Prediction of HLA-DQ3.2beta ligands: evidence of multiple registers in class II binding peptides

MOTIVATION: While processing of MHC class II antigens for presentation to helper T-cells is essential for normal immune response, it is also implicated in the pathogenesis of autoimmune disorders and hypersensitivity reactions. Sequence-based computational techniques for predicting HLA-DQ binding peptides have encountered limited success, with few prediction techniques developed using three-dimensional models. METHODS: We describe a structure-based prediction model for modeling peptide-DQ3.2beta complexes. We have developed a rapid and accurate protocol for docking candidate peptides into the DQ3.2beta receptor and a scoring function to discriminate binders from the background. The scoring function was rigorously trained, tested and validated using experimentally verified DQ3.2beta binding and non-binding peptides obtained from biochemical and functional studies. RESULTS: Our model predicts DQ3.2beta binding peptides with high accuracy [area under the receiver operating characteristic (ROC) curve A(ROC) > 0.90], compared with experimental data. We investigated the binding patterns of DQ3.2beta peptides and illustrate that several registers exist within a candidate binding peptide. Further analysis reveals that peptides with multiple registers occur predominantly for high-affinity binders.     

The HLA molecules DQA1*0501/B1*0201 and DQA1*0301/B1*0302 share an extensive overlap in peptide binding specificity

Assays to measure the binding capacity of peptides for HLA-DQA1*0501/B*0201 (DQ2.3) and DQA1*0301/B*0302 (DQ3.2) were developed using solubilized MHC molecules purified from EBV-transformed cell lines. These quantitative assays, based on the principle of the inhibition of binding of a high-affinity radiolabeled ligand, were validated by examining the binding capacity of known DQ-restricted epitopes or ligands. The availability of these assays allowed an investigation of patterns of cross-reactivity between different DQ molecules and with various common DR molecules. DQ2.3 and DQ3.2 were found to have significantly overlapping peptide binding repertoires. Specifically, of 13 peptides that bound either DQ2.3 or DQ3.2, nine (69.2%) bound both. The molecular basis of this high degree of cross-reactivity was further investigated with panels of single substitution analogs of the thyroid peroxidase 632-645Y epitope. It was found that DQ2.3 and DQ3.2 bind the same ligands by using similar anchor residues but different registers. These data suggest that in analogy to what was previously described for HLA-DR molecules, HLA-DQ supertypes characterized by largely overlapping binding repertoires can be defined. In light of the known linkage of both HLA-DQ2.3 and -DQ3.2 with insulin-dependent diabetes mellitus and celiac disease, these results might have important implications for understanding HLA class II autoimmune disease associations.     

Towards universal structure-based prediction of class II MHC epitopes for diverse allotypes

The binding of peptide fragments of antigens to class II MHC proteins is a crucial step in initiating a helper T cell immune response. The discovery of these peptide epitopes is important for understanding the normal immune response and its misregulation in autoimmunity and allergies and also for vaccine design. In spite of their biomedical importance, the high diversity of class II MHC proteins combined with the large number of possible peptide sequences make comprehensive experimental determination of epitopes for all MHC allotypes infeasible. Computational methods can address this need by predicting epitopes for a particular MHC allotype. We present a structure-based method for predicting class II epitopes that combines molecular mechanics docking of a fully flexible peptide into the MHC binding cleft followed by binding affinity prediction using a machine learning classifier trained on interaction energy components calculated from the docking solution. Although the primary advantage of structure-based prediction methods over the commonly employed sequence-based methods is their applicability to essentially any MHC allotype, this has not yet been convincingly demonstrated. In order to test the transferability of the prediction method to different MHC proteins, we trained the scoring method on binding data for DRB1*0101 and used it to make predictions for multiple MHC allotypes with distinct peptide binding specificities including representatives from the other human class II MHC loci, HLA-DP and HLA-DQ, as well as for two murine allotypes. The results showed that the prediction method was able to achieve significant discrimination between epitope and non-epitope peptides for all MHC allotypes examined, based on AUC values in the range 0.632-0.821. We also discuss how accounting for peptide binding in multiple registers to class II MHC largely explains the systematically worse performance of prediction methods for class II MHC compared with those for class I MHC based on quantitative prediction performance estimates for peptide binding to class II MHC in a fixed register.     

