title: Epidermal growth factor receptor tyrosine kinase regulates the human inward rectifier potassium K(IR)2.3 channel, stably expressed in HEK 293 cells.
abstract: OBJECTIVE
  The detailed molecular modulation of inward rectifier potassium channels (including the K(IR) 2.3 channel) is not fully understood. The present study was designed to determine whether human K(IR) 2.3 (K(IR) 2.3) channels were regulated by protein tyrosine kinases (PTKs).
  
  
  METHODS
  Whole-cell patch voltage-clamp, immunoprecipitation, Western blot analysis and site-directed mutagenesis were employed to determine the potential PTK phosphorylation of Kir2.3 current in HEK 293 cells stably expressing Kir2.3 gene.
  
  
  RESULTS
  The broad-spectrum PTK inhibitor genistein (10 µM) and the selective epidermal growth factor (EGF) kinase inhibitor AG556 (10 µM) reversibly decreased K(IR) 2.3 current and the effect was reversed by the protein tyrosine phosphatase inhibitor, orthovanadate (1 mM). Although EGF (100 ng·mL(-1) ) and orthovanadate enhanced K(IR) 2.3 current, this effect was antagonized by AG556. However, the Src-family tyrosine kinase inhibitor PP2 (10 µM) did not inhibit K(IR) 2.3 current. Tyrosine phosphorylation of K(IR) 2.3 channels was decreased by genistein or AG556, and was increased by EGF or orthovanadate. The decrease of tyrosine phosphorylation of K(IR) 2.3 channels by genistein or AG556 was reversed by orthovanadate or EGF. Interestingly, the response of K(IR) 2.3 channels to EGF or AG556 was lost in the K(IR) 2.3 Y234A mutant channel.
  
  
  CONCLUSIONS
  These results demonstrate that the EGF receptor tyrosine kinase up-regulates the K(IR) 2.3 channel via phosphorylation of the Y234 residue of the WT protein. This effect may be involved in the endogenous regulation of cellular electrical activity.
authors.affiliation: Department of Medicine, Li Ka Shing Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong SAR, China.
authors.forename: De-Yong
authors.initials: DY
authors.lastname: Zhang
authors.forename: Yan-Hui
authors.initials: YH
authors.lastname: Zhang
authors.forename: Hai-Ying
authors.initials: HY
authors.lastname: Sun
authors.forename: Chu-Pak
authors.initials: CP
authors.lastname: Lau
authors.forename: Gui-Rong
authors.initials: GR
authors.lastname: Li
mesh_terms: D015153:Blotting, Western; D004594:Electrophysiology; D066246:ErbB Receptors / Q000037:antagonists & inhibitors / Q000502:physiology*; D019833:Genistein / Q000494:pharmacology; D057809:HEK293 Cells; D006801:Humans; D047468:Immunoprecipitation; D016297:Mutagenesis, Site-Directed; D018408:Patch-Clamp Techniques; D010766:Phosphorylation; D010957:Plasmids; D024661:Potassium Channels, Inwardly Rectifying / Q000235:genetics* / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D017027:Protein Tyrosine Phosphatases / Q000037:antagonists & inhibitors; D020032:Tyrphostins / Q000494:pharmacology; D014638:Vanadates / Q000494:pharmacology
chemical_list: C099489:KCNJ4 protein, human; D024661:Potassium Channels, Inwardly Rectifying; D047428:Protein Kinase Inhibitors; D020032:Tyrphostins; C098981:AG 556; D014638:Vanadates; D019833:Genistein; C512478:EGFR protein, human; D066246:ErbB Receptors; D017027:Protein Tyrosine Phosphatases