Host range determinants of bacterial exfoliative toxins

项目摘要 病原体毒力因子的宿主范围或嗜性是感染的关键决定因素。一 需要详细了解控制物种向性的宿主和病原体突变， 评估未来人畜共患病暴发的风险，改进动物感染模型， 设计新的疗法，利用宿主特异性。 该提案的目的是确定细菌中跨物种活性的障碍， 高分辨率的毒力因子，利用葡萄球菌脱落毒素作为研究系统。 由葡萄球菌属致病菌编码的剥脱毒素导致生命- 威胁性皮肤病，包括葡萄球菌烫伤样皮肤综合征和大疱性脓疱病 以大面积起泡和表皮损伤为特征。角质毒素是 通过选择性切割皮肤桥粒钙粘蛋白桥粒芯糖蛋白-1起作用的蛋白酶，导致 表皮屏障功能丧失和水疱形成。我们的核心假设是 因子活性取决于宿主和病原体之间的遗传相容性， 探究这些相容性将揭示宿主向性的特定障碍。在初步工作中， 我们发现编码桥粒芯糖蛋白-1的基因经历了快速的进化和重复的自然进化， 在足以限制毒素活性的小蛋白质表面内跨非人灵长类动物进行选择。 我们还开发了易于处理的体外测定来测量重组蛋白的毒素切割。 desmglein-1从不同的宿主物种。 该提案的具体目标是：1）生成高分辨率的突变图， 限制毒素向性的桥粒芯糖蛋白-1，和2）测试在桥粒芯糖蛋白-1结合处的毒素突变如何 界面有助于寄主向性。在目标1中，我们将进行毒素的组合诱变 在桥粒芯糖蛋白-1的识别表面，以产生定义宿主的屏障的遗传图谱 识别.在目标2中，我们将应用结构和生物化学方法来解析桥粒芯糖蛋白- 脱落毒素结合界面与遗传学相结合，以评估毒素突变如何在此 表面控制寄主向性。总的来说，这一建议将建立一个框架，以定义遗传 高分辨率的细菌感染屏障，应用于预测和 防止未来疾病爆发。

PROJECT SUMMARY The host range, or tropism, of pathogen virulence factors is a key determinant of infection. A detailed understanding of host and pathogen mutations that control species tropism is required in order to assess the risk of future zoonotic disease outbreaks, improve animal infection models, and design new therapeutics that take advantage of host specificity. The objective of this proposal is to define barriers to cross-species activity in bacterial virulence factors at high resolution, leveraging staphylococcal exfoliative toxins as a study system. Exfoliative toxins encoded by pathogenic bacteria in the genus Staphylococcus cause life- threatening skin diseases including staphylococcal scalded skin syndrome and bullous impetigo characterized by widespread blistering and damage to the epidermis. Exfoliative toxins are proteases that act by selectively cleaving the skin desmosomal cadherin desmoglein-1, leading to loss of epidermal barrier function and blister formation. Our central hypothesis is that virulence factor activity is dependent on genetic compatibilities between hosts and pathogens, and that interrogating these compatibilities will uncover specific barriers to host tropism. In preliminary work, we found that genes encoding desmoglein-1 have undergone rapid evolution and repeated natural selection across non-human primates within a small protein surface sufficient to restrict toxin activity. We have also developed tractable in vitro assays to measure toxin cleavage of recombinant desmglein-1 from various host species. The Specific Aims of this proposal are to 1) generate a high-resolution map of mutations in desmoglein-1 that restrict toxin tropism, and 2) test how toxin mutations at the desmoglein-1 binding interface contribute to host tropism. In Aim 1 we will perform combinatorial mutagenesis of the toxin recognition surface in desmoglein-1 to produce a genetic map defining the barriers of host recognition. In Aim 2 we will apply structural and biochemical approaches to resolve the desmoglein- exfoliative toxin binding interface combined with genetics to assess how toxin mutations at this surface control host tropism. Collectively this proposal will establish a framework to define genetic barriers to bacterial infections at high-resolution, applied towards the goal of anticipating and preventing future disease outbreaks.