B. subtilis PHO REGULON SIGNAL TRANSDUCTION NETWORK

描述(申请人提供)：两组分信号转导是原核生物信号转导的主要形式，在真核生物中有独特的例子。在对数生长结束时，“过渡态”芽孢杆菌细胞接收来自环境的多个信号，这些信号同时报告温度、细胞密度、营养可获得性和氧分压等条件。这些信号被处理以确定最适合生存的基因表达和代谢反应。我们正在测试的假设是：“多重信号的处理是由涉及多个两组分系统的调节网络完成的，这些系统的功能是建立系统之间的依赖或层次结构。调节子之间的跨系统相互作用提供了一种信号整合和放大的机制，这种机制导致对给定反应的微调，以适应生物体在任何时候经历的全部信号输入。”Pho信号转导网络至少由3个双组分系统组成：Php-phor、ResD-rese和Spo0A，以及全球胁迫、分解代谢、过渡态和发育调节因子。我们将通过确定6个PhoPR启动子(S)中的哪个对AbrB的直接结合做出反应，以及AbrB是否也通过SCOC起间接作用，来研究Spo0A~P-AbrB通路在Pho信号转导网络中的作用。我们将确定在缺乏第二个PI饥饿全局调控因子SigB的情况下，哪些未知的调控电路控制着PhoPR P5启动子的调控。我们将通过分析RESA转录中绕过Phop要求的突变以及通过体外重建RESA转录来了解Phop和ResD的基本但不足的作用，来研究在磷酸盐饥饿过程中对resABCDE转录(ResD和Rese产生)至关重要的正反馈环。我们将通过正反馈环来研究跨系统的相互作用，正反馈环对于充分诱导Pho调节子是必不可少的，通过确定上游调控因子ResD在控制PI缺陷信号和/或该信号的调制中的作用。我们将确定在体外如何抑制Phor的自磷酸化的还原的菜喹酮，在体内调节磷缺乏信号，并询问氧化还原反应性半胱氨酸是否发挥作用。这些研究将有助于建立革兰氏阳性菌作为抗菌治疗靶点的双组分信号转导系统的知识库。

DESCRIPTION (provided by applicant): 2-component signaling is the principal form of signal transduction in prokaryotes with distinctive examples in eukaryotes. At the end of logarithmic growth, the "transition state" Bacillus cell receives multiple signals from the environment that are simultaneously reporting conditions such as temperature, cell density, nutrition availability and oxygen tension. These signals are processed to determine the most appropriate gene expression and metabolic response for survival. The hypothesis we are testing is that "The processing of the multiple signals is accomplished by regulatory networks involving multiple 2-component systems that function to establish dependencies or hierarchies between systems. Cross-system interaction between regulons provides a mechanism for signal integration and amplification, a mechanism which results in fine tuning of a given response that accommodates the entire signal input experienced by the organism at any 1 time." The Pho signal transduction network is comprised of at least 3 2-component systems: PhoP-PhoR, ResD-ResE and Spo0A, and global stress, catabolite, transition state and developmental regulators. We will examine the role of the Spo0A~P-AbrB pathway in the Pho signal transduction network by determining which of 6 phoPR promoter(s) respond to direct AbrB binding and if AbrB also has an indirect role via ScoC. We will determine what unknown regulatory circuits control phoPR P5 promoter regulation in the absence of a second Pi starvation global regulator, SigB. We will examine the positive feedback loop that is essential for resABCDE transcription (ResD and ResE production) during phosphate starvation by analyzing mutations that bypass the requirement for PhoP in resA transcription and via reconstruction of resA transcription in vitro to understand the essential but insufficient role of both PhoP and ResD. We will examine the cross-system interaction via the positive feedback loop that is essential for full induction of the Pho regulon by determining the role of upstream regulator, ResD, in controlling the Pi deficiency signal and/or modulation of that signal. We will determine how reduced menaquinones that inhibit the autophosphorylation of PhoR in vitro, modulate the phosphate deficiency signal in vivo and ask if redox-reactive cysteines play a role. These studies will contribute to the knowledge base of two-component signal transduction systems of gram-positive bacteria as targets for antimicrobial therapy.