The major aims of this proposal are to identify signaling mechanisms initiated by FGFs that underlie craniofacial morphogenesis. Loss of the FGFR1 receptor together with FGFR2 in neural crest cells leads to agenesis of multiple components of the midface and mandible, whereas hypomorphic mutations in Fgfr1/2 result in cleft palate. Elsewhere, FGFR2 rather than FGFR1 has a predominant role in salivary gland epithelia, and its activity is differentially regulated by various ligands. Engagement of the FGF signaling cascade leads to dimerization of the FGFRs, binding of multiple intracellular effectors and activation of cellular responses that converge on ERK1/2 and several other pathways. This application proposes: 1. To investigate how FGF signaling coordinates midface development. Combined loss of both Fgfr1 and Fgfr2 in neural crest cells leads to facial clefting that extends through the midline, agenesis of the midface, mandibular hypoplasia, and significant cell death in the lateral nasal and maxillary processes. To determine the origin of these midface defects, Fgfr1/2 mandibular and/or lateral nasal /maxillary process conditional mutants will be generated to investigate how loss of Fgfr1/2 in the mandible or lateral structures contributes to facial clefting. Furthermore, neural crest Fgfr1/2 mutants will be crossed to Bim mutants to disassociate alterations in patterning from BCL-2 family regulated cell death. 2. To identify signaling mechanisms promoted by Fgfr1 and Fgfr2 in craniofacial development. To identify signaling pathways that remain active in a previously generated Fgfr1 and Fgfr2 allelic series of point mutations that prevent the binding of single or multiple effector proteins and the initiation of specific signaling pathways, a proteomic screen will be performed in mouse embryonic palatal mesenchyme cells using endogenous epitope tagged FGFR1 and FGFR2 receptors. In a complementary approach, mice in which additional candidate tyrosine phosphorylation sites are disrupted on the receptors will be generated. 3. To characterize signaling pathways specified by ligand identity. An emerging theme in FGF signaling is that cellular responses in multiple physiological contexts can be encoded in the identity of the ligand and are not only specified at the level of the receptor. Signaling responses that are differentially encoded by FGF7 and FGF10 in submandibular salivary gland branching morphogenesis will be investigated. Critical downstream FGF pathways in this response will be further identified by transcriptional profiling following stimulation with each ligand, and morphogenetic responses that are sensitive to ERK1/2 and PI3K signaling and feedback inhibition pathways will be investigated. These proposed studies explore novel territories in the area of growth factor signaling in craniofacial biology and open new directions for the prevention of craniofacial birth defects.
Craniofacial developmental diseases including cleft lip or palate, and disorders of the salivary glands, are prevalent birth defects in the human population worldwide. The major goal of this proposal is to characterize signaling mechanisms controlled by FGFs that underlie normal craniofacial development, by investigating the functions of the FGFR1 and FGFR2 receptors, their mode of signaling, and how different FGF ligands can signal through the same receptors yet engage differential outcomes. These studies will identify mechanisms underlying craniofacial development and establish paradigms for the prevention of birth defects.
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