Outer Membrane Protein A (OmpA) of Shigella flexneri 2a Links Innate and Adaptive Immunity in a TLR2-dependent Manner and Involvement of IL-12 and Nitric Oxide

• Published in: The Journal of biological chemistry Vol. 287; no. 15; pp. 12589 - 12601
• Main Authors: Pore, Debasis, Mahata, Nibedita, Chakrabarti, Manoj K.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 06.04.2012; American Society for Biochemistry and Molecular Biology
• Subjects: Adaptive Immunity; Animals; Antigen Presentation; Bacterial Outer Membrane Proteins - immunology; Cytokine; Enzyme Activation; Female; Flow Cytometry; HEK293 Cells; Histocompatibility Antigens Class II - genetics; Histocompatibility Antigens Class II - metabolism; Host-Pathogen Interactions; Humans; IFN-γ; Immunity, Innate; Immunology; Interferon-gamma - biosynthesis; Interleukin-12 - metabolism; Interleukin-12 - physiology; Interleukin-12 Receptor beta 2 Subunit - genetics; Interleukin-12 Receptor beta 2 Subunit - metabolism; Lymphocyte Activation; Macrophages - enzymology; Macrophages - immunology; Macrophages - metabolism; Mice; Mice, Inbred BALB C; NF-kappa B - metabolism; Nitric Oxide; Nitric Oxide - metabolism; Nitric Oxide Synthase Type II - metabolism; Outer Membrane Protein A; Phosphorylation; Protein kinase C (PKC); Protein Kinase C-alpha - metabolism; Protein Transport; Receptors, Chemokine - metabolism; Shigella flexneri - immunology; Shigella flexneri - physiology; Shigella flexneri 2a; T-Lymphocytes - immunology; T-Lymphocytes - metabolism; Th1 Response; TLR2; Toll-Like Receptor 2 - genetics; Toll-Like Receptor 2 - metabolism; Toll-Like Receptor 2 - physiology; IFN-γ; Flow Cytometry; Immunology; Th1 Response; Cytokine; Outer Membrane Protein A; Nitric Oxide; Shigella flexneri 2a; Protein kinase C (PKC); TLR2
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M111.335554

We determine that OmpA of Shigella flexneri 2a is recognized by TLR2 and consequently mediates the release of proinflammatory cytokines and activates NF-κB in HEK 293 cells transfected with TLR2. We also observe that in RAW macrophages TLR2 is essential to instigate the early immune response to OmpA via NF-κB activation and secretion of cytokines and NO. Consistent with these results, TLR2 knockdown using siRNA abolishes the initiation of immune responses. Processing and presentation of OmpA depend on TLR2; MHCII presentation of the processed antigen and expression of CD80 significantly attenuated in TLR2 knockdown macrophages. The optimum production of IFN-γ by the macrophages:CD4+ T cells co-culture depends on both TLR2 activation and antigen presentation. So, TLR2 is clearly recognized as a decisive factor in initiating host innate immune response to OmpA for the development of CD4+ T cell adaptive response. Furthermore, we demonstrate in vivo that intranasal immunization of mice with OmpA selectively enhances the release of IFN-γ and IL-2 by CD4+ T cells. Importantly, OmpA increases the level of IFN-γ production in Ag-primed splenocytes. The addition of neutralizing anti-IL-12p70 mAb to cell cultures results in the decreased release of OmpA-enhanced IFN-γ by Ag-primed splenocytes. Moreover, coincubation with OmpA-pretreated macrophages enhances the production of IFN-γ by OmpA-primed CD4+ T cells, representing that OmpA may enhance IFN-γ expression in CD4+ T cells through the induction of IL-12 production in macrophages. These results demonstrate that S. flexneri 2a OmpA may play a critical role in the development of Th1 skewed adaptive immune response. Background: OmpA activates innate immunity through TLR2, but its function and mechanism of targeting adaptive immunity remains unexplored. Results: TLR2 regulates OmpA processing and presentation, NO and cytokine release, which contributes to development of type-1 adaptive immunity. Conclusion: TLR2 is crucial to link innate and adaptive responses stimulated by OmpA. Significance:S. flexneri 2a OmpA, a novel molecule, coordinates the innate and adaptive responses.