Identification of noncanonical melanoma-associated T cell epitopes for cancer immunotherapy

The identification of tumor-associated T cell epitopes has contributed significantly to the understanding of the interrelationship of tumor and immune system and is instrumental in the development of therapeutic vaccines for the treatment of cancer. Most of the known epitopes have been identified with prediction algorithms that compute the potential capacity of a peptide to bind to HLA class I molecules. However, naturally expressed T cell epitopes need not necessarily be strong HLA binders. To overcome this limitation of the available prediction algorithms we established a strategy for the identification of T cell epitopes that include suboptimal HLA binders. To this end, an artificial neural network was developed that predicts HLA-binding peptides in protein sequences by taking the entire sequence context into consideration rather than computing the sum of the contribution of the individual amino acids. Using this algorithm, we predicted seven HLA A*0201-restricted potential T cell epitopes from known melanoma-associated Ags that do not conform to the canonical anchor motif for this HLA molecule. All seven epitopes were validated as T cell epitopes and three as naturally processed by melanoma tumor cells. T cells for four of the new epitopes were found at elevated frequencies in the peripheral blood of melanoma patients. Modification of the peptides to the canonical sequence motifs led to improved HLA binding and to improved capacity to stimulate T cells.     

NetTepi: an integrated method for the prediction of T cell epitopes

Multiple factors determine the ability of a peptide to elicit a cytotoxic T cell lymphocyte response. Binding to a major histocompatibility complex class I (MHC-I) molecule is one of the most essential factors, as no peptide can become a T cell epitope unless presented on the cell surface in complex with an MHC-I molecule. As such, peptide-MHC (pMHC) binding affinity predictors are currently the premier methods for T cell epitope prediction, and these prediction methods have been shown to have high predictive performances in multiple studies. However, not all MHC-I binders are T cell epitopes, and multiple studies have investigated what additional factors are important for determining the immunogenicity of a peptide. A recent study suggested that pMHC stability plays an important role in determining if a peptide can become a T cell epitope. Likewise, a T cell propensity model has been proposed for identifying MHC binding peptides with amino acid compositions favoring T cell receptor interactions. In this study, we investigate if improved accuracy for T cell epitope discovery can be achieved by integrating predictions for pMHC binding affinity, pMHC stability, and T cell propensity. We show that a weighted sum approach allows pMHC stability and T cell propensity predictions to enrich pMHC binding affinity predictions. The integrated model leads to a consistent and significant increase in predictive performance and we demonstrate how this can be utilized to decrease the experimental workload of epitope screens. The final method, NetTepi, is publically available at www.cbs.dtu.dk/services/NetTepi.     

Staining of celiac disease-relevant T cells by peptide-DQ2 multimers.

Gluten-specific T cells in the small intestinal mucosa are thought to play a central role in the pathogenesis of celiac disease (CD). The vast majority of these T cells recognize gluten peptides when presented by HLA-DQ2 (DQA1*05/DQB1*02), a molecule which immunogenetic studies have identified as conferring susceptibility to CD. We have previously identified and characterized three DQ2-restricted gluten epitopes that are recognized by intestinal T cells isolated from CD patients, two of which are immunodominant. Because almost all of the gluten epitopes are restricted by DQ2, and because we have detailed knowledge of several of these epitopes, we chose to develop peptide-DQ2 tetramers as a reagent to further investigate the role of these T cells in CD. In the present study, stable soluble DQ2 was produced such that it contained leucine zipper dimerization motif and a covalently coupled peptide. We have made four different peptide-DQ2 staining reagents, three containing the gluten epitopes and one containing a DQ2-binding self-peptide that provides a negative control for staining. We show in this study that peptide-DQ2 when adhered to plastic specifically stimulates T cell clones and that multimers comprising these molecules specifically stain peptide-specific T cell clones and lines. Interestingly, T cell activation caused severe reduction in staining intensities obtained with the multimers and an Ab to the TCR. The problem of TCR down-modulation must be taken into consideration when using class II multimers to stain T cells that may have been recently activated in vivo.     

