Arabidopsis phytochrome A signal transduction: characterization and identifcation of cytoplasmic events and components

植物利用光敏色素作为红色/远红色感光器来监测其光环境并相应地调节发育的许多方面。光敏色素A（phyA）是拟南芥中的五种光敏色素之一，调节早期幼苗发育等反应。在其激活后，phyA易位到细胞核，在那里它调节基因表达。两个信号中间体，远红伸长的下胚轴（FHY 1）和FHY 1样（FHL），是至关重要的核转位，因为在它们的情况下，没有phyA是在核中检测。利用fhl/fhy 1双突变体中的细胞质phyA，可以揭示phyA的新的细胞质功能。在后续项目中，我们将进一步研究phyA的输入调控和FHY 1结合对phyA特性的影响，以及其他细胞质组分在早期phyA信号传导中的作用。因此，详细研究了FHY 1-phyA复合物易位到细胞核的机制。潜在的其他信号成分，已经确定在光依赖性酵母双杂交和三杂交试验与功能phyA也进行了研究。从而发现了细胞质磷酸酶对phyA极低注量反应（VLFR）的影响。此外，异源表达的phyA进行了生化和生物药理学分析，关于其与关键信号组分的相互作用。在该项目的拟议扩展中，将进一步研究易位过程中FHY 1 s二级修饰的体内状态、phyA-FHY 1相互作用的结构基础以及所发现的磷酸酶和小泛素样修饰物（SUMO）连接酶的功能。拟议的项目将是与马尔堡的埃森实验室的合作项目，该实验室将提供该项目的结构，生物物理和质谱任务，而Zeidler实验室将研究蛋白质相互作用，复杂成员的功能以及核转位的动力学和调节。该项目将促进对这种感光器功能的理解。核转位的调节在多种生物中具有普遍的兴趣，并且对受体的诱导性和定向转位的详细理解可以证明对于在许多应用中的基因表达的靶向操纵是有价值的，超出了植物领域。

Plants use phytochromes as red/far red sensing photoreceptors to monitor their light environment and regulate many aspects of development accordingly. Phytochrome A (phyA) is one of five phytochromes in Arabidopsis, regulating early seedling development among other responses. Following its activation, phyA translocates to the nucleus where it regulates gene expression. Two signaling intermediates, far red elongated hypocotol (FHY1) and FHY1-like (FHL), are crucial for nuclear translocation, as in their absence no phyA is detectable in the nucleus. Taking advantage of exclusively cytoplasmic phyA in the fhl/fhy1 double mutant, novel cytoplasmic functions of phyA could be revealed previously. In the follow-up project, the regulation of phyA import and the impact of FHY1 bindingon phyA properties, as well as the action of other cytoplasmatic components in early phyA signaling was further investigated. Thereby the mechanism by which the FHY1-phyA complex is translocated to the nucleus has been investigated in detail. Potential other signalling components which have already been identified in light-dependent yeast two- and three-hybrid assays with functional phyA have also been investigated. Thereby the influence of a cytoplasmic phosphatase on the phyA very low fluence response (VLFR) was discovered. Furthermore, heterologously expressed phyA wasbiochemically and biophysically analyzed concerning its interaction with the key signaling components. In the proposed extension of the project the in vivo status of FHY1s secondary modification during translocation, the structural basis of the phyA-FHY1 interaction and the function of the discovered phosphatase and small ubiquitin like modifier (SUMO) ligase will be investigated further. The proposed project will be a collaborative project with the Essen lab in Marburg which will provide the structural, biophysical and mass spectrometry tasks of the project while the Zeidler lab will work on Protein interaction, the function of the complex members and the kinetic and regulation of the nuclear translocation. The proposed project will advance the understanding of this photoreceptor’s function. The regulation of nuclear translocation is of general interest in a variety of organisms, and a detailed understanding of inducible and directional translocation of a receptor may prove valuable for targeted manipulations of gene expression in a number of applications,reaching beyond the plant field.