High throughput proteomics to dissect Chlamydia-host cell interactions

高通量蛋白质组学剖析衣原体-宿主细胞相互作用

• 批准号: 8491133
• 负责人: Joanne N. Engel
• 金额: $ 22.16万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-17 至 2015-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8491133
• 关键词: Affinity; Affinity Chromatography; Algorithms; Antibiotics; Bacteria; Bacterial Proteins; Binding; Biochemical Genetics; Bioinformatics; Biological; Biology; Blindness; Cancer Biology; Cell Communication; Cells; Cellular biology; Chlamydia; Chlamydia Infections; Chlamydia trachomatis; Chlamydophila pneumoniae; Chronic Disease; Communicable Diseases; Complex; Country; Data; Detection; Developing Countries; Developmental Biology; Diagnostic; Disease; Environment; Escherichia coli; Eukaryotic Cell; Event; Genes; Genetic; Genome; Heart Diseases; Human; Immune response; Immune system; Infection; Infertility; Integration Host Factors; Invaded; Lead; Life Cycle Stages; Lysosomes; Malignant neoplasm of lung; Mass Spectrum Analysis; Membrane; Microbe; Molecular; Monitor; Nutrient; Organelles; Pathogenesis; Pathway interactions; Pharmaceutical Preparations; Phosphorylation; Post-Translational Protein Processing; Prevalence; Prevention; Process; Protein-Protein Interaction Map; Proteins; Proteome; Proteomics; Recruitment Activity; Research; Respiratory Tract Infections; Role; Saccharomycetales; Sexually Transmitted Diseases; Surface; System; Transfection; Vaccines; base; cost effective; genetic manipulation; glycosylation; human disease; in vivo; innovation; insight; novel; obligate intracellular parasite; pathogen; protein function; protein protein interaction; public health relevance; response; therapeutic vaccine; trafficking; treatment strategy

DESCRIPTION (provided by applicant): Chlamydia species are an important cause of human disease for which no vaccine exists. Standard genetic approaches are not possible to employ with this obligate intracellular parasite, hampering our ability to decode its molecular pathogenesis. Recent transformative studies have revealed that despite its reduced genome size, on the order of 1000 genes, Chlamydia secretes well over 100 effector proteins into the host cell. These effectors function to selectively recruit organelles, acquire nutrients, manipulat host cell trafficking pathways, and to evade detection from the host immune system. Systematically characterizing the host proteins that are physically hijacked by pathogens that invade and replicate within the mammalian host cell is key to understanding their biology. Previous studies in simpler systems, including budding yeast and Escherichia coli, have demonstrated that such an endeavor is incredibly powerful with respect to uncovering novel biological insights. We propose two specific aims in which we will use state-of-the art high-throughput mass spectroscopy in conjunction with newly developed bioinformatic algorithms to (i) comprehensively identify host proteins that interact with secreted chlamydial effectors and to (ii) globally identify host post-translational modifications in response to chlamydial infections, including overall changes in the host proteome as well as changes in host protein phosphorylation, ubiquitylation, and glycosylation events. Aim 1. We will globally identify interactions between secreted C. trachomatis proteins and their host cell target proteins by affinity purification/mass spectrometry (AP/MS) to develop a comprehensive C. trachomatis-host protein-protein interaction (PPI) map. Aim 2. We will use an unbiased approach to globally identify changes in host post-translational modifications, including host protein phosphorylation, ubiquitylation and glycosylation, in response to C. trachomatis infection. Unraveling these complex events using these innovative approaches will yield important clues into the pathogenesis of Chlamydia respiratory infections, the role of Chlamydia infections in atherosclerotic heart disease, and the role of Chlamydia infections in lung cancer. The studies may provide new targets for diagnostics, therapeutics, and vaccines. In addition, these studies will provide novel insights into fundamental process in eukaryotic cell biology, with implications ranging from developmental biology to cancer biology.

描述（由申请人提供）：衣原体物种是不存在疫苗的人类疾病的重要原因。不可能使用标准的遗传方法来使用这种强制性细胞内寄生虫，从而阻碍了我们解码其分子发病机理的能力。最近的变革性研究表明，尽管其基因组大小降低，而衣原体在1000个基因上的阶数降低了，但分泌100多种效应蛋白，将其分泌到宿主细胞中。这些效应子在有选择地募集细胞器，获得营养，宿主细胞运输途径以及从宿主免疫系统中逃避检测。系统地表征宿主蛋白，这些宿主蛋白被侵入和复制在哺乳动物宿主细胞内的病原体劫持，这是了解其生物学的关键。先前在包括萌芽酵母和大肠杆菌在内的简单系统中的研究表明，这种努力在发现新型生物学见解方面非常强大。我们提出了两个具体目标，在其中我们将使用最先进的高通量质谱与新开发的生物信息学算法结合使用，以全面识别（i）全面识别与分泌的衣原体效应子相互作用的宿主蛋白，以及（ii）全球范围内的宿主在宿主的群体中，包括宿主的群体，包括群体的变化，包括宿主的整体变化，包括静脉内的群体变化，构成了脑层次的群体，这些蛋白质的变化是静态的，这些蛋白质的群体构成了群体的变化。磷酸化，泛素化和糖基化事件。 AIM 1。通过亲和力纯化/质谱（AP/MS），我们将在全球范围内识别分泌的沙眼蛋白蛋白及其宿主细胞靶蛋白之间的相互作用，以开发全面的沙眼型甲状腺菌 - 宿主 - 霍斯特蛋白 - 蛋白质相互作用（PPI）。 AIM 2。我们将使用一种无​​偏的方法来识别宿主翻译后修饰的变化，包括宿主蛋白磷酸化，泛素化和糖基化，以响应沙眼梭菌感染。使用这些创新方法阐明这些复杂事件将为衣原体呼吸道感染的发病机理，衣原体感染在动脉粥样硬化心脏病中的作用以及衣原体感染在肺癌中的作用。这些研究可能为诊断，治疗和疫苗提供新的靶标。此外，这些研究将为真核细胞生物学的基本过程提供新的见解，从发育生物学到癌症生物学的意义。