PHYTOCHROME A--STRUCTURE/FUNCTION AND SIGNALING PATHWAYS

描述：光敏色素是一个独特的信息家族 调节发育重要基因表达的光受体 对环境光信号的反应。的C-末端结构域 分子与递质组氨酸激酶模块具有序列相似性 细菌双组分传感器。然而，尽管有相当大的 研究努力，无论是生化机制的信号传递 光感受器，也不是早期信号中间产物的身份 已经确定了信号转导途径。私隐专员建议解决这些问题 使用光敏色素A(PhyA)的两个缺陷，最好的特征是 从实验上看，它是家族中最容易驯服的成员。具体目标 这项建议的目的是：(A)界定下列顺序和结构决定因素 负责其感光和调节的PHYA分子 活动；(B)确定信号传递的生化机制 PhyA与其初始反应伙伴(S)；和(C)识别早期信号 PhyA途径特有的中间体。实验方法 将包括：(A)分子、遗传和反向遗传分析 拟南芥定位功能活性亚区和特定残基 在光感受器分子内；(B)产生镁的量 异源宿主中重组PhyA结构域的结构 X射线结晶学测定；(C)基因座的分子克隆 编码PhyA途径特异的潜在信号中间体 在拟南芥遗传筛选中鉴定；(D)分子克隆 PhyA相互作用蛋白的体外相互作用克隆及酵母表达 双杂交筛选策略；和(E)蓝藻的开发 模型系统，聚球藻6803，最近被发现含有一个 光敏色素同系物还与双组分传感器有关，以加速 对菲亚是原核生物后代的可能性进行了分析， 光调控的组氨酸激酶。理解分子的光谱 真核细胞感知和转导胞外信号的机制 信息信号是当前生物医学研究的中心目标。 已经了解了很多关于受体激酶和磷酸化级联反应的知识 涉及丝氨酸/苏氨酸/酪氨酸类蛋白家族，以及关于G蛋白和 小分子是第二信使。相比之下，人们对此知之甚少 关于新发现的一类真核蛋白与 细菌感觉性组氨酸激酶。这些蛋白质的发现 这表明真核生物可能保留了一种感觉信号系统 对细菌双组分系统的影响，但这仍有待于 为多细胞真核生物直接演示的。

DESCRIPTION: The phytochromes are an unique family of informational photoreceptors that regulate developmentally important gene expression in response to environmental light signals. The C-terminal domain of the molecule has sequence similarity to the transmitter histidine kinase module of the bacterial two-component sensors. However, despite considerable research effort, neither the biochemical mechanism of signal transfer from the photoreceptor, nor the identity of early signaling intermediates in the transduction pathway have been determined. The PI proposes to address these two deficiencies using phytochrome A (phyA), the best characterized and experimentally most tractable member of the family. The specific objectives of this proposal are: (a) to define sequence and structural determinants of the phyA molecule responsible for its photosensory and regulatory activities; (b) to define the biochemical mechanism of signal transfer from phyA to its initial reaction partner(s); and (c) to identify early signaling intermediates specific to the phyA pathway. The experimental approaches will include: (a) molecular, genetic, and reverse genetic analysis in Arabidopsis to map functionally active sub-domains and specific residues within the photoreceptor molecule; (b) production of mg quantities of recombinant phyA structural domains in heterologous hosts for structure determination by X-ray crystallography; (c) molecular cloning of loci encoding potential signaling intermediates specific for the phyA pathway identified in genetic screens of Arabidopsis; (d) molecular cloning of phyA-interactive proteins using in vitro interaction cloning and yeast two-hybrid screening strategies; and (e) exploitation of a cyanobacterial model system, Synechocystis 6803, recently discovered to contain a phytochrome homolog also related to the two-component sensors, to accelerate analysis of the possibility that phyA is a descendent of a prokaryotic, light-regulated histidine kinase. Understanding the spectrum of molecular mechanisms by which eukaryotic cells perceive and transduce extracellular informational signals is a central goal of current biomedical research. Much has been learned about receptor kinases and phosphorylation cascades involving Ser/Thr/Tyr-class protein kinses, as well as about G-proteins and the small molecule second messengers. By contrast, very little is known about the newly discovered class of eukaryotic proteins related to the bacterial sensory histidine kinases. The discovery of these proteins suggests that eukaryotes may have retained a sensory signaling system based on that of the bacterial two-component systems, but this remains to be directly demonstrated for a multicellular eukaryote.