Endogenous Superantigens Research Articles

Deletion of Vβ3 +CD4 +T-cells by endogenous mouse mammary tumor virus 3 prevents type 1 diabetes induction by autoreactive CD8 +T-cells

Abstract In both humans and NOD mice, type 1 diabetes (T1D) develops from the autoimmune destruction of pancreatic beta cells by T-cells. Interactions between both helper CD4 +and cytotoxic CD8 +T-cells are essential for T1D development in NOD mice. Previous work has indicated that pathogenic T-cells arise from deleterious interactions between relatively common genes which regulate aspects of T-cell activation/effector function (Ctla-4, Tnfrsf9, Il2/Il21), peptide presentation (H2-A g7, B2m), and T-cell receptor signaling (Ptpn22). In this study, we used a combination of subcongenic mapping and a CRISPR/Cas9 screen to identify the NOD-encoded Mtv3 provirus as a genetic element affecting CD4 +/CD8 +T-cell interactions through an additional mechanism, altering the T-cell receptor (TCR) repertoire. Mtv3 encodes a superantigen that deletes the majority of Vβ3 +thymocytes in NOD mice. Ablating Mtv3 and restoring Vβ3 +T-cells has no effect on spontaneous T1D development in NOD mice. However, transferring Mtv3 to C57BL/6 (B6) mice congenic for the NOD H2 g7MHC haplotype (B6.H2 g7) completely blocks their normal susceptibility to T1D mediated by transferred CD8 +T-cells transgenically expressing AI4 or NY8.3 TCRs. The entire genetic effect is manifest by Vβ3 +CD4 +T-cells, which unless deleted by Mtv3, accumulate in insulitic lesions and trigger in B6 background mice the pathogenic activation of diabetogenic CD8 +T-cells. Our findings provide evidence that endogenous Mtv superantigens can influence autoimmune responses. Furthermore, since most common mouse strains have gaps in their TCR Vβ repertoire due to Mtvs, it raises questions about the role of Mtvs in other mouse models designed to reflect human disorders. Supported by grants from the American Diabetes Association (1-14-BS-051) and NIH (AI130656)

Endogenous HLA-DQ8αβ programs superantigens (SEG/SEI) to silence toxicity and unleash a tumoricidal network with long-term melanoma survival

BackgroundAs the most powerful T cell agonists known, superantigens (SAgs) have enormous potential for cancer immunotherapy. Their development has languished due to high incidence (60%–80%) of seroreactive neutralizing antibodies in...

Clonal deletion of T cell repertoires with specific T cell receptor Vβ chains by two endogenous superantigens in NC/Nga mice.

Superantigens (SAgs) are powerful T-cell stimulatory proteins. Because an atopic dermatitis (AD) model NC/Nga mice had two endogenous SAgs, namely minor lymphocyte-stimulating locus-1a (Mls-1a) and mouse mammary tumor virus (MMTV)(SHN), SAg-responsive T-cells bearing Vβ5.1, Vβ6, Vβ8.1, Vβ8.2, Vβ8.3, Vβ9, and Vβ11 should be endogenously deleted. Here, we discuss that the endogenous SAgs-expression may be involved in AD-sensitivity in NC/Nga mice.

Modest Expansion of Vß2+CD4+ T Cells and No Expansion of Vß7+CD4+ T Cells in a Subgroup of Kawasaki Disease Patients with Erythematic BCG Inoculation Site Lesions

Background: The similarities between Kawasaki disease (KD) and superantigen (SA) diseases indicate that a microbial SA might cause KD. Viral diseases can trigger an endogenous SA. Methods: We evaluated expression of Vs2 (responding to staphylococcal TSST-1) and Vs7 (responding to the endogenous SA induced by type-1 interferon or Epstein-Barr virus infection) on T cells from 70 KD patients along with the following control subjects: 18 non-vasculitic patients (NVs), 7 patients with anaphylactoid purpura (AP), and two with neonatal TSS-like exanthematous disease (NTED), a typical SA disease. We examined the correlation of clinical features of KD with Vs2 + or Vs7 + CD4 + T cell populations. Results: The Vs2 + CD4 + T cell rates were comparable between KD patients (9.9±2.9%) and NVs (9.0±1.8%), but were lower in AP patients (6.6±1.8%). However, the Vs2 + CD4 + T cell rate was significantly higher in KD patients with erythematic BCG inocula tion site lesions (10.8±3.2%) than in those without (8.8±2.1%) and NVs (9.0±1.8%), but much lower than in NTED patients (25.2%, 16.9%). Multivariate linear regression analysis with elevation of Vs2 expression as a dependent variable revealed significant correlations with BCG. In contrast, Vs7 + CD4 + T cell rates were not significantly different between KD patients and other study sub jects. Conclusion: While we were unable to find evidence supporting the involvement of the endogenous SA in the pathogenesis of KD in this study, modest expansion of the Vs2 + CD4 + T cell population in a subgroup of KD with erythematic BCG inoculation site lesions implies the involvement of a microbial agent(s) different from TSST-1 as well as immunopathological heterogeneity of KD. (249 words) ACTA MEDICA NAGASAKIENSIA 60: 13−19, 2015

