Elevated N-Linked Glycosylation of IgG V Regions in Myasthenia Gravis Disease Subtypes

Abstract

Elevated N-linked glycosylation of IgG V regions (IgG-V^N-Glyc) is an emerging molecular phenotype associated with autoimmune disorders. To test the broader specificity of elevated IgG-V^N-Glyc, we studied patients with distinct subtypes of myasthenia gravis (MG), a B cell-mediated autoimmune disease. Our experimental design focused on examining the B cell repertoire and total IgG. It specifically included adaptive immune receptor repertoire sequencing to quantify and characterize N-linked glycosylation sites in the circulating BCR repertoire, proteomics to examine glycosylation patterns of the total circulating IgG, and an exploration of human-derived recombinant autoantibodies, which were studied with mass spectrometry and Ag binding assays to respectively confirm occupation of glycosylation sites and determine whether they alter binding. We found that the frequency of IgG-V^N-Glyc motifs was increased in the total BCR repertoire of patients with MG when compared with healthy donors. The elevated frequency was attributed to both biased V gene segment usage and somatic hypermutation. IgG-V^N-Glyc could be observed in the total circulating IgG in a subset of patients with MG. Autoantigen binding, by four patient-derived MG autoantigen-specific mAbs with experimentally confirmed presence of IgG-V^N-Glyc, was not altered by the glycosylation. Our findings extend prior work on patterns of Ig V region N-linked glycosylation in autoimmunity to MG subtypes.