达REV-ERB(突变体)的小鼠模型，该突变体模仿GSK3(肝脏中的磷酸化)。这将是第一次分析确定REV-ERB(作为时钟蛋白在肝脏昼夜节律和葡萄糖代谢中的作用)。这项研究将在宾夕法尼亚大学的宾夕法尼亚糖尿病中心进行，由米切尔·拉扎尔博士监督。该中心在糖尿病和肥胖症研究方面拥有专业知识，是开展拟议项目的优秀环境。同时，该项目将为首席研究员雷茵博士提供卓越的培训，以发展糖尿病和肥胖症领域的学术生涯。 泛素-蛋白酶体途径(UPP)在新陈代谢和能量平衡过程中的作用尚未得到很好的研究。我们最近发现，基于cullin 4A的E3连接酶调节着rev-erb的蛋白质稳定性(以及对糖异生起重要作用的基因表达)，这表明cullin 4A是一种在昼夜节律和新陈代谢中发挥作用的新的调节剂。因此，在R00阶段提出的具体目标3将集中在解开cullin 4A E3泛素连接酶在调节昼夜节律、胰岛素信号和葡萄糖代谢中的作用。我们期望这项工作将为泛素-蛋白酶体系统在新陈代谢调节中的功能重要性提供一组证据。此外，这项工作可能会为确定治疗胰岛素抵抗和糖尿病的新治疗靶点提供新的曙光。

DESCRIPTION (provided by applicant): Obesity and diabetes have reached epidemic levels in the United States. Diabetes and its associated complications are becoming one of the leading causes of morbidity in this country. Although several hypotheses link obesity and insulin resistance, the role of circadian rhythm in the pathogenesis of diabetes has not been well addressed. Interestingly, there is a clear circadian variation in fasting glucose, insulin sensitivity and glucose tolerance, which is severely dampened in the condition of obesity and diabetes. The orphan nuclear receptor Rev-erb( emerged as a core circadian gene. By repressing gene expression of another clock gene, Bma1, Rev-erb( functions as a negative regulator of circadian rhythm. We showed GSK3(-dependent phosphorylation is required for maintaining the protein stability of Rev-erb( and is crucial for synchronizing the circadian oscillation of Bma1 gene in vitro. To extend our knowledge on GSK3(-dependent regulation of Rev-erb(, we propose two specific aims in the first phase of this application (K99 phase), including: Aim 1. Elucidate the signal pathways upstream of GSK3( mediating Rev-erb( degradation in hepatocytes; and Aim 2. Determine in vivo roles of Rev-erb( phosphorylation by GSK3( in circadian rhythm and metabolism. A mouse model expressing Rev-erb( mutant which mimics GSK3( phosphorylation in liver will be created to address these questions. This will be the first analysis to determine the role of Rev-erb( as a clock protein in liver circadian rhythm and glucose metabolism. This study will be carried out in the Penn Diabetes Center at University of Pennsylvania under the supervision of Dr. Mitchell Lazar. The center has expertise in diabetes and obesity research, and is an outstanding environment in which to conduct the proposed project. Meanwhile, the project will provide superb training for the principle investigator, Dr. Lei Yin, to develop an academic career in the field of diabetes and obesity. The role of the ubiquitin-proteasome pathway (UPP) has not been well-studied in the process of metabolism and energy homeostasis. We recently discovered that Cullin 4A-based E3 ligase regulates protein stability of Rev-erb( as well as gene expression important for gluconeogenesis, suggesting Cullin 4A is a novel modulator functioning in both circadian rhythm and metabolism. Therefore, the specific aim 3 proposed during the R00 phase will be focused on unraveling roles of the Cullin 4A E3 ubiquitin ligase in regulating circadian rhythm, insulin signaling and glucose metabolism. We expect this work will provide a panel of evidences for the functional importance of the ubiquitin-proteasome system in the regulation of metabolism. In addition, this work may shed the new light on identifying new therapeutic targets for treating insulin resistance and diabetes.