Mechanism of Peptide Signaling in Bacillus Subtilis

描述(申请人提供)：这项建议的长期目标是 了解革兰氏阳性菌中肽信号的一种新机制 以枯草芽孢杆菌的能力和产孢量因子(CSF)为模型 系统。脑脊液是一类新兴的信号肽的成员 在细胞内发挥作用。此前，信号肽被认为是 由于肽酶的存在，只能通过膜受体发挥作用 在牢房里。然而，我们的研究表明，脑脊液，一个未经修改的五 氨基酸肽，被运输到细胞中，在那里它发挥作用 在细胞内调节基因表达。肽信号，如脑脊液，是 由革兰氏阳性细菌用来监测其种群密度(即仲裁 感测)。细菌中的群体感应调节了这种医学上的重要性 毒力和生物膜形成等过程。因此，受监管的功能 在医学上重要的革兰氏阳性菌中通过多肽信号通路 是潜在的毒品目标。我们希望通过了解这一机制， 通过枯草杆菌中的CSF发出信号，我们将能够识别出其他 其他细菌中的细胞内功能信号肽。我们有 建议进行实验，以了解产生和响应的机制 脑脊液。脑脊液的前体蛋白是通过一般的 SEC依赖的输出途径，然后进行胞外处理。我们有 分离出一个在这种胞外加工过程中似乎有缺陷的突变体 活动。我们已经提出了基因和生物化学实验来表征 该突变体中有缺陷的蛋白质在脑脊液产生中的作用。成熟的脑脊液是 由非特异性寡肽渗透酶(OPP)导入，其同系物 存在于许多细菌中。建议进行实验来确定结合 脑脊液与Opp的亲和力及其他多肽是否与脑脊液竞争结合 敬奥普。一旦进入细胞，脑脊液似乎至少有三个细胞内 感受器。我们已经提出了识别细胞内受体的实验 并了解脑脊液如何与这些受体相互作用。脑脊液调节 使反应调节器去磷酸化的两种磷酸酶的活性。 这些磷酸酶很耐人寻味，因为它们显示出与广泛存在的 四肽重复序列(TPR)蛋白结构域家族。拟议中的实验 应该阐明这类新的信号传递机制 细胞内功能多肽。

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.