Circadian regulation of mitochondrial RNA polyadenylation

描述（由申请人提供）：昼夜节律钟是内部计时机制，其在时间上协调行为和生理的许多方面与昼夜周期。这些生物钟控制着成千上万的基因，这些基因驱动着细胞内的许多关键过程，包括许多代谢过程。此外，营养摄入和许多代谢过程也反馈调节中枢时钟机制，时钟的中断导致广泛的代谢功能障碍。生物钟的一个重要作用可能是在整个昼夜节律周期中协调代谢程序，以提高效率并避免无效反应。在这个提议中，我们提供了证据，将生物钟的影响扩展到细胞的另一个关键代谢成分--细胞色素的氧化磷酸化系统。我们提出了新的证据表明，昼夜节律蛋白Nocturnin调节细胞色素c氧化酶（复合物IV）的电子传递链的活性。Nocturnin编码一种去腺苷酶，一种负责从mRNA中去除poly A尾的酶，在这个提议中，我们提出的数据表明，Nocturnin在线粒体中工作（除了在细胞质中的已知作用之外），以转录后控制一个 这对复合物IV的组装和功能至关重要。基于这些数据，我们提出了三个具体的目标，旨在测试的一般假设，即昼夜节律钟调节氧化磷酸化通过节律性夜曲素deadenylation的mRNAs编码的。在第一个具体目标中，我们将研究Nocturnin细胞内定位是如何调节的，并测试替代翻译起始是否调节线粒体靶向信号是否被添加到Nocturnin蛋白。在第二个目标中，我们将研究Nocturnin调节复合物IV活性和线粒体氧化磷酸化的机制。我们将测试Nocturnin是否是动态调节线粒体mRNA poly A尾长的“机器”的一部分，并测试这是否会导致靶mRNA翻译的改变。最后，在第三个目标中，我们将测试这些过程是否 由生物钟直接调节，并将决定在一天的不同时间复合物IV组装的动力学。此外，我们将使用以非节律性方式表达Nocturnin的诱导型小鼠模型来测试Nocturnin的节律性对于氧化磷酸化系统的正常功能是否重要。这些拟议的研究将揭示这种新的和令人兴奋的作用背后的机制，为细胞ATP生成系统的昼夜节律系统控制。每5000人中就有一人具有影响复合物IV组装和活性的突变，导致严重疾病，包括脑肌病、神经退行性Leigh综合征、肥厚性心肌病和致命性乳酸酸中毒。因此，了解这个系统是如何调制的至关重要。

DESCRIPTION (provided by applicant): Circadian clocks are internal timekeeping mechanisms that temporally coordinate many aspects of behavior and physiology with the diurnal cycle. These clocks control thousands of genes that drive many critical processes within the cell, including many metabolic processes. Furthermore, nutrient intake and many metabolic processes also feedback to regulate the central clock mechanism and disruption of the clock results in broad metabolic dysfunction. An important role of the clock may be to temporally coordinate metabolic programs throughout the circadian cycle to increase efficiency and avoid futile reactions. In this proposal, we provide evidence that expands the influence of the clock to yet another critical metabolic component of the cell - the oxidative phosphorylation system of the mitochondrion. We present new evidence demonstrating that the circadian protein Nocturnin is regulating the activity of cytochrome c oxidase (Complex IV) of the electron transport chain. Nocturnin encodes a deadenylase, an enzyme responsible for removing poly A tails from mRNAs, and in this proposal we present data that shows that Nocturnin works in the mitochondria (in addition to its known role in the cytoplasm) to post-transcriptionally control one of the mRNAs that is critical for Complex IV assembly and function. Based on these data, we propose three specific aims designed to test the general hypothesis that the circadian clock regulates oxidative phosphorylation through rhythmic Nocturnin deadenylation of mitochondria-encoded mRNAs. In the first specific aim, we will examine how Nocturnin intracellular localization is regulated and test whether alternative translation initiation regulates whether or not a mitochondrial targeting signal is added to the Nocturnin protein. In the second aim, we will examine the mechanism by which Nocturnin regulates Complex IV activity and mitochondrial oxidative phosphorylation. We will test whether Nocturnin is part of a "machine" that dynamically regulates mitochondrial mRNA poly A tail length and test whether this results in altered translation of the target mRNA. Finally, in the third aim, we will test whether these processes are directly regulated by the circadian clock and will determine the kinetics of Complex IV assembly at different times of day. Furthermore, we will test whether Nocturnin' s rhythmicity is important for proper functioning of the oxidative phosphorylation system using an inducible mouse model that expresses Nocturnin in a non-rhythmic manner. These proposed studies will uncover the mechanism behind this new and exciting role, for the circadian system control of the ATP generating system of the cell. One in 5000 humans have mutations that affect Complex IV assembly and activity resulting in severe disease, including encephalomyopathies, neurodegenerative Leigh syndrome, hypertrophic cardiomyopathies, and fatal lactic acidosis. Therefore, understanding how this system is modulated is of critical importance.