Analysis of phosphorylation-dependent modulation of Kv1.1 potassium channels.
The voltage-gated potassium channel Kv1.1 contains phosphorylation sites for protein kinase A (PKA) and protein kinase C (PKC). To study Kv1.1 protein expression and cellular distribution in regard to its level of phosphorylation, the effects of PKA and PKC activation on Kv1.1 were investigated in HEK 293 cells stably transfected with Kv1.1 (HEK 293/1). Without kinase activation, HEK 293/1 cells carry unphosphorylated Kv1.1 protein in the plasma membranes, whereas large amounts of phosphorylated and unphosphorylated Kv1.1 protein were located intracellularly. Activation of PKA resulted in phosphorylation of intracellular Kv1.1 protein, followed by a rapid translocation of Kv1.1 into the plasma membrane. Patch-clamp analysis revealed an increase in current amplitude upon PKA activation and demonstrated differences in the voltage dependence of current activation between unphosphorylated and phosphorylated Kv1.1 channels. In contrast to PKA, even prolonged activation of PKC did not lead to direct phosphorylation of Kv1.1, but induced Kv1.1 protein synthesis. Thus, protein kinases have direct and indirect effects on the functional expression of voltage-gated potassium channels. Our data suggest that the synergistic action of protein kinases may play an important role in the fine-tuning of Kv channel function.
Winklhofer
M
M
Institute of Physiological Chemistry, Johannes Gutenberg-University, Mainz, Germany.
Matthias
K
K
Seifert
G
G
Stocker
M
M
Sewing
S
S
Herget
T
T
Steinhäuser
C
C
Saaler-Reinhardt
S
S
D000911:Antibodies, Monoclonal; D002460:Cell Line; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D004789:Enzyme Activation; D006801:Humans; D015640:Ion Channel Gating; D007700:Kinetics; D051662:Kv1.1 Potassium Channel; D018408:Patch-Clamp Techniques; D010766:Phosphorylation; D015221:Potassium Channels / Q000276:immunology / Q000378:metabolism* / Q000502:physiology; D024642:Potassium Channels, Voltage-Gated; D011493:Protein Kinase C / Q000378:metabolism; D021381:Protein Transport
D000911:Antibodies, Monoclonal; C496946:KCNA1 protein, human; D015221:Potassium Channels; D024642:Potassium Channels, Voltage-Gated; D051662:Kv1.1 Potassium Channel; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C