Molecular Mechanisms in the Arabidopsis Circadian Clock

描述（由申请人提供）：生物钟是一种内源性分子振荡器，周期约为24小时，在细菌、真菌、植物和动物物种中几乎无处不在。外部刺激，如光或温度，是确保昼夜节律振荡器与当地环境精确共振的干扰线索。时钟网络结构通过多个互锁的转录反馈回路和广泛的翻译后调节来维持自我维持的节律。强大的昼夜节律网络与环境节律协调生化、生理和行为反应，以优化资源分配和提高健康。我们建议结合遗传学、生物化学、功能基因组学和计算方法来确定构成多层时钟网络的成分和分子机制。我们将通过这项拨款中提出的工作直接了解的领域包括环境输入时钟的基本感官途径，昼夜节律网络动力学和输出的机械控制。通过筛选具有核心时钟启动子的综合转录因子库，我们已经确定了介导对时钟温度输入的感知和转录反应的假定元件。我们将探讨它们在与昼夜节律振荡器相关的温度相关现象（夹带、门控和补偿）中的作用。此外，最近在实验室的工作已经确定了关键转录因子TOC1和LUX的生化特性，这为我们理解它们在生物钟中的作用提供了重要的进展。我们建议通过生化和分子方法进一步表征它们活动的机制基础，并在基因组水平上探索它们的功能，以了解它们在控制时钟输出中的作用。最后，虽然网络的鲁棒性部分建立在冗余的基础上

DESCRIPTION (provided by applicant): The circadian clock is an endogenous molecular oscillator with a period of approximately 24 hours that is present nearly ubiquitously in bacterial fungal, plant, and animal species. External stimuli, such as light or temperature, are entrainment cues that ensure that the circadian oscillator is in precise resonance with the local environment. Self-sustained rhythms are maintained by clock network architecture through multiple, interlocked transcriptional feedback loops and extensive post-translational regulation. The robust circadian network coordinates biochemical, physiological, and behavioral responses with environmental rhythms to optimize resource allocation and increase fitness. We propose to deploy a combination of genetics, biochemistry, functional genomics, and computational approaches to identify the components and molecular mechanisms that underlie the multilayered clock network. Areas that we will gain direct insight into through the work proposed in this grant include the fundamental sensory pathways for environmental input into the clock, circadian network dynamics, and mechanistic control of outputs. By screening a comprehensive transcription factor library with core clock promoters, we have identified putative elements that mediate the perception and transcriptional responses to temperature inputs into the clock. We will explore their roles in temperature- associated phenomenon (entrainment, gating, and compensation) in relation to the circadian oscillator. Also, recent work in the laboratory has identified biochemical properties of key transcription factors, TOC1 and LUX, which provide important advances in our understanding of their roles within the clock. We propose to further characterize the mechanistic underpinnings of their activities through biochemical and molecular approaches, as well as explore their function on a genomic level to understand their role in the control of clock outputs. Finally, while robustness in networks is partially built on redundancy of components, this redundancy hinders our ability to identify new factors and understand their function through genetic perturbations. To circumvent this obstacle, we have developed a new computational approach for identifying functional specificity in multi-gene families by mining microarray data. We propose to expand this approach on all Arabidopsis transcription factor families and validate the approach on preliminary candidates affecting the circadian network. This new tool can be broadly applied to any organism with microarray expression data to identify perturbations that can separate the function of closely related homologs or members of a multi-gene family. Continued efforts such as the work proposed here and the on-going research in our laboratory to elucidate the molecular mechanisms of the plant circadian clock will complement similar analyses in other systems, ultimately translating our understanding of circadian biology to impact the treatment of human circadian disorders.