An autoimmune stem-like CD8 T cell population drives type 1 diabetes.

CD8 T cell-mediated autoimmune diseases result from the breakdown of self-tolerance mechanisms in autoreactive CD8 T cells1. How autoimmune T cell populations arise and are sustained, and the molecular programmes defining the autoimmune T cell state, are unknown. In type 1 diabetes, ß-cell-specific CD8 T cells destroy insulin-producing ß-cells. Here we followed the fate of ß-cell-specific CD8 T cells in non-obese diabetic mice throughout the course of type 1 diabetes. We identified a stem-like autoimmune progenitor population in the pancreatic draining lymph node (pLN), which self-renews and gives rise to pLN autoimmune mediators. pLN autoimmune mediators migrate to the pancreas, where they differentiate further and destroy ß-cells. Whereas transplantation of as few as 20 autoimmune progenitors induced type 1 diabetes, as many as 100,000 pancreatic autoimmune mediators did not. Pancreatic autoimmune mediators are short-lived, and stem-like autoimmune progenitors must continuously seed the pancreas to sustain ß-cell destruction. Single-cell RNA sequencing and clonal analysis revealed that autoimmune CD8 T cells represent unique T cell differentiation states and identified features driving the transition from autoimmune progenitor to autoimmune mediator. Strategies aimed at targeting the stem-like autoimmune progenitor pool could emerge as novel and powerful immunotherapeutic interventions for type 1 diabetes.

Structural and biochemical analysis of highly similar HLA-B allotypes differentially associated with type 1 diabetes.

Type 1 diabetes (T1D) is an autoimmune disease involving T cell-mediated destruction of the insulin-producing beta cells in the pancreatic islets of Langerhans. CD8+ T cells, responding to beta cell peptides presented by class I major histocompatibility complex (MHC) molecules, are important effectors leading to beta cell elimination. Human leukocyte antigen (HLA) B*39:06, B*39:01, and B*38:01 are closely related class I MHC allotypes that nonetheless show differential association with T1D. HLA-B*39:06 is the most predisposing of all HLA class I molecules and is associated with early age at disease onset. B*39:01 is also associated with susceptibility to T1D, but to a lesser extent, though differing from B*39:06 by only two amino acids. HLA-B*38:01, in contrast, is associated with protection from the disease. Upon identifying a peptide that binds to both HLA-B*39:06 and B*39:01, we determined the respective X-ray structures of the two allotypes presenting this peptide to 1.7 Å resolution. The peptide residues available for T cell receptor contact and those serving as anchors were identified. Analysis of the F pocket of HLA-B*39:06 and B*39:01 provided an explanation for the distinct peptide C-terminus preferences of the two allotypes. Structure-based modeling of the protective HLA-B*38:01 suggested a potential reason for its peptide preferences and its reduced propensity to present 8-mer peptides compared to B*39:06. Notably, the three allotypes showed differential binding to peptides derived from beta cell autoantigens. Taken together, our findings should facilitate identification of disease-relevant candidate T cell epitopes and structure-guided therapeutics to interfere with peptide binding.

Human insulin as both antigen and protector in type 1 diabetes.

Type 1 diabetes (T1D) is characterized by T-cell responses to islet antigens. Investigations in humans and the nonobese diabetic (NOD) mouse model of T1D have revealed that T-cell reactivity to insulin plays a central role in the autoimmune response. As there is no convenient NOD-based model to study human insulin (hIns) or its T-cell epitopes in the context of spontaneous T1D, we developed a NOD mouse strain transgenically expressing hIns in islets under the control of the human regulatory region. Female NOD.hIns mice developed T1D at approximately the same rate and overall incidence as NOD mice. Islet-infiltrating T cells from NOD.hIns mice recognized hIns peptides; both CD8 and CD4 T-cell epitopes were identified. We also demonstrate that islet-infiltrating T cells from HLA-transgenic NOD.hIns mice can be used to identify potentially patient-relevant hIns T-cell epitopes. Besides serving as an antigen, hIns was expressed in the thymus of NOD.hIns mice and could serve as a protector against T1D under certain circumstances, as previously suggested by genetic studies in humans. NOD.hIns mice and related strains facilitate human-relevant epitope discovery efforts and the investigation of fundamental questions that cannot be readily addressed in humans.

