title: Direct visualization by cryo-EM of the mycobacterial capsular layer: a labile structure containing ESX-1-secreted proteins.
abstract: The cell envelope of mycobacteria, a group of Gram positive bacteria, is composed of a plasma membrane and a Gram-negative-like outer membrane containing mycolic acids. In addition, the surface of the mycobacteria is coated with an ill-characterized layer of extractable, non-covalently linked glycans, lipids and proteins, collectively known as the capsule, whose occurrence is a matter of debate. By using plunge freezing cryo-electron microscopy technique, we were able to show that pathogenic mycobacteria produce a thick capsule, only present when the cells were grown under unperturbed conditions and easily removed by mild detergents. This detergent-labile capsule layer contains arabinomannan, alpha-glucan and oligomannosyl-capped glycolipids. Further immunogenic and proteomic analyses revealed that Mycobacterium marinum capsule contains high amounts of proteins that are secreted via the ESX-1 pathway. Finally, cell infection experiments demonstrated the importance of the capsule for binding to cells and dampening of pro-inflammatory cytokine response. Together, these results show a direct visualization of the mycobacterial capsular layer as a labile structure that contains ESX-1-secreted proteins.
authors.affiliation: Division of Cell Biology-B6, Netherlands Cancer Institute-Antoni van Leeuwenhoek Hospital (NKI-AVL), Amsterdam, the Netherlands.
authors.forename: Musa
authors.initials: M
authors.lastname: Sani
authors.forename: Edith N G
authors.initials: EN
authors.lastname: Houben
authors.forename: Jeroen
authors.initials: J
authors.lastname: Geurtsen
authors.forename: Jason
authors.initials: J
authors.lastname: Pierson
authors.forename: Karin
authors.initials: K
authors.lastname: de Punder
authors.forename: Maaike
authors.initials: M
authors.lastname: van Zon
authors.forename: Brigitte
authors.initials: B
authors.lastname: Wever
authors.forename: Sander R
authors.initials: SR
authors.lastname: Piersma
authors.forename: Connie R
authors.initials: CR
authors.lastname: Jiménez
authors.forename: Mamadou
authors.initials: M
authors.lastname: Daffé
authors.forename: Ben J
authors.initials: BJ
authors.lastname: Appelmelk
authors.forename: Wilbert
authors.initials: W
authors.lastname: Bitter
authors.forename: Nicole
authors.initials: N
authors.lastname: van der Wel
authors.forename: Peter J
authors.initials: PJ
authors.lastname: Peters
mesh_terms: D016667:Bacterial Capsules / Q000378:metabolism* / Q000648:ultrastructure; D001425:Bacterial Outer Membrane Proteins / Q000378:metabolism* / Q000648:ultrastructure; D020285:Cryoelectron Microscopy; D016207:Cytokines / Q000378:metabolism; D006801:Humans; D008264:Macrophages / Q000276:immunology / Q000378:metabolism / Q000382:microbiology*; D016253:Microscopy, Immunoelectron; D009161:Mycobacterium / Q000378:metabolism* / Q000648:ultrastructure*; D009163:Mycobacterium bovis / Q000378:metabolism / Q000648:ultrastructure; D019910:Mycobacterium marinum / Q000378:metabolism / Q000648:ultrastructure; D020102:Mycobacterium smegmatis / Q000378:metabolism / Q000648:ultrastructure; D009169:Mycobacterium tuberculosis / Q000378:metabolism / Q000648:ultrastructure
chemical_list: D001425:Bacterial Outer Membrane Proteins; D016207:Cytokines