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Structural, biochemical and cellular analyses show that bacterial antigens can mimic gliadin epitopes involved in celiac disease being presented by HLA-DQ2.5 and recognized by T cells derived from patients.The human leukocyte antigen (HLA) locus is strongly associated with T cell-mediated autoimmune disorders. HLA-DQ2.5-mediated celiac disease (CeD) is triggered by the ingestion of gluten, although the relative roles of genetic and environmental risk factors in CeD is unclear. Here we identify microbially derived mimics of gliadin epitopes and a parental bacterial protein that is naturally processed by antigen-presenting cells and activated gliadin reactive HLA-DQ2.5-restricted T cells derived from CeD patients. Crystal structures of T cell receptors in complex with HLA-DQ2.5 bound to two distinct bacterial peptides demonstrate that molecular mimicry underpins cross-reactivity toward the gliadin epitopes. Accordingly, gliadin reactive T cells involved in CeD pathogenesis cross-react with ubiquitous bacterial peptides, thereby suggesting microbial exposure as a potential environmental factor in CeD.

Autoreactivity to myeloperoxidase (MPO) causes anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV), with rapidly progressive glomerulonephritis. Here, we show that a Staphylococcus aureus peptide, homologous to an immunodominant MPO T-cell epitope (MPO409–428), can induce anti-MPO autoimmunity. The peptide (6PGD391–410) is part of a plasmid-encoded 6-phosphogluconate dehydrogenase found in some S. aureus strains. It induces anti-MPO T-cell autoimmunity and MPO-ANCA in mice, whereas related sequences do not. Mice immunized with 6PGD391–410, or with S. aureus containing a plasmid expressing 6PGD391–410, develop glomerulonephritis when MPO is deposited in glomeruli. The peptide induces anti-MPO autoreactivity in the context of three MHC class II allomorphs. Furthermore, we show that 6PGD391–410 is immunogenic in humans, as healthy human and AAV patient sera contain anti-6PGD and anti-6PGD391–410 antibodies. Therefore, our results support the idea that bacterial plasmids might have a function in autoimmune disease., Autoreactivity to myeloperoxidase (MPO) causes autoimmune vasculitis and severe glomerulonephritis. Here, Ooi et al. show that a Staphylococcus aureus plasmid encodes a peptide that is homologous to an immunodominant MPO epitope and induces anti-MPO autoimmunity and glomerulonephritis in mice.

Individuals expressing HLA-DR4 bearing the shared susceptibility epitope (SE) have an increased risk of developing rheumatoid arthritis (RA). Posttranslational modification of self-proteins via citrullination leads to the formation of neoantigens that can be presented by HLA-DR4 SE allomorphs. However, in T cell-mediated autoimmunity, the interplay between the HLA molecule, posttranslationally modified epitope(s), and the responding T cell repertoire remains unclear. In HLA-DR4 transgenic mice, we show that immunization with a Fibβ-74cit69-81 peptide led to a population of HLA-DR4Fibβ-74cit69-81 tetramer+ T cells that exhibited biased T cell receptor (TCR) β chain usage, which was attributable to selective clonal expansion from the preimmune repertoire. Crystal structures of pre- and postimmune TCRs showed that the SE of HLA-DR4 represented a main TCR contact zone. Immunization with a double citrullinated epitope (Fibβ-72,74cit69-81) altered the responding HLA-DR4 tetramer+ T cell repertoire, which was due to the P2-citrulline residue interacting with the TCR itself. We show that the SE of HLA-DR4 has dual functionality, namely, presentation and a direct TCR recognition determinant. Analogous biased TCR β chain usage toward the Fibβ-74cit69-81 peptide was observed in healthy HLA-DR4+ individuals and patients with HLA-DR4+ RA, thereby suggesting a link to human RA.

The HLA-DRB1 locus is strongly associated with rheumatoid arthritis (RA) susceptibility, whereupon citrullinated self-peptides bind to HLA-DR molecules bearing the shared epitope (SE) amino acid motif. However, the differing propensity for citrullinated/non-citrullinated self-peptides to bind given HLA-DR allomorphs remains unclear. Here, we used a fluorescence polarization assay to determine a hierarchy of binding affinities of 34 self-peptides implicated in RA against three HLA-DRB1 allomorphs (HLA-DRB1*04:01/*04:04/*04:05) each possessing the SE motif. For all three HLA-DRB1 allomorphs, we observed a strong correlation between binding affinity and citrullination at P4 of the bound peptide ligand. A differing hierarchy of peptide-binding affinities across the three HLA-DRB1 allomorphs was attributable to the β-chain polymorphisms that resided outside the SE motif and were consistent with sequences of naturally presented peptide ligands. Structural determination of eight HLA–DR4–self-epitope complexes revealed strict conformational convergence of the P4-Cit and surrounding HLA β-chain residues. Polymorphic residues that form part of the P1 and P9 pockets of the HLA-DR molecules provided a structural basis for the preferential binding of the citrullinated self-peptides to the HLA-DR4 allomorphs. Collectively, we provide a molecular basis for the interplay between citrullination of self-antigens and HLA polymorphisms that shape peptide–HLA-DR4 binding affinities in RA.

