The Class II Phosphatidylinositol 3 kinase C2β Is Required for the Activation of the K⁺Channel KCa3.1 and CD4 T-Cells

Abstract

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 C2␤ (PI3K-C2␤) 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␤ 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␤ siRNA-treated T-cells with PI(3)P rescued KCa3.1 channel activity. Moreover, overexpression of PI3K-C2␤ in KCa3.1-transfected Jurkat T-cells led to increased TCR-stimulated activation of KCa3.1 and Ca 2؉ influx, whereas silencing of PI3K-C2␤ inhibited both responses. Using total internal reflection fluorescence microscopy and planar lipid bilayers, we found that PI3K-C2␤ 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.

KCa3.1 channels mediate the increase of cell migration and proliferation by advanced glycation endproducts in cultured rat vascular smooth muscle cells

“…[29][30][31] Decreasing intracellular level of PI(3)P can reduce K Ca 3.1 channel activity. 32 Besides, recombinant human K Ca 3.1 channels induce HEK293 cell proliferation by a direct interaction with ERK1/2 pathways 33 and downregulating ERK reduces K Ca 3.1 channel expression. 32 These reports support the notion that K Ca 3.1 channels are the downstream target of ERK1/2, P38-MAPK and PI3K in regulating cell proliferation and migration of rat VSMCs ( Figure 6).…”

Tripartite motif containing protein 27 negatively regulates CD4 T cells by ubiquitinating and inhibiting the class II PI3K-C2β

“…Cells and Primary Human CD4 T Cells by Inhibiting PI3KC2β. Jurkat T cells that overexpress KCa3.1 (Jurkat-KCa3.1) were transfected with an siRNA to TRIM27 and KCa3.1 channel activity was assessed as previously described (19). siRNA knockdown of TRIM27 (Fig.…”

“…Recently, we found that the class II PI3KC2β plays an important and unexpected role in CD4 T-cell activation (19). These studies demonstrated that activation of PI3KC2β, but not PI3KC2α, following TCR stimulation functions to recruit PI3KC2β to the immunological synapse, leading to the generation of PI(3)P, which is subsequently required for the histidine phosphorylation and activation of KCa3.1 by nucleoside diphosphate kinase B (NDPK-B) (19,20).…”

“…These studies demonstrated that activation of PI3KC2β, but not PI3KC2α, following TCR stimulation functions to recruit PI3KC2β to the immunological synapse, leading to the generation of PI(3)P, which is subsequently required for the histidine phosphorylation and activation of KCa3.1 by nucleoside diphosphate kinase B (NDPK-B) (19,20). Activation of KCa3.1, as well as another K + channel Kv1.3, has been shown to play critical roles in CD4 T-cell activation (21)(22)(23)(24)(25).…”

The NDPK/NME superfamily: state of the art

“…For NME2/NDPK-B, at least two substrates were identified and studied in more detail: the β subunit of heterotrimeric G protein [120][121][122] and the potassium channel KCa3.1. [123][124][125] In the first case, NME2 forms a complex with β/γ dimers of heterotrimeric G proteins, in which a phospho-relay occurs from His118 of NME2 to His266 of the Gβ subunit. This ultimately leads to the formation of GTP from GDP and thus receptor-independent activation of the G protein.…”