Noncontiguous T cell epitopes in autoimmune diabetes: From mice to men and back again.

Type 1 diabetes (T1D) is a T cell-mediated autoimmune disease that affects the insulin-producing beta cells of the pancreatic islets. The nonobese diabetic mouse is a widely studied spontaneous model of the disease that has contributed greatly to our understanding of T1D pathogenesis. This is especially true in the case of antigen discovery. Upon review of existing knowledge concerning the antigens and peptide epitopes that are recognized by T cells in this model, good concordance is observed between mouse and human antigens. A fascinating recent illustration of the contribution of the nonobese diabetic mouse in the area of epitope identification is the discovery of noncontiguous CD4+ T cell epitopes. This novel epitope class is characterized by the linkage of an insulin-derived peptide to, most commonly, a fragment of a natural cleavage product of another beta cell secretory granule constituent. These so-called hybrid insulin peptides are also recognized by T cells in patients with T1D, although the precise mechanism for their generation has yet to be defined and is the subject of active investigation. Although evidence from the tumor immunology arena documented the existence of noncontiguous CD8+ T cell epitopes, generated by proteasome-mediated peptide splicing involving transpeptidation, such CD8+ T cell epitopes were thought to be a rare immunological curiosity. However, recent advances in bioinformatics and mass spectrometry have challenged this view. These developments, coupled with the discovery of hybrid insulin peptides, have spurred a search for noncontiguous CD8+ T cell epitopes in T1D, an exciting frontier area still in its infancy.

HLA-B*39:06 Efficiently Mediates Type 1 Diabetes in a Mouse Model Incorporating Reduced Thymic Insulin Expression.

Type 1 diabetes (T1D) is characterized by T cell-mediated destruction of the insulin-producing ß cells of the pancreatic islets. Among the loci associated with T1D risk, those most predisposing are found in the MHC region. HLA-B*39:06 is the most predisposing class I MHC allele and is associated with an early age of onset. To establish an NOD mouse model for the study of HLA-B*39:06, we expressed it in the absence of murine class I MHC. HLA-B*39:06 was able to mediate the development of CD8 T cells, support lymphocytic infiltration of the islets, and confer T1D susceptibility. Because reduced thymic insulin expression is associated with impaired immunological tolerance to insulin and increased T1D risk in patients, we incorporated this in our model as well, finding that HLA-B*39:06-transgenic NOD mice with reduced thymic insulin expression have an earlier age of disease onset and a higher overall prevalence as compared with littermates with typical thymic insulin expression. This was despite virtually indistinguishable blood insulin levels, T cell subset percentages, and TCR Vß family usage, confirming that reduced thymic insulin expression does not impact T cell development on a global scale. Rather, it will facilitate the thymic escape of insulin-reactive HLA-B*39:06-restricted T cells, which participate in ß cell destruction. We also found that in mice expressing either HLA-B*39:06 or HLA-A*02:01 in the absence of murine class I MHC, HLA transgene identity alters TCR Vß usage by CD8 T cells, demonstrating that some TCR Vß families have a preference for particular class I MHC alleles.

Delivery of siRNAs to Dendritic Cells Using DEC205-Targeted Lipid Nanoparticles to Inhibit Immune Responses.

Due to their ability to knock down the expression of any gene, siRNAs have been heralded as ideal candidates for treating a wide variety of diseases, including those involving "undruggable" targets. However, the therapeutic potential of siRNAs remains severely limited by a lack of effective delivery vehicles. Recently, lipid nanoparticles (LNPs) containing ionizable cationic lipids have been developed for hepatic siRNA delivery. However, their suitability for delivery to other cell types has not been determined. We have modified LNPs for preferential targeting to dendritic cells (DCs), central regulators of immune responses. To achieve directed delivery, we coated LNPs with a single-chain antibody (scFv; DEC-LNPs), specific to murine DEC205, which is highly expressed on distinct DC subsets. Here we show that injection of siRNAs encapsulated in DEC-LNPs are preferentially delivered to DEC205(+) DCs. Gene knockdown following uptake of DEC-LNPs containing siRNAs specific for the costimulatory molecules CD40, CD80, and CD86 dramatically decreases gene expression levels. We demonstrate the functionality of this knockdown with a mixed lymphocyte response (MLR). Overall, we report that injection of LNPs modified to restrict their uptake to a distinct cell population can confer profound gene knockdown, sufficient to inhibit powerful immune responses like the MLR.

