Anti-Ro/SS-A antibodies are often found in the sera of patients with the inflammatory rheumatic diseases Sjongren's syndrome (SS) and systemic lupus erythematosus (SLE). These antibodies have also been correlated with neonatal lupus and congenital heart block. The Ro/SS-A epitopes are carried by proteins which are found in the cell as part of a ribonucleoprotein (RNP) complex whose RNAs have been designated Y RNAs. To date, three protein components have been identified. The cellular function and localization of the Ro/SS-A particle remain obscure. Studies will be carried out to determine the cellular functions of the Ro/SS-A RNP and its role in autoimmune pathogenesis. To this end, recombinant DNA (rDNA) techniques will be utilized to characterize the individual components and their spatial relationship within the Ro/SS-A particle. Also, a specific and sensitive diagnostic/prognostic assay will be developed to detect anti-Ro/SS-A antibodies. cDNAs encoding the individual Ro/SS-A polypeptides will be utilized to produce these antigens by genetic engineering means. The recombinant proteins will be critical in the development of the above mentioned assay and in other aspects of this investigation. This research project will encompass the following studies for each protein: Identification of functional domains by sequence comparison as well as relatedness to other autoantigens and proteins; epitope mapping by rDNA techniques; generation of monospecific antibodies to the individual proteins/epitopes by using rDNA antigens. Genomic clones will be isolated by probing with the individual cDNAs; the genomic structure of genes encoding the Ro/SS-A antigens will be studied by Southern blot and DNA sequence analyses of the isolated genes; regulatory signals will be identified. Expression of the Ro/SS-A antigens will be studied in various tissues, species, and conditions by utilizing Northern blot analysis and gene fusions. Attempts to determine the cellular localization of the Ro/SS-A RNP will be done. Finally, the number of protein components and their interactions, both protein-RNA and protein-protein, within the Ro/SS-A complex will be characterized by reconstitution experiments using cross-linking agents.
