title: Autoantibody to DNA excision repair enzyme hMYH in a patient with rheumatic disease.
abstract: We screened a human HepG2 cell cDNA expression library using serum from a patient with rheumatic disease. This serum had immunofluorescence reactivity to nuclei with a homogeneous staining pattern and to punctuate nuclear aggregates, chromosomal metaphase plate, midbody, and cytoplasmic bridge. YT1, the longest cDNA clone isolated, has sequence identity to hMYH, the human homologue of the Escherichia coli excision repair enzyme, DNA adenine glycosylase MutY. YT1 is a truncated cDNA of 1619 bp, encoding amino acids 22-535, and contains a full-length 3'-UTR sequence. We were unable to express a bacterial malE fusion protein incorporating amino acids 22 to 535 of hMYH. Consequently, we generated two additional malE fusion proteins of hMYH encoding amino acids 1-120 (pMAL-c2:hMYH(1-120)) and amino acids 121-535 (pMAL-c2:hMYH(121-535)). The patient serum immunoblotted only pMAL-c2:hMYH(1-120), suggesting that the autoepitope(s) is restricted to amino acids 22-120 of hMYH, and detected a protein of approximately 59-kDa in total HeLa and nuclear extracts consistent with reactivity to hMYH. Affinity-purified autoantibodies to pMAL-c2:hMYH(1-120) reacted by immunoblot to pMAL-c2:hMYH(1-120), with no reactivity to pMAL-c2:hMYH(121-535). By immunofluorescence, these antibodies displayed staining of nuclei. This is the first report of autoantibodies to hMYH in a patient with rheumatic disease. We were able to identify hMYH reactivity in relatively small cohorts of sera collected from rheumatoid factor-positive patients (6 of 18) and dsDNA-positive patients (1 of 18), with no reactivity detected in serum collected from 9 healthy subjects.
authors.affiliation: Department of Pathology and Immunology, Monash Medical School, Commercial Road, Prahran, Victoria 3181, Australia.
authors.forename: F P
authors.initials: FP
authors.lastname: Lai
authors.forename: Y
authors.initials: Y
authors.lastname: Tsukada
authors.forename: H
authors.initials: H
authors.lastname: Ichikawa
authors.forename: K
authors.initials: K
authors.lastname: Dunster
authors.forename: J W
authors.initials: JW
authors.lastname: Sentry
authors.forename: B H
authors.initials: BH
authors.lastname: Toh
mesh_terms: D001323:Autoantibodies / Q000097:blood*; D002460:Cell Line; D045647:DNA Glycosylases; D004260:DNA Repair; D018076:DNA, Complementary / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology; D005260:Female; D019084:Fluorescent Antibody Technique, Indirect; D015723:Gene Library; D006367:HeLa Cells; D006801:Humans; D008875:Middle Aged; D009699:N-Glycosyl Hydrolases / Q000235:genetics / Q000276:immunology*; D012216:Rheumatic Diseases / Q000201:enzymology / Q000235:genetics / Q000276:immunology*
chemical_list: D001323:Autoantibodies; D018076:DNA, Complementary; D045647:DNA Glycosylases; D009699:N-Glycosyl Hydrolases; C061738:mutY adenine glycosylase