We have segregated DR1+ individuals into two categories according to whether or not their class II+ cells stimulated T lymphocyte clones specific for or restricted to DR1. In a majority of cases (87%), failure to stimulate was a property of cells having the B14;DR1 haplotype and/or nonclassical 21-hydroxylase deficiency. Absence of clonal proliferation could not be explained by release of an intercellular suppressor factor or by stimulator cell absorption of interleukin 2. Homozygous cells inheriting both stimulatory (DR1(n)) and nonstimulatory (DR1(x)) haplotypes did not successfully mediate clonal expansion, implying that a trans acting factor operates intracellularly to modify both DR1 alleles or their products. Other DR alleles did not appear to be affected as evidenced by normal proliferative responses of T lymphocyte clones restricted to DR2 or DR7 and stimulated by DR1(x),2 and DR1(x),7 cells, respectively. By two-dimensional gel analysis, we have further identified a 50-kD surface glycoprotein contained in anti-DR immunoprecipitates of DR1(x), but not DR1(n) or non-DR1 cellular lysates. This 50-kD structure had antigenic and peptide identity to DRα and β chains but was resistant to dissociation under conditions that normally separate DRα and β (8 M urea plus 5% 2-mercaptoethanol); boiling in sodium dodecyl sulfate was required to segregate the component polypeptides of the 50-kD heterodimer. We postulate that a product of a novel combinatorial association between constitutive chains of DR may interfere with or compete for normal T cell receptor recognition of DR1 as both an alloantigen and a restricting element. We further propose that gene abnormalities within the class III region of a haplotype associated with nonclassical 21-hydroxylase deficiency may extend into the DR subregion of the major histocompatibility complex with consequent aberrations in DR1 presentation.