Y. Fukudome et al., Characterization of a mutant E-cadherin protein encoded by a mutant gene frequently seen in diffuse-type human gastric carcinoma, INT J CANC, 88(4), 2000, pp. 579-583
The cell-cell adhesion molecule E-cadherin plays an essential role in the m
aintenance and function of epithelial tissues. Altered expression of E-cadh
erin has been implicated in tumor invasion. Analysis of mutations of the hu
man E-cadherin gene in gastric carcinoma of the diffuse type has revealed t
hat 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 o
f E-cadherin in a cell line, HSC45-M2, established from a human signet ring
-cell carcinoma, Although immunohistochemical analysis showed that the muta
nt cadherin was localized at cell-cell contact sites as usually seen with t
he wild type, these cells did not form compact colonies. HSC45-M2 cells exp
ressed 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 tran
scripts, which appeared to be caused by the splicing defect. Northern blott
ing, however, showed that the transcript lacking exon 9 was predominantly d
etected in these cells. The electrophoretic mobilities on SDS-PACE of the m
utant E-cadherin protein in HSC45-M2 cells and the protein expressed from c
DNA lacking exon 9 appeared identical. Analysis of the amino-terminal regio
n of the mutant E-cadherin protein revealed that the cadherin was capable o
f becoming a mature form by removal of its amino-terminal peptide. However,
the mutant E-cadherin was susceptible to trypsinization in the presence of
Ca2+ which is not the case for wild-type E-cadherin, suggesting that the m
utant E-cadherin frequently found in diffuse-type gastric carcinoma may hav
e lost its Ca2+-binding ability, leading to disruption of the tight cell-ce
ll association. (C) 2000 Wiley-Liss, Inc.