The major dermatomyositis-specific Mi-2 autoantigen is a presumed helicase involved in transcriptional activation.
OBJECTIVE
To characterize the complementary DNA (cDNA) and protein sequences of autoantigens recognized by anti-Mi-2 antibodies, using recombinant Mi-2 proteins for improved autoantibody detection.


METHODS
A cDNA expression library was immunoscreened, and cDNA isolation, alignment, and sequence analysis were performed. Northern blotting and in situ hybridization techniques were used. A recombinant protein (rMi-2) was synthesized. Immunoprecipitation of 35S-methionine-labeled HEp-2 cell proteins and immunoblotting of rMi-2 and natural nuclear proteins were performed. Immunofluorescence studies were done with anti-Mi-2 positive sera of dermatomyositis (DM) patients, and with human or rabbit antibodies specific for rMi-2. Antibody screening of systemic lupus erythematosus, rheumatoid arthritis, DM, and antinuclear antibody-positive human sera was performed using an rMi-2 protein enzyme-linked immunosorbent assay (ELISA).


RESULTS
A major antigen recognized by anti-Mi-2 positive sera of DM patients was found to constitute a 218-kd nuclear protein (218-kd Mi-2) encoded on chromosome 12 and to belong to the SNF2/RAD 54 helicase family. Human and rabbit antibodies that were affinity purified using the recombinant protein reacted with and precipitated a nuclear protein of similar size, which was also recognized by anti-Mi-2 sera. Anti-218-kd Mi-2 antibodies detected by rMi-2 protein ELISA seemed to be mainly restricted to sera from patients with DM.


CONCLUSIONS
The molecular characterization of the 218-kd Mi-2 antigen may contribute to our understanding of autoimmune phenomena in DM. The use of immunoreactive recombinant proteins allows structural and functional studies of the helicase and the development of sensitive and accurate antibody screening tests.
Seelig
H P
HP
Institute of Immunology and Molecular Genetics, Karlsruhe, Germany.
Moosbrugger
I
I
Ehrfeld
H
H
Fink
T
T
Renz
M
M
Genth
E
E
D000251:Adenosine Triphosphatases; D000595:Amino Acid Sequence; D000818:Animals; D000888:Antibodies, Anti-Idiotypic / Q000097:blood / Q000276:immunology; D000974:Antibodies, Antinuclear / Q000097:blood / Q000276:immunology; D000937:Antigen-Antibody Reactions; D001324:Autoantigens / Q000737:chemistry* / Q000235:genetics / Q000502:physiology; D001483:Base Sequence; D002467:Cell Nucleus / Q000276:immunology*; D011232:Chemical Precipitation; D004265:DNA Helicases / Q000502:physiology; D018076:DNA, Complementary / Q000737:chemistry*; D003882:Dermatomyositis / Q000276:immunology*; D006801:Humans; D056245:Mi-2 Nucleosome Remodeling and Deacetylase Complex; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D011817:Rabbits; D011994:Recombinant Proteins; D015533:Transcriptional Activation / Q000502:physiology
D000888:Antibodies, Anti-Idiotypic; D000974:Antibodies, Antinuclear; D001324:Autoantigens; C115323:CHD4 protein, human; D018076:DNA, Complementary; D011994:Recombinant Proteins; D056245:Mi-2 Nucleosome Remodeling and Deacetylase Complex; D000251:Adenosine Triphosphatases; D004265:DNA Helicases