Regulation of Mast Cell Development and Function

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Mast cells (MCs) are major effector cells in IgE antibody-dependent allergic disorders, such as anaphylaxis, allergic rhinitis and atopic asthma, which account for a very large burden of illness and economic costs in the developed world. MCsare also critical for the optimal expression of certain innate immune responses. While much attention has been focused on the elements which positively regulate the IgE- and specific antigen (Ag)-dependent secretion of pro-inflammatory mediators and cytokines from MCs, the molecular mechanisms which can suppress the magnitude and/or duration of such responses have been less studied. We recently reported that RabGEFI (Rajb guanine nucleotide exchange factor 1.) can negatively regulate Ras-dependent signaling pathways, and the secretion of all three classes of mediators (pre-formed, lipid and cytokine), in MCs stimulated with IgE and specific Ag. More recently, we found that RabGEFI also importantly regulates responses in MCs which are elicited by the major survival, developmental and proliferation factor for MCs, SCF (stem cell factor, the c-Kit ligand). Notably, our Rabgefl^' mice exhibit severe inflammation of the skin associated with increased numbers of MCs, evidence of dermal MC degranulation (i.e., "activation"), and increased levels of histamine and IgE in the serum. The central questions which we now wish to address are: By what molecular mechanisms does RabGEFI influence MC development, activation and function, and to what extent might these actions of RabGEFI on MCs account for some of the dramatic phenotypic abnormalities observed in Rabgefl^'mice? In Aim 1. we will investigate how RabGEFI, and its individual functional domains, can negatively regulate mouse MC activation induced by signaling via FceRI or c-Kit (the SCF receptor) in vitro or in vivo, and will identify and characterize RabGEFI-interacting proteins and their downstream effectors in MCswhich have been activated via these receptors. In Aim 2. we will define the mechanisms by which RabGEFI can regulate the survival, development, phenotype & proliferation of MCs in vitro and in vivo. The in vivo studies will take advantage of our ability to transfer in wfro-derived MCs which lack, or express mutant forms of, RabGEFI into the tissues of c-kit mutant, Kitw/w~v or Kit"-3¿"'*" genetically MC-deficient mice (which express wild type RabGEFI). We thus can study the effects of RabGEFI on MCs in mice in which only the MCs lack, or express mutant forms of, RabGEFI. Elucidating how RabGEFI negatively regulates FceRI- or c-Kit-dependent signaling in MCs will increase our understanding of the regulation of MC activation and development, which is the long-term goal of this project. Such work also may help in the development of new therapeutic approaches for the alleviation of diseases, such as asthma and atopic dermatitis, which are associated with IgE-dependent MC activation and, in many patients, with increased numbers of MCs in the affected tissues.