[Differential tyrosine phosphorylation of paxillin in human corneal epithelial cells on extracellular matrix proteins].
OBJECTIVE
To understand the interaction of corneal epithelial cells with laminin, fibronectin, and collagen type IV, major components of basement membrane, we investigated whether tyrosine phosphorylation of paxillin was increased during attachment of these cells to matrix proteins. Paxillin is one of the focal adhesion proteins and it is tyrosine phosphorylated during cell adhesion.


METHODS
SV 40-transformed human corneal epithelial (HCE) cells were plated on these extracellular matrix proteins and incubated. The cellular lysates were submitted to immunoprecipitation and western blotting to determine tyrosine phosphorylation of paxillin.


RESULTS
When the cells were plated on laminin matrix, the stained band indicating tyrosine phosphorylated paxillin increased in proportion to cultivation periods. The increase was significant at 6 to 24 hours of cultivation. When HCE cells were cultured on various concentrations of laminin, paxillin was up-regulated in phosphorylation in a dose-dependent fashion. On a fibronectin matrix, tyrosine phosphorylation of paxillin increased in a time-dependent fashion and peak time point was 6 hours of cultivation. Paxillin was up-regulated in tyrosine phosphorylation in direct relation with the fibronectin concentration. On a type IV collagen matrix, tyrosine phosphorylation of paxillin increased in relation to time, but not so rapidly as cultures on a laminin or fibronectin matrix.


CONCLUSIONS
Differential tyrosine phosphorylation of paxillin may have been caused by different extracellular matrix proteins.
Ofuji
K
K
Department of Ophthalmology, Yamaguchi University School of Medicine, Ube, Japan.
D002478:Cells, Cultured; D003094:Collagen / Q000502:physiology*; D003598:Cytoskeletal Proteins / Q000378:metabolism*; D019573:Epithelium, Corneal / Q000378:metabolism*; D005353:Fibronectins / Q000502:physiology*; D006801:Humans; D007797:Laminin / Q000502:physiology*; D051419:Paxillin; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D014443:Tyrosine / Q000378:metabolism*
D003598:Cytoskeletal Proteins; D005353:Fibronectins; D007797:Laminin; C496061:PXN protein, human; D051419:Paxillin; D010750:Phosphoproteins; D014443:Tyrosine; D003094:Collagen