This application concerns the analysis of molecular mechanisms of Wnt signaling in embryonic cells of Xenopus laevis. Wnt signaling pathways play a major role in the control of cell proliferation, cell polarity and cell fate determination in multiple developmental processes. A commonly accepted mechanism of Wnt signaling involves target gene activation by a complex of a TCF family member with B-catenin. Whereas a lot of attention has been focused on the mechanisms of B-catenin stabilization, the regulation of TCF proteins that are key mediators of Wnt signaling in the nucleus remains understudied. Genetic analysis revealed both positive and negative roles for TCF proteins in Drosophila and C. elegans embryos, although the functions of the four vertebrate TCFs appear to be more specialized. The proposal focuses on the analysis of specific Wnt-dependent phosphorylation of TCF3, a transcriptional repressor, identified in preliminary studies. Experimental data suggest that a homeodomain-interacting protein kinase (HIPK) is responsible for this phosphorylation. The main hypothesis is that TCF3-mediated transcriptional repression is relieved in the course of Wnt signaling. This hypothesis will be assessed using Xenopus embryos, an experimental system, which allows combination of biochemical and cell biological approaches with rapid functional analysis in vivo. New Wnt transcriptional targets that are negatively regulated by TCF3 in vivo will be identified and the significance of TCF3 phosphorylation for gene target activation will be evaluated. The regulation of HIPK activity, localization by Wnt signaling and its role in Wnt-mediated TCF3 phosphorylation will be assessed. These studies will help characterize the diversity of Wnt signaling mechanisms and will contribute to understanding signaling networks leading to cell fate specification during early development. Since Wnt pathways are activated in many human cancers, including colon carcinomas, melanomas and skin tumors, the proposed experiments should reveal novel molecular mechanisms, which operate during embryonic development and may be misregulated in cancer.
Wnt signaling pathways play a major role in the control of cell proliferation, cell polarity and cell fate determination in multiple developmental processes. This application concerns the analysis of specific phosphorylation of TCF proteins, key transcriptional mediators of Wnt pathways, will help characterize the diversity of Wnt signaling mechanisms and networks leading to cell fate specification during early development. Since Wnt pathways are activated in many human cancers, including colon carcinomas, melanomas and skin tumors, the proposed experiments are directly relevant to human disease.
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