Genetic and Biochemical Dissection of Protein Linkages in the Cadherin- Catenin Complex

• Published in: Proceedings of the National Academy of Sciences - PNAS 1995-05, Vol.92 (11), p.5067-5071
• Main Authors: Jou, Tzuu-Shuh, Stewart, Daniel B., Stappert, Jorg, Nelson, W. James, Marrs, James A.
• Format: Article
• Language: English
• Subjects: Adenocarcinoma; alpha Catenin; Amino Acid Sequence; Amino acids; Animals; Antibodies; Base Sequence; beta Catenin; Binding Sites; Cadherins; Cadherins - genetics; Cadherins - isolation & purification; Cadherins - metabolism; Catenins; Cell Line; Cell lines; Cells; Cellular biology; Cloning, Molecular; Colonic Neoplasms; Cytoskeletal Proteins - isolation & purification; Cytoskeletal Proteins - metabolism; DNA Primers; Dogs; Gels; Genetics; HCT116 cells; Humans; Methionine - metabolism; Molecular Sequence Data; Polymerase Chain Reaction - methods; Protein Binding; Proteins; Saccharomyces cerevisiae - metabolism; Trans-Activators; Tumor Cells, Cultured; Yeast; Yeasts
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.92.11.5067

The cadherin-catenin complex is important for mediating homotypic, calcium-dependent cell-cell interactions in diverse tissue types. Although proteins of this complex have been identified, little is known about their interactions. Using a genetic assay in yeast and an in vitro protein-binding assay, we demonstrate that β-catenin is the linker protein between E-cadherin and α-catenin and that E-cadherin does not bind directly to α-catenin. We show that a 25-amino acid sequence in the cytoplasmic domain of E-cadherin and the amino-terminal domain of α-catenin are independent binding sites for β-catenin. In addition to β-catenin and plakoglobin, another member of the armadillo family, p120 binds to E-cadherin. However, unlike β-catenin, p120 does not bind α-catenin in vitro, although a complex of p120 and endogenous α-catenin could be immunoprecipitated from cell extracts. In vitro protein-binding assays using recombinant E-cadherin cytoplasmic domain and α-catenin revealed two catenin pools in cell lysates: an ≈1000- to ≈2000-kDa complex bound to E-cadherin and an ≈220-kDa pool that did not contain E-cadherin. Only β-catenin in the ≈220-kDa pool bound exogenous E-cadherin. Delineation of these molecular linkages and the 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.