The assembly of cells into tissues depends upon formation of intercellular junctions. In adherens junctions, the catenins link transmembrane cadherin cell adhesion molecules to the actin-based cytoskeleton, and in desmosomes an analogous protein assembly links cadherins to intermediate filaments. Adherens junction assembly is an essential step in the development of cell and tissue structure, and loss of these junctions is a hallmark of metastasizing cancer cells. Likewise, cell-cell contacts liked to intermediate filaments are essential for the mechanical strength and integrity of tissues such as the skin and heart, and defects in these assemblies are responsible for a number of severe skin blistering diseases. The adherens junction protein 2-catenin also serves as a transcriptional coactivator in the Wnt signaling pathway that controls cell fate determination and in the normal renewal of tissues in the adult. In many cancers, stabilization of 2-catenin due to mutations in Wnt pathway components results in inappropriate activation of Wnt target genes. The goal of this proposal is to achieve a mechanistic understanding of cell adhesion assemblies and of 2- catenin in Wnt signaling by using purified components to reconstitute key parts of these molecular complexes. Biochemical and biophysical methods will be used to determine the structures and affinities of these interactions. 1. The assembly and structure of actin-based cell-cell contacts will be investigated in epithelia and neurons by examining the similarities and differences between the epithelial and neuronal versions of 1-catenin, in particular its 2-catenin and actin-binding properties. We will also study the interactions of EPLIN with 1-E- catenin and actin to assess its potential role in linking the cadherin-catenin complex to actin. 2. The evolutionary origins of cell adhesion assemblies will be studied by examining homologs of 1- and 2- catenin in the slime mold Dictyostelium discoideum. 3. Structures of desmosomal cadherins bound to plakoglobin will be determined, as will the N- and C-terminal portions of desmoplakin, which links the cadherin-plakoglobin complex to intermediate filaments. 4. The mechanisms by which phosphorylation and ubiquitination required for 2-catenin destruction are controlled by Wnt signaling will be studied, in particular how phosphorylation of the Wnt receptor Lrp6 inhibits phosphorylation of 2-catenin, and how the phosphorylation complex is linked to the ubiquitination machinery. 5. The molecular mechanism of how Tcf/Lef-family transcription factors control Wnt target signaling will be investigated by determining structures of Lef-1 bound to a transcriptional repressor and to a 2-catenin-activator complex.
During development, cells differentiate and form specific contacts with other cells in order to create tissues. Defects in this process give rise to developmental abnormalities, and loss of cell-cell contacts occurs in cancer metastasis, when cancer cells escape from a solid tissue, and in other diseases. This basic research project seeks to understand the molecular assemblies responsible for these processes.
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