Genetics of Monocyte Killing by Bacteria

细菌杀死单核细胞的遗传学

• 批准号: 8472434
• 负责人: HOWARD A SHUMAN
• 金额: $ 51.7万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-06-15 至 2015-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8472434
• 关键词: Actins; Acute Pneumonia; Alleles; Alveolar Macrophages; Bacteria; Binding; Biochemical; Bioinformatics; Biological; Breathing; Cell Line; Cells; Coiled-Coil Domain; Complex; Defect; Disease; Dominant-Negative Mutation; Ectopic Expression; Endoplasmic Reticulum; Ensure; Environment; Event; Exhibits; Funding; Genes; Genetic; Golgi Apparatus; Growth; Human; Individual; Infection; Legionella; Legionella pneumophila; Legionnaires' Disease; Leukocytes; Lung; Lysosomes; Mammalian Cell; Measures; Membrane; Microbial Biofilms; Modeling; Molecular; Organelles; PTPRC gene; Pathway interactions; Phagosomes; Phenotype; Phosphoric Monoester Hydrolases; Phosphotransferases; Plumbing; Property; Protein Family; Protein Tyrosine Kinase; Protein Tyrosine Phosphatase; Protein translocation; Proteins; Research; Role; SNAP receptor; Saccharomyces cerevisiae; Small Interfering RNA; Sorting - Cell Movement; Source; System; Testing; Vacuolar Protein Sorting; Vacuole; Vesicle; Water; Work; Yeasts; aerosolized; antimicrobial; bacterial genetics; base; inhibitor/antagonist; killings; macrophage; monocyte; mutant; novel; null mutation; pathogen; polymerization; tool; trafficking

PROJECT SUMMARY This proposal is focused on understanding the molecular basis for the ability of Legionella pneumophila to infect, survive within, replicate within, and eventually kill human macrophages. We propose to study a group of novel proteins that are made by L. pneumophila and translocated to host cells by the Icm/Dot translocation system. We will test the hypothesis that the translocated proteins interact with organelle trafficking pathways in the host and contribute to Legionella intracellular multiplication. One of the translocated proteins, VipA, was found to bind actin and promote polymerization of actin subunits. We will also focus on three proteins (LegC2, LegC3, LegC7) that contain coiled coil domains. VipA and the LegC proteins all cause organelle trafficking defects when expressed in the model host, Saccharomyces cerevisiae. The specific aims of this proposal are to : 1. Determine the molecular basis for the activity of VipA, an actin-binding, translocated effector that interferes with endosomal trafficking; 2. Test the hypothesis that components of the Vps/ESCRT complex are related to events during intracellular multiplication of Legionella; 3. Dominant-negative interfering alleles of effector genes and the role of effectors during intracellular multiplication; 4. Identify host cell tyrosine kinases that control the initial interactions between Legionella and host cells required for effector translocation; 5. Identify interaction partners of LegC2, LegC3 and LegC7. In order to carry out these Aims we will take advantage of a variety of cell biological tools such as depleting cells of specific organelle trafficking components by siRNA and examining the effect on Legionella infection, co-localization of known organelle markers with the Legionella -containing vacuole and ectopic expression of Legionella genes is human macrophage cell lines. We will also examine the effects of specifically targeting host tyrosine kinases and a phosphatase on Legionella intracellular multiplication and organelle trafficking. We will use bacterial genetics to isolate dominant-negative alleles of the genes encoding the translocated proteins to better understand their role during Legionella infection. All of these approaches should clarify the mechanisms that Legionella uses to avoid killing by macrophages and produce a successful infection.

