Cysteinyl leukotrienes (cys-LTs) can induce inflammatory responses and activate mast cells via their G protein-coupled receptors, CysLT1R and CysLT2R. The purinergic (P2Y) receptors have homology to the CysLTRs, and the nucleotide uridine diphosphate (UDP) can stimulate similar signaling in mast cells to that induced via CysLT1R. Jiang et al. (p. 1129) therefore hypothesized that CysLT1R and a UDP-responsive receptor might interact to modulate mast cell activity. Both human and mouse mast cells were found to express the UDP receptor P2Y6. Calcium flux and ERK activation induced by the CysLT1R agonist leukotriene D4 (LTD4) were attenuated by a P2Y6 antagonist, suggesting that LTD4 signaling involved cooperation between P2Y6 and CysLT1R. LTD4 and UDP both reduced mast cell apoptosis through a mechanism requiring P2Y6 and endogenous cys-LTs, and LTD4-induced MIP-1β production required UDP and P2Y6. Together, these data suggested that P2Y6 and CysLT1R signaling were closely interrelated; however, despite colocalization of these two receptors at the cell surface, they did not appear to directly interact. This observation that CysLT1R and P2Y6 reciprocally amplify one another’s activity to mediate mast cell activation and survival reveals a potentially important mechanism in innate immune regulation.

Although pattern recognition receptors (PRRs) have been extensively studied in microbial infection, less is known about their involvement in allergic inflammation. Previous studies suggested that cysteinyl leukotrienes (cys-LTs) regulated dendritic cell (DC) function in an autocrine manner to promote Th2-dependent allergic inflammation, but the mechanism by which this might occur was unclear. To clarify the involvement of DC PRRs in allergic inflammation, Barrett et al. (p. 1119) asked how airborne allergens from house dust mites might induce cys-LT production in DC. Dust mite extracts were shown to induce cys-LT production from pulmonary DC but not from alveolar or peritoneal macrophages. Although proteases have been proposed to be responsible for dust mite immunogenicity, the cys-LT-inducing ligand was identified to be a glycan. Dust mite-induced cys-LT production was dependent on signaling via Syk and the FcRγ-chain, and further analysis of receptors that signal through FcRγ identified the mannose-binding PRR Dectin-2 as a receptor for dust mite extracts. Further analysis confirmed that Dectin-2 was a functional receptor for a dust mite-derived glycan and was responsible for allergen-elicited cys-LT production in DC. These data present a mechanism by which indoor allergens can induce allergic inflammation that may be useful for future immunotherapy of allergic disease and asthma.

The B cell-activating factor of the TNF family (BAFF) is a cytokine important for B cell survival and activation that has been proposed to have a proinflammatory effect on T cell responses. The precise role of BAFF in T cell regulation is not clear, leading Walters et al. (p. 793) to examine T cell responses in mice overexpressing BAFF (BAFF-Tg). Such overexpression of BAFF impaired T cell responses, as demonstrated by the prolonged survival of grafts in the BAFF-Tg mice. However, BAFF-Tg T cells responded normally to TCR stimulation and effector T cells from these mice effectively rejected allografts, indicating that BAFF overexpression did not impair effector T cell function. Instead, the authors observed a dramatic increase in the frequency of CD4+CD25+FoxP3+ regulatory T cells (Treg) in BAFF-Tg mice compared with wild-type mice. These Treg were found to be directly responsible for the impaired T cell responses observed in BAFF-Tg mice. Interestingly, the Treg expansion was not caused by direct effects of BAFF on T cells but instead required B cell involvement. BAFF is therefore not a purely stimulatory molecule, as BAFF-stimulated B cells can promote Treg expansion and the subsequent suppression of T cell-mediated immune responses.

Type I IFNs can act on nearly all types of cells and are vital for the control of viral infections, but responses to these potent cytokines vary between cell types. Plasmacytoid dendritic cells (pDC) are known to produce the first wave of type I IFNs during infection with the highly cytopathic murine hepatitis virus (MHV), but the cell targets of IFN early in infection have not been elucidated. To identify these targets, Cervantes-Barragán et al. (p. 1099) first created bone marrow chimeric mice expressing the IFN-α receptor (IFNAR) on either hematopoietic or nonhematopoietic cells and determined that IFNAR expression on hematopoietic cells was required for early control of MHV infection. Within the hematopoietic compartment, macrophages and conventional DC (cDC) were most susceptible to MHV infection in vivo. Use of conditionally gene-targeted mice lacking IFNAR expression in specific cell types indicated that IFNAR expression on macrophages, and to a lesser degree on cDC, was important for survival following MHV infection. A lack of IFNAR on pDC, T cells, or B cells, however, did not significantly impact resistance to MHV. Finally, pDC-derived IFN could be shown to protect macrophages and cDC from MHV replication in vitro. These data identify the cell populations involved in early IFN-mediated antiviral defense and have implications for future attempts to control cytopathic virus infections.

