IL-17-producing NKT cells depend exclusively on IL-7 for homeostasis and survival.

Natural killer T (NKT) cells are innate-like T cells that rapidly recognize pathogens and produce cytokines that shape the ensuing immune response. IL-17-producing NKT cells are enriched in barrier tissues, such as the lung, skin, and peripheral lymph nodes, and the factors that maintain this population in the periphery have not been elucidated. Here we show that NKT17 cells deviate from other NKT cells in their survival requirements. In contrast to conventional NKT cells that are maintained by IL-15, RORγt(+) NKT cells are IL-15 independent and instead rely completely on IL-7. IL-7 initiates a T-cell receptor-independent (TCR-independent) expansion of NKT17 cells, thus supporting their homeostasis. Without IL-7, survival is dramatically impaired, yet residual cells remain lineage committed with no downregulation of RORγt evident. Their preferential response to IL-7 does not reflect enhanced signaling through STAT proteins, but instead is modulated via the PI3K/AKT/mTOR signaling pathway. The ability to compete for IL-7 is dependent on high-density IL-7 receptor expression, which would promote uptake of low levels of IL-7 produced in the non-lymphoid sites of lung and skin. This dependence on IL-7 is also reported for RORγt(+) innate lymphoid cells and CD4(+) Th17 cells, and suggests common survival requirements for functionally similar cells.

Two-way transfer of nitrogen between Dalbergia odorifera and its hemiparasite Santalum album is enhanced when the host is effectively nodulated and fixing nitrogen.

Nutrient translocation from a host plant is vital to the growth and survival of its root parasitic plant, but few studies have investigated whether a parasitic plant is also able to transfer nutrients to its host. The role of N2-fixation in nitrogen (N) transfer between 7-month-old Dalbergia odorifera T. Chen nodulated with Bradyrhizobium elkanii DG and its hemiparasite Santalum album Linn. was examined by external (15)N labeling in a pot study. Four paired treatments were used, with (15)N given to either host or hemiparasite and the host either nodulated or grown on combined N. N2-fixation supplied 41-44% of total N in D. odorifera. Biomass, N and (15)N contents were significantly greater in both nodulated D. odorifera and S. album grown with paired nodulated D. odorifera. Significantly higher total plant (15)N recovery was in N donor D. odorifera (68-72%) than in N donor S. album (42-44%), regardless of the nodulation status in D. odorifera. Nitrogen transfer to S. album was significantly greater (27.8-67.8 mg plant(-1)) than to D. odorifera (2.0-8.9 mg plant(-1)) and 2.4-4.5 times greater in the nodulated pair than in the non-nodulated pair. Irrespective of the nodulation status, S. album was always the N-sink plant. The amount of two-way N transfer was increased by the presence of effective nodules, resulting in a greater net N transfer (22.6 mg plant(-1)) from host D. odorifera to hemiparasite S. album. Our results may provide N management strategies for D. odorifera/S. album mixed plantations in the field.

Selectively expanding subsets of T cells in mice by injection of interleukin-2/antibody complexes: implications for transplantation tolerance.

The biological activity of interleukin (IL)-2 and other cytokines in vivo can be augmented by binding to certain anti-cytokine monoclonal antibodies (mAb). Here, we review evidence on how IL-2/anti-IL-2 mAb complexes can be used to cause selective stimulation and expansion of certain T-cell subsets. With some anti-IL-2 mAbs, injection of IL-2/mAb complexes leads to expansion of CD8 T effector cells but not CD4 T regulatory cells (Tregs); these complexes exert less adverse side effects than soluble IL-2 and display powerful antitumor activity. Other IL-2/mAb complexes have minimal effects on CD8 T cells but cause marked expansion of Tregs. Preconditioning mice with these complexes leads to permanent acceptance of MHC-disparate pancreatic islets in the absence of immunosuppression.

Regulatory T cells expressing granzyme B play a critical role in controlling lung inflammation during acute viral infection.

