Structure-function relationships that govern FccRI signaling by allergens

Multivalent antigens trigger robust FcϵRI-dependent signaling responses in mast cells and basophils. Most prior studies have relied on chemically heterogenous artificial ligands, rendering it difficult to predict receptor cluster size and orientation and to translate the observations to clinical relevance. This work focuses on the use of structurally defined polyvalent antigens to crosslink IgE-FcϵRI, including a symmetrical trivalent ligand (DNP3-fibritin) and natural allergen (shrimp tropomyosin). In silico docking and computer simulation methods were used to build structural models of ligand-IgE-FcϵRI ectodomain complexes and to estimate aggregation properties, including distances between individual receptors after crosslinking. Signaling responses were studied in “humanized” RBL-2H3 cells and in human basophils from allergic subjects. High resolution imaging approaches were employed to provide insight into the kinetics of crosslinking and the nanoscale organization of FcϵRI signaling. The bell-shaped secretory responses are shown to be linked to the balance of positive (Syk-mediated) and negative (SHIP-mediated) signaling, as well as to differences in aggregate size, complexity and time course. Structurally defined antigens provide an improved basis for understanding FcϵRI signal initiation and termination. These studies lay the foundation for understanding the essential structural features of natural allergens, such as valency, epitope distance and affinity.