Genetically modified human CD4(+) T cells can be evaluated in vivo without lethal graft-versus-host disease.

Adoptive cell immunotherapy for human diseases, including the use of T cells modified to express an anti-tumour T-cell receptor (TCR) or chimeric antigen receptor, is showing promise as an effective treatment modality. Further advances would be accelerated by the availability of a mouse model that would permit human T-cell engineering protocols and proposed genetic modifications to be evaluated in vivo. NOD-scid IL2rγ(null) (NSG) mice accept the engraftment of mature human T cells; however, long-term evaluation of transferred cells has been hampered by the xenogeneic graft-versus-host disease (GVHD) that occurs soon after cell transfer. We modified human primary CD4(+) T cells by lentiviral transduction to express a human TCR that recognizes a pancreatic beta cell-derived peptide in the context of HLA-DR4. The TCR-transduced cells were transferred to NSG mice engineered to express HLA-DR4 and to be deficient for murine class II MHC molecules. CD4(+) T-cell-depleted peripheral blood mononuclear cells were also transferred to facilitate engraftment. The transduced cells exhibited long-term survival (up to 3 months post-transfer) and lethal GVHD was not observed. This favourable outcome was dependent upon the pre-transfer T-cell transduction and culture conditions, which influenced both the kinetics of engraftment and the development of GVHD. This approach should now permit human T-cell transduction protocols and genetic modifications to be evaluated in vivo, and it should also facilitate the development of human disease models that incorporate human T cells.

Characterization of the peptide binding specificity of the HLA class I alleles B*38:01 and B*39:06.

B*38:01 and B*39:06 are present with phenotypic frequencies <2% in the general population, but are of interest as B*39:06 is the B allele most associated with type 1 diabetes susceptibility and 38:01 is most protective. A previous study derived putative main anchor motifs for both alleles based on peptide elution data. The present study has utilized panels of single amino acid substitution peptide libraries to derive detailed quantitative motifs accounting for both primary and secondary influences on peptide binding. From these analyses, both alleles were confirmed to utilize the canonical position 2/C-terminus main anchor spacing. B*38:01 preferentially bound peptides with the positively charged or polar residues H, R, and Q in position 2 and the large hydrophobic residues I, F, L, W, and M at the C-terminus. B*39:06 had a similar preference for R in position 2, but also well-tolerated M, Q, and K. A more dramatic contrast between the two alleles was noted at the C-terminus, where the specificity of B*39:06 was clearly for small residues, with A as most preferred, followed by G, V, S, T, and I. Detailed position-by-position and residue-by-residue coefficient values were generated from the panels to provide detailed quantitative B*38:01 and B*39:06 motifs. It is hoped that these detailed motifs will facilitate the identification of T cell epitopes recognized in the context of two class I alleles associated with dramatically different dispositions towards type 1 diabetes, offering potential avenues for the investigation of the role of CD8 T cells in this disease.

Compensatory mechanisms allow undersized anchor-deficient class I MHC ligands to mediate pathogenic autoreactive T cell responses.

