Cellular Intoxication Pathway of Cytolethal Distending Toxin

细胞致死膨胀毒素的细胞中毒途径

• 批准号: 8730187
• 负责人: Kenneth Alan Bradley
• 金额: $ 41.48万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-09-01 至 2016-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8730187
• 关键词: Address; Area; Bacterial Proteins; Bacterial Toxins; Binding; Biology; California; Carrier Proteins; Cell Cycle; Cell Cycle Arrest; Cell Line; Cell Nucleus; Cell Surface Receptors; Cell membrane; Cell physiology; Cell surface; Cells; Cellular biology; Comparative Study; Complement; Complex; Cytokinesis; Cytosol; DNA Damage; DNA Repair; Data; Deoxyribonuclease I; Destinations; Endocytosis Pathway; Event; Family; Foundations; Future; Genes; Genetic; Genetic Screening; Gram-Negative Bacteria; Hemophilus ducreyi; Illinois; Immune Sera; Integration Host Factors; Intoxication; Intracellular Transport; Knowledge; Los Angeles; Mammalian Cell; Membrane Protein Traffic; Methodology; Modeling; Molecular; Mutagenesis; Nature; Operon; Organelles; Pathway interactions; Pharmaceutical Preparations; Phosphorylation; Phosphotransferases; Process; Production; Property; Protein Subunits; Proteins; Reagent; Receptor Cell; Research; Research Personnel; Resolution; Role; Route; Signal Transduction; Structure; System; Time; Toxin; Translating; Universities; Vesicle; Vesicle Transport Pathway; Work; ataxia telangiectasia mutated protein; base; cytolethal distending toxin; design; extracellular; forward genetics; holotoxins; human H2AX protein; human disease; insight; member; molecular carrier; mutant; novel; novel therapeutics; pathogen; pathogenic bacteria; receptor; receptor binding; research study; tool; trafficking; uptake

DESCRIPTION (provided by applicant): The intracellular-acting bacterial protein toxins have evolved various means to interact with and enter mammalian cells, taking advantage of existing host cellular processes. Studies of the mechanisms by which bacterial toxins are internalized have provided fundamental advances in several areas of cell biology, including receptor biology, mechanisms and pathways of endocytosis, vesicle trafficking, and membrane translocation. Here, we propose studies focused on one member of an emerging family of toxins called the cytolethal distending toxins (CDTs), which are unusual in their intracellular localization, modulatory activities, and overall structures. CDTs are multi-subunit toxins that are generated by a diverse group of pathogenic Gram-negative bacteria and function by modulating the host cell cycle. Active CDT holotoxins are heterotrimeric complexes of three protein subunits, generally encoded by three contiguous genes (cdtA, cdtB, cdtC) in a single operon. To exert their cyclomodulatory effects on cells, CDTs must be taken up from the cell surface and transported intracellularly in a manner that ultimately results in localization of the enzymatic subunit CdtB to the nucleus. However, the molecular details and mechanism by which CDTs exploit existing uptake and transport pathways to gain access to the cytosol, and ultimately the nucleus, are poorly understood. This application addresses these current gaps in knowledge and represents a collaborative proposal between investigators at the University of California at Los Angeles (Dr. Bradley) and the University of Illinois (Dr. Blanke) to investigate the molecular basis of CDT cellular intoxication. Two Aims are proposed that address hypotheses regarding the cell surface binding (Aim 1) and intracellular transport (Aim 2) of CDTs required for cellular intoxication. Aim 1 investigates the molecular determinants of the initial CDT-cell binding contributed by both the host (Aim 1.1) and toxin (Aim 1.2). Aim 2 focuses squarely on the host cellular requirements for uptake and trafficking of CDT, utilizing hypothesis driven (Aim 2.1) and forward genetic (Aim 2.2) approaches. Importantly, the heterotrimeric nature of CDTs, as well as the localization of the catalytic CdtB subunit to the nucleus, are unique features of the CDTs. Therefore, identification of host cell determinants and trafficking pathways that are important for CDT intoxication is predicted to reveal novel host cell requirements and/or routes for transporting proteins from the cell surface to the nucleus and provide insight into toxin-host interactions.

描述（由申请人提供）：细胞内作用的细菌蛋白毒素已经利用现有的宿主细胞过程进化出多种与哺乳动物细胞相互作用并进入哺乳动物细胞的方式。对细菌毒素内化机制的研究为细胞生物学的多个领域提供了基础性进展，包括受体生物学、内吞作用、囊泡运输和膜易位的机制和途径。在这里，我们建议将研究重点放在称为细胞致死膨胀毒素（CDT）的新兴毒素家族中的一个成员，该毒素在细胞内定位、调节活性和整体结构方面都不同寻常。 CDT 是多亚基毒素，由多种致病性革兰氏阴性菌产生，通过调节宿主细胞周期发挥作用。活性 CDT 全毒素是三个蛋白质亚基的异三聚体复合物，通常由单个操纵子中的三个连续基因（cdtA、cdtB、cdtC）编码。为了对细胞发挥其环调节作用，CDT 必须从细胞表面摄取并以最终导致酶亚基 CdtB 定位到细胞核的方式在细胞内运输。然而，人们对 CDT 利用现有摄取和运输途径进入细胞质并最终进入细胞核的分子细节和机制知​​之甚少。该应用解决了当前的知识差距，并代表了加州大学洛杉矶分校（Bradley 博士）和伊利诺伊大学（Blanke 博士）研究人员之间的合作提案，以研究 CDT 细胞中毒的分子基础。提出了两个目标来解决有关细胞中毒所需的 CDT 的细胞表面结合（目标 1）和细胞内转运（目标 2）的假设。目标 1 研究由宿主（目标 1.1）和毒素（目标 1.2）共同贡献的初始 CDT 细胞结合的分子决定因素。目标 2 利用假设驱动（目标 2.1）和正向遗传（目标 2.2）方法，直接关注宿主细胞摄取和运输 CDT 的需求。重要的是，CDT 的异三聚体性质以及催化 CdtB 亚基定位于细胞核，是 CDT 的独特特征。因此，对 CDT 中毒重要的宿主细胞决定因素和运输途径的鉴定预计将揭示新的宿主细胞需求和/或将蛋白质从细胞表面转运到细胞核的途径，并提供对毒素-宿主相互作用的深入了解。

项目成果

Distinct Roles for CdtA and CdtC during Intoxication by Cytolethal Distending Toxins.

• DOI: 10.1371/journal.pone.0143977
• 发表时间: 2015
• 期刊: PloS one
• 影响因子: 3.7
• 作者: Dixon SD;Huynh MM;Tamilselvam B;Spiegelman LM;Son SB;Eshraghi A;Blanke SR;Bradley KA
• 通讯作者: Bradley KA