A powerful IFN-γ response is triggered upon infection with the protozoan parasite, Toxoplasma gondii. Several cell populations, including dendritic cells (DCs), macrophages, and neutrophils, produce IL-12, a key cytokine for IFN-γ induction. However, it is still unclear which of the above cell populations is its main source. Diphtheria toxin (DT) injection causes transient DC depletion in a transgenic mouse expressing Simian DT receptors under the control of the CD11c promoter, allowing us to investigate the role of DCs in IL-12 production. T. gondii-inoculated DT-treated and control groups were monitored for IL-12 levels and survival. We show in this study that DC depletion abolished IL-12 production and led to mortality. Furthermore, replenishment with wild-type, but not MyD88- or IL-12p35-deficient, DCs rescued IL-12 production, IFN-γ-induction, and resistance to infection in DC-depleted mice. Taken together, the results presented in this study indicate that DCs constitute the major IL-12-producing cell population in vivo during T. gondii infection.

NF-κB/Rel transcription factors are linked to innate immune responses and APC activation. Whether and how the induction of NF-κB signaling in normal CD4+ T cells regulates effector function are not well-understood. The liberation of NF-κB dimers from inhibitors of κB (IκBs) constitutes a central checkpoint for physiologic regulation of most forms of NF-κB. To investigate the role of NF-κB induction in effector T cell responses, we targeted inhibition of the NF-κB/Rel pathway specifically to T cells. The Th1 response in vivo is dramatically weakened when T cells defective in their NF-κB induction (referred to as IκBα(ΔN) transgenic cells) are activated by a normal APC population. Analyses in vivo, and IL-12-supplemented T cell cultures in vitro, reveal that the mechanism underlying this T cell-intrinsic requirement for NF-κB involves activation of the IFN-γ gene in addition to clonal expansion efficiency. The role of NF-κB in IFN-γ gene expression includes a modest decrease in Stat4 activation, T box expressed in T cell levels, and differentiation efficiency along with a more prominent postdifferentiation step. Further, induced expression of Bcl-3, a trans-activating IκB-like protein, is decreased in T cells as a consequence of NF-κB inhibition. Together, these findings indicate that NF-κB induction in T cells regulates efficient clonal expansion, Th1 differentiation, and IFN-γ production by Th1 lymphocytes at a control point downstream from differentiation.

Loss of Bruton’s tyrosine kinase (Btk) function results in mouse Xid disease characterized by a reduction in mature B cells and impaired humoral immune responses. These defects have been mainly attributed to impaired BCR signaling including reduced activation of the classical NF-κB pathway. In this study we show that Btk also couples the receptor for B cell-activating factor (BAFF) of the TNF family (BAFF-R) to the NF-κB pathway. Loss of Btk results in defective BAFF-mediated activation of both classical and alternative NF-κB pathways. Btk appears to regulate directly the classical pathway in response to BAFF such that Btk-deficient B cells exhibit reduced kinase activity of IκB kinase γ-containing complexes and defective IκBα degradation. In addition, Btk-deficient B cells produce reduced levels of NF-κB2 (p100) basally and in response to stimulation via the BCR or BAFF-R, resulting in impaired activation of the alternative NF-κB pathway by BAFF. These results suggest that Btk regulates B cell survival by directly regulating the classical NF-κB pathway under both BCR and BAFF-R, as well as by inducing the expression of the components of alternative pathway for sustained NF-κB activation in response BAFF. Thus, impaired BCR- and BAFF-induced signaling to NF-κB may contribute to the observed defects in B cell survival and humoral immune responses in Btk-deficient mice.

CD4+ T cells with a block in the NF-κB signaling pathway exhibit decreases in Th1 responses and diminished nuclear levels of multiple transactivating NF-κB/Rel/IκB proteins. To determine the lineage-intrinsic contributions of these transactivators to Th differentiation, T cells from mice deficient in specific subunits were cultured in exogenous cytokines promoting either Th1 or Th2 differentiation. RelB-deficient cells exhibited dramatic defects in Th1 differentiation and IFN-γ production, whereas no consistent defect in either Th1 or Th2 responses was observed with c-Rel-deficient cells. In sharp contrast, Bcl-3-null T cells displayed no defect in IFN-γ production, but their Th2 differentiation and IL-4, IL-5, and IL-13 production were significantly impaired. The absence of RelB led to a dramatic decrease in the expression of T-box expressed in T cells and Stat4. In contrast, Bcl-3-deficient cells exhibited decreased GATA-3, consistent with evidence that Bcl-3 can transactivate a gata3 promoter. These data indicate that Bcl-3 and RelB exert distinct and opposing effects on the expression of subset-determining transcription factors, suggesting that the characteristics of Th cell responses may be regulated by titrating the stoichiometry of transactivating NF-κB/Rel/IκB complexes in the nuclei of developing helper effector cells.