Activated CD8+ T cells induce expansion of Vβ5+ regulatory T cells via TNFR2 signaling.

Vβ5(+) regulatory T cells (Tregs), which are specific for a mouse endogenous retroviral superantigen, become activated and proliferate in response to Friend virus (FV) infection. We previously reported that FV-induced expansion of this Treg subset was dependent on CD8(+) T cells and TNF-α, but independent of IL-2. We now show that the inflammatory milieu associated with FV infection is not necessary for induction of Vβ5(+) Treg expansion. Rather, it is the presence of activated CD8(+) T cells that is critical for their expansion. The data indicate that the mechanism involves signaling between the membrane-bound form of TNF-α on activated CD8(+) T cells and TNFR2 on Tregs. CD8(+) T cells expressing membrane-bound TNF-α but no soluble TNF-α remained competent to induce strong Vβ5(+) Treg expansion in vivo. In addition, Vβ5(+) Tregs expressing only TNFR2 but no TNFR1 were still responsive to expansion. Finally, treatment of naive mice with soluble TNF-α did not induce Vβ5(+) Treg expansion, but treatment with a TNFR2-specific agonist did. These results reveal a new mechanism of intercellular communication between activated CD8(+) T cell effectors and Tregs that results in the activation and expansion of a Treg subset that subsequently suppresses CD8(+) T cell functions.

New Insight Into Early Events in Type 1 Diabetes: Role for Islet Stem Cell Exosomes

Type 1 diabetes (T1D) is an autoimmune disease in which an inappropriate self-directed immune response affects and destroys insulin-producing β-cells in pancreatic islets leading to dysregulated blood glucose levels. T1D may affect people of any age, its clinical presentation is highly variable, and its incidence is increasing worldwide (1). The initial triggering events of T1D are unknown and their elucidation is of pivotal importance. Several factors might lead to the breakdown of β-cell–specific T-cell tolerance, including genetics, exogenous infectious pathogen, noninfectious environment agents, endogenous superantigens, or physiological stress events (2). The hallmark of autoimmune diabetes is insulitis, which progresses to a destruction of β-cells that results in clinical T1D. An altered balance between proinflammatory T-helper type 1 (Th1)/Th17 cells (γ-interferon [IFN-γ], interleukin [IL]-17) and Th2 immune response (IL-4, IL-10) leads to T1D (3,4). Evidence also suggests that both genetic and environmental factors may induce local inflammatory response, where activated intraislet dendritic cells (DCs) prevent peripheral T-cell tolerance (5). Moreover, it has been recently demonstrated that the development of destructive insulitis is partly due to impaired islet-resident Foxp3+ regulatory T cells (6,7). Several animal experimental models have been used for the investigation of the pathogenesis of T1D and it appears that a NOD mouse is the model of choice. NOD mice spontaneously develop early peri-insulitis and later intraislet insulitis caused by autoreactive T cells, …

Strength of TCR signal from self‐peptide modulates autoreactive thymocyte deletion and Foxp3+Treg‐cell formation

Autoreactive CD4(+) CD8(-) (CD4SP) thymocytes can be subjected to deletion when they encounter self-peptide during their development, but they can also undergo selection to become CD4SPFoxp3(+) Treg cells. We have analyzed the relationship between these distinct developmental fates using mice in which signals transmitted by the TCR have been attenuated by mutation of a critical tyrosine residue of the adapter protein SLP-76. In mice containing polyclonal TCR repertoires, the mutation caused increased frequencies of CD4SPFoxp3(+) thymocytes. CD4SP thymocytes expressing TCR Vβ-chains that are subjected to deletion by endogenous retroviral superantigens were also present at increased frequencies, particularly among Foxp3(+) thymocytes. In transgenic mice in which CD4SP thymocytes expressing an autoreactive TCR undergo both deletion and Treg-cell formation in response to a defined self-peptide, SLP-76 mutation abrogated deletion of autoreactive CD4SP thymocytes. Notably, Foxp3(+) Treg-cell formation still occurred, albeit with a reduced efficiency, and the mutation was also associated with decreased Nur77 expression by the autoreactive CD4SP thymocytes. These studies provide evidence that the strength of the TCR signal can play a direct role in directing the extent of both thymocyte deletion and Treg-cell differentiation, and suggest that distinct TCR signaling thresholds and/or pathways can promote CD4SP thymocyte deletion versus Treg-cell formation.

