Project Summary Immunoglobulin E (IgE) binds with high affinity to its receptor, FCeR1, found on the surface of basophils and mast cells. When an antigen binds to receptor bound IgE, it can lead to IgE receptor cross-linking and activation of basophils and mast cells. In this case, the antigen is subcategorized as an allergen. Thus, IgE is a pivotal molecule in the initiation of allergic reaction in a wide range of tissue. The IgE-dependent activation of basophils leads to the release of granular mediators and, also, to the production of Th2 inflammatory cytokines, IL4/5/13. Recent work has revealed a novel, allergen independent mechanism by which IgE, bound to its receptor, can lead to basophil activation resulting in greater IL4/13 production compared to standard IgE- dependent activation. The mechanism of this allergen-independent activation involves the sugar-binding protein Galectin-3 (Gal-3), which has been shown to bind directly to IgE. The data reveal that when a Gal-3 bearing cell physically comes into contact with a basophil, it leads to strong basophil activation. However, the parameters of this interaction remain poorly defined. I hypothesize that Gal-3 interacts with IgE independently of allergen specificity due to IgE’s variable glycosylation patterns, and that such glycosylation patterns may differ between allergic and non-allergic subjects. IgE with more N-glycan sugars will interact better with Gal-3 while IgE with more sialic acid will not interact as well. My first aim is to compare Gal-3 with standard IgE- dependent activation of basophils sensitized by serum from either allergic or non-allergic patients. My second aim examines the hypothesis that the affinity of Gal-3 for IgE is based on the presence of N-glycan structures and that this binding can be masked by terminal sialic acid residues. I will treat both whole cells and purified IgE with glycan modifying enzymes and quantify the Gal-3-induced cytokine production in basophils. My preliminary data shows that neuraminidase treatment, to remove terminal sialic acid residues, of whole basophils primed with IL-3 and bearing IgE augments Th2 cytokine production when placed in co-culture with Gal-3 bearing cells. The results of the study are expected to provide insight into a novel mechanism of allergen independent IgE activation and should prompt investigation to correlate it with clinical relevance.