Deficiency of PTEN in Jurkat T cells causes constitutive localization of Itk to the plasma membrane and hyperresponsiveness to CD3 stimulation.
Pleckstrin homology (PH) domain binding to D3-phosphorylated phosphatidylinositides (PI) provides a reversible means of recruiting proteins to the plasma membrane, with the resultant change in subcellular localization playing a key role in the activation of multiple intracellular signaling pathways. Previously we found that the T-cell-specific PH domain-containing kinase Itk is constitutively membrane associated in Jurkat T cells. This distribution was unexpected given that the closely related B-cell kinase, Btk, is almost exclusively cytosolic. In addition to constitutive membrane association of Itk, unstimulated JTAg T cells also exhibited constitutive phosphorylation of Akt on Ser-473, an indication of elevated basal levels of the phosphatidylinositol 3-kinase (PI3K) products PI-3,4-P(2) and PI-3,4,5-P(3) in the plasma membrane. Here we describe a defect in expression of the D3 phosphoinositide phosphatase, PTEN, in Jurkat and JTAg T cells that leads to unregulated PH domain interactions with the plasma membrane. Inhibition of D3 phosphorylation by PI3K inhibitors, or by expression of PTEN, blocked constitutive phosphorylation of Akt on Ser-473 and caused Itk to redistribute to the cytosol. The PTEN-deficient cells were also hyperresponsive to T-cell receptor (TCR) stimulation, as measured by Itk kinase activity, tyrosine phosphorylation of phospholipase C-gamma1, and activation of Erk compared to those in PTEN-replete cells. These data support the idea that PH domain-mediated association with the plasma membrane is required for Itk activation, provide evidence for a negative regulatory role of PTEN in TCR stimulation, and suggest that signaling models based on results from Jurkat T-cell lines may underestimate the role of PI3K in TCR signaling.
Shan
X
X
Laboratory of Biological Chemistry, Gerontology Research Center, National Institute on Aging, National Institutes of Health, Baltimore, Maryland 21224-6825, USA.
Czar
M J
MJ
Bunnell
S C
SC
Liu
P
P
Liu
Y
Y
Schwartzberg
P L
PL
Wange
R L
RL
D000818:Animals; D000952:Antigens, Polyomavirus Transforming / Q000235:genetics; D001483:Base Sequence; D001665:Binding Sites; D001692:Biological Transport; D001798:Blood Proteins / Q000378:metabolism; D017252:CD3 Complex / Q000378:metabolism* / Q000494:pharmacology; D002462:Cell Membrane / Q000378:metabolism; D003600:Cytosol; D004789:Enzyme Activation; D005091:Exons; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism; D019169:Jurkat Cells; D008969:Molecular Sequence Data; D016296:Mutagenesis; D051059:PTEN Phosphohydrolase; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D018129:Phosphatidylinositol Phosphates / Q000378:metabolism; D051966:Phospholipase C gamma; D010750:Phosphoproteins / Q000378:metabolism; D010744:Phosphoric Monoester Hydrolases / Q000096:biosynthesis / Q000235:genetics / Q000502:physiology*; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D017027:Protein Tyrosine Phosphatases / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt; D011817:Rabbits; D013601:T-Lymphocytes / Q000187:drug effects / Q000276:immunology / Q000378:metabolism*; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured; D025521:Tumor Suppressor Proteins; D010738:Type C Phospholipases / Q000378:metabolism
D000952:Antigens, Polyomavirus Transforming; D001798:Blood Proteins; D017252:CD3 Complex; D007527:Isoenzymes; D051059:PTEN Phosphohydrolase; D019869:Phosphatidylinositol 3-Kinases; D018129:Phosphatidylinositol Phosphates; D051966:Phospholipase C gamma; D010750:Phosphoproteins; D010744:Phosphoric Monoester Hydrolases; D017346:Protein Serine-Threonine Kinases; D017027:Protein Tyrosine Phosphatases; D011505:Protein-Tyrosine Kinases; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D025521:Tumor Suppressor Proteins; D010738:Type C Phospholipases; C060974:phosphatidylinositol 3,4,5-triphosphate; C060980:phosphatidylinositol 3,4-diphosphate; C039077:platelet protein P47; C080813:emt protein-tyrosine kinase; C494918:AKT1 protein, human; C096335:B59 protein, human; C494929:PTEN protein, human