Arrhythmogenic calmodulin mutations impede activation of small-conductance calcium-activated potassium current

Chih Chieh Yu, Jum Suk Ko, Tomohiko Ai, Wen Chin Tsai, Zhenhui Chen, Michael Rubart-von der Lohe, Matteo Vatta, Thomas Everett, Alfred L. George, Peng-Sheng Chen

Research output: Contribution to journalArticle

7 Citations (Scopus)

Abstract

Background: Apamin-sensitive small-conductance calcium-activated potassium (SK) channels are gated by intracellular Ca2+ through a constitutive interaction with calmodulin. Objective: We hypothesize that arrhythmogenic human calmodulin mutations impede activation of SK channels. Methods: We studied 5 previously published calmodulin mutations (N54I, N98S, D96V, D130G, and F90L). Plasmids encoding either wild-type or mutant calmodulin were transiently transfected into human embryonic kidney 293 cells that stably express subtype 2 of SK protein channels (SK2 cells). Whole-cell voltage-clamp recording was used to determine apamin-sensitive current densities. We also performed optical mapping studies in normal murine hearts to determine the effects of apamin in hearts with (n=7) or without (n=3) pretreatment with sea anemone toxin. Results: SK2 cells transfected with wild-type calmodulin exhibited an apamin-sensitive current density of 33.6 pA/pF (31.4-36.5 pA/pF) (median and confidence interval 25th-75th percentile), which was significantly higher than that observed for cells transfected with N54I (17.0 pA/pF [14.0-27.7 pA/pF]; P = .016), F90L (22.6 pA/pF [20.3-24.3 pA/pF]; P = .011), D96V (13.0 pA/pF [10.9-15.8 pA/pF]; P = .003), N98S (13.7 pA/pF [8.8-20.4 pA/pF]; P = .005), and D130G (17.6 pA/pF [13.8-24.6 pA/pF]; P = .003). The decrease in SK2 current densities was not associated with a decrease in membrane protein expression or intracellular distribution of the channel protein. Apamin increased the ventricular action potential duration at 80% repolarization (from 79.6 ms [63.4-93.3 ms] to 121.8 ms [97.9-127.2 ms]; P = .010) in hearts pretreated with anemone toxin but not in control hearts. Conclusion: Human arrhythmogenic calmodulin mutations impede the activation of SK2 channels in human embryonic kidney 293 cells.

Original languageEnglish (US)
JournalHeart Rhythm
DOIs
StateAccepted/In press - 2016

Fingerprint

Apamin
Calmodulin
Potassium
Calcium
Mutation
Small-Conductance Calcium-Activated Potassium Channels
Anemone
Sea Anemones
Kidney
Action Potentials
Membrane Proteins
Proteins
Plasmids
Confidence Intervals

Keywords

  • Arrhythmias
  • Catecholaminergic polymorphic ventricular tachycardia
  • Ion channels
  • Long QT syndrome
  • Patch clamp

ASJC Scopus subject areas

  • Medicine(all)
  • Cardiology and Cardiovascular Medicine
  • Physiology (medical)

Cite this

Arrhythmogenic calmodulin mutations impede activation of small-conductance calcium-activated potassium current. / Yu, Chih Chieh; Ko, Jum Suk; Ai, Tomohiko; Tsai, Wen Chin; Chen, Zhenhui; Rubart-von der Lohe, Michael; Vatta, Matteo; Everett, Thomas; George, Alfred L.; Chen, Peng-Sheng.

In: Heart Rhythm, 2016.

Research output: Contribution to journalArticle

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title = "Arrhythmogenic calmodulin mutations impede activation of small-conductance calcium-activated potassium current",
abstract = "Background: Apamin-sensitive small-conductance calcium-activated potassium (SK) channels are gated by intracellular Ca2+ through a constitutive interaction with calmodulin. Objective: We hypothesize that arrhythmogenic human calmodulin mutations impede activation of SK channels. Methods: We studied 5 previously published calmodulin mutations (N54I, N98S, D96V, D130G, and F90L). Plasmids encoding either wild-type or mutant calmodulin were transiently transfected into human embryonic kidney 293 cells that stably express subtype 2 of SK protein channels (SK2 cells). Whole-cell voltage-clamp recording was used to determine apamin-sensitive current densities. We also performed optical mapping studies in normal murine hearts to determine the effects of apamin in hearts with (n=7) or without (n=3) pretreatment with sea anemone toxin. Results: SK2 cells transfected with wild-type calmodulin exhibited an apamin-sensitive current density of 33.6 pA/pF (31.4-36.5 pA/pF) (median and confidence interval 25th-75th percentile), which was significantly higher than that observed for cells transfected with N54I (17.0 pA/pF [14.0-27.7 pA/pF]; P = .016), F90L (22.6 pA/pF [20.3-24.3 pA/pF]; P = .011), D96V (13.0 pA/pF [10.9-15.8 pA/pF]; P = .003), N98S (13.7 pA/pF [8.8-20.4 pA/pF]; P = .005), and D130G (17.6 pA/pF [13.8-24.6 pA/pF]; P = .003). The decrease in SK2 current densities was not associated with a decrease in membrane protein expression or intracellular distribution of the channel protein. Apamin increased the ventricular action potential duration at 80{\%} repolarization (from 79.6 ms [63.4-93.3 ms] to 121.8 ms [97.9-127.2 ms]; P = .010) in hearts pretreated with anemone toxin but not in control hearts. Conclusion: Human arrhythmogenic calmodulin mutations impede the activation of SK2 channels in human embryonic kidney 293 cells.",
keywords = "Arrhythmias, Catecholaminergic polymorphic ventricular tachycardia, Ion channels, Long QT syndrome, Patch clamp",
author = "Yu, {Chih Chieh} and Ko, {Jum Suk} and Tomohiko Ai and Tsai, {Wen Chin} and Zhenhui Chen and {Rubart-von der Lohe}, Michael and Matteo Vatta and Thomas Everett and George, {Alfred L.} and Peng-Sheng Chen",
year = "2016",
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language = "English (US)",
journal = "Heart Rhythm",
issn = "1547-5271",
publisher = "Elsevier",

