Staphylocidal Mechanism of Platelet Microbicidal Protein

血小板杀菌蛋白的杀菌机制

• 批准号: 6826800
• 负责人: ARNOLD S BAYER
• 金额: $ 30.15万
• 依托单位: LUNDQUIST INSTITUTE FOR BIOMEDICAL INNOVATION AT HARBOR-UCLA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-12-01 至 2006-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6826800
• 关键词: Staphylococcus aureus; Staphylococcus infection; antibacterial agents; antibacterial antibody; bacteria infection mechanism; bacterial antigens; bacterial cytopathogenic effect; bacterial proteins; bactericidal immunity; gene mutation; host organism interaction; intracellular; microorganism culture; microorganism immunology; platelets; proteomics; site directed mutagenesis; transfection /expression vector; transposon /insertion element

DESCRIPTION (provided by the applicant): Staphylococcus aureus is a virulent pathogen which is associated with a broad-spectrum of clinical infections. Its ability to colonize host tissues, and to persist and proliferate within host tissues requires the organism to circumvent innate host defense mechanisms. We have discovered that mammalian platelets store and secrete a family of antimicrobial peptides at potential sites of endovascular damage and microbial colonization that serve to both growth inhibit and kill S. aureus. In the previous grant period, we have delineated that the principal antimicrobial peptide which is secreted from platelets (thrombin-induced platelet microbicidal protein-1 [tPMP-1]), interacts with S. aureus in vitro by initial attachment to the cytoplasmic membrane, after which a microbicidal cascade is triggered in strains intrinsically susceptible to this peptide. In contrast, those strains which were engineered to be resistant to tPMP-1 in vitro (e.g., by transposon mutagenesis) do so by changing the basic biology of their cytoplasmic membrane target for tPMP-1. In vitro susceptibility to tPMP-1 is mirrored by enhanced clearance of such strains in animal models of endovascular infection; in contrast, in vitro resistance to tPMP-1 is correlated with an augmented survival advantage in the same animal models. The overall purposes of this proposal are: i) to define the mechanisms by which tPMP-1 executes its microbicidal effects, particularly focusing on intracellular targeting and activation of stress response systems; and ii) to delineate the mechanisms, genetic pathways and membrane biochemical adaptations by which the organism is able to successfully respond to exposures to tPMP-1 for survival. For these purposes, we will utilize a series of well-characterized and isogenic strain pairs of S. aureus (including site-directed plasmid mutants, as well as mutants with plasmid reporter fusions) that will enable us to define both the mechanisms of microbicidal action of tPMP-1, as well as the homeostatic adaptive pathways used by the organism to survive tPMP-1 exposures. Moreover, we will employ proteomics approaches to divulge novel genes and metabolic pathways triggered by tPMP-1 as part of either its microbicidal cascade, or as part of the organism's adaptive strategies. These studies will provide a solid foundation for the future design of unique platelet peptide congeners which are better able to target S. aureus strains for killing, as well as to circumvent innate homeostatic mechanisms used by the organism for survival.

描述（由申请人提供）：金黄色葡萄球菌是一种剧毒菌 与广谱临床感染相关的病原体。它是 定植宿主组织并在宿主体内持续增殖的能力 组织需要生物体规避宿主的先天防御机制。我们 发现哺乳动物血小板储存和分泌一系列 抗菌肽在血管内损伤和微生物的潜在位点 定殖可抑制金黄色葡萄球菌生长并杀死金黄色葡萄球菌。在 在上一个资助期间，我们已经确定了主要的抗菌药物 血小板分泌的肽（凝血酶诱导的血小板 杀微生物蛋白-1 [tPMP-1]），在体外与金黄色葡萄球菌相互作用 附着在细胞质膜上，然后发生杀菌级联反应 在对该肽本质敏感的菌株中触发。相比之下， 那些被设计为在体外对 tPMP-1 具有抗性的菌株（例如， 通过转座子诱变）通过改变其基本生物学来做到这一点 tPMP-1 的细胞质膜靶标。体外对 tPMP-1 的敏感性为 通过在血管内注射的动物模型中此类菌株的清除率增强来反映 感染;相反，体外对 tPMP-1 的耐药性与 在相同的动物模型中增强了生存优势。总体目的 该提案是： i) 定义 tPMP-1 执行其任务的机制 杀菌作用，特别关注细胞内靶向和 激活应激反应系统； ii) 描述机制， 生物体的遗传途径和膜生化适应 能够成功地对暴露于 tPMP-1 做出反应以求生存。对于这些 为了达到目的，我们将利用一系列特征良好的同基因菌株 金黄色葡萄球菌对（包括定点质粒突变体，以及突变体 与质粒报告融合），这将使我们能够定义 tPMP-1 的杀菌作用机制以及稳态 生物体在 tPMP-1 暴露下生存的适应性途径。而且， 我们将采用蛋白质组学方法来揭示新基因和代谢 tPMP-1 触发的途径作为其杀菌级联的一部分，或者作为 有机体适应策略的一部分。这些研究将提供坚实的 为未来设计独特的血小板肽同系物奠定了基础 能够更好地针对金黄色葡萄球菌菌株进行杀死和规避 生物体为了生存而使用的先天稳态机制。