A mutation in the pore region of HERG K<sup>+</sup> channels expressed in <i>Xenopus</i> oocytes reduces rectification by shifting the voltage dependence of inactivation

Zou, Anruo; Xu, Qingyang; Sanguinetti, Michael C.

doi:10.1111/j.1469-7793.1998.129bo.x

Cited by 123 publications

(132 citation statements)

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“…8G and Supplemental Table 5). In the hERG channel, inactivation is not directly coupled to activation 15,16 , suggesting that the observed effect of the L666C S6 T (−3) peptide on the inactivation curve is not a consequence of a modification of activation. Of note, S6 T peptides bind to hERG S4-S5 L (Y542-F557) which is surrounded by regions that are involved in inactivation: double mutant cycle analysis suggests that interaction between L529, L532, V535 in S4 and I560, L564, I567 in S5 play a role in inactivation gating 17 .…”

Section: Resultsmentioning

confidence: 99%

hERG S4-S5 linker acts as a voltage-dependent ligand that binds to the activation gate and locks it in a closed state

Malak¹,

Es‐Salah‐Lamoureux²,

Loussouarn³

2017

Archives of Cardiovascular Diseases Supplements

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Delayed-rectifier potassium channels (hERG and KCNQ1) play a major role in cardiac repolarization. These channels are formed by a tetrameric pore (S5-S6) surrounded by four voltage sensor domains (S1-S4). Coupling between voltage sensor domains and the pore activation gate is critical for channel voltage-dependence. However, molecular mechanisms remain elusive. Herein, we demonstrate that covalently binding, through a disulfide bridge, a peptide mimicking the S4-S5 linker (S4-S5 L ) to the channel S6 C-terminus (S6 T ) completely inhibits hERG. This shows that channel S4-S5 L is sufficient to stabilize the pore activation gate in its closed state. Conversely, covalently binding a peptide mimicking S6 T to the channel S4-S5 L prevents its inhibiting effect and renders the channel almost completely voltage-independent. This shows that the channel S4-S5 L is necessary to stabilize the activation gate in its closed state. Altogether, our results provide chemical evidence that S4-S5 L acts as a voltagecontrolled ligand that binds S6 T to lock the channel in a closed state, elucidating the coupling between voltage sensors and the gate in delayed rectifier potassium channels and potentially other voltagegated channels.Voltage-dependent ion channels are ubiquitously expressed in human tissues. They perform a plethora of physiological functions such as generation and modulation of the electrical activity in excitable tissues, modulation of neurotransmitter and hormone release, and electrolyte transport in epithelia. Canonical voltage-gated ion channels are tetramers of subunits containing six transmembrane segments (S1 to S6). Each of the four subunits is composed of one voltage sensor domain (S1 to S4) and a pore domain (S5-S6). The four pore domains tetramerize to generate a single pore module, which is regulated by the four voltage sensor domains 1, 2 . One key question remains: how do the voltage sensor domains regulate pore gating? Several independent approaches support the idea that interaction between the S4-S5 linker (S4-S5 L ) and the S6 C-terminal (S6 T ) part plays a critical role in that regulation [2][3][4][5] . First, Lu and collaborators pinpointed the sequence complementarity between S4-S5 L and S6 T in the Shaker channel 4,5 . Second, structural data indicated the close proximity between S4-S5 L and S6 T and suggested a mechanical lever model to explain the coupling between the voltage sensor and the gate of the related K v 1.2 channel 2 . However, the mechanism may prove to be more complex. Since 2006, several works on other eukaryotic K v channels suggested state-dependent S4-S5 L and S6 T interactions 3,[6][7][8][9][10] , but the precise mechanism remains elusive and may vary from one channel to another.In a previous study, we suggested that the voltage dependence of the cardiac voltage-gated KCNQ1 channel (K v 7.1) follows a ligand/receptor model. In this model, S4-S5 L is a ligand whose interaction with S6 T is only possible at resting potentials, and this interaction locks the channel in a closed s...