The inflammatory response to lung infections must be tightly regulated, enabling pathogen elimination while maintaining crucial gas exchange. Using recently described "depletion of regulatory T cell" (DEREG) mice, we found that selective depletion of regulatory T cells (Tregs) during acute respiratory syncytial virus (RSV) infection enhanced viral clearance but increased weight loss, local cytokine and chemokine release, and T-cell activation and cellular influx into the lungs. Conversely, inflammation was decreased when Treg numbers and activity were boosted using interleukin-2 immune complexes. Unexpectedly, lung (but not draining lymph node) Tregs from RSV-infected mice expressed granzyme B (GzmB), and bone marrow chimeric mice with selective loss of GzmB in the Treg compartment displayed markedly enhanced cellular infiltration into the lung after infection. A crucial role for GzmB-expressing Tregs has not hitherto been described in the lung or during acute infections, but may explain the inability of children with perforin/GzmB defects to regulate immune responses to infection. The effects of RSV infection in mice with defective immune regulation closely parallel the observed effects of RSV in children with bronchiolitis, suggesting that the pathogenesis of bronchiolitis may involve an inability to regulate virus-induced inflammation.

In vivo anergized T cells form altered immunological synapses in vitro.

T cells contact allogeneic antigen presenting cells (APCs) and assemble, at their contact interface, a molecular platform called the immunological synapse. Synapse-based molecules provide directional signals for the T cell--either positive signals, resulting in T-cell activation, or negative signals causing T-cell inactivation or anergy. To better understand the molecular basis of in vivo T-cell anergy we analyzed the contacts made between in vivo anergized T cells and APCs, and determined which signaling molecules were included or excluded from their immunological synapses. Anergy was induced in TCR transgenic mice by the intravenous injection of semiallogeneic donor spleen cells. T cells from anergized mice were mixed with APCs, the T-cell/APC synapses imaged using deconvolution microscopy, and their molecular compositions were determined. T cells from anergic mice formed unstable immunological synapses in vitro with allogeneic APCs and failed to recruit the signaling proteins necessary to initiate T-cell activation. These findings suggest that T-cell anergy induced by exposure to semiallogeneic donor cells is associated with defects in the earliest events of T-cell activation, immunological synapse formation and recruitment of TCR-mediated signaling proteins.

A role for shoot protein in shoot-root dry matter allocation in higher plants.

BACKGROUND AND AIMS: It is stated in many recent publications that nitrate (NO3-) acts as a signal to regulate dry matter partitioning between the shoot and root of higher plants. Here we challenge this hypothesis and present evidence for the viewpoint that NO3- and other environmental effects on the shoot:root dry weight ratio (S:R) of higher plants are often related mechanistically to changes in shoot protein concentration. METHODS: The literature on environmental effects on S:R is reviewed, focusing on relationships between S:R, growth and leaf NO3- and protein concentrations. A series of experiments carried out to test the proposal that S:R is dependent on shoot protein concentration is highlighted and new data are presented for tobacco (Nicotiana tabacum). KEY RESULTS/EVIDENCE: Results from the literature and new data for tobacco show that S:R and leaf NO3- concentration are not significantly correlated over a range of environmental conditions. A mechanism involving the relative availability of C and N substrates for growth in shoots can explain how shoot protein concentration can influence shoot growth and hence root growth and S:R. Generally, results in the literature are compatible with the hypothesis that macronutrients, water, irradiance and CO2 affect S:R through changes in shoot protein concentration. In detailed studies on several species, including tobacco, a linear regression model incorporating leaf soluble protein concentration and plant dry weight could explain the greater proportion of the variation in S:R within and between treatments over a wide range of conditions. CONCLUSIONS: It is concluded that if NO3- can influence the S:R of higher plants, it does so only over a narrow range of conditions. Evidence is strong that environmental effects on S:R are often related mechanistically to their effects on shoot protein concentration.

A defect in central tolerance in NOD mice.

