The Class II Phosphatidylinositol 3 kinase C2 beta Is Required for the Activation of the K+ Channel KCa3.1 and CD4 T-Cells

The Ca2+-activated K+ channel KCa3.1 is required for Ca2+ 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 C2 beta (PI3K-C2 beta) is activated by the T-cell receptor (TCR) and functions upstream of NDPK-B to activate KCa3.1 channel activity. Decreased expression of PI3K-C2 beta 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-C2 beta siRNA-treated T-cells with PI(3) P rescued KCa3.1 channel activity. Moreover, overexpression of PI3K-C2 beta in KCa3.1-transfected Jurkat T-cells led to increased TCR-stimulated activation of KCa3.1 and Ca2+ influx, whereas silencing of PI3K-C2 beta inhibited both responses. Using total internal reflection fluorescence microscopy and planar lipid bilayers, we found that PI3K-C2 beta 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.