CDNA CLONING AND CHARACTERIZATION OF A NOVEL NUCLEOLAR PROTEIN

In an initial study of anti-nuclear antibodies in the chronic inflamma tory bladder disease interstitial cystitis, we reported that 7% of int erstitial cystitis patients studied had autoantibodies to the nucleolu s. We now report that, using an autoimmune serum from a patient with i nterstitial cystitis we have identified and partially characterized a novel protein with an M(r), of similar to 55 kDa (hereafter referred t o as No55) localized to the granular component of the nucleolus. No55 was initially characterized by diffuse nucleolar immunofluorescence st aining in interphase cells and by Western blotting as a 55-kDa doublet on whole-cell extracts. During mitosis, No55 was associated with chro mosomes and appeared in prenucleolar bodies during telophase, but it d id not colocalize with p80-coilin in coiled bodies. Immunoelectron mic roscopy revealed that No55 was localized uniformly throughout the gran ular component of the nucleolus compared with a more peripheral locali zation of nucleolar granular component protein B23. On segregation of the nucleolus with actinomycin D, No55 remained with the granular comp onent of the segregated nucleolus, whereas protein B23 was found predo minantly in the nucleoplasm. Finally, a cDNA expression library was sc reened with the human autoantibody against No55, and a 2.4-kb insert w as isolated, subcloned to homogeneity, and then sequenced. Analysis of this sequence showed an open reading frame of similar to 1.3 kb codin g for 437 amino acids with a predicted molecular weight of 50 kDa. A s earch of the gene sequence database indicated homology with SC65, a ra t synaptonemal complex protein. Therefore, on the basis of molecular w eight, nucleolar sublocalization, response to actinomycin D, and cDNA sequence determination, No55 is a novel protein of the interphase nucl eolus.