title: Differential expression of mycobacterial proteins following phagocytosis by macrophages.
abstract: Mycobacterium tuberculosis resides within the macrophages of the host, but the molecular and cellular mechanisms of survival are poorly understood. Recent evidence suggests that the attenuated vaccine strain Mycobacterium bovis BCG is both a deletion and regulatory mutant, yet retains both its immunoprotective and intra-macrophage survival potential. In an attempt to define M. bovis BCG genes expressed during interaction with macrophages, the patterns of protein synthesis were examined by both one- and two-dimensional gel electrophoresis of BCG while inside the human leukaemic macrophage cell line THP-1. This study demonstrated that BCG expresses proteins while resident inside macrophages that are not expressed during in vitro growth in culture media or under conditions of heat shock. Western blotting analysis revealed that some of the differentially expressed proteins are specifically recognized by human M. tuberculosis-infected sera. Proteome analysis by two-dimensional electrophoresis and MS identified six abundant proteins that showed increased expression inside macrophages: 16 kDa alpha-crystallin (HspX), GroEL-1 and GroEL-2, a 31.7 kDa hypothetical protein (Rv2623), InhA and elongation factor Tu (Tuf). Identification of proteins by such a strategy will help elucidate the molecular basis of the attenuation and the vaccine potential of BCG, and may provide antigens that distinguish infection with M. tuberculosis from vaccination with BCG.
authors.affiliation: Department of Medical Microbiology, St George's Hospital Medical School, Cranmer Terrace, London SW17 ORE, UK1.
authors.forename: Irene M
authors.initials: IM
authors.lastname: Monahan
authors.affiliation: Respiratory Systems, Glaxo Wellcome Research and Development, Medicines Research Centre, Gunnels Wood Road, Stevenage SG1 2NY, UK2.
authors.forename: Joanna
authors.initials: J
authors.lastname: Betts
authors.affiliation: Department of Medical Microbiology, St George's Hospital Medical School, Cranmer Terrace, London SW17 ORE, UK1.
authors.forename: Dilip K
authors.initials: DK
authors.lastname: Banerjee
authors.affiliation: Department of Medical Microbiology, St George's Hospital Medical School, Cranmer Terrace, London SW17 ORE, UK1.
authors.forename: Philip D
authors.initials: PD
authors.lastname: Butcher
mesh_terms: D000907:Antibodies, Bacterial / Q000276:immunology; D000942:Antigens, Bacterial / Q000276:immunology; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D015153:Blotting, Western; D015180:Electrophoresis, Gel, Two-Dimensional; D006801:Humans; D008264:Macrophages / Q000382:microbiology*; D009163:Mycobacterium bovis / Q000254:growth & development / Q000378:metabolism*; D009169:Mycobacterium tuberculosis / Q000276:immunology; D010587:Phagocytosis; D020543:Proteome; D014376:Tuberculosis / Q000276:immunology / Q000382:microbiology
chemical_list: D000907:Antibodies, Bacterial; D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D020543:Proteome