Novel carbohydrate binding functions of the CR3 I-domain modulate gonococcal-cervical cell interactions

CR3 I 结构域的新型碳水化合物结合功能调节淋球菌-宫颈细胞相互作用

• 批准号: 10078936
• 负责人: Jennifer L Edwards
• 金额: $ 46.5万
• 依托单位: RESEARCH INST NATIONWIDE CHILDREN'S HOSP
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-01-16 至 2022-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10078936
• 关键词: Adhesions; Affect; Agonist; Bacteria; Bacterial Proteins; Behavior; Binding; Binding Sites; Biological; C3bi; Calorimetry; Carbohydrates; Cell Communication; Cells; Centers for Disease Control and Prevention (U.S.); Cervical; Cervicitis; Cervix Uteri; Cessation of life; Coculture Techniques; Complement; Data; Development; Diagnosis; Disease; Ensure; Epithelial; Epithelial Cells; Event; Female; Fimbriae Proteins; Foundations; Frequencies; Future; Goals; Gonorrhea; HIV Infections; Health; Human; Immune; Immune response; Immune system; Immunity; Immunologic Receptors; In Vitro; Infection; Infertility; Innate Immune Response; Lectin; Ligands; Link; Literature; Macrophage-1 Antigen; Mass Spectrum Analysis; Mediating; Mediator of activation protein; Medical Care Costs; Molecular; Morbidity - disease rate; Mucosal Immunity; Mucous Membrane; Multi-Drug Resistance; Natural Immunity; Nature; Neisseria gonorrhoeae; Outcome; Pathogenesis; Pathway interactions; Pattern recognition receptor; Pelvic Inflammatory Disease; Peptides; Phagocytes; Phase; Pilum; Play; Polysaccharides; Post-Translational Protein Processing; Prevalence; Prevention; Preventive measure; Process; Proteins; Public Health; Recombinants; Reporting; Role; Seminal; Signal Transduction; Structure; Surface; Surface Plasmon Resonance; Testing; Therapeutic; United States; Ursidae Family; Variant; Woman; Work; appendage; base; experience; glycosylation; human disease; human pathogen; improved; innate immune function; innovation; insight; macrophage; microbial; neutrophil; novel; novel strategies; novel therapeutics; pathogen; pathogenic bacteria; prevent; receptor; reproductive tract; response; sugar; trafficking; vaccine development

PROJECT SUMMARY The prevalence of Neisseria gonorrhoeae (Ng), its associated morbidity, and the emergence of untreatable strains, support a critical need for new preventative and therapeutic strategies for this important human- pathogen. Such developments require a complete understanding of the pathobiology of this archetypal, host- adapted pathogen. We show that phase-variable, glycan post-translational modifications on Ng surface appendages, pili (fimbriae), play a vital role in human infection. Complement receptor 3 (CR3) is an important pattern recognition receptor that is the key receptor mediating Ng colonization of human cervical mucosa. CR3 contains an “I-domain” region, which is known as a binding site for protein ligands. However, we found that Ng binding to the I-domain is mediated by the pilin-linked glycan (PLG). This is a seminal finding in innate immunity, as all previous literature ascribe CR3 carbohydrate binding to a separate, so called "lectin" domain. This was also the first demonstration that post-translational modifications made to a bacterial protein modulate pathogenesis. These findings will impact our understanding of microbial pathogenesis and innate immune responses. The goals of the present application are to define the specific contribution of the six, naturally occurring, PLG structures in mediating the Ng-CR3 I-domain interaction and to determine the biological relevance of each of these interactions to infection in females. Guided by strong preliminary data, we hypothesize that variable PLG structures initiate key, but highly divergent, outcomes with CR3 I-domain engagement. We will resolve our hypothesis through two specific aims: Aim 1) Define the effect of variation in pilin glycan structure on direct PLG-I-domain interactions; we will define the specific molecular interactions occurring with the CR3 I-domain for the six natural Ng PLG structures. Aim 2) Define the effect of PLG-I- domain interactions on Ng pathogenesis and CR3 function; we will define targeted epithelial cell responses to CR3 I-domain engagement by Ng that bear different PLG structures and their effect on Ng pathogenesis. Our approach is innovative in using biologically relevant human primary cervical cells, both alone and in co-culture with phagocytes, combined with low passage Ng isolates. This ensures that data obtained are relevant to human processes. By defining the effector functions controlled by I-domain lectin activity on primary human epithelial cells, we will provide critical new information regarding Ng pathogenesis, CR3 function, and the fundamental mechanisms that govern human cervical mucosal immunity. Moreover, we will define the utility of targeting the PLG-CR3 I-domain interaction as a new, and improved, host-targeted approach to treat and/or prevent Ng disease in women. Our studies will very likely impact work on other human pathogens and on innate immunity, broadening the significance of our outcomes. We have extensive experience in defining the molecular mechanisms of Ng pathogenesis. We are ideally placed to do this work in having discovered CR3 as the key receptor for Ng cervical infection and the biosynthetic pathways for pili glycosylation.

项目摘要 Neiserseria Gonorrhoeae（NG）的流行，其相关的发病率和不可治疗的出现 应变，支持对这一重要人类的新预防和治疗策略的迫切需求 病原。这样的发展需要完全了解这种原型，宿主的病理学 - 改编的病原体。我们表明，NG表面上的相变，聚糖后翻译后修饰 附属物，pili（fimbriae），在人类感染中起着至关重要的作用。补体受体3（CR3）是重要的 模式识别受体是介导人宫颈粘膜NG定殖的关键受体。 CR3 包含一个“ I域”区域，该区域被称为蛋白质配体的结合位点。但是，我们发现 NG与I域的结合是由PILIN连接的聚糖（PLG）介导的。这是一个天生的半找到 免疫力，作为以前的所有文献，作为CR3碳水化合物与单独的“凝集素”结构域结合的结合。 这也是第一次对细菌蛋白进行翻译后修饰的证明 调节发病机理。这些发现将影响我们对微生物发病机理和先天性的理解 免疫反应。本应用的目标是定义六个的特定贡献， 自然发生的PLG结构在介导Ng-Cr3 I域相互作用中，并确定 这些相互作用中每一种与女性感染的生物学相关性。在强大的初步数据的指导下，我们 假设可变的PLG结构启动关键，但具有高度分歧的CR3 I域的结果 订婚。我们将通过两个具体目的来解决我们的假设：目标1）定义变化的影响 直接PLG-I域相互作用的PILIN GLYCAN结构；我们将定义特定的分子相互作用 用于六个天然NG PLG结构的CR3 I域。目标2）定义PLG-i的影响 NG发病机理和CR3功能的域相互作用；我们将定义针对的上皮细胞反应 NG的CR3 i域域的参与度具有不同的PLG结构及其对NG发病机理的影响。我们的 方法是使用与生物学相关的人类原代宫颈细胞的创新性，无论是单独还是共同培养 吞噬细胞与低通过NG分离株结合使用。这确保获得的数据与 人类过程。通过定义由I域讲座活动控制的效应子功能 上皮细胞，我们将提供有关NG发病机理，CR3功能和 控制人宫颈粘膜免疫的基本机制。而且，我们将定义实用程序 将PLG-CR3 I域相互作用作为一种新的，改进的宿主靶向方法来治疗和/或 预防女性NG疾病。我们的研究很可能会影响其他人类病原体以及 先天免疫，扩大我们的结果的重要性。我们在定义 NG发病机理的分子机制。理想情况下，我们可以在发现CR3的情况下做这项工作 NG宫颈感染的关键受体和Pili糖基化的生物合成途径。