The human leukocyte antigen (HLA) locus is strongly associated with T cell-mediated autoimmune disorders. HLA-DQ2.5-mediated celiac disease (CeD) is triggered by the ingestion of gluten, although the relative roles of genetic and environmental risk factors in CeD is unclear. Here we identify microbially derived mimics of gliadin epitopes and a parental bacterial protein that is naturally processed by antigen-presenting cells and activated gliadin reactive HLA-DQ2.5-restricted T cells derived from CeD patients. Crystal structures of T cell receptors in complex with HLA-DQ2.5 bound to two distinct bacterial peptides demonstrate that molecular mimicry underpins cross-reactivity toward the gliadin epitopes. Accordingly, gliadin reactive T cells involved in CeD pathogenesis cross-react with ubiquitous bacterial peptides, thereby suggesting microbial exposure as a potential environmental factor in CeD.

In HLA-DQ8–associated celiac disease (CD), the pathogenic T cell response is directed toward an immunodominant α-gliadin–derived peptide (DQ8-glia-α1). However, our knowledge of TCR gene usage within the primary intestinal tissue of HLA-DQ8+ CD patients is limited. We identified two populations of HLA-DQ8-glia-α1 tetramer+ CD4+ T cells that were essentially undetectable in biopsy samples from patients on a gluten-free diet but expanded rapidly and specifically after antigenic stimulation. Distinguished by expression of TRBV9, both T cell populations displayed biased clonotypic repertoires and reacted similarly against HLA-DQ8-glia-α1. In particular, TRBV9 paired most often with TRAV26-2, whereas the majority of TRBV9− TCRs used TRBV6-1 with no clear TRAV gene preference. Strikingly, both tetramer+/TRBV9+ and tetramer+/TRBV9− T cells possessed a non–germline-encoded arginine residue in their CDR3α and CDR3β loops, respectively. Comparison of the crystal structures of three TRBV9+ TCRs and a TRBV9− TCR revealed that, as a result of distinct TCR docking modes, the HLA-DQ8-glia-α1 contacts mediated by the CDR3-encoded arginine were almost identical between TRBV9+ and TRBV9− TCRs. In all cases, this interaction centered on two hydrogen bonds with a specific serine residue in the bound peptide. Replacement of serine with alanine at this position abrogated TRBV9+ and TRBV9− clonal T cell proliferation in response to HLA-DQ8-glia-α1. Gluten-specific memory CD4+ T cells with structurally and functionally conserved TCRs therefore predominate in the disease-affected tissue of patients with HLA-DQ8–mediated CD.

A comprehensive structural portrait of the association between citrullination, the HLA-DRB1 locus, and T cell autoreactivity in rheumatoid arthritis., Rheumatoid arthritis (RA) is strongly associated with the human leukocyte antigen (HLA)-DRB1 locus that possesses the shared susceptibility epitope (SE) and the citrullination of self-antigens. We show how citrullinated aggrecan and vimentin epitopes bind to HLA-DRB1*04:01/04. Citrulline was accommodated within the electropositive P4 pocket of HLA-DRB1*04:01/04, whereas the electronegative P4 pocket of the RA-resistant HLA-DRB1*04:02 allomorph interacted with arginine or citrulline-containing epitopes. Peptide elution studies revealed P4 arginine–containing peptides from HLA-DRB1*04:02, but not from HLA-DRB1*04:01/04. Citrullination altered protease susceptibility of vimentin, thereby generating self-epitopes that are presented to T cells in HLA-DRB1*04:01+ individuals. Using HLA-II tetramers, we observed citrullinated vimentin- and aggrecan-specific CD4+ T cells in the peripheral blood of HLA-DRB1*04:01+ RA-affected and healthy individuals. In RA patients, autoreactive T cell numbers correlated with disease activity and were deficient in regulatory T cells relative to healthy individuals. These findings reshape our understanding of the association between citrullination, the HLA-DRB1 locus, and T cell autoreactivity in RA.

