Regulatory Pathways in Borrelial Pathogenesis

项目摘要 莱姆病的病原体伯氏疏螺旋体（Borrelia burgdorferi，Bb）是世界上主要的节肢动物传播的感染。 美国每年确诊病例超过47万例。鉴于莱姆病是一个重要的公众 健康问题，研究Bb如何建立和维持感染对于评估其 致病潜力和发展预防或治疗策略。众所周知，Bb修改了它的 在其致病生命周期中，特别是在从节肢动物传播过程中， 传染给哺乳动物。BosR是一种重要的转录调控因子，作为一种全局性的金属调控因子发挥作用。 蛋白质，并改变哺乳动物殖民所需的基因的表达。具体而言，需要BosR 用于小鼠实验感染，并且需要激活RpoS/RpoN/Rrp2级联， 遗传上不连锁的基因的表达，包括Bb感染所需的毒力决定因子。而 毫无疑问，rpoS的BosR依赖性调节对疏螺旋体的感染性是重要的， BosR直接或间接调节的其他基因可能是Bb感染的因素。BosR如何 与这一复杂的监管网络有关的问题尚未得到彻底解决。在这里，几个新的BosR 描述了迄今未知的调节功能，包括BosR对RNA的识别。的 初步数据显示，BosR与小的非编码RNA结合，并表现出伴侣活性，表明 BosR与sRNA形成复合物，与Bb转录物结合，并靶向它们进行降解或增强， 翻译. BosR的这种转录后调控形式代表了一种新的和创新的调控机制。 该方案增加了其已知的DNA结合活性。在目标1中，BosR结合的sRNA的子集将是 评估，转录识别的BosR：：sRNA复合物确定，并与BosR相关的链接， sRNA对疏螺旋体发病机制的影响。在目标2中，将挖掘最近的ChIP-seq数据以评估新的BosR DNA结合。初步结合研究证实了两个靶点的ChIP-seq数据，并表明 BosR的氧化还原状态和金属配位可能改变DNA结合。将分析BosR DNA结合， 确定BosR的氧化还原和金属结合状态如何改变识别的靶序列。独特 将跟踪BosR结合基序，并确定共有序列。最后，实验将 确定异位产生RpoS的bosR突变体的表型， Bb感染中BosR的表达。总之，这项建议将提供新的评价如何博斯R介导的 转录和转录后调控england和新的见解如何BosR整合氧化还原 Bb状态影响核酸结合和疏螺旋体发病机制。

PROJECT SUMMARY The etiologic agent of Lyme Disease, Borrelia burgdorferi (Bb), is the leading arthropod-borne infection in the United States with over 470,000 cases diagnosed annually. Given that Lyme Disease is a significant public health concern, studies addressing how Bb establishes and maintains infection are vital to evaluating its pathogenic potential and developing prevention or therapeutic strategies. It is well known that Bb modifies its transcriptional profile throughout its pathogenic lifecycle, particularly during transmission from the arthropod vector into mammals. BosR is an important transcriptional regulator that functions as a global metalloregulatory protein and alters the expression of genes needed for mammalian colonization. Specifically, BosR is required for experimental infection in mice and is needed for the activation of the RpoS/RpoN/Rrp2 cascade that drives the expression of genetically unlinked genes, including virulence determinants required for Bb infection. While there is no doubt that the BosR-dependent regulation of rpoS is important to borrelial infectivity, there are additional genes that BosR regulates either directly or indirectly that may factor into Bb infection. How BosR is linked to this complex regulatory network has not been thoroughly addressed. Here, several novel BosR regulatory functions are described, heretofore unknown, including the recognition of RNA by BosR. The Preliminary Data shows that BosR binds to small non-coding RNA and exhibits chaperone activity suggesting that BosR forms a complex with sRNAs that binds to Bb transcripts and targets them for degradation or enhanced translation. This form of post-transcriptional regulation by BosR represents a new and innovative regulatory scheme that adds to its known DNA binding activity. In Aim 1, a subset of sRNAs that BosR binds will be evaluated, the transcripts recognized by the BosR::sRNA complex identified, and a link with the BosR-associated sRNAs to borrelial pathogenesis established. In Aim 2, recent ChIP-seq data will be mined to assess novel BosR DNA binding. Preliminary binding studies confirmed the ChIP-seq data for two targets and suggested that the redox status and metal coordination of BosR may alter DNA binding. BosR DNA binding will be analyzed to determine how the redox and metal binding status of BosR changes the target sequences recognized. Unique BosR binding motifs will be tracked, and a consensus sequence ascertained. Finally, experiments will be conducted to determine the phenotype of a bosR mutant that ectopically produces RpoS to query the direct role of BosR in Bb infection. Taken together, this proposal will provide new appreciation into how BosR-mediated transcriptional and post-transcriptional regulation ensues and novel insight into how BosR integrates the redox status of Bb to affect nucleic acid binding and borrelial pathogenesis.