The development and homeostasis of solid tissues depends upon biochemical and mechanical signals that control cell fate and the resulting organization of cells in the tissue. The conserved protein ?-catenin is a key effector of signals from both the Wnt family of secreted growth factors that specify cell fate during embryogenesis and tissue renewal in the adult, and mechanical force transmitted through cell-cell junctions in multicellular tissues. We hypothesize that mechanical force transmitted through ?-catenin links Wnt signaling and cell-cell adhesion, and our overall goal is to understand the molecular mechanisms underlying these dual roles of ?-catenin. Our strategy is to use biochemical, structural and biophysical methods to address critical knowledge gaps in these areas. In the absence of Wnts, the ?-catenin is bound in a ?destruction complex? that includes the proteins Axin and Adenomatous Polyposis Coli (APC), and kinases that phosphorylate ?-catenin; phosphorylation leads to ubiquitylation and destruction of ?-catenin by the proteasome. Wnt binding to the receptors Frizzled (Fzd) and LRP5/6 enables Fzd to recruit the cytoplasmic protein Dishevelled (Dvl), which in turn binds to Axin and thereby recruits the destruction complex to the activated receptor complex. This leads to phosphorylation of the LRP5/6 intracellular domain, which inhibits ?-catenin destruction; the stabilized ?-catenin enters the nucleus and activates target genes. We will address critical mechanistic aspects of this pathway that are not understood: 1) how secreted ligands ?activate? the Fzd-Dvl interaction needed for ?-catenin stabilization through interaction with the extracellular cysteine-rich domain of Fzd and LRP5/6; 2) how activated Dvl recruits Axin to turn off ?-catenin destruction; 3) how the ?-catenin destruction complex forms and interacts with the ubiquitylation/proteosomal machinery; 4) the essential role of APC in ?-catenin destruction. Force transmission through cell-cell adherens junctions (AJ) requires a complex of E-cadherin, ?-catenin, and ?-catenin, which binds to actin filaments and forms a minimal force-sensing unit. Tension on cadherins can release ?-catenin and cause its translocation to the nucleus independent of, but synergized by, Wnt signaling. Understanding such tension-triggered release of ?-catenin requires understanding how force is transmitted through the AJ complex. ?-Catenin additionally has a central role in organizing epithelial tissues based on its interactions with vinculin, Epithelial Protein Lost in Neoplasm (EPLIN), the tight junction (TJ) protein Zonula Occludens (ZO)-1, and afadin, all of which bind actin and recruit other scaffolding and signaling proteins. We will study: 1) The force-dependent conformational landscape and force responsivness of ?-catenin alone and bound to its partners, including how ?-catenin modifies ?-catenin force responsiveness; 2) How ?E-catenin conformation and force transmission properties are affected by its binding to its other junctional partners.
The development and maintenance of solid tissues depends upon biochemical and mechanical signals that control cell fate and the resulting organization of the tissue, and defects in these processes occur in many cancers. The protein ?-catenin is central to how cells to respond to biochemical signals called Wnt proteins, and to mechanical forces transmitted between cells. This proposal aims to define molecular mechanisms underlying these dual roles of ?-catenin, information may enable the development of novel approaches to treat cancers.