Equilibrium and kinetic analysis of the unusual binding behavior of a highly immunogenic gluten peptide to HLA-DQ2

HLA-DQ2 predisposes an individual to celiac sprue by presenting peptides from dietary gluten to intestinal CD4(+) T cells. A selectively deamidated multivalent peptide from gluten (LQLQPFPQPELPYPQPELPYPQPELPYPQPQPF; underlined residues correspond to posttranslational Q --> E alterations) is a potent trigger of DQ2 restricted T cell proliferation. Here we report equilibrium and kinetic measurements of interactions between DQ2 and (i) this highly immunogenic multivalent peptide, (ii) its individual constituent epitopes, (iii) its nondeamidated precursor, and (iv) a reference high-affinity ligand of HLA-DQ2 that is not recognized by gluten-responsive T cells from celiac sprue patients. The deamidated 33-mer peptide efficiently exchanges with a preloaded peptide in the DQ2 ligand-binding groove at pH 5.5 as well as pH 7.3, suggesting that the peptide can be presented to T cells comparably well through the endocytic pathway or via direct loading onto extracellular HLA-DQ2. In contrast, the monovalent peptides, and the nondeamidated precursor, as well as the tight-binding reference peptide show a much poorer ability to exchange with a preloaded peptide in the DQ2 binding pocket, especially at pH 7.3, suggesting that endocytosis of these peptides is a prerequisite for T cell presentation. At pH 5.5 and 7.3, dissociation of the deamidated 33-mer peptide from DQ2 is much slower than dissociation of its constituent monovalent epitopes or the nondeamidated precursor but faster than dissociation of the reference high-affinity peptide. Oligomeric states involving multiple copies of the DQ2 heterodimer bound to a single copy of the multivalent 33-mer peptide are not observed. Together, these results suggest that the remarkable antigenicity of the 33-mer gluten peptide is primarily due to its unusually efficient ability to displace existing ligands in the HLA-DQ2 binding pocket, rather than an extremely low rate of dissociation.     

Refining the rules of gliadin T cell epitope binding to the disease-associated DQ2 molecule in celiac disease: importance of proline spacing and glutamine deamidation

Celiac disease is driven by intestinal T cells responsive to proline-rich gluten peptides that often harbor glutamate residues formed by tissue transglutaminase-mediated glutamine conversion. The disease is strongly associated with the HLA variant DQ2.5 (DQA1*05, DQB1*02), and intestinal gluten-reactive T cells from DQ2.5-positive patients are uniquely restricted by this HLA molecule. In this study, we describe the mapping of two novel T cell epitopes of gamma-gliadin and the experimental identification of the DQ2.5 binding register of these and three other gamma-gliadin epitopes. The new data extend the knowledge base for understanding the binding of gluten peptides to DQ2.5. The alignment of the experimentally determined binding registers of nine gluten epitopes reveal positioning of proline residues in positions P1, P3, P6, and P8 but never in positions P2, P4, P7, and P9. Glutamate residues formed by tissue transglutaminase-mediated deamidation are found in position P1, P4, P6, P7, or P9, but only deamidations in positions P4 and P6, and rarely in P7, seem to be crucial for T cell recognition. The majority of these nine epitopes are recognized by celiac lesion T cells when presented by the related but nonassociated DQ2.2 (DQA1*0201, DQB1*02) molecule. Interestingly, the DQ2.2 presentation for most epitopes is less efficient than presentation by the DQ2.5 molecule, and this is particularly prominent for the alpha-gliadin epitopes. Contrary to previous findings, our data do not show selective presentation of DQ2.5 over DQ2.2 for gluten epitopes that carry proline residues at the P3 position.     

Cutting edge: identification of novel T cell epitopes in Lol p5a by computational prediction.

