RpoS Regulation of Borrelia burgdorferi Genes in vivo

描述（由申请人提供）：伯氏疏螺旋体（Bb），莱姆病（LD）螺旋体，通过通常涉及野生啮齿动物和蜱虫的动物循环在自然界中维持。在目前的资助期间，我们使用了不断扩大的模型系统和遗传工具来定义Bb基因在哺乳动物宿主适应中的生理和毒力相关功能，以及螺旋体从节肢动物媒介向哺乳动物宿主转变的机制。这项工作的大部分集中在描述依赖于rpos和独立于rpos的转录途径在使螺旋体从蜱虫转移到哺乳动物的过程中的相对贡献。在透析膜室（DMCs）中生长的野生型和rpoS突变螺旋体的基于微阵列的转录谱分析使我们能够定义体内rpoS调控，这是一组由备用sigma因子rpoS控制的基因，在响应哺乳动物宿主来源的信号时，既有积极的，也有消极的。通过将这些和相关的发现推断到被感染的蜱虫的情况，我们制定了我们的中心假设：从摄取血粉开始，RpoS就像一个“守门人”，协调建立哺乳动物感染所需的一组差异表达的蜱虫基因的相互上调和抑制。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地方性动物生命周期的滴答期所需的另一种发育程序。我们研究计划的主要长期目标是整合对蜱虫或小鼠内起作用的新型Bb毒力决定因素的搜索，努力破译LD螺旋体差异基因表达的机制。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调控中的基因，我们假设这些基因是由血粉诱导的，并促进哺乳动物宿主的感染；（ii）进一步表征在体内需要RpoS抑制的基因，我们假设这些基因在螺旋体生命周期的蜱虫阶段是必需的；（iii）通过描述其在地方性疾病周期中的“开”和“关”状态，并检查RpoS功能的选定方面，对RpoS规则有更广泛的了解。

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.