A NOVEL CENTROMERE MONOSPECIFIC SERUM TO A HUMAN AUTOEPITOPE ON THE HISTONE H3-LIKE PROTEIN CENP-A

Centromere autoantibodies are commonly found in the serum of patients with some systemic autoimmune diseases, Previous studies have shown th at a major human centromere autoantigen is the histone H3-like protein CENP-A. Although the human cDNA has been cloned, native CENP-A has be en neither isolated nor expressed in Escherichia coli, and specific an tibodies to this chromatin-associated centromere protein are not avail able yet. In this report, a highly charged peptide on CENP-A (residues 3-17) was used to generate a monospecific antibody that reacts by imm unoblots with the 17 kDa centromeric protein, Immunofluorescence analy sis showed reactivity of this anti-CENP-A serum in several but not all mammalian culture cells analyzed, suggesting that the sequence of thi s histone-like centromere protein could be more variable throughout ev olution than originally thought, Selective extractions of human placen ta nuclear proteins and immunoblot analysis indicated that CENP-A beha ves in a similar way to the core histone polypeptides after nuclease d igestion of chromatin, Also, immunoblot analysis demonstrated that the CENP-A peptide used as immunogen is a target region on the CENP-A mol ecule in several but not all CREST patients analyzed with high titers of autoantibodies to the centromere. Lastly, we found that in Jurkat c ells induced to apoptosis, CENP-A remains associated with the centrome re, in contrast to other human autoantigens studied during apoptosis. (C) 1998 Federation of European Biochemical Societies.