High-affinity allergen-specific human antibodies cloned from single IgE B cell transcriptomes (1)
The recognition of allergenic food proteins by IgE antibodies can lead to symptoms ranging from urticaria to potentially fatal anaphylaxis.
A plate-based single-cell RNA sequencing (scRNA-seq) on B cells isolated from peripheral blood of six food-allergic individuals was performed. The isotype identity of each B cell was determined post hoc using the bioinformatic assembly of its heavy chain sequence from scRNA-seq reads. In total, 973 B cells were analyzed, of which 89 were IgE.
Principal components analysis of normalized gene expression separated B cells into two distinct clusters identifiable as plasmablasts (PBs) and naïve/memory B cells. Circulating IgE B cells overwhelmingly belonged to the PB subset. The number of circulating IgE B cells for each individual correlated with total plasma IgE levels. A similar phenomenon has been noted in atopic individuals and individuals with hyper-IgE syndrome.
Antibody heavy chain variable region (VH) gene usage as well as mutation frequency varied widely. A host of major histocompatibility complex (MHC) genes were robustly up-regulated in IgE PBs relative to PBs of other isotypes.
Elements of classical germinal center phenomena such as somatic hypermutation, class switching, and fate determination were observed. Three were IgE PBs from individual PA12 and three were IgE PBs from individual PA13.
The antibodies produced by these six cells were highly similar in VH and VL sequences and all used the IGHV3-30*18 and IGHJ6*02 VH genes as well as the IGKV3-20*01 and IGKJ2*01 VL genes. All six antibodies were cross-reactive: They bound strongly to Ara h 2, moderately to Ara h 3, and very weakly to Ara h 1. These antibodies have high affinity; dissociation constants determined by biolayer interferometry for Ara h 2 and Ara h 3 were as low as picomolar and subnanomolar, respectively.
