Physiological Properties of hERG 1a/1b Heteromeric Currents and a hERG 1b-Specific Mutation Associated With Long-QT Syndrome

Abstract

Abstract-Cardiac I Kr is a critical repolarizing current in the heart and a target for inherited and acquired long-QT syndrome (LQTS). Biochemical and functional studies have demonstrated that I Kr channels are heteromers composed of both hERG 1a and 1b subunits, yet our current understanding of I Kr functional properties derives primarily from studies of homooligomers of the original hERG 1a isolate. Here, we examine currents produced by hERG 1a and 1a/1b channels expressed in HEK-293 cells at near-physiological temperatures. We find that heteromeric hERG 1a/1b currents are much larger than hERG 1a currents and conduct 80% more charge during an action potential. This surprising difference corresponds to a 2-fold increase in the apparent rates of activation and recovery from inactivation, thus reducing rectification and facilitating current rebound during repolarization. Kinetic modeling shows these gating differences account quantitatively for the differences in current amplitude between the 2 channel types. Drug sensitivity was also different. Compared to homomeric 1a channels, heteromeric 1a/1b channels were inhibited by E-4031 with a slower time course and a corresponding 4-fold shift in the IC 50 . The importance of hERG 1b in vivo is supported by the identification of a 1b-specific A8V missense mutation in 1/269 unrelated genotype-negative LQTS patients that was absent in 400 control alleles. Mutant 1bA8V expressed alone or with hERG 1a in HEK-293 cells dramatically reduced 1b protein levels. Thus, mutations specifically disrupting hERG 1b function are expected to reduce cardiac I Kr and enhance drug sensitivity, and represent a potential mechanism underlying inherited or acquired LQTS. (Circ Res. 2008;103:e81-e95.)Key Words: Kv11.1 Ⅲ KCNH2 Ⅲ ether-à-go-go Ⅲ arrhythmia Ⅲ potassium channels C ardiac I Kr is a potassium current contributing to ventricular repolarization in mammalian heart. 1,2 The molecular basis of cardiac I Kr was first elucidated when its unique biophysical and pharmacological properties were largely reproduced by heterologous expression of the hERG1 gene (human ether-à-go-go-related gene) (or KCNH2). 3,4 Together with the discovery of KCNH2 mutations as the pathogenic substrate in families with type 2 long-QT syndrome (LQTS), 5 these studies explained the underlying cause of disease as a loss of cardiac I Kr . They also identified hERG 1 channels as a molecular target for acquired LQTS, a much more prevalent form of the disease arising from I Kr block primarily by drugs intended for other therapeutic targets. 6 In either manifestation, LQTS is characterized by prolonged ventricular action potentials and a susceptibility to potentially life-threatening arrhythmias known as torsades de pointes (TdP). 7 Our understanding of how I Kr contributes to ventricular repolarization is based primarily on heterologous expression of the originally identified hERG 1a subunit. 3,4,8,9 Like other voltage-gated potassium channels, hERG 1a channels activate and inactivate on depolarization. However, b...

Properties of WT and mutant hERG K+ channels expressed in neonatal mouse cardiomyocytes

“…Consistent differences were also found in the deactivation rates of both WT and ⌬Y475 hERG between cardiomyocytes and HEK-293 cells, with the decay rates being 1.5-to 2-fold faster in cardiomyocytes. Possible explanations include differences in cell signaling pathways (22), membrane structure and lipid content (6), cytoskeleton interactions (10), ␣-subunit coassembly with ␤-subunits (e.g., KCNE2) (29), and that the neonatal mouse heart expresses the ERG1b (mERG1b) isoform that accelerates the deactivation kinetics of mERG1a (25,32,33). One implication of the faster deactivation kinetics found in cardiomyocyte expression is that occupancy of the open state of the hERG channel will be reduced during a cardiac action potential, and this may need to be considered in computational models of cardiac action potentials that use data from heterologous expression systems.…”

hERG 1b is critical for human cardiac repolarization

“…Surprisingly, the changes in gating also lead to differences in drug sensitivities (IC 50 values) by as much as eightfold (12). Overall, the characteristics of 1a/1b current properties appear to better resemble those of native I Kr (11,13,14).Heteromeric 1a/1b currents can be converted to 1a-like currents by coexpressing a fragment representing the PAS domain (15). The PAS domain interacts directly with the heteromeric channel, presumably occupying an empty PAS domain receptor site left available by the abbreviated hERG 1b N terminus (15, 16).…”

“…Homomeric 1a currents are unaffected by the PAS fragments, as expected if all PAS receptor sites are occupied. Thus, these differences in gating kinetics arise because the 1b N terminus lacks the PAS domain and not because of an effect conferred by the unique N-terminal sequence of 1b (15).Both 1a and 1b subunits are expressed in human ventricle (17), but to date, genetic evidence for 1b-specific disease mutations remains limited to two clinical cases (11,18). Thus, direct evidence for a functional role of hERG 1b in human cardiomyocytes is lacking.…”

“…Both 1a and 1b subunits are expressed in human ventricle (17), but to date, genetic evidence for 1b-specific disease mutations remains limited to two clinical cases (11,18). Thus, direct evidence for a functional role of hERG 1b in human cardiomyocytes is lacking.…”

“…Compared with homomeric 1a currents, hERG 1a/1b currents exhibit a twofold increase in the rates of activation, recovery from inactivation, and deactivation. During a voltage-clamp command mimicking a cardiac action potential (AP), these gating differences result in a nearly twofold increase in the repolarizing current integral (11). Surprisingly, the changes in gating also lead to differences in drug sensitivities (IC 50 values) by as much as eightfold (12).…”