The predisposition of nonobese diabetic (NOD) mice to develop autoimmune disease is usually attributed to defects in peripheral tolerance mechanisms. Here, evidence is presented that NOD mice display a defect in central tolerance (negative selection) of thymocytes. Impaired central tolerance in NOD mice was most prominent in a population of semi-mature thymocytes found in the medulla. The defect was apparent in vivo as well as in vitro, was independent of IAbetag7 expression and affected both Fas-dependent and Fas-independent pathways of apoptosis; for Fas-dependent apoptosis, the defective tolerance of NOD thymocytes correlated with the strong T cell receptor-mediated up-regulation of caspase 8-homologous FLICE (Fas-associated death-domain-like interleukin 1beta-converting enzyme)-inhibitory protein. In light of these findings, disease onset in NOD mice may reflect defects in central as well as peripheral tolerance.

IL-7 is critical for homeostatic proliferation and survival of naive T cells.

In T cell-deficient conditions, naive T cells undergo spontaneous "homeostatic" proliferation in response to contact with self-MHC/peptide ligands. With the aid of an in vitro system, we show here that homeostatic proliferation is also cytokine-dependent. The cytokines IL-4, IL-7, and IL-15 enhanced homeostatic proliferation of naive T cells in vitro. Of these cytokines, only IL-7 was found to be critical; thus, naive T cells underwent homeostatic proliferation in IL-4(-) and IL-15(-) hosts but proliferated minimally in IL-7(-) hosts. In addition to homeostatic proliferation, the prolonged survival of naive T cells requires IL-7. Thus, naïve T cells disappeared gradually over a 1-month period upon adoptive transfer into IL-7(-) hosts. These findings indicate that naive T cells depend on IL-7 for survival and homeostatic proliferation.

T cell death and memory.

In typical immune responses, contact with antigen causes naive T cells to proliferate and differentiate into effector cells. After the pathogen is destroyed, most effector T cells are eliminated-thereby preserving the primary T cell repertoire-but some cells survive and form long-lived memory cells. During each stage of this process, the life or death fate of T cells is strictly regulated.

An IFN-gamma-dependent pathway controls stimulation of memory phenotype CD8+ T cell turnover in vivo by IL-12, IL-18, and IFN-gamma.

Unlike naive T cells, memory phenotype (CD44(high)) T cells exhibit a high background rate of turnover in vivo. Previous studies showed that the turnover of memory phenotype CD8(+) (but not CD4(+)) cells in vivo can be considerably enhanced by products of infectious agents such as LPS. Such stimulation is TCR independent and hinges on the release of type I IFNs (IFN-I) which leads to the production of an effector cytokine, probably IL-15. In this study, we describe a second pathway of CD44(high) CD8(+) stimulation in vivo. This pathway is IFN-gamma rather than IFN-I dependent and is mediated by at least three cytokines, IL-12, IL-18, and IFN-gamma. As for IFN-I, these three cytokines are nonstimulatory for purified T cells and under in vivo conditions probably act via production of IL-15.

Burnet oration. T-cell survival and the role of cytokines.

Typical T cells are long-lived resting cells. Despite their quiescent appearance, there is increasing evidence that T cells are subjected to continuous stimulation through contact with various stimuli, notably by self peptide/MHC complexes and cytokines. These stimuli keep T cells alive and also cause intermittent entry into cell cycle.

The thymus and central tolerance.

