De novo synthesis of Legionella pneumophila antigens during intracellular growth in phagocytic cells.

Abstract

Legionella pneumophilia is a gram-negative rod which is able to multiply within phagocytic cells. The process of phagocytosis leads to a rapid environmental change that might require a coordinate regulation of gene expression to ensure intracellular survival. Since there is little information on up- and downregulation of genes during the early phases of phagocytosis, we radiolabeled intracellular L. pneumophila at different times after phagocytosis by macrophages of the Mono Mac 6 cell line and immunoprecipitated antigens with antilegionella sera or monoclonal antibodies. We could identify two antigens which were upregulated, one of which was the Mip protein, three antigens which were downregulated, and three antigens which were not detectable in extracellularly grown L. pneumophila. The Mip protein was stained most intensively 4 to 8 h after intracellular infection, suggesting that it is needed during intracellular multiplication rather than initiation of infection. A 44-kDa antigen which was not detectable during extracellular growth was most prominent from 2 to 4 h postinfection when Mono Mac 6 cells were used as phagocytic cells. The 44-kDa antigen was also expressed during growth with Acanthamoeba castelanii, MRC-5, and U937 cells but with different kinetics. Synthesis of this antigen was not dependent on protein synthesis of the host cell. Since the 44-kDa antigen could be precipitated by an antiserum produced against a recombinant Escherichia coli harboring a plasmid with an L. pneumophila insert which also codes for the mip gene, we believe that the corresponding gene is within the vicinity of the mip gene. We named this protein legionella intracellular growth antigen (LIGA), since it could be found exclusively in intracellularly grown L. pneumophila.