Identification of Retrieval-Trafficking of the Human Ether A-Go-Go-Related Gene Channel

Abstract

The human ether-a-go-go related gene (hERG) encodes the pore-forming subunit of the rapidly activating delayed rectifier potassium channel (IKr). A reduction in the hERG current causes long QT syndrome, predisposing affected individuals to a high risk of cardiac arrhythmias and sudden death. We previously reported that hERG channels in the plasma membrane undergo internalization under low K+ conditions. While studies have characterized the synthesis and degradation pathways that maintain the homeostasis of hERG density in the plasma membrane, whether internalization occurs under normal K+ conditions and whether internalized channels can be recycled to the plasma membrane are not addressed. Using patch clamp, Western blot and immunocytochemical analyses, we studied the retrieval trafficking of internalized hERG channels to the plasma membrane. Our data demonstrated that an enhanced internalization is accompanied by an increased rate of recovery of hERG channels to the plasma membrane. The increased recovery rate is not due to enhanced protein synthesis, since hERG mRNA expression level was not altered by low K+ exposure, and the increased recovery was not affected by the inhibition of protein synthesis using cycloheximide. Disrupting the trans-Golgi network with brefeldin A (BFA) blocked not only hERG maturation, but also the retrieval trafficking. Given these findings, we were interested to identify which Rab GTPase proteins were involved in this recycling pathway, since these proteins are known to regulate trafficking. Our study demonstrated that GTPase Rab11, but not Rab4, is involved in the retrieval trafficking of hERG channels. Furthermore, interfering with Rab11 function not only delayed hERG recovery after exposure to low K+ medium, but led to decreased hERG expression and function under normal culture conditions. We conclude that the retrieval trafficking plays an important role in the homeostasis of membrane-bound hERG channels.