Self-reactive T cells must escape thymic negative selection to mediate pathogenic autoimmunity. In the NOD mouse model of autoimmune diabetes, several ß cell-cytotoxic CD8 T cell populations are known, with the most aggressive of these represented by AI4, a T cell clone with promiscuous Ag-recognition characteristics. We identified a long-elusive ß cell-specific ligand for AI4 as an unusually short H-2D(b)-binding 7-mer peptide lacking a C-terminal anchor residue and derived from the insulin A chain (InsA14-20). Crystallography reveals that compensatory mechanisms permit peptides lacking a C-terminal anchor to bind sufficiently to the MHC to enable destructive T cell responses, yet allow cognate T cells to avoid negative selection. InsA14-20 shares two solvent-exposed residues with previously identified AI4 ligands, providing a structural explanation for AI4's promiscuity. Detection of AI4-like T cells, using mimotopes of InsA14-20 with improved H-2D(b)-binding characteristics, establishes the AI4-like T cell population as a consistent feature of the islet infiltrates of NOD mice. Our work establishes undersized peptides as previously unrecognized targets of autoreactive CD8 T cells and presents a strategy for their further exploration as Ags in autoimmune disease.

Glucagon-reactive islet-infiltrating CD8 T cells in NOD mice.

Type 1 diabetes is characterized by T-cell-mediated destruction of the insulin-producing ß cells in pancreatic islets. A number of islet antigens recognized by CD8 T cells that contribute to disease pathogenesis in non-obese diabetic (NOD) mice have been identified; however, the antigenic specificities of the majority of the islet-infiltrating cells have yet to be determined. The primary goal of the current study was to identify candidate antigens based on the level and specificity of expression of their genes in mouse islets and in the mouse ß cell line MIN6. Peptides derived from the candidates were selected based on their predicted ability to bind H-2K(d) and were examined for recognition by islet-infiltrating T cells from NOD mice. Several proteins, including those encoded by Abcc8, Atp2a2, Pcsk2, Peg3 and Scg2, were validated as antigens in this way. Interestingly, islet-infiltrating T cells were also found to recognize peptides derived from proglucagon, whose expression in pancreatic islets is associated with α cells, which are not usually implicated in type 1 diabetes pathogenesis. However, type 1 diabetes patients have been reported to have serum autoantibodies to glucagon, and NOD mouse studies have shown a decrease in α cell mass during disease pathogenesis. Our finding of islet-infiltrating glucagon-specific T cells is consistent with these reports and suggests the possibility of α cell involvement in development and progression of disease.

DEC-205-mediated antigen targeting to steady-state dendritic cells induces deletion of diabetogenic CD8⁺ T cells independently of PD-1 and PD-L1.

CD8⁺ T cells specific for islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP) have been implicated in type 1 diabetes in both humans and non-obese diabetic (NOD) mice, in which T cells specific for IGRP206₋214 are highly prevalent. We sought to manipulate these pathogenic T cells by exploiting the ability of steady-state dendritic cells (DCs) to present antigens in a tolerogenic manner. The endocytic receptor DEC-205 was utilized to deliver an IGRP206₋214 mimotope to DCs in NOD mice, and the impact of this delivery on a polyclonal population of endogenous islet-reactive cognate T cells was determined. Assessment of islet-infiltrating CD8⁺ T cells showed a decrease in the percentage, and the absolute number, of endogenous IGRP206₋214-specific T cells when the mimotope was delivered to DCs, compared with delivery of a specificity control. Employing an adoptive transfer system, deletion of CD8⁺ T cells as a result of DEC-205-mediated antigen targeting was found to occur independently of programmed death-1 (PD-1) and its ligand (PD-L1), both often implicated in the regulation of peripheral T-cell tolerance. Given its promise for the manipulation of self-reactive polyclonal T cells demonstrated here, the distinctive characteristics of this antigen delivery system will be important to appreciate as its potential as an intervention for autoimmune diseases continues to be investigated.

Beyond HLA-A*0201: new HLA-transgenic nonobese diabetic mouse models of type 1 diabetes identify the insulin C-peptide as a rich source of CD8+ T cell epitopes.

