The possible involvement of protein phosphorylation in the modulation of Kv1 channels by endothelin

It has been shown that K+ currents through Kv channels are modulated by stimulation of G-protein-coupled receptors. We previously reported that the K+ current by Kv1. 2 channel is suppressed by endothelin when cloned Kv1. 2 and ET receptors are coexpressed in Xenopus oocytes. It was also shown that the effect of endothelin is canceled in the presence of staurosporine. In this study, the mutants of Kv1. 2 were made in which putative phosphorylation sites by protein kinase C were replaced by alanine to examine the role of protein phosphorylation in the suppression of the current by endothelin. However, no currents were observed when those mutants were expressed in the oocytes. Then we made dimeric cDNAs by tandem linkage of wild and mutant Kv1. 2 to rescue the nonfunctional mutants, which showed delayed rectifier-type K+ current. We now examine the involvement of phosphorylation by protein kinase C by comparing the effects of endothelin on wild and mutant-containing channels, and have obtained preliminary results that the inhibitory effect of endothelin is more remarkable in wild than mutant-containing channels. It was suggested that suppression of Kv1. 2 currents by stimulation of M1 muscarinic cholinergic receptor could be ascribed to tyrosine phosphorylation through activation of protein kinase C. We will discuss the relative involvement of the protein kinases in modulation of Kv1. 2 by endothelin.