Defining the functions of uncharacterized genes in priority pathogens

定义优先病原体中未表征基因的功能

• 批准号: 8719930
• 负责人: Sean Crosson
• 金额: $ 85.02万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-08-13 至 2018-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8719930
• 关键词: Address; Affect; Animal Disease Models; Animal Model; Animals; Bacillus anthracis; Bacterial Proteins; Biochemical; Biochemistry; Biogenesis; Bioinformatics; Biological; Biological Assay; Biology; Biophysics; Brucella; Brucella abortus; Brucellosis; Categories; Cell physiology; Cells; Centers for Disease Control and Prevention (U.S.); Chicago; Chronic; Code; Collaborations; Communicable Diseases; Communities; Coxiella burnetii; DNA Sequence; Data; Databases; Disease; Fostering; Functional RNA; Genes; Genetic Transcription; Goals; Human; Immune; In Vitro; Inbred BALB C Mice; Individual; Infection; Instruction; Interdisciplinary Study; Investigation; Laboratories; Maintenance; Microbiology; Modeling; Molecular; Mus; National Institute of Allergy and Infectious Disease; Open Reading Frames; Operon; Oxidative Stress; PAWR protein; Pathogenesis; Physiological; Physiology; Principal Investigator; Process; Protein Family; Proteins; Publishing; Reagent; Regulation; Research; Research Infrastructure; Research Personnel; Research Project Grants; Resources; Role; Side; Sigma Factor; Signal Transduction; Stress; Structure; Technology; Testing; Transcriptional Regulation; Universities; Virulence; Yersinia pestis; acid stress; base; biological adaptation to stress; cell envelope; data management; gene function; genetic regulatory protein; genome sequencing; global health; in vivo; member; pathogen; programs; research study; success

DESCRIPTION (provided by applicant): Understanding the functions of the thousands of uncharacterized genes in sequence databases is among the most significant challenges in biology. The primary goal of this proposal is to define the biochemical and physiological functions of 235 uncharacterized genes in NIAID Category A (Bacillus anthracis and Yersinia pestis) and Category B (Brucella abortus and Coxiella burnetii) priority pathogens. These bacterial pathogens are highly infectious and have had a profound effect on global health both historically and currently. Included in this set are 185 predicted protein coding genes, 59 of which have no evidence of expression. The remaining 50 genes in our set are expressed, or are predicted to be expressed as small, non-coding RNAs (sRNAs). Our state-of-the-art research infrastructure at the Howard Taylor Ricketts Regional Biocontainment Laboratory (HTRL) and Argonne National Laboratory (ANL), and our integrated expertise in biochemistry and microbiology make us an ideal team for this interdisciplinary gene function assignment project. Our proposed research program will 1) yield important data on the function of uncharacterized genes in priority pathogens, 2) define the role of numerous uncharacterized genes in processes tied to bacterial pathogenesis, and 3) will produce useful biological and biochemical reagents for the pathogen research community. RELEVANCE: A full understanding of pathogen biology requires an understanding of how pathogen genes function at a biochemical level. Experiments proposed herein will define the biochemical function of uncharacterized genes implicated in disease processes in four NIAID priority pathogens. Project 1: Functional and Biochemical Characterization of B. abortus Stress Response Genes Project Leader: Sean Crosson DESCRIPTION: The immediate goal of this project is to define the cellular and biochemical functions of 57 uncharacterized open reading frames (ORFs), hypothetical proteins, and small non-coding RNAs in B. abortus. Our preliminary data provide evidence that these genes have a functional role in general stress adaptation and chronic mammalian infection. General stress response (GSR)-dependent transcription in B. abortus is controlled by the sigma factor SigE1. Our preliminary investigation of SigE1 and its upstream signaling partner PhyR have demonstrated that these regulatory proteins are required for adaptation to oxidative and acid stress in vitro and for maintenance of chronic infection in a BALB/c murine model. We have experimentally defined a set of 108 genes that are directly or indirectly regulated by SigE1. Within this regulated gene set are 32 uncharacterized open reading frames (ORFs) and nine small non-coding RNAs. Additionally, we have identified 16 hypothetical genes that are adjacent to or in apparent operons with SigE1-regulated ORFs but for which we have no evidence of expression. We propose to: 1) Test strains in which these genes have been deleted in oxidative and acid stress survival assays, and in cell-based and animal infection assays, 2) elucidate the biochemical functions of these hypothetical proteins and SigE1-regulated ORFs in collaboration with our Biochemical Function Technology Core, and 3) characterize the biochemical function of putative non-coding RNAs under transcriptional control of SigE1. These experiments will provide a comprehensive (in vivo and in vitro) functional understanding of conserved and non-conserved B. abortus genes that are currently uncharacterized. RELEVANCE: Brucella spp.are the causative agents of brucellosis, which is among the most common zooneses globally. Due to their high infectivity, easy aerosolization, and debilitating effects on infected individuals. Brucella spp have been classified as a category B bioterror threat by the U.S. Centers for Disease Control (CDC). Experiments proposed herein will define the biochemical function of uncharacterized genes implicated in regulation of B. abortus stress physiology and in the control of chronic brucellosis disease.

