Open AccessAutoimmunity to RNA polymerase II is focused at the carboxyl terminal domain of the large subunit
Author(s) -
Hirakata Michito,
Kanungo Jyotshna,
Suwa Akira,
Takeda Yoshihiko,
Craft Joe,
Hardin John A.
Publication year - 1996
Publication title -
arthritis & rheumatism
Language(s) - English
Resource type - Journals
eISSN - 1529-0131
pISSN - 0004-3591
DOI - 10.1002/art.1780391115
Subject(s) - ctd , protein subunit , fusion protein , microbiology and biotechnology , rna polymerase ii , immunoprecipitation , autoantibody , recombinant dna , biology , polymerase , antibody , rna , autoimmunity , biochemistry , gene expression , immunology , enzyme , gene , oceanography , promoter , geology
Objective . Previous studies have demonstrated antibodies to the large (220 kd) polypeptide subunit of RNA polymerase II (Pol II) in sera from certain patients with scleroderma. In the present study, we sought to identify the autoantigenic region on this polypeptide. Methods . A recombinant fusion protein, corresponding to the 52‐heptapeptide repeat found in the carboxyl terminal domain (CTD) of the large Pol II subunit, was used to identify 15 patient sera that contained autoantibodies. Synthetic peptides CTD7 (representing a single heptapeptide) and CTD18 (representing 2½ heptapeptide repeats) were used in a competitive inhibition assay to define the specificity of these sera and the importance of the CTD as an autoantigen. Results . All 15 sera immunoprecipitated the Pol II subunit from radiolabeled cell extracts, and 11 of them bound the CTD fusion protein in immunoblots. Immunoprecipitation of Pol II was completely inhibited by CTD18 in 5 sera and partially inhibited in 4 additional sera. Conclusion . These results indicate that the CTD heptapeptide repeat is a focal point for autoimmune responses in scleroderma. It is likely that the repetitive sequence and high content of charged residues of this structure contribute to its role as an autoantigen.