There is strong genetic association between type 1A diabetes (T1D) and autoimmune thyroid disease (AITD). T1D and AITD frequently occur together in the same individual, a condition classified as a variant of the autoimmune polyglandular syndrome type 3 (APS3). Because T1D and AITD are individually strongly associated with different HLA class II sequences, we asked which HLA class II pocket sequence and structure confer joint susceptibility to both T1D and AITD in the same individual (APS3v). We sequenced the HLA-DR gene in 105 APS3v patients and 153 controls, and identified a pocket amino acid signature, DRβ-Tyr-26, DRβ-Leu-67, DRβ-Lys-71, and DRβ-Arg-74, that was strongly associated with APS3v (P = 5.4 × 10 −14 , odds ratio = 8.38). Logistic regression analysis demonstrated that DRβ-Leu-67 (P = 9.4 × 10 −13) and DRβ-Arg-74 (P = 1.21 × 10 −13 ) gave strong independent effects on disease susceptibility. Structural modeling studies demonstrated that pocket 4 was critical for the development of T1D+AITD; all disease-associated amino acids were linked to areas of the pocket that interact directly with the peptide and, therefore, influence peptide binding. The disease-susceptible HLA-DR pocket was more positively charged (Lys-71, Arg-74) compared with the protective pocket (Ala-71, Gln-74). We conclude that a specific pocket amino acid signature confers joint susceptibility to T1D+AITD in the same individual by causing significant structural changes in the MHC II peptide binding pocket and influencing peptide binding and presentation. Moreover, Arg-74 is a major amino acid position for the development of several autoimmune diseases. These findings suggest that blocking the critical Arg-74 pocket might offer a method for treating certain autoimmune conditions. autoimmunity | gene | structure T ype 1A (autoimmune) diabetes mellitus (T1D) and autoimmune thyroid disease (AITD) are the most common autoimmune endocrine disorders (1-3). Despite affecting different tissues, T1D and AITD share a common etiology. Both are organspecific autoimmune diseases characterized by infiltration of the gland by autoreactive T and B cells and production of antibodies directed at the target organs (the pancreatic islets in T1D and the thyroid in AITD), resulting in their dysfunction or destruction. In fact, there is a well-known strong association between T1D and AITD (reviewed in ref. 1). They frequently occur within the same family (aggregation) and in the same individual (comorbidity) (1). When T1D and AITD occur in the same individual the phenotype is considered, according to a commonly accepted classification, as a variant of the autoimmune polyglandular syndrome type 3 (APS3) (4,5) [in this manuscript, we use the nomenclature which defines APS3 as AITD + another autoimmune condition excluding Addison's disease (4) for better clarity, and we refer to the combination AITD+T1D in the same individual as APS3 variant (APS3v) to emphasize that this is not the only form of APS3]. However, it should be noted that the classification of APS is...