The regulation of cell differentiation, proliferation and apoptosis is a fundamental requirement for any living multicellular organism. The fibroblast growth factor (FGF) family, in both mice and humans, consists of genes encoding 22 different ligands and four tyrosine kinase receptors (FGFR1-4). FGF stimulation has been shown to induce survival, proliferation and differentiation in cells of the ocular lens. Transgenic mice over-expressing several different FGF ligands undergo ectopic differentiation of the lens epithelium. Furthermore, conditional deletion of Fgfr genes leads to lens abnormalities depending on which receptor is deleted and what stage of lens development the deletion takes place. Deletion of Fgfr2 alone during the lens placode stage does not prevent lens fiber differentiation, but does cause a slight delay in lens cell cycle withdrawal and compromises lens cell survival. In contrast, on an Fgfr3 null background, lens fiber differentiation can be prevented by simultaneous deletion of Fgfr1 and Fgfr2 at the lens vesicle stage. Therefore, FGFs and FGFRs play a vital and fundamental role in lens biology and are absolutely required for lens fiber differentiation. The purpose of this application is to examine the relevant biochemical pathway, subsequent to Fgfr stimulation, that mediates FGF-induced responses in the lens. The hypothesis to be tested is that the major pathway by which Fgfrs mediate lens fiber differentiation is through the activation of the intermediate docking protein FRS2alpha. This hypothesis will be tested by (1) direct modification of the mouse Fgfr2 locus to create mutations designed to a) disrupt interaction with FRS2alpha, b) disrupt interaction with PLCgamma and c) create a chimeric receptor with an intracellular domain from TrkC (a receptor that also induces the activation of FRS2alpha) and testing to see if these different mutant Fgfr2 genes can or can not support a fiber differentiation response in the absence of functional Fgfr1 and Fgfr3 receptors;(2) making a conditional mutation in FRS2alpha and inducing lens-specific loss of this gene;(3) testing the ability of TrkC to induce a fiber differentiation response in lens epithelial cells in vivo. This application is relevant to public health because cataract remains the most frequent worldwide cause of blindness and FGF/FGFR signaling plays an essential role in lens development. FGF signaling also plays fundamental roles in many disease processes including cancer, but the molecular details of how FGFs elicit their effects are not fully resolved.
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