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      KChIP2 genotype dependence of transient outward current (I to) properties in cardiomyocytes isolated from male and female mice

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          Abstract

          The transient outward current (I to) in cardiomyocytes is largely mediated by Kv4 channels associated with Kv Channel Interacting Protein 2 (KChIP2). A knockout model has documented the critical role of KChIP2 in I to expression. The present study was conducted to characterize in both sexes the dependence of I to properties, including current magnitude, inactivation kinetics, recovery from inactivation and voltage dependence of inactivation, on the number of functional KChIP2 alleles. For this purpose we performed whole-cell patch-clamp experiments on isolated left ventricular cardiomyocytes from male and female mice which had different KChIP2 genotypes; i.e., wild-type (KChIP2 +/+), heterozygous knockout (KChIP2 +/-) or complete knockout of KChIP2 (KChIP2 -/-). We found in both sexes a KChIP2 gene dosage effect (i.e., a proportionality between number of alleles and phenotype) on I to magnitude, however, concerning other I to properties, KChIP2 +/- resembled KChIP2 +/+. Only in the total absence of KChIP2 (KChIP2 -/-) we observed a slowing of I to kinetics, a slowing of recovery from inactivation and a negative shift of a portion of the voltage dependence of inactivation. In a minor fraction of KChIP2 -/- myocytes I to was completely lost. The distinct KChIP2 genotype dependences of I to magnitude and inactivation kinetics, respectively, seen in cardiomyocytes were reproduced with two-electrode voltage-clamp experiments on Xenopus oocytes expressing Kv4.2 and different amounts of KChIP2. Our results corroborate the critical role of KChIP2 in controlling I to properties. They demonstrate that the Kv4.2/KChIP2 interaction in cardiomyocytes is highly dynamic, with a clear KChIP2 gene dosage effect on Kv4 channel surface expression but not on inactivation gating.

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          Most cited references56

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          Modulation of A-type potassium channels by a family of calcium sensors.

          In the brain and heart, rapidly inactivating (A-type) voltage-gated potassium (Kv) currents operate at subthreshold membrane potentials to control the excitability of neurons and cardiac myocytes. Although pore-forming alpha-subunits of the Kv4, or Shal-related, channel family form A-type currents in heterologous cells, these differ significantly from native A-type currents. Here we describe three Kv channel-interacting proteins (KChIPs) that bind to the cytoplasmic amino termini of Kv4 alpha-subunits. We find that expression of KChIP and Kv4 together reconstitutes several features of native A-type currents by modulating the density, inactivation kinetics and rate of recovery from inactivation of Kv4 channels in heterologous cells. All three KChIPs co-localize and co-immunoprecipitate with brain Kv4 alpha-subunits, and are thus integral components of native Kv4 channel complexes. The KChIPs have four EF-hand-like domains and bind calcium ions. As the activity and density of neuronal A-type currents tightly control responses to excitatory synaptic inputs, these KChIPs may regulate A-type currents, and hence neuronal excitability, in response to changes in intracellular calcium.
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            A defect in the Kv channel-interacting protein 2 (KChIP2) gene leads to a complete loss of I(to) and confers susceptibility to ventricular tachycardia.

            KChIP2, a gene encoding three auxiliary subunits of Kv4.2 and Kv4.3, is preferentially expressed in the adult heart, and its expression is downregulated in cardiac hypertrophy. Mice deficient for KChIP2 exhibit normal cardiac structure and function but display a prolonged elevation in the ST segment on the electrocardiogram. The KChIP2(-/-) mice are highly susceptible to the induction of cardiac arrhythmias. Single-cell analysis revealed a substrate for arrhythmogenesis, including a complete absence of transient outward potassium current, I(to), and a marked increase in action potential duration. These studies demonstrate that a defect in KChIP2 is sufficient to confer a marked genetic susceptibility to arrhythmias, establishing a novel genetic pathway for ventricular tachycardia via a loss of the transmural gradient of I(to).
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              Molecular determinants of cardiac transient outward potassium current (I(to)) expression and regulation.

              Rapidly activating and inactivating cardiac transient outward K(+) currents, I(to), are expressed in most mammalian cardiomyocytes, and contribute importantly to the early phase of action potential repolarization and to plateau potentials. The rapidly recovering (I(t)(o,f)) and slowly recovering (I(t)(o,s)) components are differentially expressed in the myocardium, contributing to regional heterogeneities in action potential waveforms. Consistent with the marked differences in biophysical properties, distinct pore-forming (alpha) subunits underlie the two I(t)(o) components: Kv4.3/Kv4.2 subunits encode I(t)(o,f), whereas Kv1.4 encodes I(t)(o,s), channels. It has also become increasingly clear that cardiac I(t)(o) channels function as components of macromolecular protein complexes, comprising (four) Kvalpha subunits and a variety of accessory subunits and regulatory proteins that influence channel expression, biophysical properties and interactions with the actin cytoskeleton, and contribute to the generation of normal cardiac rhythms. Derangements in the expression or the regulation of I(t)(o) channels in inherited or acquired cardiac diseases would be expected to increase the risk of potentially life-threatening cardiac arrhythmias. Indeed, a recently identified Brugada syndrome mutation in KCNE3 (MiRP2) has been suggested to result in increased I(t)(o,f) densities. Continued focus in this area seems certain to provide new and fundamentally important insights into the molecular determinants of functional I(t)(o) channels and into the molecular mechanisms involved in the dynamic regulation of I(t)(o) channel functioning in the normal and diseased myocardium. Copyright 2009 Elsevier Inc. All rights reserved.
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                Author and article information

                Contributors
                Role: Editor
                Journal
                PLoS One
                PLoS ONE
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, CA USA )
                1932-6203
                31 January 2017
                2017
                : 12
                : 1
                : e0171213
                Affiliations
                [001]Institut für Zelluläre und Integrative Physiologie, Zentrum für Experimentelle Medizin, Universitätsklinikum Hamburg-Eppendorf, Hamburg, Germany
                University of Texas Health Science Center, UNITED STATES
                Author notes

                Competing Interests: The authors have declared that no competing interests exist.

                • Conceptualization: RB.

                • Data curation: LW VJ RB.

                • Formal analysis: LW VJ RB.

                • Funding acquisition: RB.

                • Investigation: LW VJ RB.

                • Methodology: LW VJ.

                • Project administration: RB.

                • Supervision: RB.

                • Validation: RB.

                • Visualization: LW VJ RB.

                • Writing – original draft: RB.

                • Writing – review & editing: RB LW VJ.

                Article
                PONE-D-16-43430
                10.1371/journal.pone.0171213
                5283746
                28141821
                781a4231-ba3e-4855-971f-e5e694c5deab
                © 2017 Waldschmidt et al

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                History
                : 1 November 2016
                : 18 January 2017
                Page count
                Figures: 8, Tables: 2, Pages: 28
                Funding
                Funded by: Deutsche Forschungsgemeinschaft (DFG)
                Award ID: BA2055/3
                Award Recipient :
                This study was supported by grant BA2055/3 of the Deutsche Forschungsgemeinschaft (DFG) to RB. The funder had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
                Categories
                Research Article
                Biology and Life Sciences
                Cell Biology
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                Animal Cells
                Muscle Cells
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                Biological Tissue
                Muscle Tissue
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