Normal function of HERG K⁺ channels expressed in HEK293 cells requires basal protein kinase B activity

Abstract

The potential role of protein kinase B (PKB), a serine/threonine protein kinase, in regulating HERG (human ethera-go-go related gene) K + channel function was investigated. Wortmannin (a phosphoinositide 3-kinase (PI3K) inhibitor) caused V V30% reduction of HERG current (I HERG ) stably expressed in HEK293 cells. Transient transfection with the constitutively active PI3K in HERG-expressing HEK293 cells slightly increased (V V7%) I HERG while a dominant negative PI3K signi¢cantly reduced I HERG (V V25%) relative to results in vehicle-transfected cells. I HERG was V V35% greater in cells transfected with the constitutively activated PKB (caPKB), whereas it was V V47% smaller in cells transfected with dominant negative PKB (dnPKB). Basal activation of PKB was detected by immunocytochemistry. PKB activity was signi¢cantly enhanced in caPKB-transfected cells and nearly abolished in dnPKB-transfected cells. We conclude that normal HERG function in HEK293 cells requires basal activity of PKB. Our data represent the ¢rst evidence that PKB phosphorylation regulates K + channels. ß 2002 Published by Elsevier Science B.V. on behalf of the Federation of European Biochemical Societies.

Downregulation of the HERG (KCNH2) K+ channel by ceramide: evidence for ubiquitin-mediated lysosomal degradation

“…See Discussion for further details. density in stably expressing HEK cells (Zhang et al, 2003), though the underlying mechanism was not elucidated.…”

“…Similarly TNF-␣, which can induce sphingomyelinase activation and ceramide production, was recently shown to reduce the HERG current via reactive oxygen species (ROS) . In addition to ROS, ceramide signalling also results in activation of protein kinase C and alteration of protein kinase B activity (Ruvolo, 2001;Ramström et al, 2004) both of which modulate HERG channel function (Barros et al, 1998;Thomas et al, 2003;Zhang et al, 2003). However, little is known of the effects of ceramide on…”

Hypotonic activation of volume-sensitive outwardly rectifying chloride channels in cultured PASMCs is modulated by SGK

“…Previous studies have identified major roles of Akt/PKB in cell proliferation and survival and in insulin signaling (5,32). Although information regarding role of Akt/PKB in activation of ion channels is sparse (2,56), Akt/PKB was activated during hypotonic swelling of PASMCs (Fig. 6B) and hence was considered a potential regulatory protein upstream of VSOACs.…”

“…These observations independently confirmed that hypotonic swelling of PASMCs activates PI3Ks and downstream kinases, including Akt/PKB. Akt/PKB has been shown to modulate L-type Ca 2ϩ (2) and human ether-a-go-go-related gene potassium channels (56). To test whether Akt/PKB is involved in activation of VSOACs, we dialyzed through the patch pipette recombinant active Akt1 (20 U/ml) into cultured PASMCs.…”

Rapamycin inhibits oncogenic intestinal ion channels and neoplasia in APCMin/+ mice

“…APC Min/ þ animals show overexpression of several members of the Eag family of Kv channels, as well as of BK channels. Erg channels are activated by Akt (Zhang et al, 2003) and confer greater malignancy to patients with leukemia (Pillozzi et al, 2007). Moreover, mutations in the APC gene alter cell cycling (Dikovskaya et al, 2007), and as Eag family channels are cell cycle regulated (Wonderlin and Strobl, 1996), it is likely that these changes affect their activity.…”