Abstract
SS-A/Ro is a nucleocytoplasmic ribonucleoprotein (RNP) particle that is a common target of autoimmune response in Sjögren's syndrome (SS) and systemic lupus erythematosus (SLE). Previously, SS-A/Ro has been shown to be composed of at least two polypeptide antigens of 60 and 52 kD noncovalently associated with a set of small RNAs, designated Y1-Y5. A serum from an SS patient was selected to screen a lambda gt11 cDNA library constructed from human T cell lymphoblastic leukemia (MOLT-4) mRNA. An immunoreactive clone was isolated that possessed a 1.8-kb cDNA insert. In vitro transcription and translation of the cDNA resulted in the synthesis of a 57.5-kD polypeptide which was specifically immunoprecipitated by SS-A/Ro antisera. The identity of the cDNA encoded protein as the 60-kD SS-A/Ro antigen was established by proteolytic peptide mapping of the cDNA-encoded protein and the 60-kD HeLa cell antigen. The sequence of the cDNA shows that the 60-kD SS-A/Ro protein possesses both RNA binding protein consensus sequences and a single zinc-finger motif. Recombinant SS-A/Ro antigen produced in bacteria proved to be a sensitive and specific reagent for detection of anti-SS-A/Ro antibodies in patient sera. The availability of the 60-kD SS-A/Ro cDNA will enable detailed analysis of the molecular structure and function of the SS-A/Ro RNP particle and its role in autoimmune pathology.
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Selected References
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- Bachmann M., Mayet W. J., Schröder H. C., Pfeifer K., Meyer zum Büschenfelde K. H., Müller W. E. Identification of the Ro and La antigens in the endoribonuclease VII--ribonucleoprotein complex. Biochem J. 1987 Apr 1;243(1):189–194. doi: 10.1042/bj2430189. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Ben-Chetrit E., Chan E. K., Sullivan K. F., Tan E. M. A 52-kD protein is a novel component of the SS-A/Ro antigenic particle. J Exp Med. 1988 May 1;167(5):1560–1571. doi: 10.1084/jem.167.5.1560. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Chambers J. C., Kenan D., Martin B. J., Keene J. D. Genomic structure and amino acid sequence domains of the human La autoantigen. J Biol Chem. 1988 Dec 5;263(34):18043–18051. [PubMed] [Google Scholar]
- Chan E. K., Francoeur A. M., Tan E. M. Epitopes, structural domains, and asymmetry of amino acid residues in SS-B/La nuclear protein. J Immunol. 1986 May 15;136(10):3744–3749. [PubMed] [Google Scholar]
- Cleveland D. W., Fischer S. G., Kirschner M. W., Laemmli U. K. Peptide mapping by limited proteolysis in sodium dodecyl sulfate and analysis by gel electrophoresis. J Biol Chem. 1977 Feb 10;252(3):1102–1106. [PubMed] [Google Scholar]
- Dreyfuss G., Swanson M. S., Piñol-Roma S. Heterogeneous nuclear ribonucleoprotein particles and the pathway of mRNA formation. Trends Biochem Sci. 1988 Mar;13(3):86–91. doi: 10.1016/0968-0004(88)90046-1. [DOI] [PubMed] [Google Scholar]
- Earnshaw W. C., Sullivan K. F., Machlin P. S., Cooke C. A., Kaiser D. A., Pollard T. D., Rothfield N. F., Cleveland D. W. Molecular cloning of cDNA for CENP-B, the major human centromere autoantigen. J Cell Biol. 1987 Apr;104(4):817–829. doi: 10.1083/jcb.104.4.817. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Elkon K. B., Jankowski P. W. Fine specificities of autoantibodies directed against the Ro, La, Sm, RNP, and Jo-1 proteins defined by two-dimensional gel electrophoresis and immunoblotting. J Immunol. 1985 Jun;134(6):3819–3824. [PubMed] [Google Scholar]
- Evans R. M., Hollenberg S. M. Zinc fingers: gilt by association. Cell. 1988 Jan 15;52(1):1–3. doi: 10.1016/0092-8674(88)90522-3. [DOI] [PubMed] [Google Scholar]
- Kato N., Hoshino H., Harada F. Nucleotide sequence of 4.5S RNA (C8 or hY5) from HeLa cells. Biochem Biophys Res Commun. 1982 Sep 16;108(1):363–370. doi: 10.1016/0006-291x(82)91875-7. [DOI] [PubMed] [Google Scholar]
- Konarska M. M., Grabowski P. J., Padgett R. A., Sharp P. A. Characterization of the branch site in lariat RNAs produced by splicing of mRNA precursors. Nature. 1985 Feb 14;313(6003):552–557. doi: 10.1038/313552a0. [DOI] [PubMed] [Google Scholar]
