Functional Analyses of Borrelia burgdorferi Adhesins

 描述(申请人提供)：莱姆病螺旋体通过媒介扁虱传播后，可建立局部皮肤感染，然后可能进一步传播到多个组织。 与哺乳动物细胞和细胞外基质(ECM)的相互作用是不同部位定植的关键，已鉴定出许多与细胞或细胞外基质(ECM)结合的伯氏杆菌粘附素。大量的粘附素以及一些粘附素的序列变异性可能会促进慢性多系统疾病，并导致人类莱姆病的多样性。由于靶组织的多样性、粘附素的多功能性以及定植的复杂性(例如，血流存活， 血管粘连、组织侵入和逃避免疫监视)。为了将过多的受体-配体相互作用组织到一个框架中，以了解莱姆病的多系统本质，三位合作者采用了多方面的遗传、生化和成像方法来充分表征已知的伯氏杆菌粘附素，发现和验证新的粘附素，并使用体外和体内模型探索伯氏杆菌粘附素的全部谱系的功能。这些方法导致了对已知的粘附素DbpA、BBK32和p66的重要新见解，并为OSPF亚家族和之前被证明对完全传染性至关重要的OSPF亚家族和OspC定义了新的黏附功能。我们发现DbpA和OspC变异与不同的组织取向有关，因此粘附素谱系中不同菌株的差异可能导致临床表现的相应差异。我们发现BBK32和P66在血管黏附和关节侵袭中起作用，它们与血管内皮细胞的相互作用可能反映了类似于参与淋巴细胞迁移的顺序活动。我们假设伯氏杆菌利用多种具有不同活性的粘附素在多相多系统感染中协商成功的步骤。此外，粘附素的一个子集的序列变异可能会促进不同部位的定植，并导致不同菌株在组织嗜性和疾病上的差异。为了详细了解由伯氏杆菌黏附素复杂的酶系介导的不同宿主细胞相互作用如何协调血管附着、组织侵袭和不同组织的长期定植，我们将使用伯氏杆菌遗传系统和不同的小鼠感染模型。我们将扩展和充分鉴定促进与内皮细胞相互作用的伯氏杆菌粘附素，以确定小鼠组织扩散和长期定植所需的粘附素-配体相互作用。我们还将描述这些相互作用中的每一个在促进附着到血管内皮细胞、跨越这一屏障以稳定地定植组织所需的多个步骤中所起的作用。这项拟议的工作将把复杂的黏附途径组织成官能团，这些官能团介导感染过程中的离散步骤，这些步骤共同导致系统性莱姆疏螺旋体病和小鼠播散性、持续性感染。

 DESCRIPTION (provided by applicant): Upon transmission by a vector tick, Lyme disease spirochetes can establish a local skin infection, then may disseminate further to multiple tissues. Interactions with mammalian cells and extracellular matrix (ECM) are critical for colonization of diverse sites, and many B. burgdorferi adhesins that bind to cells or extracellular matrix (ECM) have been identified. The large number of adhesins, as well as the sequence variability of some, is likely to promote chronic multisystem disease and contribute to the noted diversity of human Lyme disease. Understanding this infection is complicated by the diversity of target tissues, multifunctionality of adhesins, and complexity of colonization (e.g. bloodstream survival, vascular adherence, tissue invasion, and escape from immune surveillance). To organize the plethora of receptor-ligand interactions into a framework for understanding the multisystem nature of Lyme disease, the three collaborators have taken multi-faceted genetic, biochemical and imaging approaches to fully characterize known B. burgdorferi adhesins, to discover and validate new adhesins, and to probe the functions of the full repertoire of B. burgdorferi adhesins using in vitro and in vivo models. These approaches led to important new insights into the known adhesins DbpA, BBK32, and P66, and defined new adhesive functions for the OspF subfamily and for OspC, which was previously shown to be essential for full infectivity. We showed that DbpA and OspC variants are associated with different tissue tropisms, so strain-specific differences in the repertoire of adhesins may contribute to corresponding differences in clinical manifestations. We found that BBK32 and P66 have roles in vascular adherence and joint invasion, and the interactions they mediate with the vascular endothelium may reflect sequential activities analogous to those involved in lymphocyte transmigration. We hypothesize that B. burgdorferi utilizes a multitude of adhesins with distinct activities to negotiate successie steps in a multiphasic and multisystem infection. In addition, sequence variation of a subset of adhesins may promote colonization of distinct sites and contribute to strain-specific differences in tissue tropism and disease. To gain a detailed understanding of how the diverse host cell interactions mediated by the complex repertoire of B. burgdorferi adhesins coordinate vascular attachment, tissue invasion and long-term colonization of diverse tissues, we will use the power of B. burgdorferi genetic systems and diverse mouse infection models. We will expand and fully characterize B. burgdorferi adhesins that promote interactions with endothelium to define the adhesin-ligand interactions that are required for dissemination and long-term colonization of mouse tissues. We will also characterize the role of each of these interactions in promoting the multiple steps required to attach to the vascular endothelium, to transmigrate across this barrier to stably colonize tissue. The proposed work will organize complex adhesive pathways into functional groups that mediate discrete steps in the infectious processes that together result in systemic Lyme borreliosis and disseminated, persistent infection in the mouse.