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Quantitative omics reveals that type I and II IFNs alter bacterial vacuole compositions to restrict infection in macrophages and lungs

ABSTRACT: The innate defense mechanisms that control infections with intracellular bacteria are still incompletely understood. Here we show that type I and II IFNs are key regulators of the early gene expression and the host-protective immune response during Legionella pneumophila-induced pneumonia. Using mixed bone marrow-chimeric mice and isolated cells we indicate that both IFNs protect against L. pneumophila by activating an alveolar macrophage-intrinsic antibacterial defense pathway. Quantitative mass spectrometry analysis reveals that both IFNs markedly alter the protein composition of purified Legionella-containing vacuoles, and integrated network analysis defines distinct subsets of IFN-regulated proteins. Subsequent experiments uncover Immunoresponsive gene 1 (Irg1) as a central effector that restricts the bacteria through production of itaconic acid. Collectively, we provide a comprehensive analysis of IFN-mediated effects on gene expression and the bacterial vacuole proteome, and show that L. pneumophila is restricted by an Irg1-dependent production of a bactericidal metabolite.

ORGANISM(S): Mus musculus

PROVIDER: GSE60085 | GEO | 2015/01/01

SECONDARY ACCESSION(S): PRJNA257451

REPOSITORIES: GEO

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Project description:Differential gene expression of Dictyostelium discoideum after infection with Legionella pneumophila was investigated using DNA microarrays. A detailed analysis of the 24 h time point post infection was performed in comparison to three controls, uninfected cells and co-incubation with Legionella hackeliae and L. pneumophila DeltadotA. One hundred and thirty-one differentially expressed D. discoideum genes were identified as common to all three experiments and are thought to be involved in the pathogenic response. Functional annotation of the differentially regulated genes revealed that apart from triggering a stress response Legionella apparently not only interferes with intracellular vesicle fusion and destination but also profoundly influences and exploits the metabolism of its host. The results provide the basis for a better understanding of the complex host-pathogen interactions and for further studies on the Dictyostelium response to Legionella infection. The bacterial strains used in this study were L. pneumophila Philadelphia I JR32, L. pneumophila Philadelphia I JR32 LELA 3118 (dotA3118:Tn903 DLL LacZ) and L. hackeliae (ATCC 35250). The Legionella strains were grown on buffered charcoal yeast extract agar (BCYE) at 37M-BM-0C with 5% CO2 atmosphere for 3 days. The D. discoideum wild-type strain AX2 was grown at 23M-BM-0C in 75 cm2 cell-culture flasks with 10 ml HL5 medium. For infection, Dictyostelium cells were harvested, resuspended in a 1:1 solution of HL5 medium and Soerensen buffer. Fifteen millilitres of a 1M-CM-^W10e6 cells/ml suspension were seeded into a 75 square-cm cell culture flask and the amoebae were inoculated with 10e7 bacteria/ml. Three different pairs of infection were compared: 1. AX2 infected with L. pneumophila JR32 versus uninfected cells; 19 microarrays of seven independent infections; 2. AX2 infected with L. pneumophila JR32 versus AX2 infected with L. pneumophila JR32 delta DotA; 4 microarrays of two independent infections; 3. AX2 infected with L. pneumophila JR32 versus AX2 infected with L. hackeliae; 4 microarrays of two independent infections. 24h post infection the RNA was isolated from 1.5M-CM-^W10e7 Dictyostelium cells and microarray analysis was performed as described (Farbrother et al., 2006).

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Quantitative omics reveals that type I and II IFNs alter bacterial vacuole compositions to restrict infection in macrophages and lungs

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