Background Anti-DNA antibodies have been demonstrated to play an important role in the pathogenesis of nephritis in systemic lupus erythematosus. One mechanism by which anti-DNA antibodies may contribute to nephritis is by cross-reactivity with renal antigens. However, the identity of this renal antigen remains elusive.
Objectives To identify the target antigen bound by a pathogenic anti-DNA antibody in renal tissue.
Methods Western blotting and immunoprecipitations of total cell lysate from mesangial cell lines derived from the autoimmune MRL-lpr/lpr mouse (lpr-mc) and a non-autoimmune BALB/c mouse (mc), with R4A, a pathogenic murine IgG2b anti-DNA antibody, and with 95, a non-pathogenic antibody derived from R4A by site-directed mutagenesis. The immunoprecipitated protein was identified by matrix assisted laser desorption ionisation mass spectrometry (MALDI-MS) of tryptic digests.
Results The pathogenic anti-DNA antibody R4A, but not control isotype-matched antibodies, bound to and immunoprecipitated a 100 kD protein from mesangial cell lysates. Binding was more pronounced using lpr-mc rather than non-autoimmune mc lysate. Little effect on binding was seen after treatment of the cell lysate or R4A with DNAse, while proteinase K treatment abolished binding. Preliminary MALDI-MS analysis revealed that alpha-actinin, an actin bundling protein that plays a role in experimental nephritis models, is present in the protein band immunoprecipitated by R4A. R4A, but not 95, binds to immobilised alpha-actinin by ELISA; Furthermore, double stranded DNA significantly inhibited the binding of R4A to the 100 kD mesangial cell protein, as well as to purified alpha-actinin. Serum of MRL-lpr/lpr lupus mice with active disease contain high-titer antibodies to alpha-actinin.
Conclusion The nephritogenicity of some anti-DNA antibodies may be mediated via cross-reactivity with a glomerular antigen. Variations in the display of the cross-reactive antigen in the target organ between individuals may underlie differential susceptibility to anti-DNA antibody induced renal disease.
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