Susceptibility and protection against human autoimmune diseases, including type I diabetes, multiple sclerosis, and Goodpasture disease, is associated with particular human leukocyte antigen (HLA) alleles. However, the mechanisms underpinning such HLA-mediated effects on self-tolerance remain unclear. Here we investigate the molecular mechanism of Goodpasture disease, an HLA-linked autoimmune renal disorder characterized by an immunodominant CD4(+) T-cell self-epitope derived from the α3 chain of type IV collagen (α3135-145). While HLA-DR15 confers a markedly increased disease risk, the protective HLA-DR1 allele is dominantly protective in trans with HLA-DR15 (ref. 2). We show that autoreactive α3135-145-specific T cells expand in patients with Goodpasture disease and, in α3135-145-immunized HLA-DR15 transgenic mice, α3135-145-specific T cells infiltrate the kidney and mice develop Goodpasture disease. HLA-DR15 and HLA-DR1 exhibit distinct peptide repertoires and binding preferences and present the α3135-145 epitope in different binding registers. HLA-DR15-α3135-145 tetramer(+) T cells in HLA-DR15 transgenic mice exhibit a conventional T-cell phenotype (Tconv) that secretes pro-inflammatory cytokines. In contrast, HLA-DR1-α3135-145 tetramer(+) T cells in HLA-DR1 and HLA-DR15/DR1 transgenic mice are predominantly CD4(+)Foxp3(+) regulatory T cells (Treg cells) expressing tolerogenic cytokines. HLA-DR1-induced Treg cells confer resistance to disease in HLA-DR15/DR1 transgenic mice. HLA-DR15(+) and HLA-DR1(+) healthy human donors display altered α3135-145-specific T-cell antigen receptor usage, HLA-DR15-α3135-145 tetramer(+) Foxp3(-) Tconv and HLA-DR1-α3135-145 tetramer(+) Foxp3(+)CD25(hi)CD127(lo) Treg dominant phenotypes. Moreover, patients with Goodpasture disease display a clonally expanded α3135-145-specific CD4(+) T-cell repertoire. Accordingly, we provide a mechanistic basis for the dominantly protective effect of HLA in autoimmune disease, whereby HLA polymorphism shapes the relative abundance of self-epitope specific Treg cells that leads to protection or causation of autoimmunity.

In HLA-DQ8-associated celiac disease, TRAV26-2+-TRBV9+ and TRAV8-3+-TRBV6+ T cells recognize the immunodominant DQ8-glia-α1 epitope, whereupon a non-germline-encoded arginine residue played a key role in binding HLA-DQ8-glia-α1. Whether distinct T cell receptor (TCR) recognition modes exist for gliadin epitopes remains unclear. TCR repertoire analysis revealed populations of HLA-DQ8-glia-α1 and HLA-DQ8.5-glia-γ1 restricted TRAV20+-TRBV9+ T cells that did not possess a non-germline-encoded arginine residue. The crystal structures of a TRAV20+-TRBV9+ TCR-HLA-DQ8-glia-α1 complex and two TRAV20+-TRBV9+ TCR-HLA-DQ8.5-glia-γ1 complexes were determined. This revealed that the differential specificity toward DQ8-glia-α1 and DQ8.5-glia-γ1 was governed by CDR3β-loop-mediated interactions. Surprisingly, a germline-encoded arginine residue within the CDR1α loop of the TRAV20+ TCR substituted for the role of the non-germline-encoded arginine in the TRAV26-2+-TRBV9+ and TRAV8-3+-TRBV6+ TCRs. Thus, in celiac disease, the responding TCR repertoire is driven by a common mechanism that selects for structural elements within the TCR that have convergent binding solutions in HLA-DQ8-gliadin recognition.

Celiac disease is a human leukocyte antigen (HLA)-DQ2- and/or DQ8-associated T cell-mediated disorder that is induced by dietary gluten. Although it is established how gluten peptides bind HLA-DQ8 and HLA-DQ2, it is unclear how such peptide-HLA complexes are engaged by the T cell receptor (TCR), a recognition event that triggers disease pathology. We show that biased TCR usage (TRBV9(∗)01) underpins the recognition of HLA-DQ8-α-I-gliadin. The structure of a prototypical TRBV9(∗)01-TCR-HLA-DQ8-α-I-gliadin complex shows that the TCR docks centrally above HLA-DQ8-α-I-gliadin, in which all complementarity-determining region-β (CDRβ) loops interact with the gliadin peptide. Mutagenesis at the TRBV9(∗)01-TCR-HLA-DQ8-α-I-gliadin interface provides an energetic basis for the Vβ bias. Moreover, CDR3 diversity accounts for TRBV9(∗)01(+) TCRs exhibiting differing reactivities toward the gliadin epitopes at various deamidation states. Accordingly, biased TCR usage is an important factor in the pathogenesis of DQ8-mediated celiac disease.