Although atopic allergy affects 相似文献   

Type 1 diabetes-associated HLA-DQ8 transdimer accommodates a unique peptide repertoire

HLA-DQ2 and HLA-DQ8 are strongly predisposing haplotypes for type 1 diabetes (T1D). Yet HLA-DQ2/8 heterozygous individuals have a synergistically increased risk compared with HLA-DQ2 or HLA-DQ8 homozygote subjects that may result from the presence of a transdimer formed between the α-chain of HLA-DQ2 (DQA1*05:01) and the β-chain of HLA-DQ8 (DQB1*03:02). We generated cells exclusively expressing this transdimer (HLA-DQ8trans), characterized its peptide binding repertoire, and defined a unique transdimer-specific peptide binding motif that was found to be distinct from those of HLA-DQ2 and HLA-DQ8. This motif predicts an array of peptides of islet autoantigens as candidate T cell epitopes, many of which selectively bind to the HLA transdimer, whereas others bind to both HLA-DQ8 and transdimer with similar affinity. Our findings provide a molecular basis for the association between HLA-DQ transdimers and T1D and set the stage for rational testing of potential diabetogenic peptide epitopes.     

Structural bases of T cell antigen receptor recognition in celiac disease

Celiac disease (CeD) is a human leukocyte antigen (HLA)-linked autoimmune-like disorder that is triggered by the ingestion of gluten or related storage proteins. The majority of CeD patients are HLA-DQ2.5⁺, with the remainder being either HLA-DQ8⁺ or HLA-DQ2.2⁺. Structural studies have shown how deamidation of gluten epitopes engenders binding to HLA-DQ2.5/8, which then triggers an aberrant CD4⁺ T cell response. HLA tetramer studies, combined with structural investigations, have demonstrated that repeated patterns of TCR usage underpins the immune response to some HLADQ2.5/8 restricted gluten epitopes, with distinct TCR motifs representing common landing pads atop the HLA–gluten complexes. Structural studies have provided insight into TCR specificity and cross-reactivity towards gluten epitopes, as well as cross-reactivity to bacterial homologues of gluten epitopes, suggesting that environmental factors may directly play a role in CeD pathogenesis. Collectively, structural immunology-based studies in the CeD axis may lead to new therapeutics/diagnostics to treat CeD, and also serve as an exemplar for other T cell mediated autoimmune diseases.     

T cell epitope characterization in tandemly repetitive Trypanosoma cruzi B13 protein

Proteins containing tandemly repetitive sequences are present in several immunodominant protein antigens in pathogenic protozoan parasites. The tandemly repetitive Trypanosoma cruzi B13 protein is recognized by IgG antibodies from 98% of Chagas' disease patients. Little is known about the molecular mechanisms that lead to the immunodominance of the repeated sequences, and there is limited information on T cell epitopes in such repetitive antigens. We finely characterized the T cell recognition of the tandemly repetitive, degenerate B13 protein by T cell lines, clones and PBMC from Chagas' disease cardiomyopathy (CCC), asymptomatic T. cruzi infected (ASY) and non-infected individuals (N). PBMC proliferative responses to recombinant B13 protein were restricted to individuals bearing HLA-DQA1*0501(DQ7), -DR1, and -DR2; B13 peptides bound to the same HLA molecules in binding assays. The HLA-DQ7-restricted minimal T cell epitope [FGQAAAG(D/E)KP] was identified with an overlapping combinatorial peptide library including all B13 sequence variants in T. cruzi Y strain B13 protein; the underlined small residues GQA were the major HLA contact residues. Among natural B13 15-mer variant peptides, molecular modeling showed that several variant positions were solvent (TCR)-exposed, and substitutions at exposed positions abolished recognition. While natural B13 variant peptide S15.9 seems to be the immunodominant epitope for Chagas' disease patients, S15.4 was preferentially recognized by CCC rather than ASY patients, which may be pathogenically relevant. This is the first thorough characterization of T cell epitopes of a tandemly repetitive protozoan antigen and may suggest a role for T cell help in the immunodominance of protozoan repetitive antigens.     