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Section: Resultsmentioning

confidence: 99%

hERG S4-S5 linker acts as a voltage-dependent ligand that binds to the activation gate and locks it in a closed state

Malak¹,

Es‐Salah‐Lamoureux²,

Loussouarn³

2017

Archives of Cardiovascular Diseases Supplements

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“…S631A hERG1 channels have impaired inactivation gating (22). As expected for an inactivation-deficient channel, RPR (3 M) slowed the rate of deactivation, but did not alter the magnitude of S631A channel currents (Fig.…”

Section: Paired Point Mutations Eliminate Effects Of Rpr On Inactivatmentioning

confidence: 93%

Structural basis of action for a human ether-a-go-go-related gene 1 potassium channel activator

Perry

Sachse

Sanguinetti

2007

Proc. Natl. Acad. Sci. U.S.A.

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Activation of human ether-a-go-go-related gene 1 (hERG1) K ؉ channels mediates cardiac action potential repolarization. Drugs that activate hERG1 channels represent a mechanism-based approach for the treatment of long QT syndrome, a disorder of cardiac repolarization associated with ventricular arrhythmia and sudden death. Here, we characterize the mechanisms of action and the molecular determinants for binding of RPR260243 [(3R,4R)-4-[3-(6-methoxy-quinolin-4-yl)-3-oxo-propyl]-1-[3-(2,3,5-trifluorophenyl)-prop-2-ynyl]-piperidine-3-carboxylic acid] (RPR), a recently discovered hERG1 channel activator. Channels were heterologously expressed in Xenopus laevis oocytes, and currents were measured by using the two-microelectrode voltage-clamp technique. RPR induced a concentration-dependent slowing in the rate of channel deactivation and enhanced current magnitude by shifting the voltage dependence of inactivation to more positive potentials. This mechanism was confirmed by demonstrating that RPR slowed the rate of deactivation, but did not increase current magnitude of inactivation-deficient mutant channels. The effects of RPR on hERG1 kinetics and magnitude could be simulated by reducing three rate constants in a Markov model of channel gating. Point mutations of specific residues located in the S4 -S5 linker or cytoplasmic ends of the S5 and S6 domains greatly attenuated or ablated the effects of 3 M RPR on deactivation (five residues), inactivation (one residue), or both gating mechanisms (four residues). These findings define a putative binding site for RPR and confirm the importance of an interaction between the S4 -S5 linker and the S6 domain in electromechanical coupling of voltage-gated K ؉ channels.voltage clamp ͉ Xenopus ͉ long QT syndrome H uman ether-a-go-go-related gene 1 (hERG1) ␣-subunits coassemble to form channels that conduct I Kr (1-3), the rapid delayed rectifier K ϩ current that contributes to normal repolarization of cardiac action potentials (4). Loss-of-function mutations in hERG1 cause inherited long QT syndrome (LQTS), a disorder characterized by delayed repolarization of ventricular action potentials and prolonged QT interval of the body surface electrocardiogram (5). The acquired form of LQTS is more common and is most often caused by unintended block of hERG1 channels by a plethora of common medications (6). Inherited and acquired LQTS are associated with an increased risk of torsades de pointes, an arrhythmia that can degenerate into ventricular fibrillation and cause sudden death (7).Current treatments for inherited LQTS include the administration of ␤-adrenergic receptor blockers, left cardiac sympathetic denervation, or implantation of cardiac defibrillators for the most severe cases (8). However, pharmacologic treatment is not always effective (9) and surgery or devices are expensive and require invasive procedures. Acute episodes of drug-induced LQTS are treated with magnesium sulfate administration and discontinued use of the suspect medication. Activation of hERG1 could provide an...

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“…The voltage dependence of hERG inactivation is shifted nearly Ϫ65 mV relative to channel activation (V 0.5INACT ϭ Ϫ85 mV, V 0.5ACT ϭ Ϫ20 mV). A single mutation (S631A) in the P-loop of the outer pore causes a ϩ100 mV shift in the voltage dependence of inactivation, but no shift in activation compared with wild-type (WT) channels (6,8). Furthermore, external cations have differential effects on the voltage dependence of activation and inactivation (9 -11).…”

Section: The Human Ether-a'-go-go-related Gene (Herg)mentioning

confidence: 99%

Regional Specificity of Human ether-a'-go-go-related Gene Channel Activation and Inactivation Gating