Type 1 diabetes is an autoimmune disease characterized by T cell responses to ß cell Ags, including insulin. Investigations employing the NOD mouse model of the disease have revealed an essential role for ß cell-specific CD8(+) T cells in the pathogenic process. As CD8(+) T cells specific for ß cell Ags are also present in patients, these reactivities have the potential to serve as therapeutic targets or markers for autoimmune activity. NOD mice transgenic for human class I MHC molecules have previously been employed to identify T cell epitopes having important relevance to the human disease. However, most studies have focused exclusively on HLA-A*0201. To broaden the reach of epitope-based monitoring and therapeutic strategies, we have looked beyond this allele and developed NOD mice expressing human ß(2)-microglobulin and HLA-A*1101 or HLA-B*0702, which are representative members of the A3 and B7 HLA supertypes, respectively. We have used islet-infiltrating T cells spontaneously arising in these strains to identify ß cell peptides recognized in the context of the transgenic HLA molecules. This work has identified the insulin C-peptide as an abundant source of CD8(+) T cell epitopes. Responses to these epitopes should be of considerable utility for immune monitoring, as they cannot reflect an immune reaction to exogenously administered insulin, which lacks the C-peptide. Because the peptides bound by one supertype member were found to bind certain other members also, the epitopes identified in this study have the potential to result in therapeutic and monitoring tools applicable to large numbers of patients and at-risk individuals.

Structural and functional characterization of a single-chain peptide-MHC molecule that modulates both naive and activated CD8+ T cells.

Peptide-MHC (pMHC) multimers, in addition to being tools for tracking and quantifying antigen-specific T cells, can mediate downstream signaling after T-cell receptor engagement. In the absence of costimulation, this can lead to anergy or apoptosis of cognate T cells, a property that could be exploited in the setting of autoimmune disease. Most studies with class I pMHC multimers used noncovalently linked peptides, which can allow unwanted CD8(+) T-cell activation as a result of peptide transfer to cellular MHC molecules. To circumvent this problem, and given the role of self-reactive CD8(+) T cells in the development of type 1 diabetes, we designed a single-chain pMHC complex (scK(d).IGRP) by using the class I MHC molecule H-2K(d) and a covalently linked peptide derived from islet-specific glucose-6-phosphatase catalytic subunit-related protein (IGRP(206-214)), a well established autoantigen in NOD mice. X-ray diffraction studies revealed that the peptide is presented in the groove of the MHC molecule in canonical fashion, and it was also demonstrated that scK(d).IGRP tetramers bound specifically to cognate CD8(+) T cells. Tetramer binding induced death of naive T cells and in vitro- and in vivo-differentiated cytotoxic T lymphocytes, and tetramer-treated cytotoxic T lymphocytes showed a diminished IFN-γ response to antigen stimulation. Tetramer accessibility to disease-relevant T cells in vivo was also demonstrated. Our study suggests the potential of single-chain pMHC tetramers as possible therapeutic agents in autoimmune disease. Their ability to affect the fate of naive and activated CD8(+) T cells makes them a potential intervention strategy in early and late stages of disease.

Ins2 deficiency augments spontaneous HLA-A*0201-restricted T cell responses to insulin.

Type 1 diabetes results from the autoimmune destruction of insulin-producing beta cells by T cells specific for beta cell Ags, including insulin. In humans, the non-MHC locus conferring the strongest disease susceptibility is the insulin gene, and alleles yielding lower thymic insulin expression are predisposing. We sought to incorporate this characteristic into an HLA-transgenic model of the disease and to determine the influence of reduced thymic insulin expression on CD8+ T cell responses to preproinsulin. We examined NOD.Ins2(-/-) mice, which do not express insulin in the thymus and show accelerated disease, to determine whether they exhibit quantitative or qualitative differences in CD8+ T cell responses to preproinsulin. We also generated NOD.Ins2(-/-) mice expressing type 1 diabetes-associated HLA-A*0201 (designated NOD.beta2m(-/-).HHD.Ins2(-/-)) in an effort to obtain an improved humanized disease model. We found that CD8+ T cell reactivity to certain insulin peptides was more readily detected in NOD.Ins2(-/-) mice than in NOD mice. Furthermore, the proportion of insulin-reactive CD8+ T cells infiltrating the islets of NOD.Ins2(-/-) mice was increased. NOD.beta2m(-/-).HHD.Ins2(-/-) mice exhibited rapid onset of disease and had an increased proportion of HLA-A*0201-restricted insulin-reactive T cells, including those targeting the clinically relevant epitope Ins B10-18. Our results suggest that insulin alleles that predispose to type 1 diabetes in humans do so, at least in part, by facilitating CD8+ T cell responses to the protein. We propose the NOD.beta2m(-/-).HHD.Ins2(-/-) strain as an improved humanized disease model, in particular for studies seeking to develop therapeutic strategies targeting insulin-specific T cells.

