Erk and PI-3 kinase are necessary for collagen binding and actin reorganization in corneal epithelia

PURPOSE. It was recently shown that phosphatidylinositol-(PI)3 kinase is up regulated in wounded rabbit corneal epithelia. Extracellular signal-regulat ed kinase (erk)-1 and -2 proteins and PI-3 kinase were activated in embryon ic corneal epithelia after I-hour stimulation by type I collagen. In the cu rrent investigation specific inhibitors of PI-3 kinase and mitogen-activate d kinase-kinase (MEK-1 kinase) were used to determine the role of these sig naling molecules in actin reorganization and collagen binding to isolated s heets of corneal epithelial tissue. METHODS. Effects of specific PI-3 kinase and MEK-1 inhibitors (LY294002, PD 98059, respectively) were investigated in embryonic corneal epithelial tiss ues. Avian embryonic corneal epithelia were isolated as tissue sheets, orga n cultured in the presence of these specific inhibitors, and stimulated wit h type I collagen. The tissues were evaluated for collagen-stimulated actin reorganization, erk-1 and -2 and PI-3 kinase activity, total filamentous a ctin accumulation, and collagen binding. RESULTS. The MEK-1 inhibitor PD98059 decreased erk-1 and -2 phosphorylation and blocked actin reorganization in a dose-dependent manner. The PI-3 kina se 85-kDa subunit was decreased 25% in LY294002-treated tissue, and collage n binding also decreased significantly in tissues treated with MEK-1 and PI -3 kinase inhibitors compared with control tissues. In addition, both inhib itors blocked actin cortical mat reorganization. CONCLUSIONS. PI-3 kinase and erk-1 and -2 signaling pathways are activated and necessary for collagen binding and integrin-mediated actin reorganizati on in embryonic avian corneal epithelium.