Cell phenotype in normal epithelial cell lines with high endogenous N-cadherin: comparison of RPE to an MDCK subclone.
OBJECTIVE
Unlike most monolayer epithelial cells, cultured RPE are competent to form a zonular adhesion of N- rather than E-cadherin. To determine whether other normal epithelial cells do likewise, cells with high endogenous N-cadherin were cloned from the typically E-cadherin dominant epithelial line Madin-Darby canine kidney cells (MDCK) to analyze cell and junction phenotype in the presence of N-cadherin.


METHODS
A MDCK subclonal line, clone-YH, was selected for high endogenous N-cadherin and was compared with the RPE line hTERT-RPE1 with regard to cell phenotype, cadherin gene expression and cadherin protein distribution, glycosylation state, and catenin complex composition.


RESULTS
In early cultures, hTERT-RPE1 cells are moderately epithelioid with junctional N-cadherin, but clone-YH cells are initially highly fusiform with N-cadherin in multiple sites. With time, N-cadherin in clone-YH becomes deglycosylated, resistant to detergent extraction, and zonular, and cells become epithelioid. Treatment with the N-glycosylation inhibitor tunicamycin induces an epithelioid phenotype in clone-YH, like time in culture but disrupts the hTERT-RPE1 phenotype. N-cadherin traffics to surface membranes and complexes with catenins regardless of cell type or glycosylation state, although catenin complex composition varied, showing enriched alpha-catenin under the cell-type-specific conditions in which N-cadherin was junctional. Clone-YH continued to express E-cadherin as a very minor cadherin, which trafficked to membranes but did not accumulate at junctions.


CONCLUSIONS
RPE cells are not unique in localizing N-cadherin to a zonular adhesion typical of a monolayer epithelium, because even epithelial cells derived from a typically E-cadherin dominant line (clone-YH) form a zonular N-cadherin junction if the protein is abundant. However, there are cell and cadherin differences in mechanisms of cadherin accumulation in a zonular pattern, and a previously unrecognized cell-type-specific role for protein glycosylation in epithelial phenotype development.
Youn
Yong-Ha
YH
Department of Ophthalmology, Medical College of Wisconsin, Milwaukee 53226-4812, USA.
Hong
Jeehee
J
Burke
Janice M
JM
D000818:Animals; D019921:Biotinylation; D015820:Cadherins / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D002999:Clone Cells; D004285:Dogs; D004847:Epithelial Cells / Q000166:cytology* / Q000378:metabolism; D015870:Gene Expression; D006031:Glycosylation; D006801:Humans; D007668:Kidney / Q000166:cytology; D008856:Microscopy, Fluorescence; D010641:Phenotype; D010857:Pigment Epithelium of Eye / Q000166:cytology* / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction
D015820:Cadherins