Protein:Protein Interaction Networks in the Circadian Clock

生物钟中的蛋白质：蛋白质相互作用网络

• 批准号: 8772682
• 负责人: Brian David Zoltowski
• 金额: $ 32.05万
• 依托单位: SOUTHERN METHODIST UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-08-01 至 2017-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8772682
• 关键词: Active Sites; Animal Model; Animals; Binding; Biological; Biological Process; Biophysics; C-terminal; Cell physiology; Cellular Assay; Chemicals; Chemistry; Circadian Rhythms; Complex; Couples; Degradation Pathway; Development; Diabetes Mellitus; Dimerization; Disease; Disease Progression; Environmental Risk Factor; Evaluation; F Box Domain; Family; Flavins; Genetic Transcription; Growth and Development function; Heart Diseases; Human; Hydrogen Bonding; Length; Light; Link; Maps; Masks; Measures; Mediating; Metabolic; Metabolism; Molecular; Mouse-ear Cress; Obesity; Organism; Oxidative Stress; Oxygen; Pathway interactions; Pattern; Peripheral; Photons; Photoreceptors; Physiology; Plant Physiology; Plants; Process; Protein Dynamics; Proteins; Reaction; Regulation; Regulatory Element; Resolution; Role; Sensory; Signal Pathway; Signal Transduction; Sleep; Solutions; Solvents; Stimulus; Structure; Surface; Techniques; Tertiary Protein Structure; Variant; Vertebrates; Work; X-Ray Crystallography; absorption; adduct; biological adaptation to stress; biophysical techniques; cell type; chemical kinetics; circadian pacemaker; day length; design; flexibility; in vivo; innovation; insight; photoactivation; protein complex; protein degradation; protein protein interaction; protein structure; public health relevance; quantum; receptor; response; scaffold; sensory system; tool; voltage

DESCRIPTION (provided by applicant): Here we use an innovative, combined chemical and biophysical approach to decipher the molecular mechanism circadian clocks use to integrate complex environmental sensing pathways into regulation of metabolism and development. Synchronization of cellular physiology with diurnal changes in environmental variables is a central aspect of circadian clocks. Notably, desynchronization of master and peripheral clocks due to disruption in sleep cycles or altered metabolic function have been implicated in the onset and progression of diseases ranging from obesity, diabetes and heart disease. Notably, the molecular mechanisms gating reciprocal regulation of metabolism and the core circadian oscillator have been hampered by the complexity and number of entrainment pathways in vertebrates. In contrast, the principal environmental variable regulating circadian function in plants is blue-light, enabling precise spatial and temporal interrogation of protein:protein interaction networks integrating environmental factors into circadian regulation of metabolism and development. Herein, we focus on elucidating the role of flavin- binding photoreceptors in mediating circadian function in the model organism Arabidopsis thaliana. A complete understanding of how flavin chemistry dictates activation of protein degradation pathways in a circadian manner can facilitate analysis of similar environmentally sensitive pathways in higher organisms including humans. To map a reaction trajectory beginning from initial photon absorption to alteration of organism physiology we will focus on three primary factors. 1.) Define the chemical and photochemical activation mechanisms of A. thaliana circadian clock photoreceptors. The Zeitlupe (ZTL), Flavin-Kelch-Fbox-1 (FKF1) and LOV-Kelch-Protein (LKP2) family of photoreceptors couples activation of a flavin-binding domain to alteration in protein stability though regulation of light-activated F-Box domains. Using a combination of spectroscopic techniques we will unravel chemical mechanisms regulating ZTL/FKF1/LKP2 function. 2.) Elucidate structural dynamics regulating environmental sensing. Using a combination of NMR and X-ray crystallography, we will decipher atomic resolution detail of how alteration in flavin chemistry dictates alterations in protein structure to selectively excite multple signaling pathways. 3.) Regulation of protein:protein networks. Structural characterization of the photoactivation process will guide design of protein variants to selectively disrupt formation of signaling complexes. Mapping of interaction surfaces will facilitate interrogation of cellular signaling.

描述（由申请人提供）：在这里，我们使用创新的化学和生物物理相结合的方法来破译生物钟用于将复杂的环境传感途径整合到代谢和发育调节中的分子机制。细胞生理学与环境变量的昼夜变化的同步是生物钟的核心方面。值得注意的是，由于睡眠周期中断或代谢功能改变而导致主时钟和外周时钟不同步，与肥胖、糖尿病和心脏病等疾病的发生和进展有关。值得注意的是，脊椎动物中夹带途径的复杂性和数量阻碍了代谢和核心昼夜节律振荡器相互调节的分子机制。相比之下，调节植物昼夜节律功能的主要环境变量是蓝光，能够对蛋白质：蛋白质相互作用网络进行精确的空间和时间询问，将环境因素整合到代谢和发育的昼夜节律调节中。在此，我们重点阐明黄素结合光感受器在介导模式生物拟南芥昼夜节律功能中的作用。全面了解黄素化学如何以昼夜节律方式控制蛋白质降解途径的激活，可以促进对包括人类在内的高等生物体中类似的环境敏感途径的分析。为了绘制从最初的光子吸收到生物体生理学改变的反应轨迹，我们将重点关注三个主要因素。 1.) 定义拟南芥生物钟光感受器的化学和光化学激活机制。 Zeitlupe (ZTL)、Flavin-Kelch-Fbox-1 (FKF1) 和 LOV-Kelch-Protein (LKP2) 光感受器家族通过光激活 F-Box 结构域的调节，将黄素结合结构域的激活与蛋白质稳定性的改变结合起来。通过结合光谱技术，我们将揭示调节 ZTL/FKF1/LKP2 功能的化学机制。 2.) 阐明调节环境传感的结构动力学。结合使用 NMR 和 X 射线晶体学，我们将破译原子分辨率细节，了解黄素化学的变化如何决定蛋白质结构的变化，从而选择性地激发多种信号通路。 3.) 蛋白质的调节：蛋白质网络。结构表征 光激活过程将指导蛋白质变体的设计，以选择性地破坏信号复合物的形成。相互作用表面的映射将有助于对细胞信号传导的询问。