title: Modulation of a cloned human A-type voltage-gated potassium channel (hKv1.4) by the protein tyrosine kinase inhibitor genistein.
abstract: A cloned, human, A-type, voltage-gated potassium channel (hKv1.4) was expressed transiently in Chinese hamster ovary cells and the effects of the broad-spectrum tyrosine kinase inhibitor genistein on hKv1.4 were studied using the whole-cell patch-clamp recording method. Genistein (up to 50 microM) reversibly reduced the peak currents of hKv1.4 by 44.9+/-12%. In addition, genistein markedly slowed the activation kinetics (time constant tau(a)) of hKv1.4. At +50 mV, tau(a) increased from 1.8+/-0.3 to 5.0+/-0.6 ms (P<0.01). The effect of genistein on the channel inactivation kinetics (time constant tau(i)) was more complex, in that tau(i) was increased significantly at lower step potentials but unaltered at +50 mV or more depolarized potentials. Tail current analysis showed that genistein had no effect on the kinetics of deactivation (time constant tau(d)), but shifted the steady-state activation curve significantly to the right by about 15 mV (potential for half-maximal activation, V1/2, changed from -7.4+/-4.4 to +7.7+/-2.7 mV) with a moderate change in the slope (k) of the curve (from 17.4+/-2.2 to 23+/-1.0 mV, P<0.05). Genistein slightly altered the slope of the steady-state inactivation curve from -5.5+/-0.4 to -7.5+/-0.4 mV (P<0.01). The recovery rate from inactivation was not altered by genistein. The tyrosine phosphatase inhibitor orthovanadate (1 mM) alone had little impact on current amplitude or channel kinetics. However, orthovanadate significantly, but not completely, blocked the effect of genistein on current amplitude (by 25.5%) and kinetics (by 67.1%). Daidzein (up to 50 microM), an inactive analogue of genistein, had no effect on current amplitude or kinetics. In contrast to genistein, another tyrosine kinase inhibitor, herbimycin A, had little effect on the channel peak amplitude or kinetics. In addition, genistein had a similar impact on the channel peak current amplitude and kinetics in cells with or without pre-treatment with herbimycin A (10 microM). The data suggest that genistein-induced inhibition of tyrosine phosphorylation may not be the exclusive mechanism by which hKv1.4 is down-regulated and channel gating affected. Genistein may produce a non-catalytic blockade of this channel.
authors.affiliation: Wyeth-Neuroscience, Neurodegenerative Diseases, Ion Channel Pharmacology, Princeton, New Jersey 08543, USA.
authors.forename: Z H
authors.initials: ZH
authors.lastname: Zhang
authors.forename: Q
authors.initials: Q
authors.lastname: Wang
mesh_terms: D000818:Animals; D016227:Benzoquinones; D016466:CHO Cells; D003001:Cloning, Molecular; D006224:Cricetinae; D004553:Electric Conductivity; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D019833:Genistein / Q000031:analogs & derivatives / Q000494:pharmacology*; D006801:Humans; D007529:Isoflavones / Q000494:pharmacology; D007700:Kinetics; D051665:Kv1.4 Potassium Channel; D047029:Lactams, Macrocyclic; D008954:Models, Biological; D015221:Potassium Channels / Q000187:drug effects* / Q000235:genetics / Q000502:physiology*; D024642:Potassium Channels, Voltage-Gated; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors*; D011809:Quinones / Q000494:pharmacology; D017828:Rifabutin / Q000031:analogs & derivatives; D014638:Vanadates / Q000494:pharmacology
chemical_list: D016227:Benzoquinones; D004791:Enzyme Inhibitors; D007529:Isoflavones; C496977:KCNA4 protein, human; D051665:Kv1.4 Potassium Channel; D047029:Lactams, Macrocyclic; D015221:Potassium Channels; D024642:Potassium Channels, Voltage-Gated; D011809:Quinones; D017828:Rifabutin; D014638:Vanadates; C004742:daidzein; C020754:herbimycin; D019833:Genistein; D011505:Protein-Tyrosine Kinases