Direct Visualization by cryo-EM of the Mycobacterial Capsular Layer: A Labile Structure Containing ESX-1-secreted Proteins

PLoS Pathog. 2010 Mar 5;6(3):e1000794. doi: 10.1371/journal.ppat.1000794.

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.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Bacterial Capsules / metabolism*
  • Bacterial Capsules / ultrastructure
  • Bacterial Outer Membrane Proteins / metabolism*
  • Bacterial Outer Membrane Proteins / ultrastructure
  • Cryoelectron Microscopy
  • Cytokines / metabolism
  • Humans
  • Macrophages / immunology
  • Macrophages / metabolism
  • Macrophages / microbiology*
  • Microscopy, Immunoelectron
  • Mycobacterium / metabolism*
  • Mycobacterium / ultrastructure*
  • Mycobacterium bovis / metabolism
  • Mycobacterium bovis / ultrastructure
  • Mycobacterium marinum / metabolism
  • Mycobacterium marinum / ultrastructure
  • Mycobacterium smegmatis / metabolism
  • Mycobacterium smegmatis / ultrastructure
  • Mycobacterium tuberculosis / metabolism
  • Mycobacterium tuberculosis / ultrastructure

Substances

  • Bacterial Outer Membrane Proteins
  • Cytokines