A systems level analysis of spore coat assembly in Bacillus subtilis

项目总结 几种致病芽胞形成细菌，包括炭疽芽孢杆菌的病原体， 作为孢子被吸入或摄取，并在萌发后在宿主中恢复生长和毒素产生。这个 包裹着孢子的孢子蛋白外壳是侵染循环中的一个重要因素，因为它的 种子萌发过程中的有效保护功能及其调控作用 位于孢子内膜上的受体的萌发物。孢子被是形态上的。 由大约60种不同的产孢蛋白组成的复杂结构，它们围绕着 新生的孢子在这项提案中，我们将使用一种综合的系统生物学方法来描绘蛋白质的相互作用。 网络和确定使孢子壳组装成核的关键调控枢纽-这可能作为 孢子检测和/或在反生物恐怖主义应用中作为控制孢子萌发的目标。在……里面 除了与公共卫生问题相关外，我们预计我们的项目将说明系统的价值 研究复杂结构组装的生物学方法。 我们建议使用高通量的蛋白质定位筛选来表征蛋白质的相互作用 在模式生物枯草芽孢杆菌中控制孢子壳组装的网络。具体来说，我们将使用 枯草杆菌所有外壳蛋白的荧光蛋白融合文库，以确定时空 孢子壳蛋白在孢子周围沉积的层级，以识别哪些形态发生蛋白是 所有孢子外壳蛋白的招募所需的，并学习转录和翻译后 调节涂层组装的潜在机制。两国之间建立的互动的性质 将使用大规模酵母双杂交试验和各种聚焦的方法展示成对的外壳蛋白 生化方法。最后，我们将使用计算工具来整合从这些数据中收集的数据 实验并获得了孢子外壳蛋白相互作用网络的全面表示 作为研究其他芽胞形成细菌，特别是炭疽杆菌的模板。 1个项目说明 我们的研究将得出新的抗药性假说和机制 芽孢杆菌孢子的性质和控制孢子萌发的机制。我们 预计我们的结果将对治疗方法的设计产生影响 检测和消除病原性芽胞形成细菌。

Project summary Several pathogenic spore-forming bacteria, including the causative agent of anthrax Bacillus anthracis, are inhaled or ingested as spores and resume growth and toxin production in the host after germination. The spore protein coat, which encapsulates the spore, is an important factor in the infectious cycle, because of its effective protective function and its regulatory role in the germination process by controlling access of germinants to receptors located in the inner membrane of the spore. The spore coat is a morphologically complex structure composed of approximately 60 different sporulation proteins that assemble around the nascent spore. In this proposal, we will use an integrated systems biology approach to map protein interaction networks and identify key regulatory hubs that nucleate spore coat assembly- which may serve as markers for spore detection and/or as targets for control of spore germination in applications against bioterrorism. In addition to its relevance to public health issues, we envision that our project will illustrate the value of systems biology approaches for investigating the assembly of complex structures. We propose to use high throughput protein localization screens to characterize the protein interaction networks that govern spore coat assembly in the model organism Bacillus subtilis. Specifically, we will use a library of fluorescent protein fusions to all of the coat proteins in B. subtilis, to define the spatiotemporal hierarchy of deposition of spore coat proteins around the spore, to identify which morphogenetic proteins are required for the recruitment of all spore coat proteins and learn the transcriptional and post-translational mechanisms underlying the regulation of coat assembly. The nature of the interactions established between pairs of coat proteins will be demonstrated using a large scale yeast two hybrid assay and various focused biochemical approaches. Finally, we will use computational tools to integrate the data gathered from these experiments and obtain a comprehensive representation of the spore coat protein interaction network that will serve as a template for the study of other spore-forming bacteria, particularly B. anthracis. 1 Project narrative Our research will derive new hypotheses and mechanisms for the resistance properties of Bacillus spores and the mechanisms that control spore germination. We anticipate that our results will have an impact on the design of therapeutic approaches to detect and eliminate pathogenic spore-forming bacteria.