Submitted February 2, 2006
Accepted September 5, 2006
PI3K regulates pleckstrin-2 in T-cell cytoskeletal re-organization
Tami L. Bach, Wesley Kerr, Yanfeng Wang, Eve Marie Bauman, Purnima Kine, Eileen L. Whiteman, Renell S. Morgan, Edward K. Williamson, E. Michael Ostap, Janis K. Burkhardt, Gary A. Koretzky, Morris J. Birnbaum, and Charles S. Abrams*
University of Pennsylvania School of Medicine
Department of Medicine, University of Pennsylvania School of Medicine
Children's Hospital of Philadelphia and the University of Pennsylvania
Abramson Family Cancer Institute, University of Pennsylvania School of Medicine
* Corresponding author; email: abrams{at}mail.med.upenn.edu.
Pleckstrin-2 is composed of two Pleckstrin Homology (PH) domains and a Disheveled-Egl 10-Pleckstrin (DEP) domain. A lipid-binding assay revealed that pleckstrin-2 binds with greatest affinity to D3- and D5-phosphoinositides. Pleckstrin-2 expressed in Jurkat T-cells bound to the cellular membrane and enhanced actin-dependent spreading only after stimulation of the T-cell antigen receptor or the integrin, 
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1. A pleckstrin-2 variant containing point mutations in both PH domains failed to associate with the Jurkat membrane and had no effect on spreading under the same conditions. Although still membrane-bound, a pleckstrin-2 variant containing point mutations in the DEP domain demonstrated a decreased ability to induce membrane ruffles and spread. Pleckstrin-2 also co-localized with actin at the immune synapse and integrin clusters via its PH domains. Although pleckstrin-2 can bind to purified D3- and D5-phosphoinositides, the intracellular membrane association of pleckstrin-2 and cell spreading are dependent on D3-phosphoinositides since these effects were disrupted by pharmacologic inhibition of phosphatidylinositol 3-kinase (PI3K). Our results indicate that pleckstrin-2 uses its modular domains to bind to membrane-associated phosphatidylinositols generated by PI3K whereby it co-ordinates with the actin cytoskeleton in lymphocyte spreading and immune synapse formation.
