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Journal of Experimental Medicine, Vol 165, 1172-1187, Copyright © 1987 by Rockefeller University Press
ARTICLES |
N Chiorazzi and WH Reeves
B lymphocytes from a patient with systemic lupus erythematosus (SLE) and several circulating autoantibodies (including antinucleolar antibodies) were immortalized by fusion with a hypoxanthine/guanine phosphoribosyl transferase (HGPRT)-deficient human B cell line. Multiple human monoclonal antibodies (mAb) were obtained which, in solid-phase enzyme immunoassay, were reactive with DNA. One mAb was of special interest because it reacted strongly with both single-stranded DNA and an extractable nuclear antigen found in rabbit thymus extract (RTE). In an immunofluorescent assay using fixed human cells, the latter mAb also bound predominantly to cell nucleoli. A combination of enzyme digestion and metabolic inhibitor studies of the target cells in this immunofluorescent assay suggested that the antigen(s) bound by the mAb was an RNA-associated protein or a ribonucleoprotein that is distinct from intact RNA polymerase I and not associated with the transcriptional units of the nucleolus. In other experiments, using fractions of RTE isolated by ion-exchange chromatography, the antigens bound by the mAb were shown to be highly negatively charged molecules. Immunoprecipitation and SDS-PAGE analyses of labeled cell extracts bound by the mAb revealed a doublet of 17 and 18 kD. Since the original patient's serum autoantibodies also bound to both an RNase-sensitive, acidic, extractable nuclear antigen and to nucleoli, and immunoprecipitated proteins of similar molecular masses in SDS-PAGE, it appears that the described mAb is a product of an immortalized autoantibody-producing B cell clone from the SLE patient's peripheral blood. This mAb probably defines a novel RNA-associated autoantigen residing predominantly in the nucleolus or, less likely, a variant of either RNA polymerase I or the ribosomal autoantigens (P proteins).
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