Multiple antigens versus single major antigen in type 1 diabetes: arguing for multiple antigens.

Our recent review of the literature revealed that approximately 20 antigens are now known to be targeted by T cells in the NOD mouse model of the autoimmune disease type 1 diabetes. Of these, insulin has received considerable attention and has been described by some in the research community as an 'initiating' or 'single major' antigen in the disease. Insulin may indeed be worthy of these titles, at least in NOD mice and in the context of the particular major histocompatibility complex molecules expressed in this strain. However, here we present arguments in favour of viewing type 1 diabetes as a disease in which multiple antigens should be considered, rather than just one. In our view, other antigens may prove to be more worthy of these titles in humans, and the major histocompatibility complex molecules expressed may well be a determining factor. Furthermore, even if insulin is 'the initiating antigen' in type 1 diabetes, multiple pathogenic specificities are known to exist even during the prediabetic period and it is at our peril that we ignore them. The recent discovery of novel beta-cell antigens, e.g. ZnT8 and chromogranin A, has taught us that we still have much to learn about the targets of the autoimmune response in type 1 diabetes. Increased knowledge will promote a clearer picture of disease pathogenesis and will better position the field to be successful in its translational goals of immune monitoring and disease prevention and reversal.

Predominant occupation of the class I MHC molecule H-2Kwm7 with a single self-peptide suggests a mechanism for its diabetes-protective effect.

Type 1 diabetes (T1D) is an autoimmune disease characterized by T cell-mediated destruction of insulin-producing pancreatic beta cells. In both humans and the non-obese diabetic (NOD) mouse model of T1D, class II MHC alleles are the primary determinant of disease susceptibility. However, class I MHC genes also influence risk. These findings are consistent with the requirement for both CD4(+) and CD8(+) T cells in the pathogenesis of T1D. Although a large body of work has permitted the identification of multiple mechanisms to explain the diabetes-protective effect of particular class II MHC alleles, studies examining the protective influence of class I alleles are lacking. Here, we explored this question by performing biochemical and structural analyses of the murine class I MHC molecule H-2K(wm7), which exerts a diabetes-protective effect in NOD mice. We have found that H-2K(wm7) molecules are predominantly occupied by the single self-peptide VNDIFERI, derived from the ubiquitous protein histone H2B. This unexpected finding suggests that the inability of H-2K(wm7) to support T1D development could be due, at least in part, to the failure of peptides from critical beta-cell antigens to adequately compete for binding and be presented to T cells. Predominant presentation of a single peptide would also be expected to influence T-cell selection, potentially leading to a reduced ability to select a diabetogenic CD8(+) T-cell repertoire. The report that one of the predominant peptides bound by T1D-protective HLA-A*31 is histone derived suggests the potential translation of our findings to human diabetes-protective class I MHC molecules.

Selective delivery of beta cell antigen to dendritic cells in vivo leads to deletion and tolerance of autoreactive CD8+ T cells in NOD mice.

Type 1 diabetes (T1D) is an autoimmune disease resulting from defects in central and peripheral tolerance and characterized by T cell-mediated destruction of islet beta cells. Cytotoxic CD8(+) T cells, reactive to beta cell antigens, are required for T1D development in the NOD mouse model of the disease, and CD8(+) T cells specific for beta cell antigens can be detected in the peripheral blood of T1D patients. It has been evident that in nonautoimmune-prone mice, dendritic cells (DCs) present model antigens in a tolerogenic manner in the steady state, e.g., in the absence of infection, and cause T cells to proliferate initially but then to be deleted or rendered unresponsive. However, this fundamental concept has not been evaluated in the setting of a spontaneous autoimmune disease. To do so, we delivered a mimotope peptide, recognized by the diabetogenic CD8(+) T cell clone AI4, to DCs in NOD mice via the endocytic receptor DEC-205. Proliferation of transferred antigen-specific T cells was initially observed, but this was followed by deletion. Tolerance was achieved because rechallenge of mice with the mimotope peptide in adjuvant did not induce an immune response. Thus, targeting of DCs with beta cell antigens leads to deletion of autoreactive CD8(+) T cells even in the context of ongoing autoimmunity in NOD mice with known tolerance defects. Our results provide support for the development of DC targeting of self antigens for treatment of chronic T cell-mediated autoimmune diseases.

