title: Polyketide synthase (PKS) reduces fusion of Legionella pneumophila-containing vacuoles with lysosomes and contributes to bacterial competitiveness during infection.
abstract: L. pneumophila-containing vacuoles (LCVs) exclude endocytic and lysosomal markers in human macrophages and protozoa. We screened a L. pneumophila mini-Tn10 transposon library for mutants, which fail to inhibit the fusion of LCVs with lysosomes by loading of the lysosomal compartment with colloidal iron dextran, mechanical lysis of infected host cells, and magnetic isolation of LCVs that have fused with lysosomes. In silico analysis of the mutated genes, D. discoideum plaque assays and infection assays in protozoa and U937 macrophage-like cells identified well established as well as novel putative L. pneumophila virulence factors. Promising candidates were further analyzed for their co-localization with lysosomes in host cells using fluorescence microscopy. This approach corroborated that the O-methyltransferase, PilY1, TPR-containing protein and polyketide synthase (PKS) of L. pneumophila interfere with lysosomal degradation. Competitive infections in protozoa and macrophages revealed that the identified PKS contributes to the biological fitness of pneumophila strains and may explain their prevalence in the epidemiology of Legionnaires' disease.
authors.affiliation: Institut für Mikrobiologie, Technische Universität Braunschweig, Braunschweig, Germany; Research Group Cellular Proteomics, Helmholtz Centre for Infection Research (HZI), Braunschweig, Germany.
authors.forename: Olga
authors.initials: O
authors.lastname: Shevchuk
authors.affiliation: Institut für Mikrobiologie, Technische Universität Braunschweig, Braunschweig, Germany.
authors.forename: Dennis
authors.initials: D
authors.lastname: Pägelow
authors.affiliation: Institut für Mikrobiologie, Technische Universität Braunschweig, Braunschweig, Germany.
authors.forename: Janine
authors.initials: J
authors.lastname: Rasch
authors.affiliation: Institut für Mikrobiologie, Technische Universität Braunschweig, Braunschweig, Germany.
authors.forename: Simon
authors.initials: S
authors.lastname: Döhrmann
authors.affiliation: Institut für Mikrobiologie, Technische Universität Braunschweig, Braunschweig, Germany.
authors.forename: Gabriele
authors.initials: G
authors.lastname: Günther
authors.affiliation: Institut für Mikrobiologie, Technische Universität Braunschweig, Braunschweig, Germany.
authors.forename: Julia
authors.initials: J
authors.lastname: Hoppe
authors.affiliation: Institut für Mikrobiologie, Technische Universität Braunschweig, Braunschweig, Germany; Fen Fakültesi, Türk-Alman Üniversitesi, Istanbul, Turkey.
authors.forename: Can Murat
authors.initials: CM
authors.lastname: Ünal
authors.affiliation: Phagosome Biology, Helmholtz Centre for Infection Research (HZI), Braunschweig, Germany.
authors.forename: Marc
authors.initials: M
authors.lastname: Bronietzki
authors.affiliation: Phagosome Biology, Helmholtz Centre for Infection Research (HZI), Braunschweig, Germany; MRC National Institute for Medical Research, The Ridgeway, London NW7 1AA, UK.
authors.forename: Maximiliano Gabriel
authors.initials: MG
authors.lastname: Gutierrez
authors.affiliation: Institut für Mikrobiologie, Technische Universität Braunschweig, Braunschweig, Germany. Electronic address: m.steinert@tu-bs.de.
authors.forename: Michael
authors.initials: M
authors.lastname: Steinert
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D002460:Cell Line; D004251:DNA Transposable Elements; D004023:Dictyostelium / Q000382:microbiology; D054884:Host-Pathogen Interactions; D006801:Humans; D016952:Legionella pneumophila / Q000235:genetics / Q000254:growth & development / Q000502:physiology*; D007877:Legionnaires' Disease / Q000382:microbiology*; D008247:Lysosomes / Q000378:metabolism*; D009000:Monocytes / Q000382:microbiology; D016254:Mutagenesis, Insertional; D048630:Polyketide Synthases / Q000235:genetics / Q000378:metabolism*; D014617:Vacuoles / Q000378:metabolism / Q000382:microbiology*; D037521:Virulence Factors / Q000235:genetics / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D004251:DNA Transposable Elements; D037521:Virulence Factors; D048630:Polyketide Synthases
keywords: Fitness; Legionella; PKS; Transposon mutants; Virulence