Comparative and functional genomics of Toxoplasma and Hammondia hammondi

弓形虫和 Hammondia hammondi 的比较和功能基因组学

• 批准号: 8861405
• 负责人: JON P BOYLE
• 金额: $ 37.08万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2015
• 资助国家: 美国
• 起止时间: 2015-06-15 至 2018-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8861405
• 关键词: AIDS/HIV problem; Address; Adult; Animals; Binding Proteins; Biological Assay; Biology; Candidate Disease Gene; Collaborations; Cyst; Data; Development; Disease; Eukaryota; Evolution; Fetus; Funding; Gene Deletion; Gene Expression Profile; Genes; Genetic; Genetic Transcription; Genome; Goals; Growth; Growth and Development function; Guanosine Triphosphate Phosphohydrolases; Histology; Human; Immune; Immune response; Immunocompromised Host; Immunohistochemistry; In Vitro; Individual; Infection; Kinetics; Laboratories; Molecular; Mouse Strains; Mus; Mutation; Oocysts; Orthologous Gene; Parasites; Pathogenesis; Patients; Phenotype; Play; Positioning Attribute; Publishing; Regulator Genes; Relative (related person); Resistance; Role; Serum; Sporozoites; Staging; Synteny; System; Testing; Time; Tissues; Toxoplasma; Toxoplasma gondii; Toxoplasmosis; Transplant Recipients; Vacuole; Virulence; Virulence Factors; Virulent; Work; base; comparative genomics; cytokine; experience; functional genomics; guanylate; in vivo; insight; pathogen; programs; public health relevance; research study; response; success; transcription factor; transcriptome sequencing

 DESCRIPTION (provided by applicant): Toxoplasma gondii is an important opportunistic pathogen of humans where it can cause severe disease in the developing fetus, immunocompromised individuals, and in certain cases healthy adults. In contrast the nearest extant relative of T. gondii, Hammondia hammondi, does not infect humans and is highly avirulent in mice, and experimental infections are characterized by rapid conversion to "terminally differentiated" cyst stages. These in vivo virulence differences are recapitulated in vitro, where H. hammondi spontaneously converts into a terminally differentiated cyst stage. We have extensive published and unpublished preliminary data demonstrating that T. gondii and H. hammondi share >99% of their genes in nearly perfect synteny, and that the T. gondii and H. hammondi transcriptomes during development are similar but have key differences in the transcription of multiple candidate virulence effectors and transcription factors. In the work outlined in this 3 year R01 proposal we use the T. gondii/H. hammondi system to identify the gene regulatory mechanisms that drive T. gondii pathogenesis, with a particular focus on bradyzoite cyst stages that are the leading cause of severe toxoplasmosis in HIV/AIDS and transplant patients. In Aim 1 we use ultra-deep, strand-specific RNAseq at time points that encompass clear developmental transitions during in vitro growth to identify genes and genetic networks that regulated in a species-specific manner. In doing so we will identify 1) putative secreted virulence effectors and 2) transcription factors that are uniquely expressed in one species over the other. We will then determine their role in T. gondii biology using gene deletion and cross-species complementation experiments. Success of this Aim is facilitated by extensive published and preliminary data comparing the genomes, in vitro growth rates, transcription profiles, and host responses to T. gondii and H. hammondi. In Aim 2 we address the role of the host in species-specific growth phenotypes. To do this we will compare 1) infection dynamics of T. gondii and H. hammondi in vivo using parallel histological assays to identify differences in the cellular and humoral immune response and 2) the overall infectivity and virulence of H. hammondi in innate-immune deficient mouse strains that are highly susceptible to T. gondii infection. Through these studies we will identify new T. gondii secreted effectors and transcription factors that underlie the dramatic phenotypic differences between these species, and how these effectors might impact the host response to infection. These studies should also more broadly impact our understanding of how, on a gene- by-gene basis, virulent pathogens emerge from comparatively avirulent phenotypic backgrounds.

 描述（由申请人提供）：弓形虫是人类重要的机会性病原体，它可以在发育中的胎儿、免疫功能低下的个体以及在某些情况下的健康成年人中引起严重的疾病。相比之下，弓形虫的现存近亲哈蒙菌（Hammondia hammondi）不会感染人类，并且在小鼠中具有高度无毒力，并且实验性感染的特征是快速转变为“终末分化”囊肿阶段。这些体内毒力差异在体外重现，哈蒙迪嗜血杆菌自发转变为终末分化的囊肿阶段。我们拥有大量已发表和未发表的初步数据，表明弓形虫和哈蒙迪弓形虫在近乎完美的同线性中共享超过 99% 的基因，并且弓形虫和哈蒙迪弓形虫在发育过程中的转录组相似，但在多个候选毒力效应子和转录因子的转录方面存在关键差异。在此 3 年 R01 提案中概述的工作中，我们使用弓形虫/H。 hammondi 系统来确定驱动弓形虫发病机制的基因调控机制，特别关注缓殖子包囊阶段，这是艾滋病毒/艾滋病和移植患者中严重弓形虫病的主要原因。在目标 1 中，我们在体外生长过程中的明确发育转变的时间点使用超深、链特异性 RNAseq 来识别以物种特异性方式调节的基因和遗传网络。在此过程中，我们将识别 1) 假定的分泌毒力效应子和 2) 在一个物种中相对于另一物种独特表达的转录因子。然后，我们将通过基因删除和跨物种互补实验来确定它们在弓形虫生物学中的作用。这一目标的成功得益于广泛发表的初步数据，这些数据比较了基因组、体外生长率、转录谱以及宿主对弓形虫和哈蒙迪弓形虫的反应。在目标 2 中，我们讨论了宿主在物种特异性生长表型中的作用。为此，我们将使用平行组织学测定来比较 1) 弓形虫和哈蒙迪弓形虫在体内的感染动态，以识别弓形虫和哈蒙迪弓形虫的感染动态。 细胞和体液免疫反应，2) 哈蒙迪弓形虫在先天免疫缺陷小鼠品系中的总体感染性和毒力，这些小鼠品系对刚地弓形虫感染高度敏感。通过这些研究，我们将确定弓形虫分泌的新效应子和转录因子，这些效应子和转录因子是这些物种之间显着表型差异的基础，以及这些效应子如何影响宿主对感染的反应。这些研究还应该更广泛地影响我们对逐个基因地理解剧毒病原体如何从相对无毒力的表型背景中出现的理解。