Role of cell adhesion-associated protein, pinin (DRS/memA), in corneal epithelial migration.
OBJECTIVE
To determine whether the cellular distribution of cell adhesion-associated protein, pinin, is altered during corneal epithelial migration in response to debridement wounding and to determine the effect of overexpression of pinin in cultured epithelial cells.


METHODS
Corneas from guinea pig and embryonic (day 17) chickens were excised, wounded, and placed on organ-culture rafts. At time points from 0 to 24 hours, corneas were cryosectioned and subsequently analyzed by immunofluorescence or immunoelectron microscopy for the presence and distribution of pinin. Cultured epithelial cell line MDCK (Madin Darby canine kidney) confluent monolayers were wounded by scraping and examined by immunofluorescence for pinin and desmoplakin. MDCK cells were transfected with full-length pinin cDNA. After selection in Geneticin, clones of pinin-transfected cells were isolated. Monolayers of transfected cells were scrape-wounded and assayed for their ability to migrate.


RESULTS
Within 2 hours after wounding, although morphologically identifiable desmosomes were present on migrating epithelial cells, the association of pinin to desmosomes was greatly reduced. Finally, after completion of wound closure, pinin returned to the corneal epithelial desmosome. Wounding of confluent epithelial monolayers (MDCK) in vitro demonstrated a very similar change in the distribution of pinin, whereas desmoplakin remained cell boundary-associated. Transfection of pinin into cultured epithelial cells resulted in an overexpression of pinin. Clones of cells expressing high levels of pinin exhibited marked reduction in their ability to migrate after wounding.


CONCLUSIONS
Pinin is involved in corneal epithelium migration. The localization of pinin at or near the desmosome is correlated with the epithelial quiescence. The loss of pinin from the cell boundary correlates with the transition from quiescence to actively migrating. Overexpressing pinin in cultured epithelial cells affects epithelial homeostasis and, in turn, drives the epithelial cells to a hyperstable epithelial adhesive state and inhibits the transition from quiescence to migratory.
Shi
Y
Y
Department of Anatomy and Cell Biology, University of Florida College of Medicine, Gainesville 32610-0235, USA.
Tabesh
M
M
Sugrue
S P
SP
D000818:Animals; D015815:Cell Adhesion Molecules / Q000235:genetics / Q000502:physiology*; D002465:Cell Movement / Q000502:physiology*; D002478:Cells, Cultured; D002642:Chick Embryo; D003598:Cytoskeletal Proteins / Q000378:metabolism; D003646:Debridement; D051180:Desmoplakins; D003896:Desmosomes / Q000378:metabolism; D019573:Epithelium, Corneal / Q000378:metabolism* / Q000648:ultrastructure; D019084:Fluorescent Antibody Technique, Indirect; D015870:Gene Expression; D006168:Guinea Pigs; D016253:Microscopy, Immunoelectron; D009687:Nuclear Proteins / Q000235:genetics / Q000502:physiology*; D009924:Organ Culture Techniques; D014162:Transfection; D014945:Wound Healing
D015815:Cell Adhesion Molecules; D003598:Cytoskeletal Proteins; D051180:Desmoplakins; D009687:Nuclear Proteins