Processing and major histocompatibility complex class II presentation of Legionella pneumophila antigens by infected macrophages

To better understand interactions between the intracellular pathogen Legionella pneumophila and macrophages (Mphis), host and bacterial determinants important for presentation of antigens on major histocompatibility complex class II molecules (MHC-II) were investigated. It was determined that immune CD4 T-cell responses to murine bone marrow-derived Mphis (BMphis) infected with wild-type L. pneumophila were higher than the responses to avirulent dotA mutant bacteria. Although this enhanced response by immune T cells required modulation of vacuole transport mediated by the Dot/Icm system, it did not require intracellular replication of L. pneumophila. Intracellular cytokine staining identified a population of immune CD4 T cells that produced gamma interferon upon incubation with BMphis infected with wild-type L. pneumophila that did not respond to Mphi infection with dotA mutant bacteria. Endocytic processing was required for presentation of L. pneumophila antigens on MHC-II as determined by a defect in CD4 T-cell responses when the pH of BMphi endosomes was neutralized with chloroquine. Investigation of MHC-II presentation of antigens by BMphis infected with L. pneumophila icmR, icmW, and icmS mutants indicated that these mutants have an intermediate presentation phenotype relative to those of wild-type and dotA mutant bacteria. In addition, it was found that antigens from dot and icm mutants are presented earlier than antigens from wild-type L. pneumophila. Although immune CD4 T-cell responses to proteins secreted by the L. pneumophila Lsp system were not detected, it was found that the Lsp system is important for priming L. pneumophila-specific T cells in vivo. These data indicate that optimal antigen processing and MHC-II presentation to immune CD4 T cells involves synthesis of L. pneumophila proteins in an endoplasmic reticulum-derived compartment followed by transport to lysosomes.