No allelic variation in genes with high gliadin homology in patients with celiac disease and type 1 diabetes

Celiac disease (CD) is a complex inflammatory disorder of the small intestine, induced by dietary gluten in genetically susceptible individuals. CD is strongly associated with HLA-DQ2 and it has recently been established that gut-derived DQ2-restricted T cells from patients with CD predominantly recognize gluten-derived peptides in which specific glutamine residues are deamidated to glutamic acid by tissue transglutaminase. Recently, intestinally expressed human genes with high homology to DQ2-gliadin celiac T-cell epitopes have been identified. Single or double point mutations which would increase the celiac T-cell epitope homology, and mutation in these genes, leading to the expression of glutamic acid at particular positions, could hypothetically be involved in the initiation of CD in HLA-DQ2-positive children. Six gene regions with high celiac T-cell epitope homology were investigated for single-nucleotide polymorphisms using direct sequencing of DNA from 20 CD patients, 27 type 1 diabetes mellitus (T1DM) patients with associated CD, 24 patients with T1DM without CD and 110 healthy controls, all of Caucasian origin. No variants in any of these genes in any of the investigated groups were found. We conclude that gut-expressed human celiac epitope homologous peptides are unlikely to represent non-HLA risk factors in the development of celiac disease in Caucasians.     

Antigen presentation to celiac lesion-derived T cells of a 33-mer gliadin peptide naturally formed by gastrointestinal digestion

Celiac disease is an HLA-DQ2-associated disorder characterized by intestinal T cell responses to ingested wheat gluten proteins. A peptide fragment of 33 residues (alpha(2)-gliadin 56-88) produced by normal gastrointestinal proteolysis contains six partly overlapping copies of three T cell epitopes and is a remarkably potent T cell stimulator after deamidation by tissue transglutaminase (TG2). This 33-mer is rich in proline residues and adopts the type II polyproline helical conformation in solution. In this study we report that after deamidation, the 33-mer bound with higher affinity to DQ2 compared with other monovalent peptides harboring gliadin epitopes. We found that the TG2-treated 33-mer was presented equally effectively by live and glutaraldehyde-fixed, EBV-transformed B cells. The TG2-treated 33-mer was also effectively presented by glutaraldehyde-fixed dendritic cells, albeit live dendritic cells were the most effective APCs. A strikingly increased T cell stimulatory potency of the 33-mer compared with a 12-mer peptide was also seen with fixed APCs. The 33-mer showed binding maximum to DQ2 at pH 6.3, higher than maxima found for other high affinity DQ2 binders. The 33-mer is thus a potent T cell stimulator that does not require further processing within APC for T cell presentation and that binds to DQ2 with a pH profile that promotes extracellular binding.     

HLA-DQ8-associated T cell responses to the diabetes autoantigen phogrin (IA-2 beta) in human prediabetes

Susceptibility to type 1A autoimmune diabetes is linked to expression of particular MHC class II molecules, notably HLA-DQ8 in man and the orthologous I-Ag7 in the nonobese diabetic mouse. In the present study, we analyzed two peptide epitopes (peptides 2 and 7) from the diabetes autoantigen phogrin (IA-2beta), in the context of their presentation by the I-Ag7 and HLA-DQ8 molecules and their role as potential T cell antigenic epitopes in human diabetes. Both of these peptides are targets of diabetogenic CD4+ T cell clones in the nonobese diabetic mouse. Transgenic mice expressing HLA-DQ8 as the sole class II molecule generated a robust T cell-proliferative response when primed with peptide 2 or peptide 7 in CFA. Analysis of the IL-2 secretion from peptide 2-reactive T cell hybridomas stimulated with alanine-substituted peptides identified three residues that were crucial to the response. Among 41 islet cell Ag-positive prediabetic human subjects, 36.5% showed PBMC-proliferative responses to peptide 7, 17.1% to peptide 2, and 17.1% to both peptides; no response was seen among 20 matched healthy controls. Stratification of the data based upon HLA haplotype suggested that peptide 7 could be presented by at least one HLA-DR molecule in addition to HLA-DQ8, a finding that was supported by blocking studies with monomorphic mAbs. The results indicate that common phogrin peptides are targeted by autoreactive T cells in human and murine type 1A diabetes, and that the responses may in part be associated with the similar peptide-binding specificities of I-Ag7 and HLA-DQ8.     

