Mechanism of Peptide Signaling in Bacillus Subtilis

描述（由申请人提供）：本提案的长期目标是 了解革兰氏阳性菌中肽信号传导的新机制 使用枯草芽孢杆菌的感受态和孢子形成因子（CSF）作为模型 系统CSF是一类新兴的信号肽， 在细胞内起作用。以前，信号肽被认为是 由于肽酶的存在， 在细胞内。然而，我们的研究表明，CSF，一个未经修改的五 氨基酸肽被转运到细胞中发挥作用 在细胞内调节基因表达。肽信号，如CSF， 由革兰氏阳性菌用于监测其种群密度（即群体 感测）。细菌中的群体感应调节了这种医学上重要的 毒力和生物膜的发展过程。因此，职能规定 在医学上重要的革兰氏阳性菌中通过肽信号通路 是潜在的药物靶点我们希望，通过了解 在B中通过CSF信号传导。枯草杆菌，我们将能够确定额外的 在其他细菌中的细胞内功能信号肽。我们有 提出的实验，以了解生产和响应的机制， 脑脊液。CSF的前体蛋白是通过一般的 依赖于sec的输出途径，然后在细胞外加工。我们有 我分离出一个突变体，似乎是在这种细胞外加工缺陷 活动我们已经提出了遗传和生化实验来表征 该突变体中蛋白质缺陷在CSF产生中的作用。成熟的CSF是 由非特异性寡肽通透酶（Opp）输入，其同系物 存在于许多细菌中。提出实验来确定结合 CSF对Opp的亲和力以及其他肽是否与CSF竞争结合 一旦进入细胞，CSF似乎至少有三个细胞内 受体。我们已经提出实验来鉴定细胞内受体 并了解CSF如何与这些受体相互作用。CSF调节 使反应调节剂去磷酸化的两种磷酸酶的活性。 这些磷酸酶很有趣，因为它们与广泛分布的 四方肽重复（TPR）蛋白结构域家族。拟议的实验 应该阐明这类新的信号传导机制， 胞内功能肽

DESCRIPTION (provided by applicant): The long-term goal of this proposal is to understand a novel mechanism of peptide signaling in Gram-positive bacteria using Competence and Sporulation Factor (CSF) of Bacillus subtilis as a model system. CSF is a member of an emerging class of signaling peptides that function intracellularly. Previously, signaling peptides had been thought to function only through membrane receptors, due to the presence of peptidases inside the cell. However, our studies have shown that CSF, an unmodified five amino acid peptide, is transported into the cell where it functions intracellularly to regulate gene expression. Peptide signals, like CSF, are used by Gram-positive bacteria to monitor their population density (i.e. quorum sensing). Quorum sensing in bacteria regulates such medically important processes as virulence and biofilm development. Therefore, functions regulated by peptide signaling pathways in the medically important Gram-positive bacteria are potential drug targets. We hope that, by understanding the mechanism of signaling by CSF in B. subtilis, we will be able to identify additional intracellular functioning signaling peptides in other bacteria. We have proposed experiments to understand the mechanism of production and response to CSF. The precursor protein for CSF is secreted through the general Sec-dependent export pathway and then processed extracellularly. We have isolated a mutant that appears to be defective in this extracellular processing activity. We have proposed genetic and biochemical experiments to characterize the role of protein defective in this mutant in CSF production. Mature CSF is imported by a non-specific oligopeptide permease (Opp), homologues of which exist in many bacteria. Experiments are proposed to determine the binding affinity of CSF for Opp and whether other peptides compete with CSF for binding to Opp. Once inside the cell, CSF appears to have at least three intracellular receptors. We have proposed experiments to identify the intracellular receptors of CSF and to understand how CSF interacts with these receptors. CSF regulates the activity of two phosphatases that dephosphorylate response regulators. These phosphatases are intriguing as they show similarity to the widespread tetratrico peptide repeat (TPR) protein domain family. The proposed experiments should elucidate the mechanism of signaling by this novel class of intracellular function peptides.