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Biomedical subjects

John Kappler

Publications and source records attributed to John Kappler.

At least 19 recordsLinked to original sources
PubMed2006

Ndfip1 protein promotes the function of itch ubiquitin ligase to prevent T cell activation and T helper 2 cell-mediated inflammation.

Nedd4 family interacting protein-1 (Ndfip1) is a protein whose only known function is that it binds Nedd4, a HECT-type E3 ubiquitin ligase. Here we show that mice lacking Ndfip1 developed severe inflammation of the skin and lung and died prematurely. This condition was due to a defect in Ndfip1(-/-) T cells. Ndfip1(-/-) T cells were activated, and they proliferated and adopted a T helper 2 (Th2) phenotype more readily than did their Ndfip1(+/+) counterparts. This phenotype resembled that of Itchy mutant mice, suggesting that Ndfip1 might affect the function of Itch, an E3 ubiquitin ligase. We show that T cell activation promoted both Ndfip1 expression and its association with Itch. In the absence of Ndfip1, JunB half-life was prolonged after T cell activation. Thus, in the absence of Ndfip1, Itch is inactive and JunB accumulates. As a result, T cells produce Th2 cytokines and promote Th2-mediated inflammatory disease.

Animals↗
PubMed2006

Bax does not have to adopt its final form to drive T cell death.

The introduction of antigen into animals causes antigen-specific T cells to divide and then die. Activated T cell death requires either of the death effector molecules, Bak or Bax. When T cells die, Bak and Bax change their conformations, a phenomenon that is thought to be required for Bak or Bax to drive cell death. Here we show that Bak changes conformation before activated T cells die, as detected by an antibody specific for a peptide near the NH2 terminus of Bak, but Bax does not change its shape markedly until after the cells are dead, as detected by an antibody specific for a peptide near the NH2 terminus of Bax. This latter finding is also true in activated T cells that lack Bak and are therefore dependent on Bax to die. This result suggests that Bax does not have to adopt its final, completely unfolded form until after the cells are dead.

Animals↗
PubMed2006

Loss of the proapoptotic protein, Bim, breaks B cell anergy.

Although B cells that respond with high avidity to self-antigen are eliminated early in their development, many autoreactive B cells escape elimination and are tolerized later in their lives via anergy. Anergic B cells are unresponsive to antigen and die prematurely. It has been suggested that the proapoptotic protein, Bim, controls the fate of anergic B cells. To test this idea, mice lacking Bim were crossed with mice that express soluble hen egg lysozyme and whose B cells bear receptors specific for the protein. In Bim+/+ mice these B cells are anergic and die rapidly. If the mice lack Bim, however, the B cells live longer, are more mature, respond to antigen, and secrete anti-hen egg lysozyme antibodies. This break of tolerance is not due to expression of endogenous B cell receptors, nor is it dependent on T cells. Rather, it appears to be due to a reduced requirement for the cytokine BAFF. Normal B cells require BAFF both for differentiation and survival. Bim-/- B cells, on the other hand, require BAFF only for differentiation. Therefore, autoreactive B cells are allowed to survive if they lack Bim and thus accumulate sufficient signals from differentiating factors to drive their maturation and production of autoantibodies.

Animals↗
PubMed2005

Class II major histocompatibility complex-peptide tetramer staining in relation to functional avidity and T cell receptor diversity in the mouse CD4(+) T cell response to a rheumatoid arthritis-associated antigen.

