Both EGFR kinase and Src-related tyrosine kinases regulate human ether-à-go-go-related gene potassium channels

Human ether-à-go-go-related gene (hERG or Kv11.1) encodes the rapidly activated delayed rectifier K+ current (IKr) in the human heart. Potential regulation of hERG channel by protein tyrosine kinases (PTKs) is not understood. The present study was designed to investigate whether this channel is modulated by PTKs using whole-cell patch clamp technique, and immunoprecipitation and Western blot analysis in HEK 293 cells stably expressing hERG gene. We found that the broad-spectrum PTK inhibitor genistein (30 μM), the selective EGFR (epidermal growth factor receptor) kinase inhibitor AG556 (10 μM) and the Src-family kinase inhibitor PP2 (10 μM) remarkably inhibited hERG channel current (IhERG), and the effects were significantly countered by the protein tyrosine phosphatase (PTP) inhibitor orthovanadate (1 mM). Immunoprecipitation and Western blot analysis demonstrated that membrane protein tyrosine phosphorylation of hERG channels was reduced by genistein, AG556, and PP2. The reduction of hERG channel phosphorylation level by genistein, AG556 or PP2 was antagonized by orthovanadate. Single point mutation(s) of Y475A and/or Y611A dramatically attenuated the inhibitory effect of IhERG by PP2 and/or AG556. Our results demonstrate the novel information that IhERG is modulated not only by Src-family kinases, but also by EGFR kinases. Y475 and/or Y611 are likely the preferred phosphorylation sites. Regulation of hERG channels by PTKs modifies the channel activity and thus likely alters electrophysiological properties including action potential duration and cell excitability in human heart and neurons.