IRG1 and Inducible Nitric Oxide Synthase Act Redundantly with Other Interferon-Gamma-Induced Factors To Restrict Intracellular Replication of <named-content content-type="genus-species">Legionella pneumophila</named-content>

ABSTRACT Interferon gamma (IFN-γ) restricts the intracellular replication of many pathogens, but the mechanism by which IFN-γ confers cell-intrinsic pathogen resistance remains unclear. For example, intracellular replication of the bacterial pathogen Legionella pneumophila in macrophages is potently...

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Autores principales:	Jordan V. Price, Daniel Russo, Daisy X. Ji, Roberto A. Chavez, Lucian DiPeso, Angus Yiu-Fai Lee, Jörn Coers, Russell E. Vance
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2019
Materias:	Legionella pneumophila host-pathogen interactions innate immunity interferons macrophages Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/2c4aa12c2d0e48f185f4175d317f39e4
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spelling	oai:doaj.org-article:2c4aa12c2d0e48f185f4175d317f39e42021-11-15T15:54:47ZIRG1 and Inducible Nitric Oxide Synthase Act Redundantly with Other Interferon-Gamma-Induced Factors To Restrict Intracellular Replication of <named-content content-type="genus-species">Legionella pneumophila</named-content>10.1128/mBio.02629-192150-7511https://doaj.org/article/2c4aa12c2d0e48f185f4175d317f39e42019-12-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.02629-19https://doaj.org/toc/2150-7511ABSTRACT Interferon gamma (IFN-γ) restricts the intracellular replication of many pathogens, but the mechanism by which IFN-γ confers cell-intrinsic pathogen resistance remains unclear. For example, intracellular replication of the bacterial pathogen Legionella pneumophila in macrophages is potently curtailed by IFN-γ. However, consistent with prior studies, no individual genetic deficiency that we tested completely abolished IFN-γ-mediated control. Intriguingly, we observed that the glycolysis inhibitor 2-deoxyglucose (2DG) partially rescued L. pneumophila replication in IFN-γ-treated macrophages. 2DG inhibits glycolysis and triggers the unfolded protein response, but unexpectedly, it appears these effects are not responsible for perturbing the antimicrobial activity of IFN-γ. Instead, we found that 2DG rescues bacterial replication by inhibiting the expression of two key antimicrobial factors, inducible nitric oxide synthase (iNOS) and immune-responsive gene 1 (IRG1). Using immortalized and primary macrophages deficient in iNOS and IRG1, we confirmed that loss of both iNOS and IRG1, but not individual deficiency in either gene, partially reduced IFN-γ-mediated restriction of L. pneumophila. Further, using a combinatorial CRISPR/Cas9 mutagenesis approach, we found that mutation of iNOS and IRG1 in combination with four other genes (CASP11, IRGM1, IRGM3, and NOX2) resulted in a total loss of L. pneumophila restriction by IFN-γ in primary bone marrow macrophages. Our study defines a complete set of cell-intrinsic factors required for IFN-γ-mediated restriction of an intracellular bacterial pathogen and highlights the combinatorial strategy used by hosts to block bacterial replication in macrophages. IMPORTANCE Legionella pneumophila is one example among many species of pathogenic bacteria that replicate within mammalian macrophages during infection. The immune signaling factor interferon gamma (IFN-γ) blocks L. pneumophila replication in macrophages and is an essential component of the immune response to L. pneumophila and other intracellular pathogens. However, to date, no study has identified the exact molecular factors induced by IFN-γ that are required for its activity. We generated macrophages lacking different combinations of IFN-γ-induced genes in an attempt to find a genetic background in which there is a complete loss of IFN-γ-mediated restriction of L. pneumophila. We identified six genes that comprise the totality of the IFN-γ-dependent restriction of L. pneumophila replication in macrophages. Our results clarify the molecular basis underlying the potent effects of IFN-γ and highlight how redundancy downstream of IFN-γ is key to prevent exploitation of macrophages by pathogens.Jordan V. PriceDaniel RussoDaisy X. JiRoberto A. ChavezLucian DiPesoAngus Yiu-Fai LeeJörn CoersRussell E. VanceAmerican Society for MicrobiologyarticleLegionella pneumophilahost-pathogen interactionsinnate immunityinterferonsmacrophagesMicrobiologyQR1-502ENmBio, Vol 10, Iss 6 (2019)
institution	DOAJ
collection	DOAJ
language	EN
topic	Legionella pneumophila host-pathogen interactions innate immunity interferons macrophages Microbiology QR1-502
spellingShingle	Legionella pneumophila host-pathogen interactions innate immunity interferons macrophages Microbiology QR1-502 Jordan V. Price Daniel Russo Daisy X. Ji Roberto A. Chavez Lucian DiPeso Angus Yiu-Fai Lee Jörn Coers Russell E. Vance IRG1 and Inducible Nitric Oxide Synthase Act Redundantly with Other Interferon-Gamma-Induced Factors To Restrict Intracellular Replication of <named-content content-type="genus-species">Legionella pneumophila</named-content>
