title: Genistein and tyrphostin AG556 inhibit inwardly-rectifying Kir2.1 channels expressed in HEK 293 cells via protein tyrosine kinase inhibition.
abstract: Previous studies reported the controversial effects that protein tyrosine kinase (PTK) inhibition could induce an up-regulation or down-regulation of Kir2.1 current. The present study investigates how the recombinant human Kir2.1 channels are regulated by PTKs using whole-cell patch voltage-clamp, immunoprecipitation and Western blot, and mutagenesis approaches. We found that hKir2.1 current was reversibly inhibited by the broad spectrum PTK inhibitor genistein and the highly selective EGFR (epidermal growth factor receptor) kinase inhibitor AG556 in a concentration-dependent manner. The inhibition of hKir2.1 channels by genistein or AG556 was countered by the protein tyrosine phosphatase (PTP) inhibitor orthovanadate. Immunoprecipitation and Western blot analysis revealed that tyrosine phosphorylation level of Kir2.1 channels was reduced by genistein or AG556, and the reduction was significantly antagonized by orthovanadate. The mutation of Y242 dramatically reduced the inhibitory response to AG556. The results obtained in this study demonstrate that hKir2.1 channels are down-regulated by PTK inhibition, suggesting that EGFR kinase participates in the modulation of human cardiac excitability.
authors.affiliation: Department of Medicine, LKS Faculty of Medicine, The University of Hong Kong, Pokfulam, Hong Kong, China.
authors.forename: De-Yong
authors.initials: DY
authors.lastname: Zhang
authors.forename: Wei
authors.initials: W
authors.lastname: Wu
authors.forename: Xiu-Ling
authors.initials: XL
authors.lastname: Deng
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; D004305:Dose-Response Relationship, Drug; D066246:ErbB Receptors / Q000037:antagonists & inhibitors / Q000378:metabolism; D019833:Genistein / Q000494:pharmacology*; D057809:HEK293 Cells; D006801:Humans; D047468:Immunoprecipitation; D008564:Membrane Potentials; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D018408:Patch-Clamp Techniques; D026902:Potassium Channel Blockers / Q000494:pharmacology*; D024661:Potassium Channels, Inwardly Rectifying / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D017027:Protein Tyrosine Phosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D011743:Pyrimidines / Q000494:pharmacology; D013997:Time Factors; D014162:Transfection; D020032:Tyrphostins / Q000494:pharmacology*; D014638:Vanadates / Q000494:pharmacology; D019061:src-Family Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism
chemical_list: C412373:AG 1879; C489457:KCNJ2 protein, human; D026902:Potassium Channel Blockers; D024661:Potassium Channels, Inwardly Rectifying; D047428:Protein Kinase Inhibitors; D011743:Pyrimidines; D020032:Tyrphostins; C098981:AG 556; D014638:Vanadates; D019833:Genistein; C512478:EGFR protein, human; D066246:ErbB Receptors; D011505:Protein-Tyrosine Kinases; D019061:src-Family Kinases; D017027:Protein Tyrosine Phosphatases