A Community Mycobacterial Systems Resource

描述(申请人提供)：分枝杆菌疾病，主要是结核病，每年慢性感染数十亿人，导致近200万人死亡。这场全球危机因治疗不当而加剧：卡介苗的疗效令人怀疑，结核分枝杆菌多药和极端耐药菌株的发病率正在惊人地增加。我们将应用基因组学和高通量技术来开发分枝杆菌系统资源(MSR)，这将极大地加速整个分枝杆菌社区的研究。MSR将免费提供，并将提供所有分枝杆菌共同的生物学过程的全面、集成的基因组和可视总结，这些过程可以立即应用于加深我们对分枝杆菌疾病的理解。从MSR中获得的见解将产生大量新的生物学和比较工具，并确定许多适合开发新型抗分枝杆菌疗法的新靶点。因此，MSR将刺激分枝杆菌研究社区，并促进将基本发现转化为对抗造成巨大全球健康负担的物种。这些目标将通过利用聚集在一起组成这个多学科、多机构团队的研究人员可获得的综合专业知识和尖端技术来实现。致病分枝杆菌生长极其缓慢，使用起来非常危险，因此不适合进行高通量基因组分析。我们将通过使用一个密切相关的物种来避免这个问题，耻垢分枝杆菌是一种快速生长的非致病性分枝杆菌，是公认的遗传易驯化的分枝杆菌模型。MSR的焦点将是~1,300个耻垢分枝杆菌编码和非编码基因，这些基因在结核分枝杆菌复合体成员、鸟型分枝杆菌、溃烂分枝杆菌和麻风分枝杆菌之间高度保守。这些高度保守的基因将调节基本的细胞过程，因此它们的分析将适用于所有分枝杆菌物种。MSR将有两个组成部分：物理数据库和电子数据库(或称eSource)。物理资源将包括1,300个高度保守的编码和非编码分枝杆菌基因的定向删除或敲除，以及一组编码基因与功能标签融合的表达载体，以便于未来的分析。ESource将描述在物理资源中创建的菌株和质粒的各种初始表型特征。这些包括蛋白质定位和细胞形态的显微分析，与抗分枝杆菌药物敏感性相关的基因的鉴定，以及影响细胞表面-环境界面的基因。这些研究产生的数据将汇编并整合到在线结核病数据库中，并将免费提供给科学界。总而言之，MSR将为分枝杆菌生物学提供前所未有的洞察力，并将成为分枝杆菌社区研究人员未来研究的发射台。 公共卫生相关性：分枝杆菌疾病长期感染全球数十亿人口，每年导致近200万人死亡，艾滋病毒和结核分枝杆菌之间的协同作用以及结核分枝杆菌多药耐药(MDR)和极端耐药(XDR)菌株的出现加剧了这一危机。对分枝杆菌生物学的全面了解对于确定发病机制、识别新的药物靶点和开发疫苗至关重要。这项提议将应用基因组学和其他高通量技术来建立分枝杆菌系统资源，该资源将提供所有分枝杆菌共同的生物学过程的全面、完整的基因组总和，这将极大地加速整个分枝杆菌社区的研究。

DESCRIPTION (provided by applicant): Mycobacterial diseases, predominantly tuberculosis, chronically infect billions and kill nearly two million people annually. This global crisis is exacerbated by inadequate treatments: the BCG vaccine is of questionable efficacy, and the incidence of multi-drug and extremely-drug resistant strains of M. tuberculosis is increasing alarmingly. We will apply genomics and high throughput technologies to develop a Mycobacterial Systems Resource (MSR) that will dramatically accelerate research throughout the entire mycobacterial community. The MSR will be freely available and will provide a comprehensive, integrated genomic and visual summation of biological processes common to all mycobacteria that can be immediately applied to further our understanding of mycobacterial diseases. Insights gained from the MSR will generate a plethora of new biological and comparative tools and identify many new targets suitable for the development of novel anti- mycobacterial therapeutics. Thus, the MSR will stimulate the mycobacterial research community, and facilitate the translation of basic discoveries to combat species that cause an enormous global health burden. These goals will be achieved by taking advantage of the combined expertise and cutting-edge technologies available to the researchers assembled to form this multi-disciplinary, multi-institutional team. Pathogenic mycobacteria are extremely slow-growing and hazardous to work with, making them inappropriate for high-throughput genomic analyses. We will obviate this problem by using a closely related species, M. smegmatis, which is a fast-growing, non-pathogenic mycobacterium that is the acknowledged genetically tractable model Mycobacterium. The focus of the MSR will be ~1,300 M. smegmatis coding and non-coding genes that are highly conserved between members of the M. tuberculosis complex, M. avium, M. ulcerans and M. leprae. These highly conserved genes will mediate fundamental cellular processes and therefore their analysis will be applicable to all mycobacterial species. The MSR will have two components: physical and electronic database (or eSource). The physical resource will consist of a targeted deletion or knockdown of ~1,300 highly conserved coding and non-coding mycobacterial genes, and a collection of expression vectors with the coding genes fused to functional tags to facilitate future analyses. The eSource will describe a wide variety of initial phenotypic characterizations of the strains and plasmids created in the physical resource. These include microscopic analyses of protein localization and cellular morphology, identification of genes associated with sensitivity to anti-mycobacterial drugs, and genes influencing the cell's surface-environment interface. The data generated from these studies will be assembled and integrated on the online Tuberculosis Database (TBDB) and will be freely available to the scientific community. In summary, the MSR will provide unprecedented insight into mycobacterial biology and will serve as a launch pad for future studies by researchers spanning the mycobacterial community. PUBLIC HEALTH RELEVANCE: Mycobacterial diseases chronically infect billions of the global populace and kill nearly two million annually, a crisis that is exacerbated by the synergistic association of HIV and M. tuberculosis, and the appearance of multi-drug (MDR) and extremely-drug (XDR) resistant strains of M. tuberculosis. A comprehensive understanding of the biology of mycobacteria is critical for the characterization of pathogenesis, the identification of novel drug targets, and the development of vaccines. This proposal will apply genomics and other high throughput technologies to establish a Mycobacterial Systems Resource that will provide a comprehensive, integrated genomic summation of biological processes common to all mycobacteria, which will dramatically accelerate research throughout the entire mycobacterial community.