title: The importance of E-cadherin binding partners to evaluate the pathogenicity of E-cadherin missense mutations associated to HDGC.
abstract: In hereditary diffuse gastric cancer (HDGC), CDH1 germline gene alterations are causative events in 30% of the cases. In 20% of HDGC families, CDH1 germline mutations are of the missense type and the mutation carriers constitute a problem in terms of genetic counseling and surveillance. To access the pathogenic relevance of missense mutations, we have previously developed an in vitro method to functionally characterize them. Pathogenic E-cadherin missense mutants fail to aggregate and become more invasive, in comparison with cells expressing the wild-type (WT) protein. Herein, our aim was to develop a complementary method to unravel the pathogenic significance of E-cadherin missense mutations. We used cells stably expressing WT E-cadherin and seven HDGC-associated mutations (five intracellular and two extracellular) and studied by proximity ligation assays (PLA) how these mutants bind to fundamental regulators of E-cadherin function and trafficking. We focused our attention on the interaction with: p120, β-catenin, PIPKIγ and Hakai. We showed that cytoplasmic E-cadherin mutations affect the interaction of one or more binding partners, compromising the E-cadherin stability at the plasma membrane and likely affecting the adhesion complex competence. In the present work, we demonstrated that the study of the interplay between E-cadherin and its binding partners, using PLA, is an easy, rapid, quantitative and highly reproducible technique that can be applied in routine labs to verify the pathogenicity of E-cadherin missense mutants for HDGC diagnosis, especially those located in the intracellular domain of the protein.
authors: IPATIMUP, Institute of Molecular Pathology and Immunology of the University of Porto, Porto, Portugal.
authors: Joana
authors: J
authors: Figueiredo
authors: Ola
authors: O
authors: Söderberg
authors: Joana
authors: J
authors: Simões-Correia
authors: Karin
authors: K
authors: Grannas
authors: Gianpaolo
authors: G
authors: Suriano
authors: Raquel
authors: R
authors: Seruca
mesh_terms: D000818:Animals; D015703:Antigens, CD; D001665:Binding Sites; D016466:CHO Cells; D015820:Cadherins / Q000235:genetics* / Q000378:metabolism*; D051177:Catenins / Q000378:metabolism; D006224:Cricetinae; D003593:Cytoplasm / Q000235:genetics / Q000378:metabolism; D005089:Exocytosis / Q000235:genetics; D006801:Humans; D008967:Molecular Biology / Q000379:methods; D020125:Mutation, Missense; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism; D013274:Stomach Neoplasms / Q000235:genetics* / Q000473:pathology; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism; D051176:beta Catenin / Q000378:metabolism; D000095002:Delta Catenin
chemical_list: D015703:Antigens, CD; C513384:CDH1 protein, human; D015820:Cadherins; D051177:Catenins; D051176:beta Catenin; C471638:CBLL1 protein, human; D044767:Ubiquitin-Protein Ligases; D017853:Phosphotransferases (Alcohol Group Acceptor); C020077:1-phosphatidylinositol-4-phosphate 5-kinase; D000095002:Delta Catenin