Understanding Unique Aspects of Motility and Chemotaxis in Borrelia burgdorferi

摘要 莱姆病细菌伯氏疏螺旋体(BB)具有高度的运动性，可以穿越复杂的内部环境 哺乳动物和节肢动物在其感染周期中寄主。这个应用程序的中心假设是 Bb的运动性和趋化性构成了一个独特的范式，并在宿主-媒介循环中发挥着关键作用 如在疾病过程中，包括入侵、扩散、组织趋向性和免疫逃避。在过去的几年里 在两个资金周期中，我们揭示了BB运动性和趋化性的几个独特方面；然而，它们的基础 分子机制和在疾病过程中的确切作用在很大程度上仍不清楚。建立在我们的 之前的发现，这次更新旨在通过解决三个关键问题来填补这一知识缺口：(1)BB如何 控制不对称的鞭毛旋转？由于其独特的细胞形状和几何形状，BB必须旋转其两极 周质鞭毛不对称地奔跑：前鞭毛逆时针旋转，后鞭毛逆时针旋转 PF顺时针旋转。如果没有不对称的旋转，细胞就会变形。这是的一个标志性功能 螺旋体的运动；然而，其潜在的分子机制仍然难以捉摸。目标1试图解开这一点 确定非规范鞭毛电机开关FliG1的功能和结构是一个长期存在的难题 蛋白质，使用遗传学、生物化学、冷冻电子断层扫描和结晶学的综合方法。 (2)BB是否进化出成群运动以促进其侵袭性和毒力？在地方病循环期间， 在某些情况下，BB会在高度粘稠的胶状环境中游泳，例如哺乳动物的真皮组织和 扁虱-肠道基底膜，这让人想起细菌聚集的环境，一种 允许细菌在固体和半固体表面上爬行的运动。据推测，BB已经 进化出了成群的运动能力来增强它的侵入性。Aim 2计划勾勒出 用遗传学、生物化学和生物化学等综合方法，研究群居运动及其在BB致病中的作用。 结构生物学、活体动物模型以及活体成像。(3)CheA1对BB毒力有控制作用吗 如果是这样，又是如何做到的呢？BB进化出了独特的趋化性，以适应其独特的运动性和地方性循环，例如， 它的基因组编码多种趋化蛋白，如两个CheA组氨酸激酶(HK)：CheA1和CheA2。一个 长期以来的问题是，为什么BB需要多种趋化蛋白。对BB来说，CheA2是必需的，而不是CheA1 趋化性。CheA1的作用仍不清楚。有趣的是，我们最近发现Bb需要CheA1 小鼠血行播散与BB几个关键毒力因子的表达建立在这些基础上 结果，目的3旨在阐明CheA1在BB致病中的作用和潜在的分子机制。 使用遗传学、生物化学、RNA序列和动物模型的多学科方法。实现这些目标 AIMS将有助于更好地理解BB的运动性和趋化性，以及它们在 莱姆病的发病机制。此外，将获得的基本知识具有很大的影响力，并有可能 也有助于了解其他致病螺旋体的这些过程。

SUMMARY Lyme disease bacterium Borrelia burgdorferi (Bb) is highly motile and can traverse complex environments inside mammalian and arthropod hosts during its infectious cycle. The central hypothesis of this application is that the motility and chemotaxis of Bb constitute a distinct paradigm and play a pivotal role in the host-vector cycle as well as in the disease process, including invasion, dissemination, tissue tropism, and immune evasion. During the last two funding cycles, we revealed several unique aspects of Bb motility and chemotaxis; however, their underlying molecular mechanisms and precise roles in the disease process remain largely unknown. Building upon our previous findings, this renewal aims to fill this knowledge gap by addressing three key questions: (1) How does Bb control asymmetrical flagellar rotation? Due to its unique cell shape and geometry, Bb must rotate its bipolar periplasmic flagella (PF) asymmetrically in order to run: the anterior PF rotates counterclockwise, and the posterior PF rotates clockwise. Without asymmetrical rotation, the cells become distorted. This is a hallmark feature of spirochete motility; however, its underlying molecular mechanism remains elusive. Aim 1 seeks to unravel this longstanding conundrum by determining the function and structure of FliG1, a noncanonical flagellar motor switch protein, using an integrative approach of genetics, biochemistry, cryo-electron tomography, and crystallography. (2) Has Bb evolved swarming motility to facilitate its invasiveness and virulence? During the enzootic cycle, on several occasions, Bb swims in highly viscous gel-like environments, such as mammalian dermis tissue and the tick-gut basement membrane, which are reminiscent of the environments in which bacteria swarm, a form of movement that allows bacteria to crawl over solid and semi-solid surfaces. It has been speculated that Bb has evolved swarming motility to empower its invasiveness. Aim 2 plans to delineate the underlying mechanism of swarming motility and its role in the pathogenicity of Bb, using a comprehensive approach of genetics, biochemistry, structural biology, and in vivo animal models along with intravital imaging. (3) Does CheA1 control Bb virulence and, if so, how? Bb has evolved unique chemotaxis to accommodate its distinct motility and enzootic cycle, e.g., its genome encodes multiple chemotaxis proteins such as two CheA histidine kinases (HK): CheA1 and CheA2. A longstanding question is why Bb needs multiple chemotaxis proteins. CheA2, but not CheA1, is essential for Bb chemotaxis. The role of CheA1 remains unknown. Interestingly, we recently found that CheA1 is required for Bb hematogenous dissemination in mice and expression of several key virulence factors of Bb. Building upon these results, Aim 3 proposes to elucidate the role and underlying molecular mechanism of CheA1 in Bb pathogenicity, using a multidisciplinary approach of genetics, biochemistry, RNA-seq, and animal models. Achievement of these aims will lead to a better understanding of Bb motility and chemotaxis as well as their precise roles in the pathogenesis of Lyme disease. Moreover, the fundamental knowledge to be gained is highly impactful and likely to aid understanding of these processes in other pathogenic spirochetes as well.