title: TLR4-mediated activation of dendritic cells by the heat shock protein DnaK from Francisella tularensis.
abstract: Francisella tularensis is the causative agent of tularemia, a severe, debilitating disease of humans and other mammals. As this microorganism is also classified as a "category-A pathogen" and a potential biowarfare agent, there is a need for an effective vaccine. Several antigens of F. tularensis, including the heat shock protein DnaK, have been proposed for use in a potential subunit vaccine. In this study, we characterized the innate immune response of murine bone marrow-derived dendritic cells (DC) to F. tularensis DnaK. Recombinant DnaK was produced using a bacterial expression system and purified using affinity, ion-exchange, and size-exclusion chromatography. DnaK induced the activation of MAPKs and NF-kappaB in DC and the production of the proinflammatory cytokines IL-6, TNF-alpha, and IL-12 p40, as well as low levels of IL-10. DnaK induced phenotypic maturation of DC, as demonstrated by an up-regulation of costimulatory molecules CD40, CD80, and CD86. DnaK stimulated DC through TLR4 and the adapters MyD88 and Toll/IL-1R domain-containing adaptor-inducing IFN-beta (TRIF) that mediated differential responses. DnaK induced activation of MAPKs and NF-kappaB in a MyD88- or TRIF-dependent manner. However, the presence of MyD88- and TRIF-dependent signaling pathways was essential for an optimal, DnaK-induced cytokine response in DC. In contrast, DnaK induced DC maturation in a TRIF-dependent, MyD88-independent manner. These results provide insight about the molecular interactions between an immunodominant antigen of F. tularensis and host immune cells, which is crucial for the rational design and development of a safe and efficacious vaccine against tularemia.
authors: Department of Microbiology, University of Alabama at Birmingham, 845 19th Street South, BBRB 258/5, Birmingham, AL 35294-2170, USA.
authors: Amit R
authors: AR
authors: Ashtekar
authors: Ping
authors: P
authors: Zhang
authors: Jannet
authors: J
authors: Katz
authors: Champion C S
authors: CC
authors: Deivanayagam
authors: Prasad
authors: P
authors: Rallabhandi
authors: Stefanie N
authors: SN
authors: Vogel
authors: Suzanne M
authors: SM
authors: Michalek
mesh_terms: D033942:Adaptor Proteins, Vesicular Transport / Q000378:metabolism; D000818:Animals; D018122:B7-1 Antigen / Q000378:metabolism; D051940:B7-2 Antigen / Q000378:metabolism; D015153:Blotting, Western; D002478:Cells, Cultured; D003713:Dendritic Cells / Q000378:metabolism*; D004731:Endotoxins / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D005604:Francisella tularensis / Q000201:enzymology*; D006360:Heat-Shock Proteins / Q000302:isolation & purification / Q000494:pharmacology*; D006801:Humans; D007371:Interferon-gamma / Q000378:metabolism; D018664:Interleukin-12 / Q000378:metabolism; D007668:Kidney / Q000166:cytology / Q000276:immunology / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism; D053594:Myeloid Differentiation Factor 88 / Q000502:physiology; D016328:NF-kappa B / Q000378:metabolism; D010641:Phenotype; D051195:Toll-Like Receptor 2 / Q000502:physiology; D051197:Toll-Like Receptor 4 / Q000502:physiology*; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
chemical_list: D033942:Adaptor Proteins, Vesicular Transport; D018122:B7-1 Antigen; D051940:B7-2 Antigen; D004731:Endotoxins; D006360:Heat-Shock Proteins; D007371:Interferon-gamma; D018664:Interleukin-12; D008070:Lipopolysaccharides; D020929:Mitogen-Activated Protein Kinase Kinases; D053594:Myeloid Differentiation Factor 88; D016328:NF-kappa B; D051195:Toll-Like Receptor 2; D051197:Toll-Like Receptor 4; D014409:Tumor Necrosis Factor-alpha; C507780:Myd88 protein, mouse; C476286:Ticam1 protein, mouse; C493486:Tlr2 protein, mouse; C493487:Tlr4 protein, mouse