Phytochromes: Structural Perspectives on Photoactivation and Signaling

光敏色素：光活化和信号传导的结构视角

• 批准号: 10242010
• 负责人: RICHARD DAVID VIERSTRA
• 金额: $ 29.26万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-09-01 至 2022-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10242010
• 关键词: Advanced Development; Agriculture; Arabidopsis; Architecture; Back; Behavior; Bilin; Binding; Biochemical; Biological; Biology; Biotechnology; Collection; Coupled; Cryoelectron Microscopy; Deuterium; Disease; Drops; Ecosystem; Engineering; Environment; Event; Evolution; Family; Goals; Growth and Development function; Health; Human; Hydrogen; Influentials; Kinetics; Knowledge; Length; Life; Life Cycle Stages; Light; Mass Spectrum Analysis; Measures; Medical; Membrane Proteins; Methods; Mission; Modeling; Molecular Conformation; N-terminal; Nature; Organism; Output; Paint; Pathway interactions; Perception; Performance; Photochemistry; Photons; Photoreceptors; Physiological Processes; Phytochrome; Plant Components; Plants; Process; Protein Isoforms; Proteins; Reagent; Recombinants; Research; Resolution; Seeds; Signal Transduction; Source; Structure; Surface; Synchrotrons; Temperature; Temperature Sense; Time; Tissue imaging; Transcription Repressor; United States National Institutes of Health; Variant; Work; X-Ray Crystallography; absorption; base; chromophore; conformer; dimer; driving force; fluorophore; human pathogen; improved; light intensity; member; microbial; microorganism; nano; novel; optogenetics; oxidation; particle; pathogen; photoactivation; phyA phytochrome; plant growth/development; protein-histidine kinase; reaction rate; response; three dimensional structure; three-dimensional modeling; tool; transcription factor; x-ray free-electron laser

PROJECT SUMMARY Most organisms employ an array of photoreceptors to detect their light environment. Arguably the most influential are the phytochromes (Phys), a diverse group essential for plant growth and development, and widely distributed in many bacterial, fungal, and algal genera. By reversible photointerconversion of their bilin chromophores between a red light- absorbing Pr state and a far-red light-absorbing Pfr state, Phys act as photoswitches in various signaling cascades responsive to light intensity, direction, duration, and spectral quality. Moreover, through the thermal reversion of Pfr back to Pr, some Phys sense temperature through enthalpic effects on the rate of this reaction, and possibly perceive time via the nighttime depletion of Pfr. The cumulative effects of this Pr/Pfr interconversion impact numerous physiological processes important to agriculture and the biology of harmful plant and human pathogens. In addition, their unique photochemistries have recently provided invaluable optogenetic tools, including novel fluorophores for tissue imaging, and engineered photoswitches that can regulate cellular events with remarkable temporal and spatial precision. Recently, we and others have made great strides in understanding how Phys signal through studies on the photosensing region. An emerging toggle model posits that a light-triggered isomerization of the bilin yields angstrom-scale rearrangements within the bilin-binding pocket that is ultimately transduced into large-scale conformational changes in the dimeric photoreceptor. While the model helps clarify gross changes required for endstate conversion, the intermediates of photoexcitation and ensuing structural changes necessary for a signaling-competent Pfr state are uncertain. It is also unclear how well the model applies to plant Phys given their distinctive modular architectures. The objective of this proposal is to complete this picture through continued structural and biochemical analyses of representative Phys in their Pr and Pfr states, and in combination with their downstream effectors. Specific aims are to: (1) use x-ray crystallography and cryo-electron microscopy to develop more comprehensive structures of plant and bacterial Phys, including models of full-length dimeric photoreceptors with theirs signal output modules; (2) define how Phys transduce the light signal through association with their downstream partners; (3) exploit serial femtosecond x-ray crystallography to structurally define the intermediates generated after photon absorption; (4) use steady-state and surface mapping methods to better understand the protein surface dynamics during photoconversion; and (5) appreciate how diversity within the plant Phy family is used to enhance thermal perception through the biochemical and structural analyses of the PhyB isoform that employs a predicted intrinsically disorder region at its N-terminus to sense temperature. Taken together, this project will provide an essential framework to better appreciate the structure, allosteric mechanism, and evolution of the Phy superfamily. Its anticipated results should help elucidate how microorganisms and plants sense light, temperature, and possibly time, which could have important ramifications for improving the agricultural performance of crop plants, understanding microbial ecosystems, controlling the life cycle of medically-relevant pathogens, and enhancing the application of Phys as optogenetic reagents.

项目摘要 大多数生物体采用一系列光感受器来检测其光环境。可以说最有影响力的是 植物色素（物理），这是一个对植物生长和发育至关重要的多元化群体，在许多人中分布广泛 细菌，真菌和藻类属。通过可逆的光接转换其在红色光之间的bilin发色团 吸收PR状态和远红色的光吸收PFR状态，物理在各种信号级联 响应光强度，方向，持续时间和光谱质量。此外，通过PFR背部的热归还 PR，通过焓影响该反应速率，并可能通过 PFR的夜间耗尽。该PR/PFR互连的累积效应影响了许多生理 对农业和有害植物和人类病原体的生物学重要的过程。此外，他们的独特 光化学最近提供了无价的光遗传学工具，包括用于组织成像的新型荧光团， 以及可以以显着的时间和空间精度来调节细胞事件的工程照片开关。 最近，我们和其他人在了解如何通过对光感的研究来理解物理信号方面取得了长足的进步 地区。一个新兴的切换模型认为，bilin的光触发异构化产生了Angstrom尺度 Bilin结合口袋内的重排，最终被转换为大规模构象的变化 二聚体光感受器。虽然该模型有助于阐明端州转换所需的总更改，但 光激发的中间体和信号功能能力PFR状态所需的结构性变化是 不确定。鉴于其独特的模块化体系结构，该模型还适用于植物物理，这也尚不清楚该模型如何适用。 该提案的目的是通过持续的结构和生化分析来完成此图片 代表性物理在其PR和PFR状态中，并结合其下游效应子。具体目的是： （1）使用X射线晶体学和冷冻电子显微镜开发植物和 细菌性物理，包括具有其信号输出模块的全长二聚体光感受器的模型； （2）定义如何 物理通过与下游伴侣的关联来传递光信号； （3）利用串行飞秒X射线 晶体学以在结构上定义光子吸收后产生的中间体； （4）使用稳态和表面 映射方法可以更好地理解光转换过程中蛋白质表面动力学； （5）感谢 PHY家族中的多样性用于通过生化和结构来增强热感知 对PHYB同工型的分析，该工型采用了N末端的预测本质上的疾病区域来感知温度。 综上所述，该项目将提供一个基本框架，以更好地欣赏结构，变构机制， 和PHY超家族的演变。它的预期结果应有助于阐明微生物和植物的感觉 光，温度和可能的时间，这可能会产生重要的影响以改善农业 作物植物的性能，了解微生物生态系统，控制与医学相关的生命周期 病原体，并增强物理作为光遗传试剂的应用。