A comprehensive integrated post-GWAS analysis of Type 1 diabetes reveals enhancer-based immune dysregulation.

Type 1 diabetes (T1D) is an organ-specific autoimmune disease, whereby immune cell-mediated killing leads to loss of the insulin-producing ß cells in the pancreas. Genome-wide association studies (GWAS) have identified over 200 genetic variants associated with risk for T1D. The majority of the GWAS risk variants reside in the non-coding regions of the genome, suggesting that gene regulatory changes substantially contribute to T1D. However, identification of causal regulatory variants associated with T1D risk and their affected genes is challenging due to incomplete knowledge of non-coding regulatory elements and the cellular states and processes in which they function. Here, we performed a comprehensive integrated post-GWAS analysis of T1D to identify functional regulatory variants in enhancers and their cognate target genes. Starting with 1,817 candidate T1D SNPs defined from the GWAS catalog and LDlink databases, we conducted functional annotation analysis using genomic data from various public databases. These include 1) Roadmap Epigenomics, ENCODE, and RegulomeDB for epigenome data; 2) GTEx for tissue-specific gene expression and expression quantitative trait loci data; and 3) lncRNASNP2 for long non-coding RNA data. Our results indicated a prevalent enhancer-based immune dysregulation in T1D pathogenesis. We identified 26 high-probability causal enhancer SNPs associated with T1D, and 64 predicted target genes. The majority of the target genes play major roles in antigen presentation and immune response and are regulated through complex transcriptional regulatory circuits, including those in HLA (6p21) and non-HLA (16p11.2) loci. These candidate causal enhancer SNPs are supported by strong evidence and warrant functional follow-up studies.

T-cell autoantigens in the non-obese diabetic mouse model of autoimmune diabetes.

The non-obese diabetic (NOD) mouse model of autoimmune (type 1) diabetes has contributed greatly to our understanding of disease pathogenesis and has facilitated the development and testing of therapeutic strategies to combat the disease. Although the model is a valuable immunological tool in its own right, it reaches its fullest potential in areas where its findings translate to the human disease. Perhaps the foremost example of this is the field of T-cell antigen discovery, from which diverse benefits can be derived, including the development of antigen-specific disease interventions. The majority of NOD T-cell antigens are also targets of T-cell autoimmunity in patients with type 1 diabetes, and several of these are currently being evaluated in clinical trials. Here we review the journeys of these antigens from bench to bedside. We also discuss several recently identified NOD T-cell autoantigens whose translational potential warrants further investigation.

Assembly and structural properties of glucocorticoid-induced TNF receptor ligand: Implications for function.

Glucocorticoid-induced TNF receptor ligand (GITRL), a recently identified member of the TNF family, binds to its receptor GITR on both effector and regulatory T cells and generates positive costimulatory signals implicated in a wide range of T cell functions. Structural analysis reveals that the human GITRL (hGITRL) ectodomain self-assembles into an atypical expanded homotrimer with sparse monomer-monomer interfaces. Consistent with the small intersubunit interfaces, hGITRL exhibits a relatively weak tendency to trimerize in solution and displays a monomer-trimer equilibrium not reported for other TNF family members. This unique assembly behavior has direct implications for hGITRL-GITR signaling, because enforced trimerization of soluble hGITRL ectodomain results in an approximately 100-fold increase in its receptor binding affinity and also in enhanced costimulatory activity. The apparent reduction in affinity that is the consequence of this dynamic equilibrium may represent a mechanism to realize the biologically optimal level of signaling through the hGITRL-GITR pathway, as opposed to the maximal achievable level.