Determination of glutamic acid decarboxylase 65 peptides presented by the type I diabetes-associated HLA-DQ8 class II molecule identifies an immunogenic peptide motif

Particular HLA class II allelic sequences are associated with susceptibility to type I diabetes. To understand the mechanism, knowledge of the molecular nature of the specific TCR/peptide/class II interactions involved in the disease process is required. To this end, we have introduced the diabetes-associated human class II HLA-DQ8 allele (DQA1*0301/DQB1*0302) as a transgene into mice and analyzed T cell responses restricted by this molecule to an important Ag in human diabetes, human glutamic acid decarboxylase 65. Hybridomas were used to determine the particular peptides from this Ag presented by HLA-DQ8 to T cells and to map the core minimal epitopes required for T cell stimulation. Analysis of these core epitopes reveals a motif and relevant features for peptides that are immunogenic to T cells when presented by HLA-DQ8. The major immunogenic epitopes of glutamic acid decarboxylase 65 do not contain a negatively charged residue that binds in the P9 pocket of the HLA-DQ8 molecule. PBMC from HLA-DQ8+ diabetic and nondiabetic individuals respond to these peptides, confirming that the mouse model is a useful tool to define epitopes of autoantigens that are processed by human APC and recognized by human T cells.     

Acetylcholine receptor peptide recognition in HLA DR3-transgenic mice: in vivo responses correlate with MHC-peptide binding.

HLA DR3 is an MHC molecule that reportedly predisposes humans to myasthenia gravis (MG). Though MG is an Ab-mediated autoimmune disease, CD4+ T cells are essential for the generation of high-affinity Abs; hence the specificities of autoreactive CD4+ T cells are important. In this study we report the HLA DR3-restricted T cell determinants on the extracellular region sequence of human acetylcholine receptor alpha subunit. We find two promiscuous determinants on this region 141-160 and 171-190 as defined by their immunogenicity in HLA DR3-, HLA DQ8-, and HLA DQ6-transgenic mice in the absence of endogenous mouse class II molecules. We also studied the minimal determinants of these two regions by truncation analysis, and the MHC binding affinity of a set of overlapping peptides spanning the complete sequence region of human acetylcholine receptor alpha subunit. One of the peptide sequences strongly immunogenic in HLA DR3-transgenic mice also had the highest binding affinity to HLA DR3. Identification of T cell determinants restricted to an MHC molecule known to predispose to MG may be an important step toward the development of peptide-based immunomodulation strategies for this autoimmune disease.     

HLA-DQ6 (DQB1*0601)-restricted T cells protect against experimental autoimmune encephalomyelitis in HLA-DR3.DQ6 double-transgenic mice by generating anti-inflammatory IFN-gamma

The human MHC class II genes are associated with genetic susceptibility to multiple sclerosis (MS), a chronic inflammatory demyelinating disease of the CNS of presumed autoimmune origin. These genes encode for proteins responsible for shaping immune response. The exact role of HLA-DQ and -DR genes in disease pathogenesis is not well-understood due to the high polymorphism, linkage disequilibrium, and heterogeneity of human populations. The advent of HLA class II-transgenic (Tg) mice has helped in answering some of these questions. Previously, using single-Tg mice (expressing the HLA-DR or -DQ gene), we showed that proteolipid protein (PLP)(91-110) peptide induced classical experimental autoimmune encephalomyelitis only in DR3.Abeta degrees mice, suggesting that DR3 (DRB1*0301) is a disease susceptible gene in the context of PLP. Human population studies have suggested that HLA-DQ6 (DQB1*0601) may be a protective gene in MS. To test this disease protection in an experimental model, we generated double-Tg mice expressing both HLA-DR3 and -DQ6. Introduction of DQ6 onto DR3-Tg mice led to a decrease in disease incidence on immunization with PLP(91-110) peptide indicating a dominant protective role of DQ6. This protective effect is due to high levels of IFN-gamma produced by DQ6-restricted T cells, which suppressed proliferation of encephalitogenic DR3-restricted T cells by inducing apoptosis. Our study indicates that DQ6 modifies the PLP(91-110)-specific T cell response in DR3 through anti-inflammatory effects of IFN-gamma, which is protective for experimental autoimmune encephalomyelitis. Thus, our double-Tg mouse provides a novel model in which to study epistatic interactions between HLA class II molecules in MS.     

