Upregulation of the large conductance voltage- and Ca²⁺-activated K⁺ channels by Janus kinase 2

Abstract

The iberiotoxin-sensitive large conductance voltage- and Ca2+-activated potassium (BK) channels (maxi-K+-channels) hyperpolarize the cell membrane thus supporting Ca2+ entry through Ca2+-release activated Ca2+ channels. Janus kinase-2 (JAK2) has been identified as novel regulator of ion transport. To explore whether JAK2 participates in the regulation of BK channels, cRNA encoding Ca2+-insensitive BK channels (BKM513I+Δ899–903) was injected into Xenopus oocytes with or without cRNA encoding wild-type JAK2, gain-of-function V617FJAK2, or inactive K882EJAK2. K+ conductance was determined by dual electrode voltage clamp and BK-channel protein abundance by confocal microscopy. In A204 alveolar rhabdomyosarcoma cells, iberiotoxin-sensitive K+ current was determined utilizing whole cell patch clamp. A204 cells were further transfected with JAK2 and BK-channel transcript, and protein abundance was quantified by RT-PCR and Western blotting, respectively. As a result, the K+ current in BKM513I+Δ899–903-expressing oocytes was significantly increased following coexpression of JAK2 or V617FJAK2 but not K882EJAK2. Coexpression of the BK channel with V617FJAK2 but not K882EJAK2 enhanced BK-channel protein abundance in the oocyte cell membrane. Exposure of BK-channel and V617FJAK2-expressing oocytes to the JAK2 inhibitor AG490 (40 μM) significantly decreased K+ current. Inhibition of channel insertion by brefeldin A (5 μM) decreased the K+ current to a similar extent in oocytes expressing the BK channel alone and in oocytes expressing the BK channel and V617FJAK2. The iberiotoxin (50 nM)-sensitive K+ current in rhabdomyosarcoma cells was significantly decreased by AG490 pretreatment (40 μM, 12 h). Moreover, overexpression of JAK2 in A204 cells significantly enhanced BK channel mRNA and protein abundance. In conclusion, JAK2 upregulates BK channels by increasing channel protein abundance in the cell membrane.

Biophysical Characterization and Expression Analysis of Kv1.3 Potassium Channel in Primary Human Leukemic B Cells

“…Several kinases have been shown to be implicated in regulation of potassium channel protein abundance at the level of the PM. For example Janus kinase in the case of calcium-dependent potassium channels [35], mTOR kinase in the case of Kv1.3 and Kv1.5 [36] and the serine/threonine protein kinase B-RAF in the case of HERG [26] impact channel expression. Here we show that B-RAF signalling contributes to the observed increase of Kv1.3 expression in the pathologic B cells.…”

BK channel activators and their therapeutic perspectives

“…Moreover co-assembly with auxillary subunits including β1-4 and leucine-rich repeat-containing proteins (LRRC) changes the channels calcium- and voltage-sensitivity, as well as pharmacological properties (Knaus et al, 1994 ; McManus et al, 1995 ; Valverde et al, 1999 ; Wallner et al, 1999 ; Meera et al, 2000 ; Yan and Aldrich, 2010 , 2012 ). On the post-translational level, channel activity is regulated by a number of endogenous mediators such as arachidonic acids, NO, pH (Avdonin et al, 2003 ), zinc (Hou et al, 2010 ), and phosphorylation of the channel (Yan et al, 2008 ) by protein kinase A, C, G, and CaMKII (Hou et al, 2009 ; Wang et al, 2013 ; Hosseinzadeh et al, 2014 ; Toro et al, 2014 ). Other modifications such as palmitoylation favor cell surface expression, while myristoylation appear to favor endocytosis (Jeffries et al, 2010 ; Alioua et al, 2011 ; Tian et al, 2012 ).…”

Up-Regulation of the Large-Conductance Ca2+-Activated K+ Channel by Glycogen Synthase Kinase GSK3β

“…The Ca 2+ -insensitive BK mutant was utilized because the activity of wild-type BK requires an increase in the intracellular Ca 2+ level in oocytes, which leads to likely side effects interfering with the measurement [41,42].…”