The role of MHC class II polymorphisms for DM-mediated peptide editing and autoimmunity

Abstract

Abstract Polymorphisms in MHC class II (MHCII) genes are the strongest risk factors for the development of autoimmune diseases, such as type I diabetes (T1D). T1D is associated with allelic variants of the MHCII molecule HLA-DQ. In the nonobese diabetic (NOD) mouse, the orthologous molecule, I-Ag7, also predisposes mice to spontaneous diabetes development. To investigate the functional impact of natural MHCII polymorphisms on DM binding and T1D we introduced selected mutations in the genome of the NOD mouse by CRISPR/Cas9 genomic editing. Our approach was to first identify unique polymorphisms in the putative interface of I-Ag7, and then assess the impact of these polymorphisms on DM-mediated peptide editing in vitro. Several polymorphisms increased the interaction between I-Ag7 and DM and augmented peptide binding. The substitution of lysine (K) 40 in I-Aα with glutamic acid (E) had the strongest effect on DM editing and accelerated peptide binding more than 9-fold. Lysine at position α40 is unique to the diabetes associated I-Ag7 molecule and was therefore selected for in vivo evaluation. Homozygous and heterozygous K40E mutant mice showed increased H2-DM protein expression compared to WT littermate controls. Increased H2-DM levels were accompanied by an increase in intra- and extracellular I-Ag7 expression. This suggests that the K40E mutation promotes the interaction between H2-DM and I-Ag7, and likely, represents increased intracellular dimerization and more stable I-Ag7/peptide complexes on the cell surface. Currently, we are evaluating whether increased editing of the MHCII peptide repertoire mediated by the K40E mutation influences spontaneous diabetes development.