GPS-MBA: computational analysis of MHC class II epitopes in type 1 diabetes

As a severe chronic metabolic disease and autoimmune disorder, type 1 diabetes (T1D) affects millions of people world-wide. Recent advances in antigen-based immunotherapy have provided a great opportunity for further treating T1D with a high degree of selectivity. It is reported that MHC class II I-A(g7) in the non-obese diabetic (NOD) mouse and human HLA-DQ8 are strongly linked to susceptibility to T1D. Thus, the identification of new I-A(g7) and HLA-DQ8 epitopes would be of great help to further experimental and biomedical manipulation efforts. In this study, a novel GPS-MBA (MHC Binding Analyzer) software package was developed for the prediction of I-A(g7) and HLA-DQ8 epitopes. Using experimentally identified epitopes as the training data sets, a previously developed GPS (Group-based Prediction System) algorithm was adopted and improved. By extensive evaluation and comparison, the GPS-MBA performance was found to be much better than other tools of this type. With this powerful tool, we predicted a number of potentially new I-A(g7) and HLA-DQ8 epitopes. Furthermore, we designed a T1D epitope database (TEDB) for all of the experimentally identified and predicted T1D-associated epitopes. Taken together, this computational prediction result and analysis provides a starting point for further experimental considerations, and GPS-MBA is demonstrated to be a useful tool for generating starting information for experimentalists. The GPS-MBA is freely accessible for academic researchers at: http://mba.biocuckoo.org.     

Physiological induction of regulatory Qa-1-restricted CD8+ T cells triggered by endogenous CD4+ T cell responses

T cell-dependent autoimmune diseases are characterized by the expansion of T cell clones that recognize immunodominant epitopes on the target antigen. As a consequence, for a given autoimmune disorder, pathogenic T cell clones express T cell receptors with a limited number of variable regions that define antigenic specificity. Qa-1, a MHC class I-like molecule, presents peptides from the variable region of TCRs to Qa-1-restricted CD8+ T cells. The induction of Vß-specific CD8+ T cells has been harnessed in an immunotherapeutic strategy known as the “T cell vaccination” (TCV) that comprises the injection of activated and attenuated CD4+ T cell clones so as to induce protective CD8+ T cells. We hypothesized that Qa-1-restricted CD8+ regulatory T cells could also constitute a physiologic regulatory arm of lymphocyte responses upon expansion of endogenous CD4+ T cells, in the absence of deliberate exogenous T cell vaccination. We immunized mice with two types of antigenic challenges in order to sequentially expand antigen-specific endogenous CD4+ T cells with distinct antigenic specificities but characterized by a common Vß chain in their TCR. The first immunization was performed with a non-self antigen while the second challenge was performed with a myelin-derived peptide known to drive experimental autoimmune encephalomyelitis (EAE), a mouse model of multiple sclerosis. We show that regulatory Vß-specific Qa-1-restricted CD8+ T cells induced during the first endogenous CD4+ T cell responses are able to control the expansion of subsequently mobilized pathogenic autoreactive CD4+ T cells. In conclusion, apart from the immunotherapeutic TCV, Qa-1-restricted specialized CD8+ regulatory T cells can also be induced during endogenous CD4+ T cell responses. At variance with other regulatory T cell subsets, the action of these Qa-1-restricted T cells seems to be restricted to the immediate re-activation of CD4+ T cells.