The class II phosphatidylinositol 3 kinase C2beta is required for the activation of the K+ channel KCa3.1 and CD4 T-cells.
The Ca(2+)-activated K(+) channel KCa3.1 is required for Ca(2+) influx and the subsequent activation of T-cells. We previously showed that nucleoside diphosphate kinase beta (NDPK-B), a mammalian histidine kinase, directly phosphorylates and activates KCa3.1 and is required for the activation of human CD4 T lymphocytes. We now show that the class II phosphatidylinositol 3 kinase C2beta (PI3K-C2beta) is activated by the T-cell receptor (TCR) and functions upstream of NDPK-B to activate KCa3.1 channel activity. Decreased expression of PI3K-C2beta by siRNA in human CD4 T-cells resulted in inhibition of KCa3.1 channel activity. The inhibition was due to decreased phosphatidylinositol 3-phosphate [PI(3)P] because dialyzing PI3K-C2beta siRNA-treated T-cells with PI(3)P rescued KCa3.1 channel activity. Moreover, overexpression of PI3K-C2beta in KCa3.1-transfected Jurkat T-cells led to increased TCR-stimulated activation of KCa3.1 and Ca(2+) influx, whereas silencing of PI3K-C2beta inhibited both responses. Using total internal reflection fluorescence microscopy and planar lipid bilayers, we found that PI3K-C2beta colocalized with Zap70 and the TCR in peripheral microclusters in the immunological synapse. This is the first demonstration that a class II PI3K plays a critical role in T-cell activation.
Srivastava
Shekhar
S
Division of Nephrology, Department of Pharmacology, The Helen L. and Martin S. Kimmel Center for Biology and Medicine at the Skirball Institute for Biomolecular Medicine, New York University Langone Medical Center, New York, NY 10016.
Di
Lie
L
Zhdanova
Olga
O
Li
Zhai
Z
Vardhana
Santosha
S
Wan
Qi
Q
Yan
Ying
Y
Varma
Rajat
R
Backer
Jonathan
J
Wulff
Heike
H
Dustin
Michael L
ML
Skolnik
Edward Y
EY
D015496:CD4-Positive T-Lymphocytes / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D020013:Calcium Signaling / Q000502:physiology; D058541:Class II Phosphatidylinositol 3-Kinases; D006801:Humans; D051660:Intermediate-Conductance Calcium-Activated Potassium Channels / Q000235:genetics / Q000378:metabolism*; D019169:Jurkat Cells; D008213:Lymphocyte Activation; D009701:Nucleoside-Diphosphate Kinase / Q000235:genetics / Q000378:metabolism; D018408:Patch-Clamp Techniques; D019869:Phosphatidylinositol 3-Kinases / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics / Q000378:metabolism; D011948:Receptors, Antigen, T-Cell / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D051746:ZAP-70 Protein-Tyrosine Kinase / Q000235:genetics / Q000378:metabolism
D051660:Intermediate-Conductance Calcium-Activated Potassium Channels; C496923:KCNN4 protein, human; D034741:RNA, Small Interfering; D011948:Receptors, Antigen, T-Cell; D011993:Recombinant Fusion Proteins; D019869:Phosphatidylinositol 3-Kinases; D058541:Class II Phosphatidylinositol 3-Kinases; C502616:PIK3C2B protein, human; D051746:ZAP-70 Protein-Tyrosine Kinase; D009701:Nucleoside-Diphosphate Kinase; C542596:NDPK-B protein, human; D002118:Calcium