Abstract Major histocompatibility complex (MHC) genes, also known as human leukocyte antigen (HLA) genes in humans, are the prevailing risk factors in multiple autoimmune diseases, including multiple sclerosis (MS), autoimmune diabetes (type 1 diabetes or T1D), and rheumatoid arthritis (RA), yet MHC-linked susceptibility is not fully understood. In T1D, HLA-DQ8trans (DQ8trans), a trans-complementary molecule encoded by DQA1*0501 (HLA-DQ2α) and DQB1*0302 (HLA-DQ8β), confers exceptionally high risk. In contrast to the notion that DQ8trans expands a unique pathogenic T cell repertoire due to its trans-complementary nature, here we report a high degree of epitope overlap between DQ8trans and another high-risk allele, HLA-DQ8 (DQ8; DQA1*0301/DQB1*0302).T cells specific for shared islet-epitopes are cross-reactive to both molecules. Cross-reactivity is driven mainly by the shared β-chain and is observed ex vivo in polyclonal T cell populations in T1D patients. Stronger disease-associated responses to cross-reactive epitopes are elicited by DQ8trans, as the HLA-DQ2 α-chain enhances peptide-MHC-TCR interaction.These results suggest that a functional epistatic interaction between DQ8 and DQ8trans promotes the pathogenesis of T1D and provide an important general mechanism whereby disease risk may be altered primarily byT cell promiscuity among susceptible HLA haplotypes.