A long existent medical puzzle is why some patients with certain rhematic and connective tissue diseases produce antibodies against their own cellular components. An understanding of the pathogenesis of these diseases would be facilitated if one could critically define the structure and function of the target antigens. The subjects of this proposal are two such antigens, La (or SS-B/Sjogren's syndrome-B) and Ro (or SS-A). In each case, the immunoreactive species have been identified as protein in nature. These polypeptides will be isolated and described in terms of any unique or shared physical properties utilizing a number of techniques including affinity chromatography, one- and two-dimensional gel electrophoresis and protein blots. Both La and Ro function within the context of larger ribonucleoprotein complexes designated snRNPs (small nuclear RNPs) or scRNPs (small cytoplasmic RNPs) and these will be isolated, aided by the use of differential immunoaffinity chromatography. Each particle will be identified as to its characteristic protein moieties with particular emphasis on the possible discrimination of individual La and Ro snRNPs (scRNPs) that contain unique RNA species (including those of viral origin) and on the delineation of those particles that contain both La and Ro polypeptides. The end result of these experiments will be to describe discrete, characterized entities that willhave relevance in the development of more defined and sensitive diagnostic assays than are currently available, and that will facilitate functional studies of these antigens and their associated complexes. To the latter end, an RNA polymerase III transcription factor has been associated with the La snRNP by this laboratory; this factor will be characterized as to its interactions with DNA templates and with the other components of the active transcription complex. Lastly, anti-La and Ro-monoclonal antibodies will be produced to assist in these studies; they will be made using the hybridoma technology and implemented by different protocols of in vivo and in vitro immunization.
