Human endogenous retrovirus (HERV) long terminal repeat (LTR) elements contain regulatory sequences that can influence the expression of adjacent cellular genes, which may contribute to breakdowns of the immune function leading to autoimmune disease. Rheumatoid arthritis (RA) is associated with particular HLA-DR/DQ haplotypes that modulate the pathogenesis of this autoimmune disease. We have therefore studied a solitary LTR element (DQ-LTR3) of the HERV-K family at the HLA-DQB1 locus for a possible disease association among 228 RA patients and 311 unrelated blood donors. The DQ-LTR3 was significantly more frequent among patients (76% vs 33%, OR = 5.07,p < 0.0001), with the majority of patients being heterozygous for the DQ-LTR3 (61% vs 22%, p < 0.0001). HLA-DRB1*04 positive patients did still differ for the presence of the DQ-LTR3 (88% vs 70%, OR = 3.03, p < 0.001), with an increase of both DQ-LTR3 homozygous and heterozygous patients, when compared to DRB1*04 positive controls (p = 0.0015). HLA-DR/DQ genotype analysis among HLA-DRB1*04 positive individuals revealed significantly more DQ-LTR3 homozygotes among HLA-DRB1*04-DQBI*03 homozygous patients (72% vs 27%, P = 0.015), and the number of DQ-LTR3 homozygous (23% vs 19%) and heterozygous (66% vs 53%) individuals was also increased among HLA-DRB1*04 heterozygous patients (p = 0.034). The presence of the DQ-LTR3 element increased both the relative risk and the positive predictive value for either DRB1*04-DQB1*03 positive/negative individuals when compared to the presence of HLA-DRB1*04-DQB1*03 alone. In conclusion, these data suggest that this DQ-LTR3 enhances susceptibility to RA.