IL-2–Independent and TNF-α–Dependent Expansion of Vβ5+ Natural Regulatory T Cells during Retrovirus Infection

Friend virus infection of mice induces the expansion and activation of regulatory T cells (Tregs) that dampen acute immune responses and promote the establishment and maintenance of chronic infection. Adoptive transfer experiments and the expression of neuropilin-1 indicate that these cells are predominantly natural Tregs rather than virus-specific conventional CD4(+) T cells that converted into induced Tregs. Analysis of Treg TCR Vβ chain usage revealed a broadly distributed polyclonal response with a high proportionate expansion of the Vβ5(+) Treg subset, which is known to be responsive to endogenous retrovirus-encoded superantigens. In contrast to the major population of Tregs, the Vβ5(+) subset expressed markers of terminally differentiated effector cells, and their expansion was associated with the level of the antiviral CD8(+) T cell response rather than the level of Friend virus infection. Surprisingly, the expansion and accumulation of the Vβ5(+) Tregs was IL-2 independent but dependent on TNF-α. These experiments reveal a subset-specific Treg induction by a new pathway.

The role of germinal centers in T cell receptor revision (P1421)

Abstract Vβ5 transgenic CD4 T cells are tolerized to the endogenous superantigen Mtv-8 by either deletion or TCR revision. Revision occurs when a peripheral CD4 T cell downregulates Vβ5 and undergoes RAG-mediated recombination and surface expression of an endogenous TCR. Given that revision occurs in germinal centers, and that follicular helper T cells, or Tfh, in Vβ5 Tg mice increase in an Mtv-8-dependent manner, we are studying whether revising cells have a Tfh phenotype and whether germinal center interactions are required for revision. Revising cells have a transcription factor and surface phenotype resembling that of Tfh, with low Blimp-1 and elevated Bcl-6, CXCR5, PD-1, IL-21 receptor, and OX-40 expression. The frequency of post-revision T cells increases in mice heterozygous for Blimp-1, a transcriptional repressor of Tfh formation, but revision intermediates are unaffected. The adaptor molecule SAP is required for prolonged B and T cell interactions in the germinal center, and SAP null mice have reduced numbers of post-revision T cells, but no reduction in revision intermediates. These results suggest that revising T cells share some traits with Tfh and that Tfh formation and prolonged B-T cell interactions are required for the completion, but not initiation, of TCR revision. Ongoing studies will analyze revision in the absence of Bcl-6, a transcription factor required for Tfh formation, and TNFR1, a receptor required for the formation of germinal center structures.

A new method for quantitative analysis of the T cell receptor V region repertoires in healthy common marmosets by microplate hybridization assay

The common marmoset, Callithrix jacchus, is one of the smallest primates and is increasingly used for an experimental nonhuman primate model in many research fields. Analysis of T cell receptor (TCR) repertoires is a powerful tool to investigate T cell immunity in terms of antigen specificity and variability of TCR expression. However, monoclonal antibodies specific for many TCR Vα or Vβ chains have not been created. We have recently identified a large number of TCRα chain variable (TRAV) and TCRβ chain variable (TRBV) sequences from a cDNA library of common marmosets. The purpose of this study is to develop a new method for analysis of TCR repertoires in the common marmoset using this sequence information. This method is based on a microplate hybridization technique using 32 TRAV-specific and 32 TRBV-specific oligoprobes following an adaptor-ligation PCR. This enables the easy quantitation of the respective TRAV and TRBV expression levels. No cross-hybridization among specific-oligoprobes and very low variances in repeated measures of the same samples was found, demonstrating high specificity and reproducibility. Furthermore, this method was validated by an antihuman Vβ23 antibody which specifically bound to marmoset Vβ23. Using this method, we analyzed TCR repertoires from various tissue samples (PBMCs, spleen, lymph node and thymus) and isolated T cell subpopulations (CD4+CD8+, CD4+CD8− and CD4−CD8+) from the thymus of 10 common marmosets. Neither tissue-specific nor T cell subpopulation-specific differences was found in TRAV and TRBV repertoires. These results suggest that, unlike mice, TCR repertoires in the common marmoset are not affected by endogenous superantigens and are conserved among individuals, among tissues, and among T cell subpopulations. Thus, TCR repertoire analysis with high specificity and reproducibility is a very useful technique, with the potential to replace flow cytometric analysis using a panel of TRV-specific antibodies, many of which remain unavailable.

