Analysis of Mammalian Circadian Mechanism with Cell-permeant Clock Proteins

描述(申请人提供)：人类和大多数其他生物表现出昼夜(每日)节律，由调节睡眠时间、心血管功能和新陈代谢的内源性生化振荡器控制。这些“生物钟”对人类生理学很重要。例如，精神病学和医学研究表明，昼夜节律与某些形式的抑郁症、时差、药物耐受性/疗效、记忆力、失眠和其他睡眠障碍有关。因此，了解生物钟的生化机制可能有助于诊断和治疗与睡眠、心理健康和药理学相关的疾病。生物钟的显著特性--24小时时间恒定、高精度、温度补偿--目前还无法用生物化学来解释。目前的昼夜节律转录和翻译反馈模型认为，节律时钟蛋白的丰度对时钟功能至关重要。在模型系统中，通过在特定阶段诱导时钟蛋白质合成来重置昼夜节律的阶段是可能的。这种方法在哺乳动物系统中很难实现。目前的项目将测试有关哺乳动物节律时钟蛋白丰度意义的假设，方法是使用新的方法，通过多肽介导的跨细胞膜转导将蛋白质直接引入细胞。这项技术将使我们能够调节细胞、组织切片和完整动物中时钟蛋白的细胞内浓度。该项目适合美国国立卫生研究院探索/发展助学金(R21)计划，因为它满足以下所有主要标准：(1)创新研究方向，(2)探索对实质性领域新的方法，以及(3)开发新技术和方法。 与公共健康相关：对目前的昼夜节律振荡器模型进行严格测试，对于我们理解这些控制着人类生理学如此之多的生物钟至关重要。这些研究将产生具有理论重要性的结果，但最近的证据也表明，当正常的生物钟功能被打乱时，睡眠障碍、肥胖、心血管疾病和癌症。因此，这些研究也有可能设计治疗睡眠障碍、时差、失眠、心血管疾病和其他与时钟相关的疾病的方法。

DESCRIPTION (provided by applicant): Humans and most other organisms manifest circadian (daily) rhythms that are controlled by an endogenous biochemical oscillator that regulates the timing of sleep, cardiovascular functions, and metabolism. These "biological clocks" are important to human physiology. For example, psychiatric and medical studies have shown that circadian rhythmicity is involved in some forms of depressive illness, "jet lag," drug tolerance/efficacy, memory, insomnia, and other sleep disorders. Therefore, understanding the biochemical mechanism of circadian clocks may lead to procedures which will be useful in the diagnosis and treatment of disorders that are relevant to sleep, mental health, and pharmacology. The salient properties of circadian clocks-24 hour time constant, high precision, temperature compensation-are presently impossible to explain biochemically. The current Transcription and Translation Feedback Model for circadian rhythms posits that rhythmic clock protein abundance is critical for clock function. In model systems, it has been possible to reset the phase of circadian rhythms by induction of clock protein synthesis at specific phases. This approach has been difficult to accomplish in mammalian systems. The current project will test hypotheses concerning the significance of rhythmic clock protein abundance in mammals by using new methods to introduce proteins directly into cells by peptide-mediated transduction across cell membranes. This technology will allow us to modulate the intracellular concentration of clock proteins in cells, tissue slices, and intact animals. This project is appropriate for the NIH Exploratory/Developmental Grant (R21) Program because it fulfills all of the following primary criteria: (1) innovative research directions, (2) exploration of approaches that are new to a substantive area, and (3) development of new technologies and methods. PUBLIC HEALTH RELEVANCE: Rigorous testing of the current model for circadian oscillators is crucial for our understanding of these biological clocks that control so much of human physiology. These studies will yield results of theoretical importance, but recent evidence also implicates sleep disorders, obesity, cardiovascular disease, and cancer when normal circadian clock function is disrupted. Therefore, these studies also have the potential for designing treatments for sleep disorders, jet lag, insomnia, cardiovascular disease, and other clock-related disorders.