title: Characterization of a mutant E-cadherin protein encoded by a mutant gene frequently seen in diffuse-type human gastric carcinoma.
abstract: The cell-cell adhesion molecule E-cadherin plays an essential role in the maintenance and function of epithelial tissues. Altered expression of E-cadherin has been implicated in tumor invasion. Analysis of mutations of the human E-cadherin gene in gastric carcinoma of the diffuse type has revealed that deletion of exon 8 or 9 in its cDNA appears to be predominant. In this study, we carried out structural and functional analyses of a mutant form of E-cadherin in a cell line, HSC45-M2, established from a human signet ring-cell carcinoma. Although immunohistochemical analysis showed that the mutant cadherin was localized at cell-cell contact sites as usually seen with the wild type, these cells did not form compact colonies. HSC45-M2 cells expressed aberrant E-cadherin with an m.w. larger than that of the wild type. In these cells, we found deletion of the exon 9-intron 9 boundary including the splicing donor site in E-cadherin genomic DNA. RT-PCR indicated 2 transcripts, which appeared to be caused by the splicing defect. Northern blotting, however, showed that the transcript lacking exon 9 was predominantly detected in these cells. The electrophoretic mobilities on SDS-PAGE of the mutant E-cadherin protein in HSC45-M2 cells and the protein expressed from cDNA lacking exon 9 appeared identical. Analysis of the amino-terminal region of the mutant E-cadherin protein revealed that the cadherin was capable of becoming a mature form by removal of its amino-terminal peptide. However, the mutant E-cadherin was susceptible to trypsinization in the presence of Ca(2+), which is not the case for wild-type E-cadherin, suggesting that the mutant E-cadherin frequently found in diffuse-type gastric carcinoma may have lost its Ca(2+)-binding ability, leading to disruption of the tight cell-cell association.
authors.affiliation: Department of Biochemistry, Faculty of Pharmaceutical Sciences, Setsunan University, Hirakata, Osaka, Japan.
authors.forename: Y
authors.initials: Y
authors.lastname: Fukudome
authors.forename: K
authors.initials: K
authors.lastname: Yanagihara
authors.forename: M
authors.initials: M
authors.lastname: Takeichi
authors.forename: F
authors.initials: F
authors.lastname: Ito
authors.forename: S
authors.initials: S
authors.lastname: Shibamoto
mesh_terms: D000818:Animals; D001483:Base Sequence; D015820:Cadherins / Q000032:analysis / Q000737:chemistry / Q000235:genetics*; D018279:Carcinoma, Signet Ring Cell / Q000235:genetics*; D005091:Exons; D006801:Humans; D007150:Immunohistochemistry; D007365:Intercellular Junctions / Q000473:pathology; D007438:Introns; D007739:L Cells; D051379:Mice; D008970:Molecular Weight; D009154:Mutation; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017384:Sequence Deletion; D013274:Stomach Neoplasms / Q000235:genetics*; D014162:Transfection; D014407:Tumor Cells, Cultured
chemical_list: D015820:Cadherins; D011994:Recombinant Proteins