Inability of Viral Superantigens to Induce CD4-mediated Islet Transplant Rejection

Superantigens have the ability to bypass the specific interactions of the MHC class II and T-cell receptor by binding outside of the peptide binding region and onto the Vβ chain. This ability allows superantigens to stimulate a wide array of T-cell populations, irrespective of T-cell receptor (TCR) specificity. Research on bacterial and viral superantigens have demonstrated various outcomes ranging from superantigen dependent cellular cytoxicity (SDCC), rheumatoid arthritis, to superantigen stimulated T-cell clonal deletion and anergy. Due to its ability to proliferate a large population of T-cells, this paper asks whether superantigens have a role in islet transplant rejection. We used transgenic Vβ6 TCR mice specific for male ‘H-Y’ antigen as recipients to islet transplants. Donors comprised mice expressing endogenous superantigen specific for the Vβ6 chain. Transplantation of these donor islets did not induce rejection. Recipients were also primed with ‘H-Y’ antigen to induce a CD4 effector memory T-cell population prior to islet transplantation. Even with primed recipients, donor islet transplants did not induce rejection by recipient transgenic mice.

Continuous Activation of the CD122/STAT-5 Signaling Pathway during Selection of Antigen-Specific Regulatory T Cells in the Murine Thymus

Signaling events affecting thymic selection of un-manipulated polyclonal natural CD25+foxp3+ regulatory T cells (nTreg) have not been established ex vivo. Here, we report a higher frequency of phosphorylated STAT-5 (pSTAT-5) in nTreg cells in the adult murine thymus and to a lesser extent in the periphery, compared to other CD4+CD8− subsets. In the neonatal thymus, the numbers of pSTAT-5+ cells in CD25+foxp3− and nTreg cells increased in parallel, suggesting that pSTAT-5+CD25+foxp3− cells might represent the precursors of foxp3+ regulatory T cells. This “specific” pSTAT-5 expression detected in nTreg cells ex vivo was likely due to a very recent signal given by IL-2/IL-15 cytokines in vivo since (i) it disappeared rapidly if cells were left unstimulated in vitro and (ii) was also observed if total thymocytes were stimulated in vitro with saturating amounts of IL-2 and/or IL-15 but not IL-7. Interestingly, STAT-5 activation upon IL-2 stimulation correlated better with foxp3 and CD122 than with CD25 expression. Finally, we show that expression of an endogenous superantigen strongly affected the early Treg cell repertoire but not the proportion of pSTAT-5+ cells within this repertoire. Our results reveal that continuous activation of the CD122/STAT-5 signaling pathway characterize regulatory lineage differentiation in the murine thymus.

CD30L null SJL mice exhibit reduced lymph node and spleen weights but support growth of transplantable SJL lymphoma RCS-X (48.29)

Abstract The process by which endogenous retroviral superantigen (vSAg29) expressing SJL lymphomas (RCS) stimulate host CD4+Vβ16+ T cells, in order to elicit help (notably IL-4 and -5) for their growth, has been characterized as “reverse immune surveillance”. This response is facilitated by the high expression of an array of co-stimulating molecules on RCS cells, including B7.1, B7.2, 41BBL, CD40 and CD30. On the basis of their germinal center derivation, SJL lymphomas bear close resemblance to a subpopulation of human germinal center B cell lymphomas, such as CD30+ Hodgkin’s lymphomas. In our earlier studies, growth of transplantable SJL lymphoma (RCS-X) in vivo was inhibited by anti-CD30 mAb. While the evidence shows that this effect is indirectly on the lymphoma-responsive CD4 T cells, the mechanism of action was unclear. To shed light on the role of CD30-CD30L interactions in SJL lymphoma development, CD30L-/- were bred onto SJL background and tested for their support of transplantable lymphoma growth. A significant reduction in lymph node (17.4-54.7%) and spleen weights (21.5-38.2%) was observed for CD30 null mice, compared to wild type SJL mice. Despite this reduction in the lymphoid compartment, no reduction in lymphoma growth was observed. The results suggest that while CD30-CD30L interactions might be important for SJL lymphoma growth, other costimulating molecules might contribute significantly to the process of reverse immune surveillance in RCS lymphoma.

