OBJECTIVE: To investigate which members of the heterogeneous nuclear RNP (hnRNP) family are targeted by autoantibodies from patients with systemic rheumatic diseases. METHODS: Using a semipurified preparation of natural hnRNP proteins, 365 sera from patients with rheumatic diseases and control subjects were screened by immunoblotting for the presence of autoantibodies. Bacterially expressed recombinant hnRNP D (AUF1) proteins were used for confirming the data obtained. Binding of RNA and autoantibody to AUF1 was investigated by gel retardation assays. Expression of AUF1 in cultivated cells and synovial tissue was analyzed by indirect immunofluorescence and immunohistochemistry. RESULTS: Autoantibodies to AUF1 proteins were detected in 33% of patients with systemic lupus erythematosus, 20% of patients with rheumatoid arthritis, 17% of patients with mixed connective tissue disease, and <10% of patients with other rheumatic disorders. Epitope mapping studies showed the autoantibodies to be directed to conformational epitopes in the N-terminal RNA-binding part of AUF1. However, autoantibody binding did not interfere with RNA binding as assessed by gel-shift assays. Immunohistochemical studies revealed AUF1 to be expressed in the cytoplasm of RA synovial tissue as compared with nuclear staining in osteoarthritis and normal synovium, particularly in macrophages of the lining layer and in fibroblasts of the sublining areas. CONCLUSION: These data identify AUF1 proteins as novel autoantigens in SLE and related autoimmune disorders. Because AUF1 proteins are major components of messenger RNA stability complexes, our findings suggest that these complexes form a novel macromolecular target structure for autoantibodies in rheumatic autoimmune diseases.
OBJECTIVE: To investigate which members of the heterogeneous nuclear RNP (hnRNP) family are targeted by autoantibodies from patients with systemic rheumatic diseases. METHODS: Using a semipurified preparation of natural hnRNP proteins, 365 sera from patients with rheumatic diseases and control subjects were screened by immunoblotting for the presence of autoantibodies. Bacterially expressed recombinant hnRNP D (AUF1) proteins were used for confirming the data obtained. Binding of RNA and autoantibody to AUF1 was investigated by gel retardation assays. Expression of AUF1 in cultivated cells and synovial tissue was analyzed by indirect immunofluorescence and immunohistochemistry. RESULTS: Autoantibodies to AUF1 proteins were detected in 33% of patients with systemic lupus erythematosus, 20% of patients with rheumatoid arthritis, 17% of patients with mixed connective tissue disease, and <10% of patients with other rheumatic disorders. Epitope mapping studies showed the autoantibodies to be directed to conformational epitopes in the N-terminal RNA-binding part of AUF1. However, autoantibody binding did not interfere with RNA binding as assessed by gel-shift assays. Immunohistochemical studies revealed AUF1 to be expressed in the cytoplasm of RA synovial tissue as compared with nuclear staining in osteoarthritis and normal synovium, particularly in macrophages of the lining layer and in fibroblasts of the sublining areas. CONCLUSION: These data identify AUF1 proteins as novel autoantigens in SLE and related autoimmune disorders. Because AUF1 proteins are major components of messenger RNA stability complexes, our findings suggest that these complexes form a novel macromolecular target structure for autoantibodies in rheumatic autoimmune diseases.
Authors: Katy A Lloyd; Gustaf Wigerblad; Peter Sahlström; Manasa G Garimella; Karine Chemin; Johanna Steen; Philip J Titcombe; Bianka Marklein; Diana Zhou; Ragnhild Stålesen; Elena Ossipova; Christina Lundqvist; Olov Ekwall; Johan Rönnelid; Daniel L Mueller; Mikael C I Karlsson; Mariana J Kaplan; Karl Skriner; Lars Klareskog; Fredrik Wermeling; Vivianne Malmström; Caroline Grönwall Journal: Front Immunol Date: 2019-01-04 Impact factor: 7.561