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

Abstract

Phosphotyrosine proteins involvement, particularly cortactin, was studied in cell-cell contacts of cultured bovine corneal endothelial (BCE) cells. These proteins, including alpha-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.

Extending the knowledge in histochemistry and cell biology

“…In experiments using S1P receptor subtype 1 siRNA or dominant-negative-Rho family GTPases, these eVects were abrogated, leading the authors to hypothesize that activation of integrin v 3 via S1P-and Rho GTPasemediated signaling is important for S1P-stimulated chemotactic responses of endothelial cells. Kredy-Farhan et al (2008) studied the localization of cortactin in cell-cell contacts of cultured bovine corneal endothelial cells. Cortactin is a phosphotyrosine protein and expressed as two known isoforms, p80 and p85 (CosenBinker and Kapus 2006).…”

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“…The authors concluded from their Wndings that Rac1 activation, which might partly be mediated by Epac/Rap1 pathway, contributes to cAMP-induced barrier stabilization. The involvement of cortactin in intercellular contact formation in cultured bovine corneal endothelial cell was the subject of a study by Kredy-Farhan et al (2008). Two isoforms of cortactin, p80 and p85, phosphorylated at diVerent tyrosine residues, were localized in subconXuent and conXuent endothelial cell layers, and distinct localization patterns and associations with Triton-soluble and -insoluble cellular fractions were found, The Wndings were interpreted as indicating that various cortactin isoforms phosphorylated at speciWc tyrosines may play diVerent roles and have diVerent preferential localization and associations with the cytoskeleton in bovine corneal endothelial cells.…”