HUMAN AUTOANTIBODIES STABILIZE THE QUATERNARY STRUCTURE OF KU ANTIGEN

Objective. To examine humoral immune responses to the native Ku antige n and to evaluate the role of autoantibodies in stabilizing intermolec ular contacts between the p70 and p80 Ku subunits. Methods. Recombinan t free human p70 and p80 Ku subunits and p70/p80 heterodimers were exp ressed in Sf9 (insect) cells using baculovirus vectors. Affinity-purif ied recombinant human p70, p80, and p70/p80 dimer were studied by enzy me-linked immunosorbent assay (ELISA) and immunoprecipitation to evalu ate autoantibody specificities in sera from 58 patients with systemic autoimmune disease. Results. Anti-Ku antibodies were detected by ELISA or immunoprecipitation using K562 cell Ku antigen. All of the sera we re reactive with the native recombinant p70, p80, or p70/p80 antigens: 47% were anti-p70+,anti-p80+ and 32% were anti-p70-,anti-p80 +, but o nly 3% were anti-p70+,anti-p80-. Unexpectedly, 18% of the sera recogni zed the p70/p80 dimer but did not recognize native p70 or p80 alone. A subset of sera containing autoantibodies that prevent the dissociatio n of p70 from p80 by high salt and detergent treatment was identified; monoclonal antibody (MAb) 162, a murine anti-Ku MAb, displays the sam e property. Autoantibodies that stabilize the p70-p80 interaction were found most frequently in sera containing both anti-p70 and anti-p80 a ntibodies. Conclusion. Autoantibodies to the native p80 subunit of Ku are more common than are anti-p70 antibodies. When anti-p70 antibodies were detected, they generally were found together with anti-p80. A no vel type of autoantibody capable of stabilizing the p70/p80 heterodime r was identified in human sera for the first time. These ''stabilizing '' autoantibodies are found in sera containing both anti-p70 and anti- p80 antibodies, and also are produced by mice immunized with human Ku antigen. Autoimmunity to Ku may be initiated with an immune response t o p80, followed by spreading to p70. We hypothesize that stabilizing a ntibodies could facilitate the spreading of autoimmunity from one subu nit of Ku to another by altering the processing of p70 or p80 by antig en-presenting cells.