Piper

Hinz

Tallurri

et al. 2005

Journal of Biological Chemistry

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Slow activation and rapid C-type inactivation produce inward rectification of the current-voltage relationship for human ether-a'-go-go-related gene (hERG) channels. To characterize the voltage sensor movement associated with hERG activation and inactivation, we performed an Ala scan of the 32 amino acids (Gly 514 -Tyr 545 ) that comprise the S4 domain and the flanking S3-S4 and S4 -S5 linkers. Gating and ionic currents of wild-type and mutant channels were measured using cut-open oocyte Vaseline gap and two microelectrode voltage clamp techniques to determine the voltage dependence of charge movement, activation, and inactivation. Mapping the position of the charge-perturbing mutations (defined as ͦ⌬⌬Gͦ > 1.0 kcal/mol) on a threedimensional S4 homology model revealed a spiral pattern. As expected, mutation of these residues also altered activation. However, mutation of residues in the S3-S4 and S4 -S5 linkers and the C-terminal end of S4 perturbed activation (ͦ⌬⌬Gͦ > 1.0 kcal/mol) without altering charge movement, suggesting that the native residues in these regions couple S4 movement to the opening of the activation gate or stabilize the open or closed state of the channel. Finally, mutation of a distinct set of residues impacted inactivation and mapped to a single face of the S4 helix that was devoid of activation-perturbing residues. These results define regions on the S4 voltage sensor that contribute differentially to hERG activation and inactivation gating. The human ether-a'-go-go-related gene (hERG)1 channels are primarily expressed in the brain and the heart (1) but are also up-regulated in tumors from a variety of tissues (2). In the heart, hERG channels conduct the rapidly activating, delayed rectifier potassium current, I Kr (3, 4). Unlike most voltagegated K ϩ (Kv) channels, the fully activated current-voltage relationship of hERG channels exhibits inward rectification, a property that limits efflux of K ϩ during the plateau phase of the cardiac action potential.Inward rectification of hERG channels results from a rapid and voltage-dependent C-type inactivation that proceeds at a rate much faster than activation. At membrane potentials between Ϫ20 and ϩ20 mV, channels activate over hundreds of milliseconds but inactivate in milliseconds (5-7). The voltage dependence of hERG inactivation is shifted nearly Ϫ65 mV relative to channel activation (V 0.5INACT ϭ Ϫ85 mV, V 0.5ACT ϭ Ϫ20 mV). A single mutation (S631A) in the P-loop of the outer pore causes a ϩ100 mV shift in the voltage dependence of inactivation, but no shift in activation compared with wild-type (WT) channels (6, 8). Furthermore, external cations have differential effects on the voltage dependence of activation and inactivation (9 -11). Together, these findings led to the suggestion that distinct voltage-sensing mechanisms underlie activation and inactivation gating in hERG channels.The highly charged S4 transmembrane helix functions as the primary voltage sensor in voltage-gated channels. Neutralization of basic S4 residues shifts the vol...

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A mutation in the pore region of HERG K⁺ channels expressed in Xenopus oocytes reduces rectification by shifting the voltage dependence of inactivation

Cited by 123 publications

References 32 publications

hERG S4-S5 linker acts as a voltage-dependent ligand that binds to the activation gate and locks it in a closed state

hERG S4-S5 linker acts as a voltage-dependent ligand that binds to the activation gate and locks it in a closed state

Structural basis of action for a human ether-a-go-go-related gene 1 potassium channel activator

Regional Specificity of Human ether-a'-go-go-related Gene Channel Activation and Inactivation Gating

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A mutation in the pore region of HERG K+ channels expressed in Xenopus oocytes reduces rectification by shifting the voltage dependence of inactivation

Cited by 123 publications

References 32 publications

hERG S4-S5 linker acts as a voltage-dependent ligand that binds to the activation gate and locks it in a closed state

hERG S4-S5 linker acts as a voltage-dependent ligand that binds to the activation gate and locks it in a closed state

Structural basis of action for a human ether-a-go-go-related gene 1 potassium channel activator

Regional Specificity of Human ether-a'-go-go-related Gene Channel Activation and Inactivation Gating

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A mutation in the pore region of HERG K⁺ channels expressed in Xenopus oocytes reduces rectification by shifting the voltage dependence of inactivation