T-cell differentiation in the thymus generates a peripheral repertoire of mature T cells that mounts strong responses to foreign antigens but is largely unresponsive to self-antigens. This state of specific immunological tolerance to self-components involves both central and peripheral mechanisms. Here we review the process whereby many T cells with potential reactivity for self-antigens are eliminated in the thymus during early T-cell differentiation. This process of central tolerance (negative selection) reflects apoptosis and is a consequence of immature T cells receiving strong intracellular signalling through T-cell receptor (TCR) recognition of peptides bound to major histocompatibility complex (MHC) molecules. Central tolerance occurs mainly in the medullary region of the thymus and depends upon contact with peptide-MHC complexes expressed on bone-marrow-derived antigen-presenting cells (APCs); whether tolerance also occurs in the cortex is still controversial. Tolerance induction requires a combination of TCR ligation and co-stimulatory signals. Co-stimulation reflects interaction between complementary molecules on T cells and APCs and probably involves multiple molecules acting in consort, which may account for why deletion of individual molecules with known or potential co-stimulatory function has little or no effect on central tolerance. The range of self-antigens that induce central tolerance is considerable and, via low-level expression in the thymus, may also include tissue-specific antigens; central tolerance to these latter antigens, however, is likely to be limited to high-affinity T cells, leaving low-affinity cells to escape. Tolerance to alloantigens and the possibility of using central tolerance to promote acceptance of allografts are discussed.

Role of the actin cytoskeleton in T cell absorption and internalization of ligands from APC.

A feature of T-APC interaction is that, via either TCR or CD28, T cells can absorb molecules from APC on to the cell surface and then internalize these molecules. Here, using both normal and TCR-transgenic T cells, we investigated the mechanism of T cell absorption of molecules from APC and the role of the cytoskeleton. The results show that although activated T cells could absorb APC molecules in the form of cell fragments, uptake of molecules by resting T cells required direct T-APC interaction. Based on studies with latrunculin B, surface absorption of molecules by resting T cells was crucially dependent upon the actin cytoskeleton for both CD28- and TCR-mediated absorption. Significantly, however, TCR-mediated absorption became strongly resistant to latrunculin B when the concentration of MHC-bound peptide on APC was raised to a high level, implying that the actin cytoskeleton is only important for absorption when the density of receptor/ligand interaction is low. By contrast, in all situations tested, the actin cytoskeleton played a decisive role in controlling T cell internalization of ligands from the cell surface.

The dalbergioid legumes (Fabaceae): delimitation of a pantropical monophyletic clade.

A monophyletic pantropical group of papilionoid legumes, here referred to as the "dalbergioid" legumes, is circumscribed to include all genera previously referred to the tribes Aeschynomeneae and Adesmieae, the subtribe Bryinae of the Desmodieae, and tribe Dalbergieae except Andira, Hymenolobium, Vatairea, and Vataireopsis. This previously undetected group was discovered with phylogenetic analysis of DNA sequences from the chloroplast trnK (including matK) and trnL introns, and the nuclear ribosomal 5.8S and flanking internal transcribed spacers 1 and 2. All dalbergioids belong to one of three well-supported subclades, the Adesmia, Dalbergia, and Pterocarpus clades. The dalbergioid clade and its three main subclades are cryptic in the sense that they are genetically distinct but poorly, if at all, distinguished by nonmolecular data. Traditionally important taxonomic characters, such as arborescent habit, free stamens, and lomented pods, do not provide support for the major clades identified by the molecular analysis. Short shoots, glandular-based trichomes, bilabiate calyces, and aeschynomenoid root nodules, in contrast, are better indicators of relationship at this hierarchical level. The discovery of the dalbergioid clade prompted a re-analysis of root nodule structure and the subsequent finding that the aeschynomenoid root nodule is synapomorphic for the dalbergioids.

Generation and maintenance of memory T cells.

Typical immune responses lead to the prominent clonal expansion of antigen-specific T cells followed by their differentiation into effector cells. Most effector cells die at the end of the immune response but some of the responding cells survive and form long-lived memory cells. The factors controlling the formation and survival of memory T cells are discussed. Recent evidence suggests that T memory cells arise from a subset of effector cells. The longevity of T memory cells may require continuous contact with cytokines, notably IL-15 for CD8(+) cells.

Multiple effects of immunostimulatory DNA on T cells and the role of type I interferons.

