GENETIC AND BIOCHEMICAL DISSECTION OF PROTEIN LINKAGES IN THE CADHERIN-CATENIN COMPLEX

The cadherin-catenin complex is important for mediating homotypic, cal cium-dependent cell-cell interactions in diverse tissue types. Althoug h proteins of this complex have been identified, little is known about their interactions. Using a genetic assay in yeast and an in vitro pr otein-binding assay, we demonstrate that beta-catenin is the linker pr otein between E-cadherin and alpha-catenin and that E-cadherin does no t bind directly to alpha-catenin. We show that a 25-amino acid sequenc e in the cytoplasmic domain of E-cadherin and the amino-terminal domai n of alpha-catenin are independent binding sites for beta-catenin. In addition to beta-catenin and plakoglobin, another member of the armadi llo family, p120 binds to E-cadherin. However, unlike beta-catenin, p1 20 does not bind alpha-catenin in vitro, although a complex of p120 an d endogenous alpha-catenin could be immunoprecipitated from cell extra cts. In vitro protein-binding assays using recombinant E-cadherin cyto plasmic domain and alpha-catenin revealed two catenin pools in cell ly sates: an approximate to 1000- to approximate to 2000-kDa complex boun d to E-cadherin and an approximate to 220-kDa pool that did not contai n E-cadherin. Only beta-catenin in the approximate to 220-kDa pool bou nd exogenous E cadherin. Delineation of these molecular linkages and t he demonstration of separate pools of catenins in different cell lines provide a foundation for examining regulatory mechanisms involved in the assembly and function of the cadherin-catenin complex.