OBJECTIVE: Although studies have suggested that human cartilage (HC) gp-39 may be an antigen recognized by autoreactive CD4(+) T cells in rheumatoid arthritis, we previously failed to identify specific CD4(+) T cells in patients' synovial fluid or blood using a class II major histocompatibility complex-peptide tetramer composed of the immunodominant HC gp-39(263-275) epitope covalently linked to DR4. We undertook this study to better understand the parameters for specific binding of this tetramer. METHODS: DR4-transgenic mice were immunized with the HC gp-39 peptide, and a set of peptide-responsive hybridomas was derived. Hybridomas were stained with the DR4-gp-39 tetramer and cultured with increasing amounts of peptide in the presence of DR4-expressing antigen-presenting cells to determine functional avidity. RESULTS: Great variability was apparent in the ability of the tetramer to stain the hybridomas, and there was a strong correlation between the intensity of tetramer staining and functional avidity. Importantly, nearly 30% of the hybridomas did not stain with tetramer, and these cells exhibited relatively low functional avidity. Although the addition of an anti-T cell receptor (anti-TCR) monoclonal antibody during the staining procedure enhanced binding of the tetramer to a number of the hybridomas, a significant percentage remained unstainable. Analysis of TCR expression showed that >90% of the hybridomas expressed the same TCR beta-chain variable region (V(beta)10), and sequencing of the TCR junctional regions showed diversity in the third complementarity-determining region. CONCLUSION: These results suggest that immune responses dominated by relatively low-affinity TCR interactions, such as those that may occur in autoimmune disease, will be difficult to detect using standard tetramer techniques.

Animals↗
PubMed2004

Loss of Bim allows precursor B cell survival but not precursor B cell differentiation in the absence of interleukin 7.

Interleukin (IL)-7 is a stromal cell-derived cytokine required for the survival, proliferation, and differentiation of B cell precursors. Members of the Bcl-2 family of proteins are known to have profound effects on lymphocyte survival, but not lymphocyte differentiation. To distinguish the relative dependence on IL-7 of B cell precursor survival versus B cell differentiation, the combined effects of lack of IL-7 and lack of the proapoptotic Bcl-2 relative, Bim, were studied. Bim is expressed to varying degrees in all B cell precursors and B cells. Lack of Bim compensated for lack of IL-7 in the survival of pro-, pre-, and immature B cells; however, lack of Bim did not substitute for the requirement for IL-7 in B cell precursor differentiation or B cell precursor proliferation. Precursor B cell survival is more dependent on sufficient levels of IL-7 than precursor B cell differentiation because the number of B cells and their precursors were reduced by half in mice heterozygous for IL-7 expression, but were restored to normal numbers in mice also lacking Bim. Hence, Bim and IL-7 work together to control the survival of B cell precursors and the number of B cells that exist in animals.

Animals↗
PubMed2004

Promotion of B cell immune responses via an alum-induced myeloid cell population.

Exposure of naïve B cells to the cytokine interleukin-4 (IL-4) and/or antigen leads to a state of "priming," in which subsequent aggregation of major histocompatibility complex class II molecules induces the mobilization of calcium ions and cell proliferation. However, it is not clear how critical this priming is for immune responses or how it is normally induced in vivo. Injection of mice with the commonly used adjuvant alum led to priming of splenic B cells and to the accumulation in the spleen of a previously unknown population of IL-4-producing, Gr1+ cells. These cells and IL-4 were both required for in vivo priming and expansion of antigen-specific B cells, as well as for optimal production of antibody. These studies reveal a key role for a previously unknown accessory myeloid cell population in the generation of humoral immune responses.

Adjuvants, Immunologic↗
PubMed2004

Constitutive association of the proapoptotic protein Bim with Bcl-2-related proteins on mitochondria in T cells.

Apoptosis in activated T cells in vivo requires the proapoptotic Bcl-2 family member Bim. We show here that, despite its ability to bind LC8, a component of the microtubule dynein motor complex, most of the Bim in both healthy and apoptotic T cells is associated with mitochondria, not microtubules. In healthy resting T cells Bim is bound to the antiapoptotic proteins Bcl-2 and Bcl-x(L). In activated T cells, levels of Bcl-2 fall, and Bim is associated more with Bcl-x(L) and less with Bcl-2. Our results indicate that, in T cells, Bim function is regulated by interaction with Bcl-2 family members on mitochondria rather than by sequestration to the microtubules.

Animals↗
PubMed2004

An animal model of hemophagocytic lymphohistiocytosis (HLH): CD8+ T cells and interferon gamma are essential for the disorder.

