Differential Regulation of hERG Current and Expression by Activation of Protein Kinase C Using Phorbol Ester Treatment
The human ether-à-go-go-related gene (hERG) encodes the pore-forming alpha subunit of the channel that conducts the rapidly activating delayed rectifier potassium current (IKr) in the heart. Reductions in IKr cause long QT syndrome (LQTS), which predisposes individuals to potentially fatal arrhythmias that can be triggered by stress. One potential link between stress and hERG function is protein kinase C (PKC) activation. However, PKC regulation of hERG is complex and seemingly conflicting results have been reported. In the present study, both acute and chronic effects of PKC activation were investigated using phorbol 12-myristate 13-acetate (PMA) on hERG channels expressed in human embryonic kidney (HEK) 293 cells. Western blot analyses demonstrate that chronic PKC activation increases expression of intracellular and membrane-bound hERG protein. However, the increased channel abundance is accompanied by a decrease in hERG current (IhERG) after chronic PMA treatment. Furthermore, patch clamp data reveal that acute PKC activation reduces IhERG, and this effect is dependent on the presence of the N-terminus of the channel. Upon truncation of the N-terminus of hERG, chronic activation of PKC increases both hERG protein expression and current. The increase in hERG protein is partially mediated by reduced degradation of mature hERG channels. This results from increased phosphorylation of neural precursor cell expressed developmentally down-regulated protein 4 subtype 2 (Nedd4-2), an E3 ubiquitin ligase that mediates hERG degradation. These findings demonstrate that PKC regulates hERG in a balanced manner, increasing expression while decreasing current.