- Kozak M. An analysis of 5'-noncoding sequences from 699 vertebrate messenger RNAs. Nucleic Acids Res. 1987 Oct 26;15(20):8125–8148. doi: 10.1093/nar/15.20.8125. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Leff S. E., Rosenfeld M. G., Evans R. M. Complex transcriptional units: diversity in gene expression by alternative RNA processing. Annu Rev Biochem. 1986;55:1091–1117. doi: 10.1146/annurev.bi.55.070186.005303. [DOI] [PubMed] [Google Scholar]
- Lieu T. S., Newkirk M. M., Capra J. D., Sontheimer R. D. Molecular characterization of human Ro/SS-A antigen. Amino terminal sequence of the protein moiety of human Ro/SS-A antigen and immunological activity of a corresponding synthetic peptide. J Clin Invest. 1988 Jul;82(1):96–101. doi: 10.1172/JCI113607. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Miller J., McLachlan A. D., Klug A. Repetitive zinc-binding domains in the protein transcription factor IIIA from Xenopus oocytes. EMBO J. 1985 Jun;4(6):1609–1614. doi: 10.1002/j.1460-2075.1985.tb03825.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Olmsted J. B. Affinity purification of antibodies from diazotized paper blots of heterogeneous protein samples. J Biol Chem. 1981 Dec 10;256(23):11955–11957. [PubMed] [Google Scholar]
- Proudfoot N. J., Brownlee G. G. 3' non-coding region sequences in eukaryotic messenger RNA. Nature. 1976 Sep 16;263(5574):211–214. doi: 10.1038/263211a0. [DOI] [PubMed] [Google Scholar]
- Sanger F. Determination of nucleotide sequences in DNA. Science. 1981 Dec 11;214(4526):1205–1210. doi: 10.1126/science.7302589. [DOI] [PubMed] [Google Scholar]
- Shine J., Dalgarno L. The 3'-terminal sequence of Escherichia coli 16S ribosomal RNA: complementarity to nonsense triplets and ribosome binding sites. Proc Natl Acad Sci U S A. 1974 Apr;71(4):1342–1346. doi: 10.1073/pnas.71.4.1342. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Sillekens P. T., Habets W. J., Beijer R. P., van Venrooij W. J. cDNA cloning of the human U1 snRNA-associated A protein: extensive homology between U1 and U2 snRNP-specific proteins. EMBO J. 1987 Dec 1;6(12):3841–3848. doi: 10.1002/j.1460-2075.1987.tb02721.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Spindler K. R., Rosser D. S., Berk A. J. Analysis of adenovirus transforming proteins from early regions 1A and 1B with antisera to inducible fusion antigens produced in Escherichia coli. J Virol. 1984 Jan;49(1):132–141. doi: 10.1128/jvi.49.1.132-141.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Sturgess A. D., Peterson M. G., McNeilage L. J., Whittingham S., Coppel R. L. Characteristics and epitope mapping of a cloned human autoantigen La. J Immunol. 1988 May 1;140(9):3212–3218. [PubMed] [Google Scholar]
- Sullivan K. F., Machlin P. S., Ratrie H., 3rd, Cleveland D. W. Sequence and expression of the chicken beta 3 tubulin gene. A vertebrate testis beta-tubulin isotype. J Biol Chem. 1986 Oct 5;261(28):13317–13322. [PubMed] [Google Scholar]
- Tan E. M. Autoantibodies to nuclear antigens (ANA): their immunobiology and medicine. Adv Immunol. 1982;33:167–240. doi: 10.1016/s0065-2776(08)60836-6. [DOI] [PubMed] [Google Scholar]
- Theissen H., Etzerodt M., Reuter R., Schneider C., Lottspeich F., Argos P., Lührmann R., Philipson L. Cloning of the human cDNA for the U1 RNA-associated 70K protein. EMBO J. 1986 Dec 1;5(12):3209–3217. doi: 10.1002/j.1460-2075.1986.tb04631.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Theofilopoulos A. N., Dixon F. J. The biology and detection of immune complexes. Adv Immunol. 1979;28:89–220. doi: 10.1016/s0065-2776(08)60800-7. [DOI] [PubMed] [Google Scholar]
- Towbin H., Staehelin T., Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. doi: 10.1073/pnas.76.9.4350. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Wolin S. L., Steitz J. A. The Ro small cytoplasmic ribonucleoproteins: identification of the antigenic protein and its binding site on the Ro RNAs. Proc Natl Acad Sci U S A. 1984 Apr;81(7):1996–2000. doi: 10.1073/pnas.81.7.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Young R. A., Davis R. W. Yeast RNA polymerase II genes: isolation with antibody probes. Science. 1983 Nov 18;222(4625):778–782. doi: 10.1126/science.6356359. [DOI] [PubMed] [Google Scholar]