Hemophagocytic lymphohistiocytosis (HLH) is a rare disorder with familial and acquired forms. The familial form is associated with mutations in the perforin gene and both forms are associated with severe defects in lymphocyte cytotoxic function. We examined perforin-deficient mice as a model of HLH in order to gain insight into this poorly understood disorder. While these mice do not spontaneously develop HLH-like symptoms, we found that they manifest all of the features of HLH after infection with lymphocytic choriomeningitic virus (LCMV). Following LCMV infection, perforin-deficient mice develop fever, splenomegaly, pancytopenia, hypertriglyceridemia, hypofibrinogenemia, and elevation of multiple serum cytokine levels, and hemophagocytosis is evident in many tissues. Investigation into how this phenotype develops has revealed that CD8+ T cells, but not natural killer (NK) cells, are necessary for the development of this disorder. Cytokine neutralization studies have revealed that interferon gamma (IFNgamma) is uniquely essential as well. Finally, the excessive amount of IFNgamma seen in affected mice appears to be driven by increased antigen presentation to CD8+ T cells. These studies provide insight into the pathophysiology of HLH, and provide new targets for specific therapeutic intervention in this fatal disorder.

Animals↗
PubMed2004

TCR-MHC/peptide interactions: kissing-cousins or a shotgun wedding?

The purpose of this Commentary is to put into modern-day perspective Jerne's hypothesis that antigen receptors encoded in the genome have been evolutionarily selected for their ability to react with major histocompatibility proteins and that the process of eliminating self reactivity is the catalyst for the generation of diversity of antigen receptors. In writing his hypothesis Jerne was trying to deal with the obsession of the immune system with the MHC, an obsession that was manifest in his days by the strong reactions of the immune system with allogeneic MHC proteins. However, Jerne's hypothesis also took on other issues that were not understood at the time--issues that included lymphocyte selection and tolerance, the generation of somatic diversity and the ability of the MHC to control responses to other antigens. In so doing, Jerne generated a hypothesis that accounted remarkably satisfactorily for what was known in 1971. Whilst the details of much of the hypothesis have since turned out to be incorrect, in his ideas Jerne did anticipate many of the most interesting and surprising findings of the subsequent 33 years.

Animals↗
PubMed2004

Control of T cell viability.

The factors affecting T cell viability vary depending on the type and status of the T cell involved. Naive T cells die via a Bcl-2/Bim dependent route. Their deaths are prevented in animals by IL-7 and contact with MHC. Activated T cells die in many different ways. Among these is a pathway involving signals that come from outside the T cell and affect it via surface receptors such as Fas. Activated T cells also die through a pathway driven by signals generated within the T cell itself, a cell autonomous route. This pathway involves members of the Bcl-2 family, in particular Bcl-2, Bcl-xl, Bim, and probably Bak. The viability of CD8+ and CD4+ memory T cells is controlled in different ways. CD8+ memory T cells are maintained by IL-15 and IL-7. The control of CD4+ memory T cells is more mysterious, with roles reported for IL-7 and/or contact via the TCR.

Animals↗
PubMed2003

Control of Bcl-2 expression by reactive oxygen species.

Reactive oxygen species (ROS) mediate apoptosis in many different cell types. We have previously shown that the antioxidant Mn(III) tetrakis(5,10,15,20-benzoic acid)porphyrin (MnTBAP) decreased intracellular ROS and prevented the apoptosis of activated T cells in vitro. To determine the mechanism(s) by which MnTBAP afforded such protection, we used Affymetrix (Santa Clara, CA) gene arrays to compare gene expression in T cells activated with staphylococcal enterotoxin B in vivo then cultured with or without MnTBAP. This analysis showed that the antioxidant increased the expression of Bcl-2, an antiapoptotic molecule whose levels are normally decreased by T cell activation. Culture with MnTBAP revealed a tight inverse correlation between the levels of Bcl-2 and ROS within T cells. In vivo, production of ROS in activated T cells occurred before Bcl-2 down-regulation. Furthermore, MnTBAP's ability to prevent death required the expression of Bcl-2 in most T cells. Finally, neither ROS production nor the effects on Bcl-2 expression required Bim, the Bcl-2 antagonist that mediates the death of activated T cells in vivo. Taken together, our results suggest that ROS sensitize T cells to apoptosis by decreasing expression of Bcl-2.

