Host Pathways Important for <named-content content-type="genus-species">Coxiella burnetii</named-content> Infection Revealed by Genome-Wide RNA Interference Screening

ABSTRACT Coxiella burnetii is an intracellular pathogen that replicates within a lysosome-like vacuole. A Dot/Icm type IVB secretion system is used by C. burnetii to translocate effector proteins into the host cytosol that likely modulate host factor function. To identify host determinants required...

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Autores principales:	Justin A. McDonough, Hayley J. Newton, Scott Klum, Rachel Swiss, Hervé Agaisse, Craig R. Roy
Formato:	article
Lenguaje:	EN
Publicado:	American Society for Microbiology 2013
Materias:	Microbiology QR1-502
Acceso en línea:	https://doaj.org/article/a179843a94dc4a3b9faff6d6e6760f39
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spelling	oai:doaj.org-article:a179843a94dc4a3b9faff6d6e6760f392021-11-15T15:40:24ZHost Pathways Important for <named-content content-type="genus-species">Coxiella burnetii</named-content> Infection Revealed by Genome-Wide RNA Interference Screening10.1128/mBio.00606-122150-7511https://doaj.org/article/a179843a94dc4a3b9faff6d6e6760f392013-03-01T00:00:00Zhttps://journals.asm.org/doi/10.1128/mBio.00606-12https://doaj.org/toc/2150-7511ABSTRACT Coxiella burnetii is an intracellular pathogen that replicates within a lysosome-like vacuole. A Dot/Icm type IVB secretion system is used by C. burnetii to translocate effector proteins into the host cytosol that likely modulate host factor function. To identify host determinants required for C. burnetii intracellular growth, a genome-wide screen was performed using gene silencing by small interfering RNA (siRNA). Replication of C. burnetii was measured by immunofluorescence microscopy in siRNA-transfected HeLa cells. Newly identified host factors included components of the retromer complex, which mediates cargo cycling between the endocytic pathway and the Golgi apparatus. Reducing the levels of the retromer cargo-adapter VPS26-VPS29-VPS35 complex or retromer-associated sorting nexins abrogated C. burnetii replication. Several genes, when silenced, resulted in enlarged vacuoles or an increased number of vacuoles within C. burnetii-infected cells. Silencing of the STX17 gene encoding syntaxin-17 resulted in a striking defect in homotypic fusion of vacuoles containing C. burnetii, suggesting a role for syntaxin-17 in regulating this process. Lastly, silencing host genes needed for C. burnetii replication correlated with defects in the translocation of Dot/Icm effectors, whereas, silencing of genes that affected vacuole morphology, but did not impact replication, did not affect Dot/Icm translocation. These data demonstrate that C. burnetii vacuole maturation is important for creating a niche that permits Dot/Icm function. Thus, genome-wide screening has revealed host determinants involved in sequential events that occur during C. burnetii infection as defined by bacterial uptake, vacuole transport and acidification, activation of the Dot/Icm system, homotypic fusion of vacuoles, and intracellular replication. IMPORTANCE Q fever in humans is caused by the bacterium Coxiella burnetii. Infection with C. burnetii is marked by its unique ability to replicate within a large vacuolar compartment inside cells that resembles the harsh, acidic environment of a lysosome. Central to its pathogenesis is the delivery of bacterial effector proteins into the host cell cytosol by a Dot/Icm type IVB secretion system. These proteins can interact with and manipulate host factors, thereby leading to creation and maintenance of the vacuole that the bacteria grow within. Using high-throughput genome-wide screening in human cells, we identified host factors important for several facets of C. burnetii infection, including vacuole transport and membrane fusion events that promote vacuole expansion. In addition, we show that maturation of the C. burnetii vacuole is necessary for creating an environment permissive for the Dot/Icm delivery of bacterial effector proteins into the host cytosol.Justin A. McDonoughHayley J. NewtonScott KlumRachel SwissHervé AgaisseCraig R. RoyAmerican Society for MicrobiologyarticleMicrobiologyQR1-502ENmBio, Vol 4, Iss 1 (2013)
institution	DOAJ
collection	DOAJ
language	EN
topic	Microbiology QR1-502
spellingShingle	Microbiology QR1-502 Justin A. McDonough Hayley J. Newton Scott Klum Rachel Swiss Hervé Agaisse Craig R. Roy Host Pathways Important for <named-content content-type="genus-species">Coxiella burnetii</named-content> Infection Revealed by Genome-Wide RNA Interference Screening
