Functional and Mechanistic Studies of the VlsE-mediated Immune Avoidance System in the Lyme Disease Spirochete

莱姆病螺旋体 VlsE 介导的免疫回避系统的功能和机制研究

• 批准号: 10371053
• 负责人: Troy Michael Bankhead
• 金额: $ 22.95万
• 依托单位: WASHINGTON STATE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-03-13 至 2024-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10371053
• 关键词: Animals; Antibodies; Antibody Response; Antibody titer measurement; Antigenic Variation; Antigens; Area; Arthritis; Binding; Biological Assay; Borrelia; Borrelia burgdorferi; Carditis; Cell Surface Proteins; Cell surface; Data; Enzyme-Linked Immunosorbent Assay; Epitopes; Exhibits; Future; Goals; Health; Human; Immune; Immune Evasion; Immunoglobulin G; Infection; Investigation; Knowledge; Lipoproteins; Longitudinal Studies; Lyme Disease; Masks; Mediating; Membrane Proteins; Mission; Mus; Neurologic; North America; Outcome; Passive Immunization; Process; Proteins; Public Health; Publishing; Quantitative Reverse Transcriptase PCR; Research; Surface; Surface Antigens; System; Techniques; Testing; Tick-Borne Diseases; United States National Institutes of Health; Variant; Western Blotting; Work; base; chronic infection; crosslink; design; disability; human pathogen; immunogenic; in vivo; innovation; mutant; novel; pathogen; prevent; protein function; protein protein interaction; tool

Project Summary A key mechanism for immune evasion and persistent infection by the Lyme disease spirochete, Borrelia burgdorferi, is antigenic variation of the VlsE surface protein. Despite the presence of a substantial number of additional proteins residing on the bacterial surface, VlsE is the only antigen that exhibits ongoing variation of its exposed epitopes. Recent work in our lab has identified a VlsE-mediated immune avoidance system that allows non-VlsE surface antigens to escape the killing effects of host antibodies. Moreover, we have identified the Arp lipoprotein as one such surface antigen that benefits from VlsE-mediated immune protection. Despite this important evidence, certain functional and mechanistic aspects involved in this system are still unknown. Our long-term goals are to determine the mechanism and overall implications of surface antigen protection promoted by the VlsE lipoprotein during host infection by B. burgdorferi. The objective of this application is to decipher the functional details of factors involved in VlsE-mediated immune avoidance, and identify any additional B. burgdorferi surface antigens that are shielded by VlsE. Based on published studies and preliminary findings, our central hypothesis is that VlsE exists establishes protein-protein interactions with Arp and other proteins on the B. burgdorferi cell surface. Additionally, we hypothesize that the presence of VlsE functions to dampen the antibody response to Arp. The rationale for the proposed research is that identifying the functional details of this system will provide the knowledge required to design downstream studies targeted at dissecting the overall mechanism. Together, the proposed research is relevant to NIH’s mission that pertains to developing fundamental knowledge that will potentially help to reduce the burdens of human illness and disability. Guided by preliminary findings, our hypotheses will be tested by pursuing two specific aims: 1) Determine whether VlsE directly interacts with Arp and other B. burgdorferi cell surface proteins; and 2) Determine whether the presence of VlsE modulates the antibody response to Arp. Under the first aim, a specialized in vivo crosslinking/elution technique will be used to detect protein-protein interactions between mutant VlsE and Arp, and to identify novel VlsE binding partners. Under the second aim, qRT-PCR will be utilized to quantitate the relative arp expression levels during host infection by a VlsE-deficient clone compared to a wild type control, and anti-Arp antibody titers in mice infected with clones expressing or lacking VlsE will be assessed by ELISA. The proposed work is innovative, because it utilizes a new VlsE/Arp over-expresser clone to analyze potential VlsE-Arp interactions, and uses the Arp lipoprotein as a readout for VlsE protection assays. Overall, these studies will significantly advance our knowledge of immune evasion by B. burgdorferi, and provide more useful strategies to prevent and treat Lyme disease in humans.

项目摘要 莱姆病螺旋体，渗透症的免疫进化和持续感染的关键机制 Burgdorferi是VLSE表面蛋白的抗原变异。尽管存在大量 驻留在细菌表面上的其他蛋白质，VLSE是唯一表现出持续变化的抗原 它暴露的表位。我们实验室的最新工作已经确定了VLSE介导的免疫避免系统 允许非VLSE表面抗原逃脱宿主抗体的杀伤作用。而且，我们已经确定了 ARP脂蛋白是一种受益于VLSE介导的免疫保护的表面抗原。尽管 该系统中涉及的某些功能和机械方面的这一重要证据仍然未知。 我们的长期目标是确定表面抗原保护的机制和整体含义 B. burgdorferi在宿主感染期间由VLSE脂蛋白促进。该应用的目的是 破译VLSE介导的免疫避免涉及因素的功能细节，并确定任何 由VLSE屏蔽的其他B. burgdorferi表面抗原。根据已发表的研究和 初步发现，我们的中心假设是，VLSE存在与ARP建立蛋白质 - 蛋白质相互作用 和墨芽孢杆菌细胞表面上的其他蛋白质。此外，我们假设VLSE的存在 功能可以使抗体对ARP的反应跳舞。拟议研究的理由是确定 该系统的功能细节将提供设计针对下游研究所需的知识 解剖整体机制。拟议的研究共同与NIH的使命有关 发展基本知识，这有可能有助于减少人类疾病的伯纳斯和 残疾。 在初步发现的指导下，我们的假设将通过追求两个具体目标来检验：1） 确定VLSE是否直接与ARP和其他B. burgdorferi细胞表面蛋白相互作用；和2） 确定VLSE的存在是否调节抗体对ARP的反应。在第一个目标下， 专门的体内交联/洗脱技术将用于检测蛋白质 - 蛋白质相互作用 突变的VLSE和ARP，并确定新颖的VLSE结合伙伴。在第二个目标下，QRT-PCR将是 与VLSE缺陷克隆相比，用于定量宿主感染期间相对ARP表达水平 对野生型控制和抗ARP抗体滴度，感染了表达或缺乏VLSE的克隆的小鼠 由Elisa评估。拟议的工作具有创新性，因为它使用了新的VLSE/ARP过表达者 克隆分析潜在的VLSE-ARP相互作用，并使用ARP脂蛋白作为VLSE保护的读数 测定。总体而言，这些研究将大大提高我们对B. burgdorferi的免疫进化的了解， 并提供更有用的策略来预防和治疗人类的莱姆病。