Involvement of cortactin and phosphotyrosine proteins in cell–cell contact formation in cultured bovine corneal endothelial cells

摘要

Phosphotyrosine proteins involvement, particularly cortactin, was studied in cell–cell contacts of cultured bovine corneal endothelial (BCE) cells. These proteins, including α-catenin, vinculin and cortactin, are localized at cell–cell contacts separate from the cortical actin ring. Approximately 50% of cortactin isoforms p80 and p85 were associated with the Triton-insoluble fraction while phosphotyrosine proteins were in the soluble fraction. Disruption of cell–cell contacts by EDTA treatment was associated with a decrease in cortactin isoforms p80 (26%) and p85 (57%). Cortactin isoform p85 was phosphorylated at Y466, expressed in reattaching cells and associated with the Triton-soluble fraction, whereas cortactin isoform p80 was phosphorylated at Y421 and associated with the Triton-insoluble fraction. In sub-confluent cultures, pY421-cortactin was localized at the leading edge and pY466-cortactin at a perinuclear area. In confluent cultures both pY466- and pY421-cortactin isoforms were localized at the cell–cell contacts. In conclusion, in BCE cells, the most prominent appearance of cortactin was at the cell–cell contacts separate from the cortical actin ring. Isoform p80 was phosphorylated at Y421 and associated with the Triton-insoluble fraction and isoform p85 was phosphorylated at Y466 and associated with the Triton-insoluble fraction.