The role of germinal centers in T cell receptor revision (160.10)

Abstract CD4+ T cells in Vβ5 transgenic mice become tolerant to an endogenous superantigen through either deletion or T cell receptor (TCR) revision. In the revision process, T cells downregulate surface Vβ5 expression and undergo RAG-mediated rearrangement and expression of endogenous TCRs. Revision occurs in germinal centers (GCs), suggesting that revising T cells may be follicular helper T cells (Tfh). We are studying whether revising T cells have a Tfh phenotype and whether GC interactions are required for revision. Preliminary data indicate that revising T cells have an RNA phenotype closely resembling that of Tfh and distinct from that of post-revision T cells. Studies on the intracellular and surface phenotype of revising and post-revision T cells are ongoing. The importance of GCs to TCR revision is being analyzed using SAP and CXCR5 null mice. SAP is required for prolonged interactions of B and T cells in the GC, and CXCR5 is required for cellular migration into the follicle. Analyses of mixed chimeras generated using CXCR5 null and wildtype bone marrow donors are ongoing. CXCR5 and SAP null mice have reduced numbers of post-revision T cells, but no reduction in revision intermediates. These results suggest that migration into the follicle and prolonged B-T cell interactions are both required for completion, but not initiation, of TCR revision.

Exogenous and endogenous virus interactions in head and neck cancer immunity (105.34)

Abstract We report that two common tumor viruses etiologically associated with head and neck cancer, Epstein-Barr virus (EBV) with undifferentiated nasopharyngeal carcinoma, and human papillomavirus (HPV) with oropharyngeal squamous cell carcinoma, transactivate an endogenous retrovirus (HERV-K18), which encodes a superantigen that activates TCRBV13 T cells. EBV LMP-2 transactivated the HERV-K18 superantigen in nasopharyngeal epithelial cells, and EBV-associated nasopharyngeal carcinomas expressed the superantigen. HPV type-16 E6 and E7 proteins also transactivated the HERV-K18 superantigen in oropharyngeal epithelial cells, and superantigen expression was significantly increased in HPV16-associated carcinomas. Serum cytokine levels in head and neck carcinoma patients indicated that there were distinct inflammatory profiles associated with exogenous and endogenous viruses. HPV-associated patients had increased levels of serum IL-8, while HPV-negative patients had higher serum IL-6 and G-CSF levels. Interestingly, HERV-K18 superantigen levels correlated independently with serum levels of the Th2 cytokine IL-13. Among HPV+ tumors, gene expression array analyses indicated that B cell specific genes were significantly upregulated in the superantigen high group. Our results suggest that endogenous superantigen expression in HPV+ tumors was associated with a Th2 immune response, demonstrating that chronic and endogenous viruses comprising the virome affect both host and tumor immunity.

Regulatory T-cell expansion during chronic viral infection is dependent on endogenous retroviral superantigens

Regulatory T cells (Treg) play critical roles in the modulation of immune responses to infectious agents. Further understanding of the factors that control Treg activation and expansion in response to pathogens is needed to manipulate Treg function in acute and chronic infections. Here we show that chronic, but not acute, infection of mice with lymphocytic choriomeningitis virus results in a marked expansion of Foxp3(+) Treg that is dependent on retroviral superantigen (sag) genes encoded in the mouse genome. Sag-dependent Treg expansion was MHC class II dependent, CD4 independent, and required dendritic cells. Thus, one unique mechanism by which certain infectious agents evade host immune responses may be mediated by endogenous Sag-dependent activation and expansion of Treg.

Bcl-2–Interacting Mediator of Cell Death Influences Autoantigen-Driven Deletion and TCR Revision

Peripheral CD4(+)Vβ5(+) T cells are tolerized to an endogenous mouse mammary tumor virus superantigen either by deletion or TCR revision. Through TCR revision, RAG reexpression mediates extrathymic TCRβ rearrangement and results in a population of postrevision CD4(+)Vβ5(-) T cells expressing revised TCRβ chains. We have hypothesized that cell death pathways regulate the selection of cells undergoing TCR revision to ensure the safety and utility of the postrevision population. In this study, we investigate the role of Bcl-2-interacting mediator of cell death (Bim)-mediated cell death in autoantigen-driven deletion and TCR revision. Bim deficiency and Bcl-2 overexpression in Vβ5 transgenic (Tg) mice both impair peripheral deletion. Vβ5 Tg Bim-deficient and Bcl-2 Tg mice exhibit an elevated frequency of CD4(+) T cells expressing both the transgene-encoded Vβ5 chain and a revised TCRβ chain. We now show that these dual-TCR-expressing cells are TCR revision intermediates and that the population of RAG-expressing, revising CD4(+) T cells is increased in Bim-deficient Vβ5 Tg mice. These findings support a role for Bim and Bcl-2 in regulating the balance of survival versus apoptosis in peripheral T cells undergoing RAG-dependent TCR rearrangements during TCR revision, thereby ensuring the utility of the postrevision repertoire.

