title: A polymeric protein induces specific cytotoxicity in a TLR4 dependent manner in the absence of adjuvants.
abstract: Lumazine synthase from Brucella spp. (BLS) is a highly immunogenic decameric protein. It is possible to insert foreign peptides or proteins at its ten-amino acid termini. These chimeras elicit systemic and oral immunity without adjuvants, which are commonly needed in the formulation of subunit-based vaccines. Here, we show that BLS induces the cross presentation of a covalently attached peptide OVA(257-264) and a specific cytotoxic response to this peptide in the absence of adjuvants. Unlike other subunit-based vaccines, this chimera induces rapid activation of CTLs and a specific cytotoxic response, making this polymeric protein an ideal antigen carrier for vaccine development. Adoptive transfer of transgenic OT-I T cells revealed efficient cross presentation of BLS-OVA(257-264)in vivo. BLS-OVA(257-264) immunization induced the proliferation of OVA(257-264)-specific CD8+ lymphocytes and also increased the percentage of OVA(257-264)-specific CD8+ cells expressing the early activation marker CD69; after 5 days, the percentage of OVA(257-264)-specific CD8+ cells expressing high levels of CD44 increased. This cell subpopulation showed decreased expression of IL-7Rα, indicating that BLS-OVA(257-264) induced the generation of CD8+ effector cells. BLS-OVA(257-264) was cross presented in vitro independently of the presence of a functional TLR4 in the DCs. Finally, we show that immunization of wild type mice with the chimera BLS-OVA(257-264) without adjuvants induced a strong OVA(257-264)-specific effector cytotoxic response. This cytotoxicity is dependent on TLR4 as is not induced in mice lacking a functional receptor. These data show that TLR4 signaling is necessary for the induction of a cytotoxic response but not for antigen cross presentation.
authors.affiliation: Fundación Instituto Leloir, IIBBA, Consejo Nacional de Investigaciones Científicas y Técnicas (CONICET), Buenos Aires, Argentina. pau_berguer@yahoo.com
authors.forename: Paula M
authors.initials: PM
authors.lastname: Berguer
authors.forename: Vanina A
authors.initials: VA
authors.lastname: Alzogaray
authors.forename: Andrés Hugo
authors.initials: AH
authors.lastname: Rossi
authors.forename: Juliana
authors.initials: J
authors.lastname: Mundiñano
authors.forename: Isabel
authors.initials: I
authors.lastname: Piazzon
authors.forename: Fernando A
authors.initials: FA
authors.lastname: Goldbaum
mesh_terms: D000276:Adjuvants, Immunologic / Q000494:pharmacology; D000818:Animals; D001704:Biopolymers; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology; D045142:Cross-Priming / Q000276:immunology; D003602:Cytotoxicity, Immunologic / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D005456:Fluorescent Dyes; D008213:Lymphocyte Activation; D051379:Mice; D008810:Mice, Inbred C57BL; D009097:Multienzyme Complexes / Q000276:immunology*; D010047:Ovalbumin / Q000276:immunology; D016133:Polymerase Chain Reaction; D051197:Toll-Like Receptor 4 / Q000502:physiology*
chemical_list: D000276:Adjuvants, Immunologic; D001704:Biopolymers; D005456:Fluorescent Dyes; D009097:Multienzyme Complexes; D051197:Toll-Like Receptor 4; C044822:6,7-dimethyl-8-ribityllumazine synthase; D010047:Ovalbumin