Collaborative Research: Investigations of the Plant Photoreceptor Kinase Family of Phototropins

植物光感受器在整个植物发育过程中介导广泛的反应，从种子萌发到营养生长的各个阶段，再到开花和衰老。这些光感受器包括光敏色素、隐色素和光感受器。另外三种蛋白质，FKF1， LKP2和ZTL，参与调节植物节律，可能形成了植物光感受器的第四个家族。趋光素（phot1和phot2）介导植物对蓝光的反应，包括：趋光性、叶绿体重新定位、气孔打开、叶片扩张和对黄化幼苗生长的快速抑制。它们包含两个光感应结构域（LOV1和LOV2），每个都是非共价结合一个FMN发色团和一个激酶结构域。光刺激LOV结构域导致激酶活化和自磷酸化。拟南芥趋光蛋白LOV结构域和更复杂的结构域可以在转基因植物和异源系统中表达。这些结构体具有光活性，但它们的光化学可以被位点特异性突变干扰或选择性地失活。本项目主要研究包括发色团结合结构域和全长蛋白在内的构建体的生化和生物物理特性，以及它们在植物中补充功能的能力。这些研究将探讨光激活的机制，包括最初的光化学和随后的分子内信号传播事件。这项研究将涉及使用时间分辨光学和振动光谱，标准和时间分辨荧光和圆二色光谱，时间分辨偏振光光谱，顺磁和光谱探针的分子结构变化。对趋光蛋白的研究发现了一个发色团模块，LOV结构域作为一个光驱动的分子开关。这些LOV结构域与在多种生物的蛋白质中发现的PAS结构域超家族密切相关。这些研究应该为了解它们的功能提供一个新的层面。研究和教育的结合是这个项目的重要组成部分。这种整合包括对博士后研究员和研究生的培训，以及对本科生的指导，包括那些来自代表性不足群体的学生。将继续向高中学生和教师伸出援手。

Plant photoreceptors mediate a broad spectrum of responses throughout plant development, from seed germination through all phases of vegetative growth to flowering and senescence. These photoreceptors include phytochromes, cryptochromes, and phototropins. Three additional proteins, FKF1, LKP2 and ZTL, involved in regulation of plant rhythms, likely form a fourth family of plant photoreceptors. The Phototropins (phot1 and phot2) mediate several blue light responses in plants including: phototropism, chloroplast relocation, stomatal opening, leaf expansion and the rapid inhibition of growth of etiolated seedlings. They contain two light-sensing domains (LOV1 and LOV2), each non-covalently binding an FMN chromophore, and a kinase domain. Light stimulation of the LOV domains leads to activation the kinase, and autophosphorylation. Arabidopsis phototropin LOV domains and more complex constructs can be expressed in transgenic plants and heterologous systems. These constructs are photoactive, but their photochemistry can be perturbed or selectively inactivated with site-specific mutations. This project focuses on the biochemical and biophysical characterization of constructs including the chromophore-binding domains and the full-length protein, and their ability to complement function in plants. These studies will probe the mechanism of light activation including the initial photochemistry and subsequent intramolecular signal-propagation events. This research will involve the use of time-resolved optical and vibrational spectroscopy, standard and time-resolved fluorescence and circular dichroism spectroscopy, time-resolved polarized light spectroscopy, and paramagnetic and spectroscopic probes of molecular structural changes. Studies on the phototropins have uncovered a chromophore module, the LOV domain that functions as a light-driven molecular switch. These LOV domains are closely related to the superfamily of PAS domains found in proteins in a diverse range of organisms. These studies should provide a new dimension in the understanding of their functions.Integration of research and education is an essential component of this project. This integration involves training of post-doctoral fellows and graduate students and mentoring of undergraduate students, including those from underrepresented groups. Outreach to high school students and teachers will continue.