The Role of Kinesin I and a Small Gtpase in the Forward Trafficking of Kv1.5 Channels

Kv channels play important roles in the repolarization phase of the action potential in cardiac cells. The regulation of functional Kv1.5 surface expression has been reported to be modulated by retrograde trafficking through dynein motor but little is known about regulation by forward trafficking. Here, we use electrophysiological and immunocytochemical methods to investigate the mechanisms and regulation of anterograde trafficking of newly synthesized Kv1.5 channel proteins in cultured cells and in adult cardiomyocytes. Over-expression of a kinesin I isoform (Kif5b) increased outward K+ current by two fold in cultured cells stably expressing Kv1.5. This enhancement of Kv1.5 current by Kif5b was blocked by a six hour treatment with Brefeldin A. Over-expression of Kif5b increased Kv1.5 current additively with inhibition of endocytosis by p50 over-expression and dynamin inhibitory peptide. Deletion of a specific SH3-binding domain in Kv1.5 that is essential for internalization of the channel similarly enhanced Kif5b-induced Kv1.5 current. Expression of a dominant negative Kif5b mutant prior to induction of Kv1.5 in a tetracycline-inducible system almost completely blocked Kv1.5 current. These results demonstrate that Kif5b is required for the forward trafficking of newly synthesized Kv1.5 channel to the plasma membrane. This work has been extended to adult rat cardiomyocytes transfected with Kif5b constructs and wild-type and dominant negative Rab-type small GTPases. Results indicate that newly synthesized Kv1.5 traffics via a non-conventional pathway and on to the plasmalemma in a Kif5b-dependent process.