Pleckstrin-2 membrane association and cell spreading are PI3K dependent. (A) Jurkat cells expressing GFP fused to the amino-terminus of wild-type pleckstrin-2 were plated on fibronectin. Real-time confocal imaging of live cells was performed. Shown is a cell expressing pleckstrin-2 before and after inhibition of PI3K with 100 nM wortmannin. This figure shows that wild-type pleckstrin-2 associates with membrane projections in cells spread on fibronectin, and this membrane association is rapidly (about 2 minutes) disrupted by the addition of wortmannin. (B) Jurkat cells expressing GFP (GFP) alone or GFP fused to the amino-terminus of wild-type pleckstrin-2 (WT pleckstrin-2) were plated on fibronectin. Where indicated, 100 nM wortmannin was added for 5 minutes to inhibit PI3K. Images were captured at 100× magnification and cell-footprint size was quantified using IP Lab imaging software. Shown is the mean ± SEM of 3 independent experiments.