AUTOIMMUNITY TO RNA-POLYMERASE-II IS FOCUSED AT THE CARBOXYL-TERMINALDOMAIN OF THE LARGE SUBUNIT

Objective. Previous studies have demonstrated antibodies to the large (220 kd) polypeptide subunit of RNA polymerase II (Pol II) in sera fro m certain patients with scleroderma. In the present study, we sought t o identify the autoantigenic region on this polypeptide. Methods. A re combinant fusion protein, corresponding to the 52-heptapeptide repeat found in the carboxyl terminal domain (CTD) of the large Pol II subuni t, was used to identify 15 patient sera that contained autoantibodies, Synthetic peptides CTD7 (representing a single heptapeptide) and CTD1 8 (representing 2 1/2 heptapeptide repeats) mere used in a competitive inhibition assay to define the specificity of these sera and the impo rtance of the CTD as an autoantigen. Results. All 15 sera immunoprecip itated the Pol II subunit from radiolabeled cell extracts, and 11 of t hem bound the CTD fusion protein in immunoblots, Immunoprecipitation o f Pol II was completely inhibited by CTD18 in 5 sera and partially inh ibited in 4 additional sera. Conclusion. These results indicate that t he CTD heptapeptide repeat is a focal point for autoimmune responses i n scleroderma. It is likely that the repetitive sequence and high cont ent of charged residues of this structure contribute to its role as an autoantigen.