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Cytoskeletal Regulation of IgE-Mediated Degranulation in Mast Cells

J.R. Apgar, L.G. Frigeri, T. Kawakami*

*La Jolla Institute of Allergy and Immunology, La Jolla, CA

The goal of our research is to determine the mechanisms by which actin microfilaments regulate the signaling pathways involved in degranulation in mast cells. The relationship between microfilaments and these signaling pathways is complex. Actin filaments can both upregulate and downregulate the response, and different sets of filaments are probably involved. Mast cells express high-affinity IgE receptors on the cell surface, which bind IgE in a 1:1 ratio. In this state, the cells are sensitized but inactive. Binding of multivalent antigen to the IgE molecule causes cross-linking of the receptors, the crucial step leading to activation of the cells. Cross-linking of the receptors leads to a signaling cascade that results in degranulation and the release of preformed mediators such as histamine and in the de novo production of eicosanoids and cytokines. These mediators are responsible for a variety of allergic inflammatory diseases.

Rat basophilic leukemia cells, which are of mucosal mast cell origin, are a major model for studying IgE-mediated allergic reactions. These cells adhere and spread on extracellular matrix proteins such as fibronectin. Spreading of the cells depends on actin polymerization and the rearrangement of microfilaments. Adhesion and spreading do not cause any degranulation on their own, but they do prime the cells, because antigen-stimulated spread cells degranulate to a much greater extent than do activated cells in suspension. Studies have indicated that spreading of the cells, not simply adhesion, is important in the increased activation of the cells. We are investigating the role of the tyrosine kinases Lyn, Syk, and Btk in mediating this increased degranulation. Of particular interest is Btk, which contains a plecstrin homology domain that allows it to bind actin filaments.

Cross-linking of the receptors activates several signaling pathways, an event that leads to degranulation and further polymerization of actin. Inhibition of actin polymerization leads to increased degranulation in antigen-activated cells. In addition, increases occur in the activity of phospholipases and of Lyn and Syk. These 2 tyrosine kinases are the first signaling enzymes to be activated, and they are associated with the cross-linked receptor. This finding suggests that these newly formed microfilaments may be involved in downregulating the responses by uncoupling the cross-linked receptors from Lyn. Thus, in a negative feedback loop, these microfilaments downregulate the same pathways that signaled for the creation of the filaments.