PHYTOCHROME A--STRUCTURE/FUNCTION AND SIGNALING PATHWAYS

描述：光敏色素是一个独特的信息家族， 光感受器调节发育重要的基因表达， 响应环境光信号。 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，尽管相当大的 研究工作，既不是信号传递的生化机制， 光感受器，也不是早期信号中间体的身份， 转导途径已经确定。 PI建议解决这些问题 使用光敏色素A（phyA）的两个缺陷，最好的特点， 实验上最听话的家庭成员 的具体目标 （a）确定生物多样性的顺序和结构决定因素， phyA分子负责其光敏和调节 （B）定义信号传递的生化机制， phyA与其初始反应伴侣的结合;以及（c）鉴定早期信号传导 phyA途径特异性的中间体。 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结构域和特定残基 （B）产生mg量的 异源宿主中的重组phyA结构域 X射线晶体学测定;（c）基因座的分子克隆 编码对phyA途径特异的潜在信号传导中间体 在拟南芥的遗传筛选中鉴定;（d） 使用体外相互作用克隆和酵母的phyA相互作用蛋白 双杂交筛选策略;和（e）利用蓝藻 模式系统，集胞藻6803，最近发现含有一个 光敏色素同系物也涉及到双组分传感器，以加速 分析phyA是原核生物后代的可能性， 光调节组氨酸激酶。 了解分子光谱 真核细胞感知和吸收细胞外的机制 信息信号是当前生物医学研究的中心目标。 人们对受体激酶和磷酸化级联反应已经有了很多了解 涉及Ser/Thr/Tyr类蛋白激酶，以及关于G蛋白和 小分子第二信使。 相比之下， 关于新发现的真核生物蛋白质类， 细菌感觉组氨酸激酶。 这些蛋白质的发现 这表明真核生物可能保留了一个基于 对细菌双组分系统，但这仍然是 直接证明了它是多细胞真核生物。

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.