Molecular analysis of circadian regulation in Arabidopsis

描述（由申请人提供）：昼夜节律为生物体提供了适应性优势，通过确保在一天中最合适的时间发生不同的生理过程来增强个体的健康和健身。这项提案的长期目标是了解真核细胞昼夜节律的分子基础。我们已经在以前的研究中确定了生物钟的关键调节因子，现在将使用基因组，生物化学，遗传和数学建模方法来适当地将这些蛋白质放置在昼夜节律系统中。我们将在拟南芥中进行我们的研究，拟南芥是一种模式植物，由于其紧凑的基因组，广泛的遗传和基因组资源，以及耐受对其他复杂真核生物致命的染色质调控途径突变的能力，因此非常适合这些实验。我们将首先使用基因组，生物化学和遗传学的方法来表征在真核生物中保守的蛋白质（但未知的生化功能）在染色质调节中的作用。我们预计这项工作将产生的机制，在不同的真核生物的染色质组织，从而基因表达的见解。接下来，我们将使用遗传和生物化学技术来研究一个相关转录因子家族在昼夜节律运作中的作用。 振荡器在这个过程中，我们将测试由数学模型做出的预测，评估这个模型如何描述驱动时钟的监管关系。最后，我们将修改现有的竞争性染色质免疫沉淀协议，以研究如何结合动态的拮抗性转录因子染色质形状的动态调节基因表达在体内。这些实验将揭示基因表达的时间调控的一般原则，因此将适用于许多生物体和过程。我们对控制不同生物体生物钟功能的共享机制的探索将增加我们对生物钟如何在人类中发挥作用的理解，并揭示它们如何促进人类健康和福利。

DESCRIPTION (provided by applicant): Circadian rhythms provide organisms with an adaptive advantage, enhancing the health and fitness of individuals by ensuring that diverse physiological processes occur at the most appropriate times of day. The long-term objective of this proposal is to understand the molecular basis of circadian rhythms in eukaryotic cells. We have identified key regulators of the circadian clock in our previous studies, and will now use genomic, biochemical, genetic, and mathematical modeling approaches to appropriately place these proteins in the circadian system. We will conduct our studies in Arabidopsis thaliana, a model plant that is uniquely well-suited for these experiments due to its compact genome, extensive genetic and genomic resources, and ability to tolerate mutations in chromatin regulatory pathways that are lethal to other complex eukaryotes. We will first use genomic, biochemical, and genetic approaches to characterize the role of a protein conserved across eukaryotes (but of unknown biochemical function) in the regulation of chromatin. We anticipate this work will generate insights into mechanisms governing chromatin organization and thus gene expression in diverse eukaryotes. Next, we will use genetic and biochemical techniques to investigate the roles of a family of related transcription factors in the workings of the circadian oscillator. In the process, we will test predictions made by a mathematical model, evaluating how well this model describes the regulatory relationships that drive the clock. Finally, we will modify an existing competitive chromatin immunoprecipitation protocol to investigate how the binding dynamics of antagonistic transcription factors to chromatin shape the dynamic regulation of gene expression in vivo. These experiments will reveal general principles governing the temporal regulation of gene expression and thus will be applicable to many organisms and processes. Our exploration of shared mechanisms that control circadian clock function in diverse organisms will increase our understanding of how clocks function in humans and shed light on how they promote human health and welfare.