Epidermal growth factor receptor tyrosine kinase regulates the human inward rectifier potassium K IR2.3 channel, stably expressed in HEK 293 cells

Abstract

Background and Purpose The detailed molecular modulation of inward rectifier potassium channels (including the K IR2.3 channel) is not fully understood. The present study was designed to determine whether human K IR2.3 (K IR2.3) channels were regulated by protein tyrosine kinases (PTKs). Experimental Approach 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. Key Results The broad-spectrum PTK inhibitor genistein (10 ÂμM) and the selective epidermal growth factor (EGF) kinase inhibitor AG556 (10 ÂμM) reversibly decreased K IR2.3 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 IR2.3 current, this effect was antagonized by AG556. However, the Src-family tyrosine kinase inhibitor PP2 (10 ÂμM) did not inhibit K IR2.3 current. Tyrosine phosphorylation of K IR2.3 channels was decreased by genistein or AG556, and was increased by EGF or orthovanadate. The decrease of tyrosine phosphorylation of K IR2.3 channels by genistein or AG556 was reversed by orthovanadate or EGF. Interestingly, the response of K IR2.3 channels to EGF or AG556 was lost in the K IR2.3 Y234A mutant channel. Conclusion and Implications These results demonstrate that the EGF receptor tyrosine kinase up-regulates the K IR2.3 channel via phosphorylation of the Y234 residue of the WT protein. This effect may be involved in the endogenous regulation of cellular electrical activity. © 2011 The British Pharmacological Society.