Dectin-1, a pattern recognition receptor, mediates phagocytosis of pathogenic fungi and cooperates with TLR2 to initiate the antifungal response. Signaling through Dectin-1 was shown to directly activate NF-κB in dendritic cells (DC) via the caspase recruitment domain (CARD)-containing adaptor protein CARD9. However, as previous studies also showed that Dectin-1 signals were insufficient to activate NF-κB in macrophages, Goodridge et al. (p. 1146) reassessed Dectin-1-mediated signaling in macrophages and DC. Consistent with the literature, Dectin-1-CARD9 signaling directly induced NF-κB activation and subsequent TNF-α production in bone marrow-derived DC but not in bone marrow-derived macrophages, although the addition of GM-CSF or IFN-γ could overcome this deficiency in macrophages. Other pathways of Dectin-1 signaling were active in macrophages, as CARD9 was recruited to Dectin-1-containing phagosomes following stimulation with the fungal cell wall preparation zymosan, and Dectin-1-CARD9 signaling activated p38 MAP kinase phosphorylation and enhanced TLR2-mediated TNF-α production. Interestingly, different populations of macrophages and DC varied in their ability to produce TNF-α in response to Dectin-1 stimulation, suggesting that the ability of Dectin-1-CARD9 signaling to activate NF-κB is dependent upon myeloid cell lineage combined with the cytokine environment. These data indicate that antifungal responses mediated by Dectin-1 signaling are more complex than previously appreciated.

Mycobacterial infections induce the rapid expansion of Vγ2Vδ2 T cells, which are only present in primates and protect against tuberculosis in rhesus monkeys. To investigate cytokines that might be involved in Vγ2Vδ2 T cell regulation, Yuan et al. (p. 811) analyzed cytokine mRNA production in mycobacterially infected macaques and identified a novel IL-4 splice variant that they dubbed variant IL-4 (VIL-4). VIL-4 consisted of an N-terminal 97-aa region identical to that of IL-4 and a unique 96-aa C-terminal tail containing a proline-rich motif. This variant cytokine stimulated expansion of (E)-4-hydroxy-3-methyl-but-2-enyl pyrophosphate (HMBPP)-stimulated Vγ2Vδ2 T cells to a much greater extent than did IL-4 and appeared to act via the IL-4R. Vγ2Vδ2 T cells produced IL-4, IFN-γ, and TNF-α after stimulation with HMBPP and VIL-4, but not after stimulation with HMBPP and IL-4. Thus, this identification of a novel IL-4 splice variant that induces expansion and effector cytokine production by Vγ2Vδ2 T cells suggests a mechanism by which these cells may be activated during mycobacterial infection and may lead to new ideas for therapeutic augmentation of antimicrobial responses.

The role of plasmacytoid dendritic cells (pDC) in transplantation and graft-vs-host disease (GVHD) is somewhat controversial, leading Banovic et al. (p. 912) to attempt to clarify the effects of pDC on GVHD. Host pDC were found to be completely depleted by total body irradiation before transplantation, which allowed the authors to focus on the activities of donor pDC. Depletion of pDC from bone marrow grafts led to exacerbated GVHD compared with transfer of control grafts, but depletion of mature pDC from splenic grafts did not affect GVHD development. It therefore appeared that bone marrow pDC could attenuate GVHD but lost this immunomodulatory ability after maturation. Donor pDC failed to reconstitute in animals with GVHD; however, a pre-DC population reconstituted quickly and transiently. These pre-DC lacked B220 expression and failed to produce IFN-α but otherwise demonstrated a pDC-like phenotype. In addition, these cells showed characteristics of common DC precursors and could differentiate into both pDC and conventional DC in the absence of GVHD. The reconstituting pre-DC strongly inhibited allogeneic T cell proliferation in vitro via a mechanism requiring cell contact and suppressed GVHD in vivo. Thus, bone marrow-derived pDC and pre-DC suppress GVHD, whereas GVHD inhibits pDC maturation and thereby somewhat paradoxically promotes immunosuppression.

The integrin LFA-1 mediates binding of NK cells to their targets and induces signaling pathways important for cytotoxic activity. Talin, a cytoskeletal adaptor protein, is thought to be important for integrin activation and is found in the NK cell:target cell immune synapse. Mace et al. (p. 948) sought to determine what role talin might play in LFA-1-mediated NK cell activity by generating talin−/− NK cells from talin-deficient embryonic stem cells. As predicted, talin−/− NK cells did not bind to ICAM-1, indicating that talin was required for LFA-1-mediated adhesion. Interestingly, talin−/− NK cells also showed severely impaired cytotoxicity, even if LFA-1 adhesion was enhanced by the presence of manganese, suggesting that talin might be involved in LFA-1 outside-in signaling in addition to the inside-out signaling required for adhesion. Analysis of this outside-in signaling indicated that LFA-1 interaction with ICAM-1 led to talin recruitment and talin-dependent actin accumulation at the site of adhesion. Coligation of LFA-1 and the activating NK cell receptor NKG2D led to polarization of granzyme-containing cytotoxic granules that also required the presence of talin. Taken together, these data clarify some of the signals required for successful NK cell-mediated cytotoxicity and the involvement of talin in LFA-1 function.

Summaries written by Jennifer Hartt Meyers, Ph.D.