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T1 - Arrhythmogenic calmodulin mutations impede activation of small-conductance calcium-activated potassium current

AU - Yu, Chih Chieh

AU - Ko, Jum Suk

AU - Ai, Tomohiko

AU - Tsai, Wen Chin

AU - Chen, Zhenhui

AU - Rubart-von der Lohe, Michael

AU - Vatta, Matteo

AU - Everett, Thomas

AU - George, Alfred L.

AU - Chen, Peng-Sheng

PY - 2016

Y1 - 2016

N2 - Background: Apamin-sensitive small-conductance calcium-activated potassium (SK) channels are gated by intracellular Ca2+ through a constitutive interaction with calmodulin. Objective: We hypothesize that arrhythmogenic human calmodulin mutations impede activation of SK channels. Methods: We studied 5 previously published calmodulin mutations (N54I, N98S, D96V, D130G, and F90L). Plasmids encoding either wild-type or mutant calmodulin were transiently transfected into human embryonic kidney 293 cells that stably express subtype 2 of SK protein channels (SK2 cells). Whole-cell voltage-clamp recording was used to determine apamin-sensitive current densities. We also performed optical mapping studies in normal murine hearts to determine the effects of apamin in hearts with (n=7) or without (n=3) pretreatment with sea anemone toxin. Results: SK2 cells transfected with wild-type calmodulin exhibited an apamin-sensitive current density of 33.6 pA/pF (31.4-36.5 pA/pF) (median and confidence interval 25th-75th percentile), which was significantly higher than that observed for cells transfected with N54I (17.0 pA/pF [14.0-27.7 pA/pF]; P = .016), F90L (22.6 pA/pF [20.3-24.3 pA/pF]; P = .011), D96V (13.0 pA/pF [10.9-15.8 pA/pF]; P = .003), N98S (13.7 pA/pF [8.8-20.4 pA/pF]; P = .005), and D130G (17.6 pA/pF [13.8-24.6 pA/pF]; P = .003). The decrease in SK2 current densities was not associated with a decrease in membrane protein expression or intracellular distribution of the channel protein. Apamin increased the ventricular action potential duration at 80% repolarization (from 79.6 ms [63.4-93.3 ms] to 121.8 ms [97.9-127.2 ms]; P = .010) in hearts pretreated with anemone toxin but not in control hearts. Conclusion: Human arrhythmogenic calmodulin mutations impede the activation of SK2 channels in human embryonic kidney 293 cells.

AB - Background: Apamin-sensitive small-conductance calcium-activated potassium (SK) channels are gated by intracellular Ca2+ through a constitutive interaction with calmodulin. Objective: We hypothesize that arrhythmogenic human calmodulin mutations impede activation of SK channels. Methods: We studied 5 previously published calmodulin mutations (N54I, N98S, D96V, D130G, and F90L). Plasmids encoding either wild-type or mutant calmodulin were transiently transfected into human embryonic kidney 293 cells that stably express subtype 2 of SK protein channels (SK2 cells). Whole-cell voltage-clamp recording was used to determine apamin-sensitive current densities. We also performed optical mapping studies in normal murine hearts to determine the effects of apamin in hearts with (n=7) or without (n=3) pretreatment with sea anemone toxin. Results: SK2 cells transfected with wild-type calmodulin exhibited an apamin-sensitive current density of 33.6 pA/pF (31.4-36.5 pA/pF) (median and confidence interval 25th-75th percentile), which was significantly higher than that observed for cells transfected with N54I (17.0 pA/pF [14.0-27.7 pA/pF]; P = .016), F90L (22.6 pA/pF [20.3-24.3 pA/pF]; P = .011), D96V (13.0 pA/pF [10.9-15.8 pA/pF]; P = .003), N98S (13.7 pA/pF [8.8-20.4 pA/pF]; P = .005), and D130G (17.6 pA/pF [13.8-24.6 pA/pF]; P = .003). The decrease in SK2 current densities was not associated with a decrease in membrane protein expression or intracellular distribution of the channel protein. Apamin increased the ventricular action potential duration at 80% repolarization (from 79.6 ms [63.4-93.3 ms] to 121.8 ms [97.9-127.2 ms]; P = .010) in hearts pretreated with anemone toxin but not in control hearts. Conclusion: Human arrhythmogenic calmodulin mutations impede the activation of SK2 channels in human embryonic kidney 293 cells.

KW - Arrhythmias

KW - Catecholaminergic polymorphic ventricular tachycardia

KW - Ion channels

KW - Long QT syndrome

KW - Patch clamp

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DO - 10.1016/j.hrthm.2016.05.009

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