description	ABSTRACT Interferon gamma (IFN-γ) restricts the intracellular replication of many pathogens, but the mechanism by which IFN-γ confers cell-intrinsic pathogen resistance remains unclear. For example, intracellular replication of the bacterial pathogen Legionella pneumophila in macrophages is potently curtailed by IFN-γ. However, consistent with prior studies, no individual genetic deficiency that we tested completely abolished IFN-γ-mediated control. Intriguingly, we observed that the glycolysis inhibitor 2-deoxyglucose (2DG) partially rescued L. pneumophila replication in IFN-γ-treated macrophages. 2DG inhibits glycolysis and triggers the unfolded protein response, but unexpectedly, it appears these effects are not responsible for perturbing the antimicrobial activity of IFN-γ. Instead, we found that 2DG rescues bacterial replication by inhibiting the expression of two key antimicrobial factors, inducible nitric oxide synthase (iNOS) and immune-responsive gene 1 (IRG1). Using immortalized and primary macrophages deficient in iNOS and IRG1, we confirmed that loss of both iNOS and IRG1, but not individual deficiency in either gene, partially reduced IFN-γ-mediated restriction of L. pneumophila. Further, using a combinatorial CRISPR/Cas9 mutagenesis approach, we found that mutation of iNOS and IRG1 in combination with four other genes (CASP11, IRGM1, IRGM3, and NOX2) resulted in a total loss of L. pneumophila restriction by IFN-γ in primary bone marrow macrophages. Our study defines a complete set of cell-intrinsic factors required for IFN-γ-mediated restriction of an intracellular bacterial pathogen and highlights the combinatorial strategy used by hosts to block bacterial replication in macrophages. IMPORTANCE Legionella pneumophila is one example among many species of pathogenic bacteria that replicate within mammalian macrophages during infection. The immune signaling factor interferon gamma (IFN-γ) blocks L. pneumophila replication in macrophages and is an essential component of the immune response to L. pneumophila and other intracellular pathogens. However, to date, no study has identified the exact molecular factors induced by IFN-γ that are required for its activity. We generated macrophages lacking different combinations of IFN-γ-induced genes in an attempt to find a genetic background in which there is a complete loss of IFN-γ-mediated restriction of L. pneumophila. We identified six genes that comprise the totality of the IFN-γ-dependent restriction of L. pneumophila replication in macrophages. Our results clarify the molecular basis underlying the potent effects of IFN-γ and highlight how redundancy downstream of IFN-γ is key to prevent exploitation of macrophages by pathogens.
format	article
author	Jordan V. Price Daniel Russo Daisy X. Ji Roberto A. Chavez Lucian DiPeso Angus Yiu-Fai Lee Jörn Coers Russell E. Vance
author_facet	Jordan V. Price Daniel Russo Daisy X. Ji Roberto A. Chavez Lucian DiPeso Angus Yiu-Fai Lee Jörn Coers Russell E. Vance
author_sort	Jordan V. Price
title	IRG1 and Inducible Nitric Oxide Synthase Act Redundantly with Other Interferon-Gamma-Induced Factors To Restrict Intracellular Replication of <named-content content-type="genus-species">Legionella pneumophila</named-content>
title_short	IRG1 and Inducible Nitric Oxide Synthase Act Redundantly with Other Interferon-Gamma-Induced Factors To Restrict Intracellular Replication of <named-content content-type="genus-species">Legionella pneumophila</named-content>
title_full	IRG1 and Inducible Nitric Oxide Synthase Act Redundantly with Other Interferon-Gamma-Induced Factors To Restrict Intracellular Replication of <named-content content-type="genus-species">Legionella pneumophila</named-content>
title_fullStr	IRG1 and Inducible Nitric Oxide Synthase Act Redundantly with Other Interferon-Gamma-Induced Factors To Restrict Intracellular Replication of <named-content content-type="genus-species">Legionella pneumophila</named-content>
title_full_unstemmed	IRG1 and Inducible Nitric Oxide Synthase Act Redundantly with Other Interferon-Gamma-Induced Factors To Restrict Intracellular Replication of <named-content content-type="genus-species">Legionella pneumophila</named-content>
title_sort	irg1 and inducible nitric oxide synthase act redundantly with other interferon-gamma-induced factors to restrict intracellular replication of <named-content content-type="genus-species">legionella pneumophila</named-content>
publisher	American Society for Microbiology
publishDate	2019
url	https://doaj.org/article/2c4aa12c2d0e48f185f4175d317f39e4
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IRG1 and Inducible Nitric Oxide Synthase Act Redundantly with Other Interferon-Gamma-Induced Factors To Restrict Intracellular Replication of <named-content content-type="genus-species">Legionella pneumophila</named-content>

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