title: Enterococcus faecalis Glycolipids Modulate Lipoprotein-Content of the Bacterial Cell Membrane and Host Immune Response.
abstract: In this study, we investigated the impact of the cell membrane composition of E. faecalis on its recognition by the host immune system. To this end, we employed an E. faecalis deletion mutant (ΔbgsA) that does not synthesize the major cell membrane glycolipid diglycosyl-diacylglycerol (DGlcDAG). Proteomic analysis revealed that 13 of a total of 21 upregulated surface-associated proteins of E. faecalis ΔbgsA were lipoproteins. This led to a total lipoprotein content in the cell membrane of 35.8% in ΔbgsA compared to only 9.4% in wild-type bacteria. Increased lipoprotein content strongly affected the recognition of ΔbgsA by mouse macrophages in vitro with an increased stimulation of TNF-α production by heat-fixed bacteria and secreted antigens. Inactivation of the prolipoprotein diacylglycerol transferase (lgt) in ΔbgsA abrogated TNF-α induction by a ΔbgsA_lgt double mutant indicating that lipoproteins mediate increased activation of mouse macrophages by ΔbgsA. Heat-fixed ΔbgsA bacteria, culture supernatant, or cell membrane lipid extract activated transfected HEK cells in a TLR2-dependent fashion; the same was not true of wild-type bacteria. In mice infected intraperitoneally with a sublethal dose of E. faecalis we observed a 70% greater mortality in mice infected with ΔbgsA compared with wild-type-infected mice. Increased mortality due to ΔbgsA infection was associated with elevated plasma levels of the inflammatory cytokines TNF-α, IL-6 and MIP-2. In summary, our results provide evidence that an E. faecalis mutant lacking its major bilayer forming glycolipid DGlcDAG upregulates lipoprotein expression leading to increased activation of the host innate immune system and virulence in vivo.
authors.affiliation: Center for Chronic Immunodeficiency (CCI), University Medical Center Freiburg, Freiburg, Germany; Center for Infectious Disease and Travel Medicine, University Medical Center Freiburg, Freiburg, Germany.
authors.forename: Christian
authors.initials: C
authors.lastname: Theilacker
authors.affiliation: Center for Infectious Disease and Travel Medicine, University Medical Center Freiburg, Freiburg, Germany; Division of Pediatric Infectious Diseases, Dr. von Hauner Children's Hospital, Ludwig-Maximilians-University, Munich, Germany; Department of Microbiology, Faculty of Biology, Albert-Ludwigs-University, Freiburg, Germany.
authors.forename: Ann-Kristin
authors.initials: AK
authors.lastname: Diederich
authors.affiliation: Institute for Microbiology, Department of Microbial Physiology, Ernst-Moritz-Arndt-University, Greifswald, Germany.
authors.forename: Andreas
authors.initials: A
authors.lastname: Otto
authors.affiliation: Research Center for Nutrition and Food Science, Technical University Munich, Freising, Germany.
authors.forename: Irina G
authors.initials: IG
authors.lastname: Sava
authors.affiliation: Center for Infectious Disease and Travel Medicine, University Medical Center Freiburg, Freiburg, Germany.
authors.forename: Dominique
authors.initials: D
authors.lastname: Wobser
authors.affiliation: Center for Infectious Disease and Travel Medicine, University Medical Center Freiburg, Freiburg, Germany.
authors.forename: Yinyin
authors.initials: Y
authors.lastname: Bao
authors.affiliation: Center for Chronic Immunodeficiency (CCI), University Medical Center Freiburg, Freiburg, Germany.
authors.forename: Katrin
authors.initials: K
authors.lastname: Hese
authors.affiliation: Center for Infectious Disease and Travel Medicine, University Medical Center Freiburg, Freiburg, Germany.
authors.forename: Melanie
authors.initials: M
authors.lastname: Broszat
authors.affiliation: Center for Chronic Immunodeficiency (CCI), University Medical Center Freiburg, Freiburg, Germany; Center for Paediatrics and Adolescent Medicine, University Medical Center Freiburg, Freiburg, Germany.
authors.forename: Philipp
authors.initials: P
authors.lastname: Henneke
authors.affiliation: Institute for Microbiology, Department of Microbial Physiology, Ernst-Moritz-Arndt-University, Greifswald, Germany.
authors.forename: Dörte
authors.initials: D
authors.lastname: Becher
authors.affiliation: Center for Infectious Disease and Travel Medicine, University Medical Center Freiburg, Freiburg, Germany; Division of Pediatric Infectious Diseases, Dr. von Hauner Children's Hospital, Ludwig-Maximilians-University, Munich, Germany.
authors.forename: Johannes
authors.initials: J
authors.lastname: Huebner
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000276:immunology; D002460:Cell Line; D002462:Cell Membrane / Q000276:immunology*; D054426:Chemokine CXCL2 / Q000097:blood; D013293:Enterococcus faecalis / Q000276:immunology*; D005260:Female; D006017:Glycolipids / Q000276:immunology*; D057809:HEK293 Cells; D054884:Host-Pathogen Interactions / Q000276:immunology*; D006801:Humans; D007113:Immunity, Innate / Q000276:immunology; D015850:Interleukin-6 / Q000097:blood; D008074:Lipoproteins / Q000276:immunology*; D008264:Macrophages; D008563:Membrane Lipids / Q000276:immunology; D008565:Membrane Proteins / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D051195:Toll-Like Receptor 2 / Q000276:immunology; D014409:Tumor Necrosis Factor-alpha / Q000097:blood / Q000276:immunology; D014774:Virulence / Q000276:immunology
chemical_list: D001426:Bacterial Proteins; D054426:Chemokine CXCL2; D006017:Glycolipids; D015850:Interleukin-6; D008074:Lipoproteins; D008563:Membrane Lipids; D008565:Membrane Proteins; D051195:Toll-Like Receptor 2; D014409:Tumor Necrosis Factor-alpha; C025142:diglucosyl diacylglycerol