Epitope mapping of the cloned human autoantigen, histidyl-tRNA synthetase. Analysis of the myositis-associated anti-Jo-1 autoimmune response

Abstract

Autoantibodies that bind aminoacyl-tRNA synthetases are strongly associated with the human inflammatory myopathies polymyositis and dermatomyositis, but their molecular origins and relationship to pathogenesis are not known. To address these issues, we wished to identify the autoantigenic epitopes which react with these autoantibodies and to this end, we previously isolated a full length cDNA clone encoding the target Ag recognized most frequently by myositis sera, histidyl-tRNA synthetase (HRS). In the present study, we have analyzed the HRS autoepitopes by two amino acid insertion linker mutagenesis of HRS proteins expressed in Cos 1 cells. A series of mutant HRS cDNA were constructed and the expressed proteins were tested for enzyme activity and for immune reactivity with a panel of sera with anti-Jo-1 antibodies. Immunoblotting and immunoprecipitation analyses revealed that anti-Jo-1 antibodies recognize multiple conformation-dependent and independent epitopes on HRS and that the autoepitopes vary among different myositis patients.