项目摘要 该建议的重点是理解肺炎军团菌的能力的分子基础 感染，在内部生存，在内部复制，并最终杀死人类巨噬细胞。我们建议研究一组 新型蛋白质是由肺炎乳杆菌制成的，并通过ICM/DOT易位转移至宿主细胞 系统。我们将检验以下假设：易位蛋白与细胞器运输途径相互作用 宿主并有助于军团菌的细胞内乘法。易位的蛋白质之一是 发现结合肌动蛋白并促进肌动蛋白亚基的聚合。我们还将专注于三种蛋白质（legc2， LEGC3，LEGC7）包含盘绕线圈域。 vipa和legc蛋白都会导致细胞器贩运 在模型宿主酿酒酵母中表达的缺陷。该提议的具体目的是 至：1。确定VIPA活性的分子基础，VIPA是肌动蛋白结合的，转运效应子 干扰内体贩运； 2。检验VPS/ESCRT复合物的组成部分的假设是 与军团菌细胞内繁殖期间的事件有关； 3。主要阴性干扰等位基因 效应子基因和效应子在细胞内繁殖过程中的作用； 4。识别宿主细胞酪氨酸激酶 控制效应子易位所需的军团菌与宿主细胞之间的初始相互作用； 5。 确定LEGC2，LEGC3和LEGC7的互动伙伴。为了实现这些目标，我们将采取 各种细胞生物学工具的优势，例如耗尽特定细胞器运输的细胞 siRNA的组件并检查对军团菌感染的影响，已知细胞器的共定位 具有军团菌的液泡和军团菌基因异位表达的标记是人 巨噬细胞系。我们还将研究专门靶向宿主酪氨酸激酶和A的影响 磷酸酶的细胞内繁殖和细胞器运输。我们将使用细菌遗传学 隔离编码转运蛋白的基因的显性阴性等位基因，以更好地了解其 在军团菌感染中的作用。所有这些方法都应阐明Legionella用于的机制 避免被巨噬细胞杀死并成功感染。

项目成果

The Legionella pneumophila effector VipA is an actin nucleator that alters host cell organelle trafficking.

• DOI: 10.1371/journal.ppat.1002546
• 发表时间: 2012-02
• 期刊: PLoS pathogens
• 影响因子: 6.7
• 作者: Franco IS;Shohdy N;Shuman HA
• 通讯作者: Shuman HA

Legionella eukaryotic-like type IV substrates interfere with organelle trafficking.

类军团菌真核 IV 型底物干扰细胞器运输。

• DOI: 10.1371/journal.ppat.1000117
• 发表时间: 2008-08-01
• 期刊: PLOS PATHOGENS
• 影响因子: 6.7
• 作者: de Felipe, Karim Suwwan;Glover, Robert T.;Charpentier, Xavier;Anderson, O. Roger;Reyes, Moraima;Pericone, Christopher D.;Shuman, Howard A.
• 通讯作者: Shuman, Howard A.

A new heterolobosean amoeba Solumitrus palustris n. g., n. sp. isolated from freshwater marsh soil.

一新异叶阿米巴Solumitrus palustris n.

• DOI: 10.1111/j.1550-7408.2010.00520.x
• 发表时间: 2011
• 期刊: The Journal of eukaryotic microbiology
• 作者: Anderson,ORoger;Wang,Wen;Faucher,SebastienP;Bi,Keran;Shuman,HowardA
• 通讯作者: Shuman,HowardA

Analysis of the transcriptome of Legionella pneumophila hfq mutant reveals a new mobile genetic element.

对嗜肺军团菌 hfq 突变体转录组的分析揭示了一种新的可移动遗传元件。

• DOI: 10.1099/mic.0.067983-0
• 发表时间: 2013
• 期刊: Microbiology (Reading, England)
• 作者: Trigui,Hana;Dudyk,Paulina;Sum,Janet;Shuman,HowardA;Faucher,SebastienP
• 通讯作者: Faucher,SebastienP

Genomic analysis of 38 Legionella species identifies large and diverse effector repertoires.

• DOI: 10.1038/ng.3481
• 发表时间: 2016-02
• 期刊: NATURE GENETICS
• 影响因子: 30.8
• 作者: Burstein, David;Amaro, Francisco;Zusman, Tal;Lifshitz, Ziv;Cohen, Ofir;Gilbert, Jack A.;Pupko, Tal;Shuman, Howard A.;Segal, Gil
• 通讯作者: Segal, Gil