Different binding motifs of the celiac disease-associated HLA molecules DQ2.5, DQ2.2, and DQ7.5 revealed by relative quantitative proteomics of endogenous peptide repertoires

Abstract

Celiac disease is caused by intolerance to cereal gluten proteins, and HLA-DQ molecules are involved in the disease pathogenesis by presentation of gluten peptides to CD4+ T cells. The α- or β-chain sharing HLA molecules DQ2.5, DQ2.2, and DQ7.5 display different risks for the disease. It was recently demonstrated that T cells of DQ2.5 and DQ2.2 patients recognize distinct sets of gluten epitopes, suggesting that these two DQ2 variants select different peptides for display. To explore whether this is the case, we performed a comprehensive comparison of the endogenous self-peptides bound to HLA-DQ molecules of B-lymphoblastoid cell lines. Peptides were eluted from affinity-purified HLA molecules of nine cell lines and subjected to quadrupole orbitrap mass spectrometry and MaxQuant software analysis. Altogether, 12,712 endogenous peptides were identified at very different relative abundances. Hierarchical clustering of normalized quantitative data demonstrated significant differences in repertoires of peptides between the three DQ variant molecules. The neural network-based method, NNAlign, was used to identify peptide-binding motifs. The binding motifs of DQ2.5 and DQ7.5 concurred with previously established binding motifs. The binding motif of DQ2.2 was strikingly different from that of DQ2.5 with position P3 being a major anchor having a preference for threonine and serine. This is notable as three recently identified epitopes of gluten recognized by T cells of DQ2.2 celiac patients harbor serine at position P3. This study demonstrates that relative quantitative comparison of endogenous peptides sampled from our protein metabolism by HLA molecules provides clues to understand HLA association with disease.Electronic supplementary materialThe online version of this article (doi:10.1007/s00251-014-0819-9) contains supplementary material, which is available to authorized users.

T-cell and B-cell immunity in celiac disease

“…To bind to HLA molecules, peptides need to conform to certain binding motifs. In celiac disease, the different HLA types that are associated with the disease haven been shown to have distinct peptide binding preferences, corresponding with different T-cell repertoires that are found in these patients [39,40]. The HLA-DQ2.5, HLA-DQ8 and HLA-DQ2.2 molecules all prefer negatively charged anchor residues that are introduced into gluten peptides through deamidation.…”

“…This difference in epitope selection is governed by distinct binding specificity of the two HLA-DQ2 variants. Despite the fact that only one of the ten residues that differ between HLA-DQ2.2 and HLA-DQ2.5 makes contact with the peptide in the crystal structure of HLA-DQ2.5 with a bound gliadin epitope [41], the two molecules have widely different repertoires of endogenous peptides [39]. Peptides binding to HLA-DQ2.2 use P3 as a major anchor where there is preference for serine, threonine and asparatic acid.…”

“…Peptides binding to HLA-DQ2.2 use P3 as a major anchor where there is preference for serine, threonine and asparatic acid. Interestingly, the three characterized DQ2.2 restricted gluten epitopes to date all have serine at P3 [7, 26,39]. The remarkable difference in risk between the two variants is likely explained by quantitative differences in the amount of gluten peptides that are able to form stable complexes with each of the two HLA-DQ molecules.…”

“…This led to the prediction that different gluten epitopes would be presented in HLA-DQ2.2-expressing patients. Indeed, the two HLA-DQ2 variants select for distinct sets of gluten peptides for presentation to CD4 þ T cells [7, 26,39]. This difference in epitope selection is governed by distinct binding specificity of the two HLA-DQ2 variants.…”

Small Bowel, Celiac Disease and Adaptive Immunity

“…Moreover, peptide binding experiments indicated that Ser and Thr are anchor residues for binding of peptides to DQ2.2 but not to DQ2.5 [28,29]. This notion was further established by elution of endogenous peptides from B-lymphoblastoid cell lines [36]. This analysis revealed big differences in the repertoires of peptides eluted from DQ2.2 compared to DQ2.5, with a minority of peptides being presented by both HLA-DQ molecules.…”