In addition to stimulating antigen-specific immune responses, infectious agents cause nonspecific activation of the innate immune system, notably up-regulation of costimulatory/adhesion molecules on APCs and cytokine production. In recent years it has become apparent that stimulation of the immune system by microorganisms is a property of a number of different cellular components, including DNA. As discussed earlier and elsewhere in this volume, the DNA of infectious agents--and indeed of all non-vertebrates tested--differs from mammalian DNA in being enriched for unmethylated CpG motifs. With appropriate flanking sequences, CpG DNA and synthetic CpG ODNs cause strong activation of APCs and other cells. In this article we have focussed on the capacity of CpG DNA/ODNs to alter T cell function. Whether these compounds act directly on T cells or function indirectly by activating other cells, especially APCs, is controversial [7, 8, 13, 14]. In contrast to other workers [8], we have yet to find definitive evidence that CpG DNA/ODNs can provide a co-stimulatory signal for purified T cells subjected to TCR ligation ([14] and unpublished data of authors). For this reason we lean to the notion that CpG DNA/ODNs modulate T cell function by inducing activation of APC rather than by acting directly on T cells. When injected in vivo in the absence of specific antigen, CpG DNA/ODNs have two striking effects on T cells, namely (1) induction of overt activation (proliferation) of memory-phenotype CD8+ cells, and (2) partial activation of all T cells, including naïve-phenotype T cells. Both actions of CpG DNA/ODNs are heavily dependent on the production of IFN-I by APC. For memory-phenotype (CD44hi) CD8+ cells, neither CpG DNA nor IFN-I can cause proliferation of purified APC-depleted T cells in vitro. Hence, under in vivo conditions, CpG DNA-induced proliferation of CD44hi CD8+ cells is probably mediated through the production of a secondary cytokine, i.e., by a cytokine that is directly stimulatory for CD44hi CD8+ cells. Based on the available evidence, it is highly likely that the effector cytokine is IL-15. With this assumption, our current model is that proliferation of CD44hi CD8+ cells induced by injection of CpG DNA/ODNs reflects production of IFN-I which, in turn, leads to synthesis of IL-15. Which particular cell types produce these two cytokines is unclear, although APCs are probably of prime importance. In addition to inducing proliferation of memory-phenotype CD8+ cells via IL-15, the IFN-I induced by CpG DNA/ODNs can also induce partial activation of naive T cells. This form of activation leads to up-regulation of CD69 and other molecules but does not cause entry into cell cycle. It is of interest that the partial activation of naive T cells induced by IFN-I is associated with decreased T proliferative responses. Thus, proliferation of purified naïve T cells elicited by combined TCR/CD28 ligation in vitro is greatly reduced by addition of IFN-I. This inhibitory effect of IFN-I does not influence cytokine production and probably reflects production of cell cycle inhibitors. Surprisingly, except at high doses, IFN-I fails to exert an anti-proliferative effect when T proliferative responses are driven by viable APCs. Indeed, in this situation, IFN-I enhances antigen-specific T proliferative responses, both in vivo and in vitro. This adjuvant effect of IFN-I is presumably a reflection of APC activation, but direct evidence on this issue is still lacking. In this article we have emphasized that contact with CpG DNA/ODNs has multiple effects on T cell function in vivo. Many of these effects seem to be related to the production of certain cytokines by APCs, notably IFN-I and IL-15. It should be stressed, however, that CpG DNA/ODNs probably lead to the production of many other cytokines. Hence, our current models of how CpG DNA/ODNs influence T cell function are undoubtedly oversimplified.

The thymus and negative selection.

Self-tolerance induction is largely a reflection of negative selection (deletion) of autoreactive T cells in the thymus. Evidence is presented that negative selection occurs at a relatively late stage of thymocyte differentiation and affects a population of semimature HSA(hi) CD4+8- cells found in the medulla. Negative selection involves a number of cell-surface molecules on T cells, including Fas, CD28, CD5, and CD43. These molecules appear to act in consort, thereby ensuring that negative selection is highly efficient.