Molecular mechanisms of mammalian circadian clock function

摘要 整个身体的生物钟驱动着成千上万个基因的有节奏的表达， 生物化学、生理学和行为的节奏。通过遗传学或生物钟的破坏 环境干扰，如时差或轮班工作，可能会产生深远的负面影响 并与肥胖、糖尿病、癌症、心血管疾病和精神疾病有关。我们 工作主要集中在了解哺乳动物的详细分子机制 生物钟机制以及这些时钟控制节律基因的机制 表情根据目前的模型，这个时钟机制的核心部分是一个负 反馈环，从而转录因子异二聚体CLOCK/BMAL 1驱动转录的 “时钟”蛋白PERIOD（PER）1、PER 2、色氨酸（CRY）1和CRY 2，它们与 彼此抑制CLOCK/BMAL 1的活性，从而抑制它们自身的合成。我们已经解决 CLOCK/BMAL 1和BMAL 2/PER 2复合物的晶体结构，这些数据允许 鉴定了整个蛋白质中进化上保守的功能结构域， 对这些蛋白质运作和设定昼夜节律周期的机制的额外见解。 在接下来的五年里，我们将扩大这一信息，以确定原子的细节， 这个钟走得准。这些核心生物钟转录因子在驱动节律性变化中的作用， 转录是有据可查的，但最近的数据表明转录后控制， 尽管了解得少得多，但对正常节律蛋白表达谱也是至关重要的。 一种类型的转录后控制是mRNA poly（A）尾长度的调节，其影响mRNA的转录。 mRNA稳定性和翻译调节。我们已经鉴定了数百种小鼠肝脏mRNA 在poly（A）尾的长度上表现出强大的昼夜节律。在许多情况下， 节律性尾长是节律性胞质多腺苷酸化和去腺苷酸化的结果 节律和细胞质多聚腺苷酸化和去腺苷酸化机制的许多组成部分， 在昼夜节律的控制下。此外，节律性poly（A）尾与 有节奏的蛋白质表达。因此，生物钟调节动态 许多mRNA的多聚腺苷酸化状态可以驱动蛋白质的节律性表达， 消息的稳态水平。Nocturnin是一种去除poly（A）尾的强节奏蛋白 从mRNA我们已经证明，小鼠中该基因的缺失会导致对饮食诱导的 肥胖和改变的节奏，胆固醇和甘油三酯代谢，暗示它是一个重要的 昼夜节律转录后介质。在未来五年，我们将重点确定 细胞质和线粒体中Nocturnin的mRNA底物。

Abstract Circadian clocks throughout the body drive rhythmic expression of thousands of genes, resulting in rhythms in biochemistry, physiology and behavior. Disruption of circadian clocks through genetics or environmental perturbations such as jet lag or shift-work, can have profound negative consequences and has been linked to obesity, diabetes, cancer, cardiovascular disease and mental illness. Our work is focused generally on understanding the detailed molecular mechanisms of the mammalian circadian clock machinery and the mechanisms by which these clocks control rhythmic gene expression. According to the current model, the core part of this clock mechanism is a negative feedback loop whereby the transcription factor heterodimer CLOCK/BMAL1 drives transcription of the “clock” proteins PERIOD (PER) 1, PER 2, CRYPTOCHROME (CRY) 1 and CRY 2 which interact with each other to repress the activity of CLOCK/BMAL1, and thus their own synthesis. We have solved crystal structures for the CLOCK/BMAL1 and CRY2/PER2 complexes and these data have allowed the identification of evolutionarily conserved functional domains throughout the proteins and revealed additional insights into the mechanisms by which these proteins operate and set the circadian period. Over the next five years, we will expand on this information to determine the atomic details of how this clock keeps time. The roles of these core circadian clock transcription factors in driving rhythmic transcription is well-documented, but recent data have demonstrated that post-transcriptional control, although much less well understood, is also critical for normal rhythmic protein expression profiles. One type of post-transcriptional control is regulation of mRNA poly(A) tail length, which impacts the stability and translational regulation of mRNA. We have identified hundreds of mouse liver mRNAs that exhibit robust circadian rhythms in the length of their poly(A) tails. In many of these cases, the rhythmic tail lengths are the result of rhythmic cytoplasmic polyadenylation and deadenylation rhythms and many components of the cytoplasmic polyadenylation and deadenylation machinery are themselves under circadian control. Furthermore, the rhythmic poly(A) tails are closely correlated with the rhythmic protein expression. Therefore, the circadian clock regulates dynamic polyadenylation status of many mRNAs that can drive rhythmic protein expression independent of the steady-state levels of the message. Nocturnin is a robustly rhythmic protein that removes poly(A) tails from mRNAs. We have shown that loss of this gene in mice causes resistance to diet-induced obesity and altered rhythms in cholesterol and triglyceride metabolism, implicating it as an important circadian post-transcriptional mediator. Over the next five years, we will focus on identifying the mRNA substrates of Nocturnin both in the cytosol and in the mitochondria.