Degradation of Trafficking-defective Long QT Syndrome Type II Mutant Channels by the Ubiquitin-Proteasome Pathway

Qiuming Gong,David R Keeney,Maurizio Molinari,Zhengfeng Zhou

doi:10.1074/jbc.m502327200

Abstract

Mutations in the human ether-a-go-go-related gene (hERG) cause chromosome 7-linked long QT syndrome type II (LQT2). We have shown previously that LQT2 mutations lead to endoplasmic reticulum (ER) retention and rapid degradation of mutant hERG proteins. In this study we examined the role of the ubiquitin-proteasome pathway in the degradation of the LQT2 mutation Y611H. We showed that proteasome inhibitors N-acetyl-L-leucyl-L-leucyl-L-norleucinal and lactacystin but not lysosome inhibitor leupeptin inhibited the degradation of Y611H mutant channels. In addition, ER mannosidase I inhibitor kifunensine and down-regulation of EDEM (ER degradation-enhancing alpha-mannosidase-like protein) also suppressed the degradation of Y611H mutant channels. Proteasome inhibition but not mannosidase inhibition led to the accumulation of full-length hERG protein in the cytosol. The hERG protein accumulated in the cytosol was deglycosylated. Proteasome inhibition also resulted in the accumulation of polyubiquitinated hERG channels. These results suggest that the degradation of LQT2 mutant channels is mediated by the cytosolic proteasome in a process that involves mannose trimming, polyubiquitination, and deglycosylation of mutant channels.

Highlights

Long QT syndrome is a cardiac disorder characterized by prolongation of QT intervals on the electrocardiogram and a high risk of sudden death due to ventricular arrhythmias
We have previously shown that the upper band is the fully glycosylated, mature form of the channel protein located in the plasma membrane, and the lower band is a core-glycosylated, immature form of the channel protein located in the endoplasmic reticulum (ER) [5]
The lysosome inhibitor leupeptin had no effect on protein levels of wild type human ether-a-go-go-related gene (hERG) or Y611H. These results strongly suggest that the proteasome is involved in the degradation of the immature form of hERG channels

Summary

Introduction

Long QT syndrome is a cardiac disorder characterized by prolongation of QT intervals on the electrocardiogram and a high risk of sudden death due to ventricular arrhythmias. These results suggest that the degradation of LQT2 mutant channels is mediated by the cytosolic proteasome in a process that involves mannose trimming, polyubiquitination, and deglycosylation of mutant channels. Our results suggest that the mutant hERG protein is targeted to the proteasome for degradation by dislocation from the ER membrane to the cytosol.

Results

Conclusion