Protocol

Legionella

Volume 954 of the series Methods in Molecular Biology pp 333-344

Date:

Subcellular Localization of Legionella Dot/Icm Effectors

  • Adam J. VogrinAffiliated withDepartment of Microbiology and Immunology, University of Melbourne
  • , Aurelie MousnierAffiliated withDivision of Cell and Molecular Biology, Centre for Molecular Microbiology and Infection, Imperial College of London
  • , Gad FrankelAffiliated withDivision of Cell and Molecular Biology, Centre for Molecular Microbiology and Infection, Imperial College London
  • , Elizabeth L. HartlandAffiliated withDepartment of Microbiology and Immunology, University of Melbourne Email author 

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Abstract

The translocation of effector proteins by the Dot/Icm type IV secretion system is central to the ability of Legionella pneumophila to persist and replicate within eukaryotic cells. The subcellular localization of translocated Dot/Icm proteins in host cells provides insight into their function. Through co-staining with host cell markers, effector proteins may be localized to specific subcellular compartments and membranes, which frequently reflects their host cell target and mechanism of action. In this chapter, we describe protocols to (1) localize effector proteins within cells by ectopic expression using green fluorescent protein fusions and (2) localize effector proteins within infected cells using epitope-tagged effector proteins and immuno-fluorescence microscopy.

Key words

Legionella pneumophila HEK293T cells Epitope tagging Laser scanning confocal microscopy Transfection Infection Type IV secretion