描述（申请人提供）：在序列数据库中了解数千个未表征基因的功能是生物学中最重要的挑战之一。该提案的主要目的是定义NIAID类别中235个未表征基因的生化和生理功能A（炭疽芽孢杆菌和鼠疫）和B类（Brucella abortus and Coxiella burnetii）优先病原体。这些细菌病原体具有很高的感染力，并且在历史上和目前都对全球健康产生了深远的影响。该组中包括185个预测的蛋白质编码基因，其中59个没有表达的证据。我们集合中的其余50个基因被表达，或者被预测为小的非编码RNA（SRNA）。 我们在霍华德·泰勒·里基茨（Howard Taylor Ricketts 实验室（HTRL）和Argonne国家实验室（ANL），以及我们在生物化学和微生物学方面的综合专业知识使我们成为该跨学科基因功能分配项目的理想团队。我们提出的研究计划将1）产生有关优先病原体中未表征基因功能的重要数据，2）确定许多未表征基因在与细菌发病机理相关的过程中的作用，而3）将产生有用的生物学和生物化学试验，用于病原体研究社区。 相关性：对病原体生物学的充分理解需要了解病原体基因在生化水平上的功能。本文提出的实验将定义与四个NIAID优先病原体中疾病过程有关的未表征基因的生化功能。 项目1：B. b. bortus应激反应基因的功能和生化表征 项目负责人：肖恩·克罗森（Sean Crosson） 描述：该项目的直接目标是定义57个未表征的开放式阅读框（ORF），假设蛋白质和小型非编码RNA的细胞和生化功能。我们的初步数据提供了证据，表明这些基因在一般应激适应和慢性哺乳动物感染中具有功能作用。 abortus中的一般应力反应（GSR）依赖性转录受Sigma因子SIGE1控制。我们对SIGE1及其上游信号合作伙伴Phyr的初步研究表明，这些调节蛋白是在体外适应氧化和酸应激所必需的，并在BALB/C鼠模型中维持慢性感染。我们在实验上定义了一组108个基因，这些基因是由SIGE1直接或间接调节的。在这个受调节的基因集中，有32个未表征的开放式阅读框（ORF）和9个小型非编码RNA。此外，我们已经确定了16种与SIGE1调节的ORF相邻或明显操纵子中的假设基因，但我们没有表达的证据。 We propose to: 1) Test strains in which these genes have been deleted in oxidative and acid stress survival assays, and in cell-based and animal infection assays, 2) elucidate the biochemical functions of these hypothetical proteins and SigE1-regulated ORFs in collaboration with our Biochemical Function Technology Core, and 3) characterize the biochemical function of putative non-coding RNAs under transcriptional control of SIGE1。这些实验将对当前未表征的保守和未经保守的Abortus基因提供全面的（体内和体外）功能理解。 相关性：Brucella spp。是布鲁氏菌病的致病药物，这是全球最常见的世界之一。由于它们的高感染力，易于气化和对感染者的衰弱作用。 Brucella SPP已被归类为B类BiotError威胁 美国疾病控制中心（CDC）。本文提出的实验将定义涉及对Abortus B. bortus胁迫生理学和控制慢性布鲁氏菌病疾病的调节基因的生化功能。