Among the heterogeneous family of Fcgamma receptors, we have determined that only FcgammaRIIA is expressed on human platelets where it induces signal transduction leading to platelet activation. FcgammaRIIA is also expressed on macrophages which are important in the clearance of IgG-coated platelets and RBCs in immune thrombocytopenia and hemolytic anemia.Considerable data from us and others have helped elucidate the number of Fcgamma RIIA on platelets and macrophages, its affinity for IgC, its response to regulatory signals, and its role in the induction of serotonin release from platelets and in the mediation of a phagocytic signal in macrophages, megakaryocyte-like cell lines and transfected COS-1 cells. FcgammaRIIA is unusual among the human Fcgamma receptors in that it induces signal transduction in the absence of an associated subunit. Rather, signaling requires YXXL sequences within its unique cytoplsmic domain (CYT). Our goals in this research proposal are to delineate the structure/function relationships of FcgammaRIIA in signal transduction and to define the sequences responsible for the release of serotonin and for the phagocytosis of IgG coated cells. We will use two established model system: one in rat basophilic leukemia (RBL) cells and a second in COS1 cells. Specifically, we will explore the following: 1) The sequences of FcgammaRIIA necessary for signaling. We will define the contrivution of the inernal amino acids of the CYT conserved YXXL sequences and the unique intervening region which separates the two YXXLs. Our hypothesis is that the unique FcgammaRIIA CYT features are advantageious for signaling. We will also define FcgammaRIIA sequences necessary for its interaction with Syk, a tyrosine kinse required for the FcgammaRIIA phagocytic signal. Our hypothesis is that the distinct interaction with Syk sequences absent from ZAP-70, a Syk family kinase inefficient in phagocytic signaling 2) The requirements for FcgammaRIIA mediated serotonin release. RBL cells, which release serotonin in response to Fc receptor crosslinking, will be transfected with FcgammaRIIA to determine structure/function relationships relationships important for FcgammaRIIA induced serotonin release and to determine the role of Syk and PI-3 kinase. The hypothesis to be tested is that serotonin release medicated by Fcgamma RIIA has requirements distinct from those for FcgammaRIIA mediated phagocytosis. 3) Finally, we will examine in human monocytes and COS-1 cell transfectants the inhibition of FcgammaRIIA function by another FcgammaRIIA gene product, FcgammaRIIA. Our hypothesis is that FcgammaRIIB. Our hypothesis is that FcgammaRIIA regulates phagocytic signaling and does so by decreasing tyrosine phosphorylation of FcgammaRIIA and Syk.
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