Several cDNA fragments encoding parts of the (U1)RNP specific 68-kDa autoantigen were expressed in Escherichia coli and the fusion proteins were used as substrate for localization of the autoreactive epitopes. We have identified a region of approximately 30 amino acids reacting with more than 90% (16 of 17) of all human anti-p68 sera tested, regions which carry only a few and a region with no autoepitopes. Comparative analysis of epitopes recognized on partially degraded fusion proteins indicated that the anti-p68 autoimmune response is polyclonal. It involves generation of antibodies to several epitopes including one in a region with retroviral gag protein homology speculated to play a role in the initiation of the autoimmune response. Each of the 17 sera tested contained a different set of autoantibody specificities. These data are not consistent with random mutation as a sole mechanism of anti-p68 autoantibody induction and argue for an Ag-driven autoimmune response.