title: Anti-dsDNA antibodies cross-react with ribosomal P proteins expressed on the surface of glomerular mesangial cells to exert a cytostatic effect.
abstract: Affinity-purified human polyclonal anti-double-stranded DNA antibodies (anti-dsDNA) exerted a cytostatic effect towards human and rat glomerular mesangial cells (MC). In order to identify the cognate antigens for anti-dsDNA on the surface of MC, we used these autoantibodies to probe a human renal lambda gt11 cDNA expression library. Two cDNA clones encoding the cognate proteins for the autoantibodies were isolated. Sequencing analysis of the two cDNA showed that they had 98.6% homology with the gene of the P0 and 99.2% homology with the gene of the P1 human acidic ribosomal phosphoproteins (P protein). Two galactosidase fusion proteins (125,000 and 150,000 MW) derived from the two cDNA inserts expressed in lysogenic Escherichia coli Y1089 could react with the original screening antibodies in an immunoblotting analysis. After transformation and expression of the full-length P1 clone in prokaryotic cells, the purified P1 protein was able to react with anti-dsDNA. In a cross-inhibition experiment, the dsDNA binding activity of anti-dsDNA was inhibited by a synthetic polypeptide corresponding to the carboxyl-terminal 20 amino acids of P protein and purified P1 protein in a dose-dependent manner, but this was less potent than the inhibition caused by calf thymus dsDNA. By use of well-defined systemic lupus erythematosus (SLE) sera, we found only sera containing a high titre of anti-dsDNA activity (> 300 IU/ml) reacted with P1 of rat MC lysate. Furthermore, the 38,000 and 19,000 MW macromolecules were proved to be the cognate antigens for anti-dsDNA expression on the surface of the MC, by Western blot of the MC plasma membrane lysates. These results suggest that anti-dsDNA may cross-react with ribosomal P proteins expressed on the surface of the MC and exert cytostasis towards these cells.
authors.affiliation: Faculty of Medical Technology, Institute of Microbiology and Immunology, Veterans General Hospital-Taipei, National Yang-Ming University, Taiwan, China.
authors.forename: K H
authors.initials: KH
authors.lastname: Sun
authors.forename: W T
authors.initials: WT
authors.lastname: Liu
authors.forename: C Y
authors.initials: CY
authors.lastname: Tsai
authors.forename: S J
authors.initials: SJ
authors.lastname: Tang
authors.forename: S H
authors.initials: SH
authors.lastname: Han
authors.forename: C L
authors.initials: CL
authors.lastname: Yu
mesh_terms: D000818:Animals; D000974:Antibodies, Antinuclear / Q000378:metabolism*; D001483:Base Sequence; D015153:Blotting, Western; D002455:Cell Division; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D017931:DNA Primers / Q000235:genetics; D005920:Glomerular Mesangium / Q000378:metabolism*; D006801:Humans; D008181:Lupus Nephritis / Q000378:metabolism; D008969:Molecular Sequence Data; D010750:Phosphoproteins / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D012269:Ribosomal Proteins / Q000378:metabolism*
chemical_list: D000974:Antibodies, Antinuclear; D017931:DNA Primers; D010750:Phosphoproteins; D012269:Ribosomal Proteins; C035690:ribosomal phosphoprotein P1; C054675:ribosomal protein P0