PI3K regulates pleckstrin-2 in T-cell cytoskeletal reorganization.
Pleckstrin-2 is composed of 2 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 alpha4beta1. 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 colocalized 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, because 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 coordinates with the actin cytoskeleton in lymphocyte spreading and immune synapse formation.
Bach
Tami L
TL
Department of Medicine, University of Pennsylvania School of Medicine, and Department of Pathology, Children's Hospital of Philadelphia 19104, USA.
Kerr
Wesley T
WT
Wang
Yanfeng
Y
Bauman
Eve Marie
EM
Kine
Purnima
P
Whiteman
Eileen L
EL
Morgan
Renell S
RS
Williamson
Edward K
EK
Ostap
E Michael
EM
Burkhardt
Janis K
JK
Koretzky
Gary A
GA
Birnbaum
Morris J
MJ
Abrams
Charles S
CS
D000199:Actins / Q000378:metabolism; D001665:Binding Sites; D002448:Cell Adhesion; D003599:Cytoskeleton / Q000378:metabolism*; D006801:Humans; D019169:Jurkat Cells; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism / Q000494:pharmacology / Q000502:physiology*; D019869:Phosphatidylinositol 3-Kinases / Q000502:physiology*; D010716:Phosphatidylinositols / Q000378:metabolism; D017354:Point Mutation; D013601:T-Lymphocytes / Q000166:cytology*; D014161:Transduction, Genetic
D000199:Actins; D008565:Membrane Proteins; C120651:PLEK2 protein, human; D010716:Phosphatidylinositols; D019869:Phosphatidylinositol 3-Kinases