AUTOIMMUNE ANTIGENS AND SYSTEMIC SCLEROSIS

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Autoantibodies are characteristic of the rheumatic disease, systemic sclerosis or scleroderma. A major specificity is designated anti-centromere, and encompasses at least six target (CENP) antigens. This proposal focuses on the centromere polypeptides, with particular emphasis on two of the primary antigen targets, CENP-A and CENP-C. Using molecular and biochemical techniques, this research is directed toward the characterization of these centromere antigens, both as immunoreactive polypeptides and as nuclear kinetochore-associated components. These antigens will be isolated using a variety of chromatographic techniques. Included in their characterization will be the determination of polypeptide sequences to design oligonucleotide probes to facilitate cDNA cloning. A continuing emphasis in this laboratory is the use of recombinant DNA technology for the cloning and expression of genes encoding relevant autoantigens. The availability of cDNA clones and recombinant polypeptides will allow for a comprehensive assessment of the centromere antigens at various levels. These studies will include at the DNA level: Northern blot analysis to characterize specific mRNA species and heterogeneity; genomic analysis to determine organization of the intact gene. The centromere polypeptides will be described as to physicochemical properties based on: cDNA encoded sequence analysis; assessment of posttranslational modifications; exploration of nucleic acid binding properties; determination of any oligomeric centromere structures. Delineation of properties essential to the centromere polypeptides as antigens will include: detailed epitope mapping; analysis of shared epitopes among the various antigens; production of monoclonal and polyclonal antibodies to be used as probes of centromere immunoreactive domains. A critical end point of these studies will be the development of quantitative diagnostic/prognostic assays for scleroderma based on the individual polypeptide antigens. Overall, these studies are asking what features make the centromere antigens targets in the autoimmune sequence of events, while relating these same polypeptides to the native cellular environment in which they function.