Central to adaptive immunity is the interaction between the αβ T cell receptor (TCR) and peptide presented by the major histocompatibility complex (MHC) molecule. Presumably reflecting TCR-MHC bias and T cell signaling constraints, the TCR universally adopts a canonical polarity atop the MHC. We report the structures of two TCRs, derived from human induced T regulatory (iT(reg)) cells, complexed to an MHC class II molecule presenting a proinsulin-derived peptide. The ternary complexes revealed a 180° polarity reversal compared to all other TCR-peptide-MHC complex structures. Namely, the iT(reg) TCR α-chain and β-chain are overlaid with the α-chain and β-chain of MHC class II, respectively. Nevertheless, this TCR interaction elicited a peptide-reactive, MHC-restricted T cell signal. Thus TCRs are not 'hardwired' to interact with MHC molecules in a stereotypic manner to elicit a T cell signal, a finding that fundamentally challenges our understanding of TCR recognition.

Celiac disease is a T cell-mediated disease induced by dietary gluten, a component of which is gliadin. 95% of individuals with celiac disease carry the HLA (human leukocyte antigen)-DQ2 locus. Here we determined the T-cell receptor (TCR) usage and fine specificity of patient-derived T-cell clones specific for two epitopes from wheat gliadin, DQ2.5-glia-α1a and DQ2.5-glia-α2. We determined the ternary structures of four distinct biased TCRs specific for those epitopes. All three TCRs specific for DQ2.5-glia-α2 docked centrally above HLA-DQ2, which together with mutagenesis and affinity measurements provided a basis for the biased TCR usage. A non-germline encoded arginine residue within the CDR3β loop acted as the lynchpin within this common docking footprint. Although the TCRs specific for DQ2.5-glia-α1a and DQ2.5-glia-α2 docked similarly, their interactions with the respective gliadin determinants differed markedly, thereby providing a basis for epitope specificity.

Rheumatoid arthritis (RA) is a chronic and debilitating autoimmune disease, characterized by inflammation of synovial tissue, joint pannus and bone erosion. The human leukocyte antigen (HLA) locus plays a vital role in immunity, encoding highly polymorphic molecules that present peptides to T cell lymphocytes. RA has a strong association with a region of the HLA-DRB1 locus known as the `shared epitope' (SE) and the presence of autoantibodies specific for citrullinated proteins. The SE maps to a highly polymorphic N-terminal region of the HLA-DRβ chain around amino acids 70-74. This region encodes a positively charged residue at position 71 that is thought to dictate the amino acid that is accommodated in the P4 pocket of the antigen-binding groove. Citrullination, the conversion of positively charged arginine to polar citrulline, is a physiological process catalyzed by peptidyl arginine deiminase. Previous studies have shown that citrullination of self-antigens can significantly increase the affinity of epitopes for SE alleles. Here we provide a molecular basis for how citrullinated vimentin and aggrecan epitopes can be presented by the SE alleles, HLA-DRB1*0401 and HLA-DRB1*0404. Citrulline was accommodated in the electropositive P4 pocket of HLA-DRB1*0401/04, whilst arginine was not. In addition, the RA resistant HLA-DRB1*0402 allomorph was capable of binding both arginine and citrulline in its electronegative P4 pocket. Peptide elution studies revealed that arginine was presented by HLA-DRB1*0402 but not by HLA-DRB1*0401/04. Moreover, citrullinated vimentin showed a greater sensitivity to proteolysis by cathepsin L, when compared to unmodified vimentin, indicating that citrullination can impact the repertoire of self-antigens presented. Using HLA Class II tetramers, we identified citrullinated vimentin and aggrecan specific CD4+ T cells from both HLA-DRB1*0401+ RA patients and healthy controls. In RA patients, the number of autoreactive T cells correlated with disease activity and were deficient in regulatory T cells compared to healthy controls. Together these findings provide significant insight into the role citrullination plays in the pathogenesis of RA[1].