IN THIS ISSUE

Polymorphisms (L263P, M313V, and S331P) in an intracellular loop of the G protein-coupled receptor (GPCR) murine histamine receptor H₁ (H₁R) increase susceptibility to experimental allergic encephalomyelitis (EAE) and experimental allergic orchitis. Previous work identified an EAE-associated phenotype of Bordetella pertussis toxin-induced histamine sensitization controlled by the autosomal dominant Bphs gene locus that encodes H₁R. Noubade et al. (p. 7471 ) tested the expression of two haplotypes in T cells: the P-V-P susceptible haplotype (H₁R^S) and the L-M-S resistant haplotype (H₁R^R). This group had previously shown that the selective re-expression of H₁R^S in the T cells of H₁R-deficient animals was sufficient to restore EAE and produce IFN-γ while expression of the resistant allele was not. This dichotomy was attributed to the difference in surface expression between the two receptor alleles; H₁R^R was expressed at lower levels than H₁R^S and was found to be retained within the endoplasmic reticulum. Both forms were equivalently able to activate downstream G proteins. This group has elegantly demonstrated that structural polymorphisms in a T cell GPCR can alter autoimmune susceptibility by determining surface expression.

Regulatory T cells (Treg) that express CD4 and CD25 are increasingly known to be a heterogenous population. Chen et al. (p. 7327 ) have expanded on that heterogeneity by identifying a CD4⁺CD25⁺ Treg subset that has surface expression of the latency-associated peptide (LAP) and potent suppressive activity in a model of experimental autoimmune encephalomyelitis (EAE). These CD4⁺CD25⁺LAP⁺ cells expressed elevated levels of FoxP3, GITR, CTLA-4, and TGF-β receptors in addition to membrane-bound and secreted TGF-β. In contrast to CD4⁺CD25⁺LAP⁻ cells, whose in vitro suppressive activity was cell contact dependent, this novel subset of CD4⁺CD25⁺LAP⁺ cells had both soluble and contact-dependent suppressive effects. In vivo, the suppressive activity of CD4⁺CD25⁺LAP⁺ cells was TGF-β dependent and more potent than that exerted by CD4⁺CD25⁺LAP⁻ cells, as adoptive transfer of CD4⁺CD25⁺LAP⁺ cells but not CD4⁺CD25⁺LAP⁻ cells was able to ameliorate EAE. Neutralizing Ab to TGF-β blocked this amelioration. CD4⁺CD25⁺LAP⁺ cells suppressed the effects of myelin oligodendrocyte protein-specific effector T cells by converting them into FoxP3-expressing cells and inhibiting IFN-γ and IL-17 production. Thus, the authors have identified another population of regulatory T cells, identified LAP as a marker defining this subset, and helped to clarify the somewhat controversial role of TGF-β in Treg suppression.

During Bacillus anthracis infection the vascular endothelium, which expresses high levels of anthrax toxin receptors, undergoes disruption of normal function and integrity. This causes vascular leakage, vasculitis, and impaired immune responses to infection. Warfel et al. (p.7516 ) have demonstrated that the anthrax lethal toxin (LT) contributes to this endothelial dysfunction by transcriptionally enhancing cytokine-induced VCAM-1 expression. Endothelial cells treated with TNF-α and LT showed enhanced VCAM-1 expression over TNF-α or LT treatment alone. This enhancement correlated with NF-κB phosphorylation and nuclear translocation and delayed reaccumulation of Iκ-Bα, and also increased STAT1-mediated IFN regulatory factor-1 (IRF-1) induction. However, LT did not increase TNF-α-mediated expression of ICAM-1 and E-selectin. Because these molecules are also controlled by NF-κB but not by IRF-1, the authors concluded that LT is capable of differentially influencing NF-κB target genes. Thus, these data create an insight into how LT influences the host response to anthrax infection and describe a role for IRF-1 in influencing VCAM-1 expression.

Whereas cytokines have long been investigated for their potential therapeutic uses, one of the major problems has been increasing the biological activity of the molecule. Previous work with IL-2 indicates that complexing the cytokine with an Ab dramatically increases its activity, and IL-15 has been shown to be most effective when transpresented by its cell surface receptor IL-15Rα. Boyman et al. (p. 7265 ) have now tested the efficacy of IL-7:IL-7mAb complexes for their biological activity in thymopoiesis. In vivo, treatment with IL-7:IL-7mAb caused a 50- to 100-fold increase in thymic cellularity over that seen with IL-7 treatment alone. A massive expansion of pre-B cells was also seen in these mice. The cytokine:antibody complexes were also able to affect the expansion of mature cells, increasing the homeostatic proliferation of naive and memory CD4⁺ and CD8⁺ T cells and enhancing the primary Ag-specific naive CD8⁺ T cell response. The authors have demonstrated the potential therapeutic efficacy of complexing cytokines with Ab, increased the understanding of IL-7 in vivo function, and raised important concerns about administering Abs in vivo to block cytokine function.

