Secreted autoantibody repertoires in Sjögren's syndrome and systemic lupus erythematosus: A proteomic approach

• 2-DE/MS sequencing workflow was developed to characterise autoantibody proteomes.
• SS/SLE V-region autoantibody signatures reveal clonal restriction/public clonotypes.
• A combination of NGS/MS sequencing is required for complete autoantibody sequence.
• Targeted MRM should allow novel multiplex diagnostic assays for autoimmunity.

The structures of epitopes bound by autoantibodies against RNA–protein complexes have been well-defined over several decades, but little is known of the clonality, immunoglobulin (Ig) variable (V) gene usage and mutational status of the autoantibodies themselves at the level of the secreted (serum) proteome. A novel proteomic workflow is presented based on affinity purification of specific Igs from serum, high-resolution two-dimensional gel electrophoresis, and de novo and database-driven sequencing of V-region proteins by mass spectrometry. Analysis of anti-Ro52/Ro60/La proteomes in primary Sjögren's syndrome (SS) and anti-Sm and anti-ribosomal P proteomes in systemic lupus erythematosus (SLE) has revealed that these antibody responses are dominated by restricted sets of public (shared) clonotypes, consistent with common pathways of production across unrelated individuals. The discovery of shared sets of specific V-region peptides can be exploited for diagnostic biomarkers in targeted mass spectrometry platforms and for tracking and removal of pathogenic clones.