Animals↗
PubMed2003

Negative selection imparts peptide specificity to the mature T cell repertoire.

The T cell alphabeta receptor (TCR) recognizes foreign peptide antigens bound to proteins encoded in the MHC. The MHC portion of this complex contributes much to the footprint of the TCR on the ligand, yet T cells are usually very specific for individual foreign peptides. Here, we show that the development of peptide-specific T cells is not intrinsic to thymocytes that undergo thymic-positive selection but is an outcome of eliminating, through negative selection, thymocytes bearing TCRs with extensive peptide cross-reactivity. Hence, thymic-negative selection imposes peptide specificity on the mature T cell repertoire.

Amino Acid Sequence↗
PubMed2003

Mast cell-dependent migration of effector CD8+ T cells through production of leukotriene B4.

Studies in both humans and rodents indicate that CD8+ T cells may be important in allergic inflammation. However, neither the mechanisms that mediate CD8+ T cell recruitment to inflamed tissues nor the relative participation of effector and central memory CD8+ T cells is known. Here we report that activated mast cells induced chemotaxis of effector, but not central memory, CD8+ T cells through production of leukotriene B4 (LTB4). These studies indicate that LTB4 production by activated peripheral leukocytes could be important for the recruitment of effector CD8+ T cells to sites of inflammation.

Animals↗
PubMed2003

Ligand-receptor binding revealed by the TNF family member TALL-1.

The tumour necrosis factor (TNF) ligand TALL-1 and its cognate receptors, BCMA, TACI and BAFF-R, were recently identified as members of the TNF superfamily, which are essential factors contributing to B-cell maturation. The functional, soluble fragment of TALL-1 (sTALL-1) forms a virus-like assembly for its proper function. Here we determine the crystal structures of sTALL-1 complexed with the extracellular domains of BCMA and BAFF-R at 2.6 and 2.5 A, respectively. The single cysteine-rich domain of BCMA and BAFF-R both have saddle-like architectures, which sit on the horseback-like surface formed by four coil regions on each individual sTALL-1 monomer. Three novel structural modules, D2, X2 and N, were revealed from the current structures. Sequence alignments, structural modelling and mutagenesis revealed that one disulphide bridge in BAFF-R is critical for determining the binding specificity of the extracellular domain eBAFF-R to TALL-1 instead of APRIL, a closely related ligand of TALL-1, which was confirmed by binding experiments in vitro.

Amino Acid Sequence↗
PubMed2003

Components of the ligand for a Ni++ reactive human T cell clone.

The major histocompatibility complex (MHC) restriction element for a human Ni(2+) reactive T cell, ANi-2.3, was identified as DR52c. A series of experiments established that the functional ligand for this T cell was a preformed complex of Ni(2+) bound to the combination of DR52c and a specific peptide that was generated in human and mouse B cells, but not in fibroblasts nor other antigen processing-deficient cells. In addition, ANi-2.3 recognition of this complex was dependent on His81 of the MHC beta chain, suggesting a role for this amino acid in Ni(2+) binding to MHC. We propose a general model for Ni(2+) recognition in which betaHis81 and two amino acids from the NH(2)-terminal part of the MHC bound peptide coordinate Ni(2+) which then interacts with some portion of the Valpha CDR1 or CDR2 region.

Animals↗
PubMed2002

Staging and resetting T cell activation in SMACs.

During the productive interaction of T cells with antigen-presenting cells (APCs), engaged receptors, including the T cell antigen receptors and their associated tyrosine kinases, assemble into spatially segregated supramolecular activation clusters (SMACs) at the area of cell contact. Here, we studied intracellular signaling in SMACs by three-dimensional immunofluorescence microscopic localization of CD3, CD45, talin, phosphotyrosine, Lck and phosphorylated ZAP-70 in T cell-APC conjugates. Two distinct phases of spatial-temporal activation, one before and one after SMAC formation, which were separated by a brief state of inactivation caused by CD45, were observed at the T cell-APC contact area. We propose that pre-SMAC signals are sufficient to activate cell adhesion, but not productive T cell responses, which require orchestrated signaling in SMACs.

Animals↗