RpoS Regulation of Borrelia burgdorferi Genes in vivo

描述（由申请人提供）：莱姆病螺旋体（Bb）通过通常涉及野生啮齿动物和硬蜱的地方病循环在自然界中维持。在目前的资金间隔期间，我们已经使用了不断扩大的模型系统和遗传工具，以确定参与哺乳动物宿主适应的Bb基因的生理和毒力相关功能，以及螺旋体从节肢动物载体到哺乳动物宿主的过渡机制。这项工作的大部分集中在描绘的相对贡献的RPOS依赖和独立的转录途径的过程，使螺旋体从蜱到哺乳动物的过境。基于微阵列的转录谱的野生型和rpoS突变体的螺旋体生长在透析膜室（DMC），使我们能够定义在体内RpoS调节子，交替西格玛因子RpoS控制的基因的队列，积极和消极的，在响应哺乳动物宿主来源的信号。通过从这些和相关发现外推到感染的若虫蜱的情况，我们已经制定了我们的中心假设：从血餐开始，RpoS充当“看门人”，协调建立哺乳动物感染所需的差异表达疏螺旋体基因的子集的相互上调和抑制。作为这一假设的推论，我们提出，“RpoS关闭”状态代表了螺旋体建立蜱阶段的地方性生活史所需的替代发展计划。我们的研究计划的主要长期目标是整合寻找新的Bb毒力决定因素，在蜱或小鼠的功能，努力破译的机制，根据LD螺旋体的差异基因表达。为了实现这一目标，我们制定了三个具体目标：（i）进一步表征体内RpoS调节子内的基因，我们假设这些基因由血粉诱导并促进哺乳动物宿主的感染;（ii）进一步表征需要RpoS进行体内抑制的基因，我们假设这些基因是螺旋体生命周期的蜱阶段所需的;和（iii）通过描绘其在地方性流行病周期中的“开”和“关”状态，并检查RpoS功能的选定方面，来发展对RpoS调节子的更广泛的理解。

DESCRIPTION (provided by applicant): Borrelia burgdorferi (Bb), the Lyme disease (LD) spirochete, is maintained in nature via an enzootic cycle which typically involves wild rodents and Ixodes ticks. During the current funding interval, we have used an expanding armamentarium of model systems and genetic tools to define the physiological and virulence-related functions of Bb genes involved in mammalian host adaptation as well as the mechanisms that underlie the spirochete's transition from arthropod vector to mammalian host. Much of this work has centered about delineating the relative contributions of the RpoS-dependent and -independent transcriptional pathways to the processes which enable spirochetes to transit from tick to mammal. Microarray-based transcriptional profiling of wild-type and rpoS mutant spirochetes grown in dialysis membrane chambers (DMCs) has enabled us to define the in vivo RpoS regulon, the cohort of genes that the alternate sigma factor RpoS controls, both positively and negatively, in response to mammalian host-derived signals. By extrapolating from these and related findings to the scenario of the infected nymphal tick, we have formulated our central hypothesis: beginning with the taking of the blood meal, RpoS acts as a "gatekeeper" that coordinates the reciprocal upregulation and repression of a subset of differentially expressed borrelial genes required to establish mammalian infection. As a corollary to this hypothesis, we propose that the "RpoS off' state represents an alternate developmental program required by the spirochete to establish the tick-phase of the enzootic life cycle. The principal long-term objective of our Research Plan is to integrate the search for novel Bb virulence determinants that function within the tick or the mouse with efforts to decipher the mechanisms that underlie differential gene expression by the LD spirochete. To accomplish this objective, we have formulated three Specific Aims: (i) to further characterize genes within the in vivo RpoS regulon which we hypothesize are induced by the blood meal and promote infection of the mammalian host; (ii) to further characterize genes which require RpoS for repression in vivo and which we hypothesize are required for the tick- phase of the spirochete's life cycle; and (iii) to develop a broader understanding of the RpoS regulon by delineating its "on" and "off" states during the enzootic cycle and examining selected facets of RpoS function.