Background: The R582C mutation is one of many Long-QT Syndrome type 2 (LQT2)-causing mutations localized to the human ether-a-go-go related gene (hERG) channel's S5-P linker subdomain, yet its specific mechanism of dysfunction has not been examined.
Objective: This study sought to characterize the biophysical properties of the congenital LQT2-causing mutation, R582C, and utilize this mutation to provide the first report of voltage-dependent fluorescence from the S5-P linker.
Methods: Properties of the R582C channels were characterized by heterologous expression in both HEK293 cells and Xenopus oocytes using a combination of patch-clamp, 2-electrode voltage-clamp, immunoblot assay, and voltage-clamp fluorimetry.
Results: Expression of hERG R582C was found to be deficient in HEK293 cells, yet was amenable to rescue by incubation at reduced temperature or by treatment with dofetilide. Rescued channels expressed at levels comparable to wild type (WT) channels. Kinetic differences result in decreased outward repolarizing current evoked by an action potential clamp protocol. Voltage-clamp fluorimetry experiments utilized the introduced cysteine to covalently attach a fluorescent probe (tetramethylrhodamine-5-maleimide) to the S5-P linker to directly observe conformational changes occurring due to inactivation.
Conclusion: The major mechanism underlying pathogenicity of the R582C mutation is a trafficking deficiency, although channels also exhibit kinetic deficiencies, perhaps reflecting the position of the mutation in the pore turret. Voltage clamp fluorescence signals from R582C channels provide evidence that the hERG turret undergoes distinct conformational changes during inactivation.
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http://dx.doi.org/10.1016/j.hrthm.2011.02.035 | DOI Listing |
Mol Pharmacol
October 2023
Department of Biomedical and Molecular Sciences (N.C., E.W., K.L., J.G., W.L., T.Y., S.Z.); Division of Cardiology, Department of Medicine (A.B.); and Division of Cardiac Surgery, Department of Surgery (M.E.-D.), Queen's University, Kingston, Ontario, Canada
The encodes for the pore-forming subunit of the channel that conducts the rapidly activating delayed K current (I) in the heart. The hERG channel is important for cardiac repolarization, and reduction of its expression in the plasma membrane due to mutations causes long QT syndrome type 2 (LQT2). As such, promoting hERG membrane expression is a strategy to rescue mutant channel function.
View Article and Find Full Text PDFPhysiol Rep
July 2022
Natural and Medical Sciences Research Center, University of Nizwa, Nizwa, Oman.
Long QT syndrome type II (LQT2) is caused by loss-of-function mutations in the hERG K channel, leading to increased incidence of cardiac arrest and sudden death. Many genetic variants have been reported in the hERG gene with various consequences on channel expression, permeation, and gating. Only a small number of LQT2 causing variants has been characterized to define the underlying pathophysiological causes of the disease.
View Article and Find Full Text PDFFront Physiol
December 2021
Department of Anatomy and Embryology, Leiden University Medical Center, Leiden, Netherlands.
While rare mutations in ion channel genes are primarily responsible for inherited cardiac arrhythmias, common genetic variants are also an important contributor to the clinical heterogeneity observed among mutation carriers. The common single nucleotide polymorphism (SNP) KCNH2-K897T is associated with QT interval duration, but its influence on the disease phenotype in patients with long QT syndrome type 2 (LQT2) remains unclear. Human induced pluripotent stem cells (hiPSCs), coupled with advances in gene editing technologies, are proving an invaluable tool for modeling cardiac genetic diseases and identifying variants responsible for variability in disease expressivity.
View Article and Find Full Text PDFBiomolecules
August 2020
Department of Physiology, Cardiovascular Research Center, Center for Muscle Biology, University of Kentucky, Lexington, KY 40536, USA.
Significant advances in our understanding of the molecular mechanisms that cause congenital long QT syndrome (LQTS) have been made. A wide variety of experimental approaches, including heterologous expression of mutant ion channel proteins and the use of inducible pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) from LQTS patients offer insights into etiology and new therapeutic strategies. This review briefly discusses the major molecular mechanisms underlying LQTS type 2 (LQT2), which is caused by loss-of-function (LOF) mutations in the gene (also known as the human ether-à-go-go-related gene or ).
View Article and Find Full Text PDFBiophys J
March 2015
Department of Anesthesiology, Pharmacology and Therapeutics, University of British Columbia, Vancouver, British Columbia, Canada. Electronic address:
KV11.1 voltage-gated K(+) channels are noted for unusually slow activation, fast inactivation, and slow deactivation kinetics, which tune channel activity to provide vital repolarizing current during later stages of the cardiac action potential. The bulk of charge movement in human ether-a-go-go-related gene (hERG) is slow, as is return of charge upon repolarization, suggesting that the rates of hERG channel opening and, critically, that of deactivation might be determined by slow voltage sensor movement, and also by a mode-shift after activation.
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