Position β57 of I-A controls early anti-insulin responses in NOD mice, linking an MHC susceptibility allele to type 1 diabetes onset.

The class II region of the major histocompatibility complex (MHC) locus is the main contributor to the genetic susceptibility to type 1 diabetes (T1D). The loss of an aspartic acid at position 57 of diabetogenic HLA-DQβ chains supports this association; this single amino acid change influences how TCRs recognize peptides in the context of HLA-DQ8 and I-A using a mechanism termed the P9 switch. Here, we built register-specific insulin peptide MHC tetramers to examine CD4 T cell responses to Ins and Ins peptides during the early prediabetic phase of disease in nonobese diabetic (NOD) mice. A single-cell analysis of anti-insulin CD4 T cells performed in 6- and 12-week-old NOD mice revealed tissue-specific gene expression signatures. TCR signaling and clonal expansion were found only in the islets of Langerhans and produced either classical T1 differentiation or an unusual T phenotype, independent of TCR usage. The early phase of the anti-insulin response was dominated by T cells specific for Ins, the register that supports a P9 switch mode of recognition. The presence of the P9 switch was demonstrated by TCR sequencing, reexpression, mutagenesis, and functional testing of TCRαβ pairs in vitro. Genetic correction of the I-Aβ57 mutation in NOD mice resulted in the disappearance of D/E residues in the CDR3β of anti-Ins T cells. These results provide a mechanistic molecular explanation that links the characteristic MHC class II polymorphism of T1D with the recognition of islet autoantigens and disease onset.