The classical and lectin pathways have been demonstrated to be necessary for complement-mediated clearance of apoptotic bodies. Xu et al. (p. 7613 ) have now established that properdin, a positive regulator of complement, binds and promotes clearance of late apoptotic and necrotic cells through the alternative pathway. Although properdin is predominantly known for its ability to bind and stabilize C3b on the target surface, the authors identified a function that is independent of C3b. Properdin bound directly to apoptotic cells, amplifying alterative pathway complement activation. This binding was confined to late apoptotic and necrotic cells and not to early apoptotic cells. Properdin also bound directly to exposed DNA that was on the surface of apoptotic cells; this binding was confirmed in vitro and did not compete with other complement molecules such C1q and mannose-binding lectin, suggesting recognition of a unique structure. These data demonstrate that properdin mediates clearance of apoptotic and necrotic material through a C3b-independent mechanism, substantiating a novel role for both the molecule and the alternative complement pathway.

The transcription factor NF-κB controls the expression of many immunologically relevant genes and in turn is controlled by modification to IκB kinase (IKK). Previous work indicated that ubiquitination of the IKK regulatory subunit NEMO is necessary for Ag receptor-stimulated NF-κB and MAPK signaling. In this issue’s Cutting Edge section, Ni et al. (p. 7107 ) have conducted an elegant study elucidating the function served by ubiquitinating NEMO. The authors introduced a point mutation into the mouse germline that disrupted K63-linked ubiquitination of NEMO by changing lysine 392 to arginine. The mice generated (NEMO-KR) developed normal numbers of lymphocytes and showed no defect in T cell signaling after TCR crosslinking or PMA/ionomycin treatment. Anti-IgM treatment of B cells from NEMO-KR animals also induced normal proliferation when compared with wild type. However, LPS-stimulated NEMO-KR macrophages were defective in IL-6 and IL-12p40 cytokine production, indicating a defect in TLR signaling. NEMO-KR mice were also more resistant than wild-type animals to LPS-induced endotoxic shock. Thus, the authors provide evidence for the surprising conclusion that ubiquitination of NEMO does not affect NF-κB or MAPK signaling but does act to control TLR-dependent innate immune responses.

Granulysin is the cytotoxic molecule that CD4⁺ T cells use to kill Cryptococcus neoformans. However, CD4⁺ T cells do not produce granulysin until 5 days after stimulation with IL-2, and the mechanism of the delay is unknown. Previously, Zheng et al. observed that the CD4⁺ T cells of HIV-infected patients did not produce granulysin after IL-2 stimulation and have now (p. 7221 ) used this knowledge to determine the mechanism responsible for delayed production of this cytotoxic molecule. Stimulation with IL-2 led to activation of STAT5 and PI3K in normal CD4⁺ T cells and increased expression of IL-2Rβ. This, in turn, led to the production of granulysin and mediated killing of C. neoformans. Blocking expression of IL-2Rβ by small interfering RNA or IL-2Rβ blocking Abs abrogated the expression of granulysin. The CD4⁺ T cells of HIV-infected patients were defective in STAT5 and PI3K signaling, failed to express IL-2Rβ, and could not produce granulysin or mediate Cryptococcal killing. Not only did the authors elucidate the IL-2-STAT5-PI3K- IL-2Rβ-granulysin pathway and its delay, but they have also demonstrated in part why HIV-infected individuals are susceptible to C. neoformans infections.

Blocking the CD28 and CD154 costimulation pathways does not consistently control rejection of skin grafts. To find out why, Ford et al. (p. 7203 ) tested the hypothesis that following transplantation, the frequency of CD4⁺ T cell precursors could affect CD4⁺ T cell proliferation, cytokine production, and help for CD8⁺ T cells and B cells. They used TCR-transgenic OT-I (CD8⁺) and OT-II (CD4⁺) T cells adoptively transferred into naive B6 mice to control the precursor frequency of Ag-specific cells in the presence of OVA-expressing skin grafts (Act-mOVA). They found that following blockade of the CD28 and CD154 costimulatory pathways, increasing the CD4⁺ T cell precursor frequency caused graft rejection by providing help to donor-specific B cells and CD8⁺ T cells. CD4⁺ T cells were able to provide this help in the presence of costimualtory blockade despite an inhibited ability to proliferate or produce IFN-γ and TNF-α. Thus, the authors conclude that CD4⁺ T cell precursor frequency can determine the provision of T cell help independently of proliferation and cytokine production and explains why rejection can occur in the presence of costimulatory blockade.

Summaries written by Kira R. Gantt, Ph.D.