LIGHT-RESPONSIVE GENES

植物依赖于光作为调节两者的信号 发育和代谢过程，而光敏色素被认为是 调节这些反应的主要植物感光器。长的- 我们实验室的学期目标是确定 光敏色素信号通路，并了解这些通路是如何 受监管的。通过开发基于微注射的实验系统，我们 已经证明了三条信号转导通路的存在 是一种光敏色素(PhyA)的下游，它调节 编码叶绿体成分和花青素基因的表达 生物合成酶。一种途径依赖于钙，第二种依赖于钙。 在cGMP上，第三个在这两个效应器上。我们有 已建立CAB、CHS和FNR作为该活动的特异性报告基因 三条PhyA信号通路中的每一条。我们现在提议 使用四种方法进一步研究这些途径：a) 确定CAB、CHS和FNR启动子中的单个顺式元件 对钙和/或cGMP有反应的细胞。这将由以下人员执行 利用不同启动子缺失和合成的显微注射实验 包含一个或多个先前识别的顺式元件的启动子。我们的 人们的注意力将集中在转录因子的结合位点上 GT-1在CAB启动子、G盒和Myb因子结合部位 在CHS启动子内。B)除了PhyA信号外 中间体已经澄清，我们希望使用我们的微量注射 刻画他人的技巧。特别是，我们将研究 钙/钙调蛋白依赖性蛋白激酶II的可能参与 和鸟苷酸环化酶。C)在拟南芥中使用突变隔离筛选 基于光调控报告基因(CAB-1)的异常表达 LUC)作为对光的反应，我们已经分离出一些突变体，它们可能 在PHYA信号转导中发生改变。光生物学与菲亚 突变表型的依赖性将被表征，以及那些明确的 将对影响PHYA的信号转导进行分析，以确定 信号通路内的病变部位。D)根据最近 实验e现在对个体Phya如何发出信号有了一些了解 通路是受调控的，以及每条通路之间的串扰机制 做手术吧。我们已经设计了新的筛选策略来分离突变株 这种基于CAB异常表达的调控机制-- LUC报告基因或CHS-LUC报告基因最近被 插入到我们实验室的拟南芥基因组中。使用这些 我们希望不仅通过以下方式来扩展我们对过程的了解 哪些植物通过光敏色素感知光，但也通过调节 调控基因表达的协调过程 光敏色素。

Plants are dependent upon light as a signal for regulating both developmental and metabolic processes, and phytochrome is held to be the principal plant photoreceptor that regulates these responses. The long- term objective of our laboratory is to identify the components of phytochrome signaling pathways and to understand how these pathways are regulated. By developing a microinjection-based experimental system, we have demonstrated the existence of three signal transduction pathways that are downstream of one of the phytochromes (PHYA), and which regulate the expression of genes encoding chloroplast components and anthocyanin biosynthetic enzymes. One pathway is dependent upon calcium, the second upon cGMP, and the third upon both of these effectors. We have established CAB, CHS, and FNR, as specific reporter genes for the activity of each of the three PHYA signaling pathways respectively. We now propose to investigate these pathways further using four approaches: A) To identify individual cis-elements within the CAB, CHS and FNR promoters that can respond to calcium and/or cGMP. This will be carried out by microinjection experiments using various promoter deletions and synthetic promoters containing one or more previously identified cis-elements. Our attention will be focused on binding sites for the transcription factor GT-1 in the CAB promoter, and on G-boxes and Myb-factor binding sites within the CHS promoter. B) In addition to the PHYA signaling intermediates already elucidated, we hope to use our microinjection techniques to characterize others. In particular, we will examine the possible participation of calcium/calmodulin-dependent protein kinase II and guanylyl cyclase. C) Using a mutant isolation screen in Arabidopsis based upon the aberrant expression of a photoregulated reporter gene (CAB- LUC) in response to light, we have isolated a number of mutants which may be altered in PHYA signal transduction. The photobiology and PHYA dependency of mutant phenotypes will be characterized and those clearly affected in PHYA signal transduction will be analyzed to determine the sites of lesions within the signaling pathways. D) Based upon recent experiments e now have some understanding of how individual PHYA signaling pathways are regulated, and how cross-talk mechanisms between each pathway operate. We have devised new screening strategies to isolate mutants in such regulatory mechanisms, based upon the aberrant expression of the CAB- LUC reporter gene or a CHS-LUC reporter gene which has recently been inserted into the Arabidopsis genome in our laboratory. Using these approaches we hope to expand our knowledge not only of the processes by which plants perceive light via phytochrome, but also the regulatory processes which operate to coordinate the expression of genes regulated by phytochrome.