The role of small non-coding RNA in borrelial pathogenesis

 描述（由申请人提供）：莱姆病或莱姆病疏螺旋体病的病原体，伯氏疏螺旋体，在美国产生比任何其他病原体更多的节肢动物传播的感染，每年诊断出超过30万例。在B.伯氏菌感染是普遍的，与这种感染相关的发病率很高，并且主要以心脏、神经或关节炎后遗症的形式出现。虽然B.相对于大多数生命系统，burgdorferi具有简单的基因组，当病原体从节肢动物载体和哺乳动物物种移动时，它善于改变其基因调控模式。虽然我们了解几种转录调节因子在这种适应性反应中的作用，但对于一些疏螺旋体基因还有额外的调节层（例如，bosR和dbpA）。考虑到它们在许多细菌系统中作为重要的调节效应物的出现，包括与许多病原体中的毒力基因表达和蛋白质产生的联系，我们假设小的非编码RNA（sRNA）在B中需要类似的调节功能。burgdorferi。使用协作方法，B。 利用与Tn-seq偶联的burgdorferi Tn文库以及RNA-seq技术，鉴定了14种在实验感染后减毒的推定sRNA突变体。这些相同的位点编码先前通过总转录物谱的RNA-seq以及sRNA特异性文库鉴定的sRNA。随后感染个别Tn突变体在这些网站表明，4突变体的能力受损，他们的殖民小鼠。这项建议旨在 在sRNA与它们通过改变靶mRNA调节基因表达和/或蛋白质产生的作用之间建立联系。为了解决这个问题，我们提出了以下具体目标：（1）将sRNA种类与疏螺旋体感染性联系起来。将测试被设计为消除特定sRNA的精确突变体，以确定它们在空间和时间方式的体内疏螺旋体感染性中的作用;和（2）确定选择疏螺旋体sRNA消耗的全局调节作用。将使用全局转录物和蛋白质谱来跟踪特定sRNA的损失的影响，以确定sRNA如何影响未连接基因的转录或翻译。本文概述的工作提供了sRNA调控途径和B之间的联系。伯氏感染迄今为止，这种联系尚未深入分析，因此，这代表了确定sRNA如何调节影响疏螺旋体生理学和发病机制的转录或转录后调节途径的机会。

 DESCRIPTION (provided by applicant): The agent of Lyme disease or Lyme borreliosis, Borrelia burgdorferi, yields more arthropod-borne infections than any other pathogen in the United States with over 300,000 cases diagnosed each year. In areas where B. burgdorferi infections are prevalent, morbidity associated with this infection is high and is seen mostly in th form of cardiac, neurologic, or arthritic sequelae. Although B. burgdorferi has a simplistic genome relative to most living systems, it is adept at altering its gene regulatory pattern as the pathogen moves from the arthropod vector and mammalian species it can occupy during its lifecycle. Although we understand the role of several transcriptional regulators in this adaptive response, there are additional layers of regulation for some borrelial genes (e.g., bosR and dbpA) that have not been elucidated. Given their emergence as important regulatory effectors in many bacterial systems, including a linkage to virulence gene expression and protein production in a number of pathogens, we hypothesized that small, non-coding RNAs (sRNAs) were required for similar regulatory functions in B. burgdorferi. Using a collaborative approach, the B. burgdorferi Tn library coupled with Tn-seq, as well as RNA-seq technologies, identified 14 putative sRNA mutants that were attenuated following experimental infection. These same sites encoded sRNA previously identified via RNA- seq of total transcript profiles as well as libraries specific for sRNAs. Subsequent infections with individual Tn mutants at these sites indicated that 4 of the mutants were impaired in their ability to colonize mice. This proposal is designed to establish a link between the sRNAs and their role in regulating gene expression and/or protein production via alteration of target mRNAs. To address this we propose the following Specific Aims: (1) Link sRNA species to borrelial infectivity. Precise mutants, designed to eliminate specific sRNAs, will be tested to determine their role in borrelial infectivity in vivo in both a spatial and temporal manner; and (2) Determine the global regulatory effect of select borrelial sRNA depletion. The effect of the loss of specific sRNAs will be tracked using global transcript and protein profiles to determine how the sRNA affects transcription or translation of unlinked genes. The work outlined herein provides a link between sRNA regulatory pathways and B. burgdorferi infection. To date, this connection has not analyzed in depth and, as such, represents an opportunity to determine how sRNA modulate either transcriptional or post- transcriptional regulatory pathways that impact borrelial physiology and pathogenesis.