description	ABSTRACT Coxiella burnetii is an intracellular pathogen that replicates within a lysosome-like vacuole. A Dot/Icm type IVB secretion system is used by C. burnetii to translocate effector proteins into the host cytosol that likely modulate host factor function. To identify host determinants required for C. burnetii intracellular growth, a genome-wide screen was performed using gene silencing by small interfering RNA (siRNA). Replication of C. burnetii was measured by immunofluorescence microscopy in siRNA-transfected HeLa cells. Newly identified host factors included components of the retromer complex, which mediates cargo cycling between the endocytic pathway and the Golgi apparatus. Reducing the levels of the retromer cargo-adapter VPS26-VPS29-VPS35 complex or retromer-associated sorting nexins abrogated C. burnetii replication. Several genes, when silenced, resulted in enlarged vacuoles or an increased number of vacuoles within C. burnetii-infected cells. Silencing of the STX17 gene encoding syntaxin-17 resulted in a striking defect in homotypic fusion of vacuoles containing C. burnetii, suggesting a role for syntaxin-17 in regulating this process. Lastly, silencing host genes needed for C. burnetii replication correlated with defects in the translocation of Dot/Icm effectors, whereas, silencing of genes that affected vacuole morphology, but did not impact replication, did not affect Dot/Icm translocation. These data demonstrate that C. burnetii vacuole maturation is important for creating a niche that permits Dot/Icm function. Thus, genome-wide screening has revealed host determinants involved in sequential events that occur during C. burnetii infection as defined by bacterial uptake, vacuole transport and acidification, activation of the Dot/Icm system, homotypic fusion of vacuoles, and intracellular replication. IMPORTANCE Q fever in humans is caused by the bacterium Coxiella burnetii. Infection with C. burnetii is marked by its unique ability to replicate within a large vacuolar compartment inside cells that resembles the harsh, acidic environment of a lysosome. Central to its pathogenesis is the delivery of bacterial effector proteins into the host cell cytosol by a Dot/Icm type IVB secretion system. These proteins can interact with and manipulate host factors, thereby leading to creation and maintenance of the vacuole that the bacteria grow within. Using high-throughput genome-wide screening in human cells, we identified host factors important for several facets of C. burnetii infection, including vacuole transport and membrane fusion events that promote vacuole expansion. In addition, we show that maturation of the C. burnetii vacuole is necessary for creating an environment permissive for the Dot/Icm delivery of bacterial effector proteins into the host cytosol.
format	article
author	Justin A. McDonough Hayley J. Newton Scott Klum Rachel Swiss Hervé Agaisse Craig R. Roy
author_facet	Justin A. McDonough Hayley J. Newton Scott Klum Rachel Swiss Hervé Agaisse Craig R. Roy
author_sort	Justin A. McDonough
title	Host Pathways Important for <named-content content-type="genus-species">Coxiella burnetii</named-content> Infection Revealed by Genome-Wide RNA Interference Screening
title_short	Host Pathways Important for <named-content content-type="genus-species">Coxiella burnetii</named-content> Infection Revealed by Genome-Wide RNA Interference Screening
title_full	Host Pathways Important for <named-content content-type="genus-species">Coxiella burnetii</named-content> Infection Revealed by Genome-Wide RNA Interference Screening
title_fullStr	Host Pathways Important for <named-content content-type="genus-species">Coxiella burnetii</named-content> Infection Revealed by Genome-Wide RNA Interference Screening
title_full_unstemmed	Host Pathways Important for <named-content content-type="genus-species">Coxiella burnetii</named-content> Infection Revealed by Genome-Wide RNA Interference Screening
title_sort	host pathways important for <named-content content-type="genus-species">coxiella burnetii</named-content> infection revealed by genome-wide rna interference screening
publisher	American Society for Microbiology
publishDate	2013
url	https://doaj.org/article/a179843a94dc4a3b9faff6d6e6760f39
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Host Pathways Important for <named-content content-type="genus-species">Coxiella burnetii</named-content> Infection Revealed by Genome-Wide RNA Interference Screening

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