Light responsive genes

描述(申请人提供)：光为光合作用提供能量，也为植物的生长和发育提供信号。拟南芥含有几个光感受器，它们监测不同波长的光子。其中，被远红光激活的光敏色素A(PhyA)在幼苗脱黄化过程中起着重要作用。我们的长期目标是了解各种信号中间体之间的相互关系，以及它们的相互作用如何导致下游发育反应的PhyA信号转导。为此，我们将分析携带不同PhyA信号因子组合中损伤的多个突变的突变体，以建立两个同源蛋白FHY1/FHL和三个转录激活因子LAF1、HFR1和HY5之间的等级关系。相互作用区域将通过体外下拉试验进行绘制。免疫共沉淀法将确定在体内对黑暗和FR反应的因子关联，并将在幼苗去黄化过程中分析对PhyA反应的生理影响。控制PHYA信号因子的丰度已成为一种基本的调控机制，而COP1是PHYA信号的负调控因子，起着重要作用。COP1对LAF1和HFR1具有E3连接酶活性，这两种连接酶在体外相互作用。利用表达这两种因子的转基因植物，我们将确定它们在体内的关联是否会导致FR超敏反应。COP1的另一个靶标是bZIP因子HY5，它似乎在下胚轴和子叶对FR光的反应中调节相反的细胞扩张效应。我们将研究HY5如何差异控制膜类固醇结合蛋白(MSBP)1在这两个器官中的表达，MSBP 1是已知的细胞扩张的负调控因子，表达适当结构的转基因植物将被用于确定调控是在转录、转录后还是翻译后水平上进行的。最后，我们将探讨miRNA和siRNAs是否参与了PhyA信号的调节。我们期望这些实验的结果能揭示PHYA信号转导的未知方面，并提供有关真核细胞信号转导一般机制的更多信息。

DESCRIPTION (provided by applicant): Light provides energy for photosynthesis and also signals for plants to grow and develop. Arabidopsis contains several photoreceptors that monitor photons at different wavelengths. Amongst these, phytochrome A (phyA), which is activated by far-red (FR) light, plays a major role in seedling deetiolation. Our long-term objective is to understand the inter-relationship between the various signaling intermediates and how their interactions lead to transduction of phyA signals for downstream developmental responses. To this end, we will analyze mutants carrying multiple mutations with lesions in various combinations of phyA signaling factors to establish the hierarchical relationship between the two homologous proteins, FHY1/FHL, and three transcription activators, LAF1, HFR1 and HY5. Interaction domains will be mapped by in vitro pull- down assays. Factor association in vivo in response to darkness and FR will be determined by co-immunoprecipitation and the physiological impact on phyA responses will be assayed during seedling deetiolation. Control of phy A signaling factor abundances has emerged to be an essential regulatory mechanism and COP1, a negative regulator of phYA signaling, plays a major role. COP1 shows E3 ligase activity toward LAF1 and HFR1, which interact with one another in vitro. Using transgenic plants expressing both factors we will determine whether their association in vivo would lead to FR-hypersensitivity. Another COP1 target is the bZIP factor, HY5, which appears to mediate opposing effects on cell expansion in hypocotyls and cotyledons in response to FR light. We will investigate how HY5 differentially controls expression of the Membrane Steroid Binding Protein (MSBP) 1 in these two organs, which is known to be a negative regulator of cell expansion, Transgenic plants expressing the appropriate constructs will be used to determine whether regulation is at the transcriptional, post-transcriptional or post-translational level. Finally, we will explore whether miRNA and siRNAs are involved in the regulation of phyA signaling. We expect results from these experiments to uncover previously unknown aspects of phyA signaling and also provide additional information on general mechanisms of eukaryotic cell signaling.