IgE Occupancy and Antigen Valency Cooperate to Control FcεRI Aggregation Geometry and Signaling Efficiency

The crosslinking of IgE-bound FcεRI by multivalent allergens initiates mast cell and basophil signaling underlying Type 1 allergy. Yet, how allergen properties and IgE occupancy impact receptor aggregation and downstream signaling remains unclear. We used Phl p 1-specific IgE and recombinant fusion proteins presenting a Phl p 1-derived peptide in defined valencies and positions (MB1N, MB1N1C, MB2N, and MB4N) to probe antigen-dependent signaling. Tetravalent MB4N evoked stronger degranulation and Ca2+ response than bivalent antigens MB1N1C and MB2N. MB4N was also capable of signaling at low IgE occupancy and in Lyn-deficient cells. Monte Carlo simulations predicted that MB4N forms larger, complex receptor aggregates, while MB1N1C and MB2N produce linear chains. Consistently, the addition of MB4N showed larger aggregates by electron microscopy and slower mobility by single-particle tracking, compared to bivalent antigens. Thus, allergen valency and epitope spatial arrangement dictate FcεRI aggregate organization and subsequent effector cell activation.