TCR Revision Generates Functional CD4+ T Cells

CD4(+)Vβ5(+) peripheral T cells in C57BL/6 mice respond to encounter with a peripherally expressed endogenous superantigen by undergoing either deletion or TCR revision. In this latter process, cells lose surface Vβ5 expression and undergo RAG-dependent rearrangement of endogenous TCRβ genes, driving surface expression of novel TCRs. Although postrevision CD4(+)Vβ5(-)TCRβ(+) T cells accumulate with age in Vβ5 transgenic mice and bear a diverse TCR Vβ repertoire, it is unknown whether they respond to homeostatic and antigenic stimuli and thus may benefit the host. We demonstrate in this study that postrevision cells are functional. These cells have a high rate of steady-state homeostatic proliferation in situ, and they undergo extensive MHC class II-dependent lymphopenia-induced proliferation. Importantly, postrevision cells do not proliferate in response to the tolerizing superantigen, implicating TCR revision as a mechanism of tolerance induction and demonstrating that TCR-dependent activation of postrevision cells is not driven by the transgene-encoded receptor. Postrevision cells proliferate extensively to commensal bacterial Ags and can generate I-A(b)-restricted responses to Ag by producing IFN-γ following Listeria monocytogenes challenge. These data show that rescued postrevision T cells are responsive to homeostatic signals and recognize self- and foreign peptides in the context of self-MHC and are thus useful to the host.

A Superantigen Interacts with Leishmanial Infection in Antigen‐Presenting Cells to Regulate Cytokine Commitment of Responding CD4 T Cells

Germ-line retroviral insertions in vertebrate genomes are implicated in the modulation of host immune responses. We demonstrate that CBA/J mice, which carry the proviral integrants mammary tumor virus locus 6 (Mtv6) and mammary tumor virus locus 7 (Mtv7), are less resistant to infection with the protozoan pathogen Leishmania major compared with closely related but Mtv6-negative and Mtv7-negative CBA/CaJ mice. Although both strains generated comparable L. major-specific CD4 T cell frequencies, T cells from CBA/J mice made much less interferon γ (IFN-γ). L. major-infected CBA/CaJ dendritic cells primed L. major-specific and allospecific IFN-γ-producing CD4 T cells better in vivo and in vitro, respectively, than CBA/J dendritic cells did. L. major susceptibility appeared to be associated with Mtv7, and v-Sag-7 superantigen expression and L. major infection together reduced the ability of an antigen-presenting cell line to prime alloresponder CD4 T cells for IFN-γ commitment. These data show that an endogenous superantigen can interact with L. major infection to alter antigen-presenting cell properties and modulate T cell cytokine commitment, with implications for human susceptibility to infectious diseases.

Cutting Edge: Rag Deletion in Peripheral T Cells Blocks TCR Revision

Mature CD4(+)Vbeta5(+) T cells that recognize a peripherally expressed endogenous superantigen are tolerized either by deletion or TCR revision. In Vbeta5 transgenic mice, this latter tolerance pathway results in the appearance of CD4(+)Vbeta5(-)TCRbeta(+) T cells, coinciding with Rag1, Rag2, and TdT expression and the accumulation of V(beta)-DJ(beta) recombination intermediates in peripheral CD4(+) T cells. Because postthymic RAG-dependent TCR rearrangement has remained controversial, we sought to definitively determine whether TCR revision is an extrathymic process that occurs in mature peripheral T cells. We show in this study that Rag deletion in post-positive selection T cells in Vbeta5 transgenic mice blocks TCR revision in vivo and that mature peripheral T cells sorted to remove cells bearing endogenous TCRbeta-chains can express newly generated TCRbeta molecules in adoptive hosts. These findings unambiguously demonstrate postthymic, RAG-dependent TCR rearrangement and define TCR revision as a tolerance pathway that targets mature peripheral CD4(+) T cells.