Sca-1 expression defines developmental stages of mouse pDCs that show functional heterogeneity in the endosomal but not lysosomal TLR9 response.

Plasmacytoid dendritic cells (pDCs) play an important role in innate and adaptive immunity and were shown to be identical to previously described natural interferon (IFN)-α-producing cells. Here, we describe two functionally distinct pDC subpopulations that are characterized by the differential expression of stem cell antigen-1 (Sca-1; Ly-6A/E). Sca-1(-) pDCs are mainly found in the BM, appear first during development, show a higher proliferative activity, and represent the more precursor phenotype. Sca-1(+) pDCs are mostly located in secondary lymphoid organs and represent a later developmental stage. Sca-1(-) pDCs give rise to an Sca-1(+) subset upon activation or in response to endogenous type I IFN. Interestingly, in contrast to Sca-1(-) pDCs, Sca-1(+) pDCs are defective in IFN-α production upon endosomal TLR9 stimulation, whereas lysosomal signaling via TLR9 is functional in both subsets. Gene expression analysis revealed that osteopontin is strongly upregulated in Sca-1(-) pDCs. These data provide evidence for the molecular basis of the observed functional heterogeneity, as the intracellular isoform of osteopontin couples TLR9 signaling to IFN-α expression. Taken together, our results indicate that Sca-1(-) pDCs are an early developmental stage of pDCs with distinct innate functions representing the true murine natural IFN-α-producing cells.

Enhanced dendritic cell-induced immune responses mediated by the novel C-type lectin receptor mDCAR1.

The dendritic cell (DC) immunoreceptors (DCIR) and DC-immunoactivating receptors (DCAR) represent a subfamily of cell surface C-type lectin receptors (CLR), whose multifunctional capacities range from classical Ag uptake and immunoregulatory mechanisms to the involvement in DC ontogeny. On the basis of the generation of specific mAbs, we functionally characterized mouse DCAR1 (mDCAR1) as a member of the DCIR/DCAR family. Expression of mDCAR1 was strongly tissue dependent. mDCAR1 expression on DCs was restricted to the CD8(+) DC subset in spleen and thymus and on subpopulations of CD11b(+) myeloid cells in bone marrow and spleen, whereas the molecule was not detectable on both cell types in lymph nodes and peripheral blood. With respect to the function of CLRs as pattern recognition receptors, Ag delivered via mDCAR1 was internalized, was trafficked to early and late endosomes/lysosomes and, as a consequence, induced cellular and humoral responses in vivo even in the absence of CD40 stimulation. Intriguingly, upon triggering mDCAR1, CD8(+) DCs increased the secretion of bioactive IL-12, whereas IL-10 release is markedly reduced, thereby indicating that Ag recognized by mDCAR1 induces enhanced proinflammatory responses. These data indicate that mDCAR1 is a functional receptor on cells of the immune system and provides further insights into the regulation of immune responses by CLRs.

Extensive characterization of IFN-induced GTPases mGBP1 to mGBP10 involved in host defense.

IFN-gamma orchestrates a potent antimicrobial host response. However, the underlying molecular basis for this immunological defense system is largely unknown. In a systematic approach to identify IFN-gamma-regulated host effector molecules, a notable number of transcripts with consensus GTP-binding motives were obtained. Further extensive transcriptome and genome analyses identified five novel family members of murine guanylate-binding proteins (mGBPs) now designated mGBP6, 7, 8, 9, and 10. Moreover, in this study, all 10 mGBP members (mGBP1-10) were extensively characterized. mGBPs are selectively up-regulated in vitro by a set of proinflammatory cytokines and TLR agonists as well as in vivo after Listeria monocytogenes and Toxoplasma gondii infection. After IFN-gamma stimulation, mGBP1, 2, 3, 6, 7, and 9 are associated with intracellular Toxoplasma parasites and, interestingly, virulent Toxoplasma interfere with mGBP recruitment. Taken together, mGBPs comprise an important set of host defense molecules.