The Role of PI3K in Ephrin A1‐Induced Actin Remodeling

The receptor EphA2 is one of fourteen highly conserved Eph receptor tyrosine kinases, which along with their cognate ligands, the ephrins, confer contact‐mediated inhibition of cellular migration. Ephrin A1 stimulation of cultured cells has been shown to activate the small GTPase Rho to cause marked retraction of fillipodia and lamellapodia following detachment of the cell from its substrate, resulting in cellular rounding via actin reorganization. Our biochemical and morphologic studies on NIH 3T3 cells have identified phosphatidyl inositol 3′ kinase (PI3K) as a downstream effector protein of the EphA2 receptor following Ephrin A1 stimulation, which plays a critical role in mediating the cellular retraction/actin reorganization responses. The regulatory subunit of PI3K (p85) associates with the activated EphA2 receptor following ephrin A1 stimulation, leading to a rapid increase in PI3K activity as assayed by phosphorylation of Akt on serine 473. Through the use of the PI3K‐specific inhibitor LY294002 we have shown that PI3K's catalytic activity is necessary for the phosphorylation of myosin light chain 2 (MLC2) on threonine 18 and serine 19. Imaging of the actin architecture has shown that ephrin A1‐induced actin reorganization and fillipodia/lamellapodia retraction are also blocked following the inhibition of PI3K. These results indicate that PI3K activity mediates the ephrin A1‐induced cellular retraction through the phosporylation of the contractile protein MLC2 and the reorganization of actin microfilaments. This work was supported by NIH research grants (HL64382, HL43026).