Genetic Analysis of Circadian Oscillator Hierarchy

描述（由申请人提供）：昼夜节律存在于整个动物王国的物种中，昼夜节律定时障碍导致基于昼夜节律的睡眠障碍、轮班工人和时差期间的适应不良，并可能导致神经精神障碍，包括抑郁症和季节性情感障碍。转录-翻译反馈环是生物钟机制的中心。在时钟基因中具有显性负突变的小鼠或在Bmal 1基因中具有无效突变的小鼠的表型表明，这些转录因子对于生物钟功能是必不可少的。在这个提议中，我们将产生有条件破坏时钟基因的小鼠，其中时钟基因可以以组织特异性方式被破坏。这项工作将使用分子，生物化学和行为方法来定义时钟基因在核心转录机制中的贡献。此外，组织特异性基因靶向策略将首次使我们能够解决有关振荡器在外周组织中的功能重要性的重要问题。我们的假设是，时钟是一个不可替代的转录激活因子，分子和生理节律性所必需的。我们预计，大脑中的行为和分子节奏将在大脑特异性破坏时钟基因后被破坏。在肝脏特异性破坏Clock基因的小鼠中，我们预计基于大脑的节律性将持续存在，但肝脏中的局部节律性将被破坏。这些小鼠将用于评估肝脏固有的节律性负责药物和毒素的节律性代谢的假设。拟议的研究将验证一种破坏特定组织昼夜节律功能的新方法，使我们能够确定外周振荡器的生理意义。这些研究将作为一个“原则的证明”建立方法的遗传解剖的昼夜节律振荡器的层次结构和评估振荡器的功能，在特定的组织。

DESCRIPTION (provided by applicant): Circadian rhythms are present in species throughout the animal kingdom, and disorders of circadian timing contribute to circadian-based sleep disorders, maladjustment of shift workers and during jet lag, and may contribute to neuropsychiatric disorders including depression and seasonal affective disorder. A transcriptional-translational feedback loop is at the center of the circadian clock mechanism. The phenotype of mice with a dominant-negative mutation in the Clock gene, or of mice with a null mutation in the Bmal1 gene, suggest that these transcription factors are essential for circadian clock function. In this proposal, we will generate mice with conditional disruption of the Clock gene, in which the Clock gene can be disrupted in a tissue specific manner. The work will use molecular, biochemical and behavioral approaches to define the contribution of the Clock gene in the core transcriptional machinery. Furthermore, the tissue-specific gene targeting strategy will for the first time allow us to address important issues regarding the functional importance of oscillators in peripheral tissues. Our hypothesis is that CLOCK is an irreplaceable transcriptional activator, necessary for molecular and physiological rhythmicity. We expect that behavioral and molecular rhythms in brain will be disrupted following a brain-specific disruption of the Clock gene. In mice with a liver-specific disruption of the Clock gene, we expect that brain-based rhythmicity will persist, but local rhythmicity in the liver will be disrupted. These mice will be used to assess the hypothesis that rhythmicity intrinsic to the liver is responsible for rhythmic metabolism of drugs and toxins. The proposed studies will validate a novel method for disruption of circadian function in specific tissues, allowing us to determine the physiological significance of peripheral oscillators. These studies will serve as a "proof of principle" establishing methods for genetic dissection of the circadian oscillator hierarchy and assessment of oscillator function in specific tissues.