\beta-catenin (BC) is a bifunctional protein that controls two cellular activities altered in colon tumorigenesis. BC is required for the establishment of adherens junctions and it is aiso a transcriptional cofactor. E-cadherin-unbound BC can translocate to the nucleus and associate to hTcf-4 transcriptional factor; this BC activity depends on the cytosolic levels of this protein. Degradation of BC is mediated by APC, a gen frequently mutated in colon cancer. This project consists in characterising the structure of the association between BC (throughout the domain denominated "armadillo") and hTcf-4, and to determine the requirements that distinguish this association from those established by BC with E-cadherin or APC, that bind to the same arm domain. The domains of interaction of these proteins will be purified, and the minimal zone of interaction will be delimited using binding assays. In parallel, the structure of the BC "arm" domain will be compared to "arm" repeats present in other proteins, as APC or p120ctn that do not bind hTcf-4. We will design a model compatible with all the structural requirements and that explains the specificity determined with the functional binding assays. The model will be refined by the study of mutants in aminoadds, presumed key in the structure. Once the model has been validated, we will design peptides that inhibit hTcf-4/BC binding, without affecting the interaction of BC with APC or E-cadherin. The specificity of these inhibitors will be analysed by functional binding assays. Optimal inhibitors will be analyzed\i in vivo\i0 determining their ability to block: a) BC/hTcf-4 transcriptional activity in intestinal cell lines; and b) the growth of tumors spontaneously generated by \i min\i0 mice. The structural characterization of the binding hTcf-4/BC and the obtention of specific inhibitors will be especially useful for designing new anticancer drugs for patients not only with colon tumors but also with other tumors
|Effective start/end date||14/11/00 → 14/11/03|
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