Role of protein kinase C in metabolic regulation of the cardiac Na+ channel

Man Liu, Guangbin Shi, Kai Chien Yang, Lianzhi Gu, Anumantha G. Kanthasamy, Vellareddy Anantharam, Samuel C. Dudley

Research output: Contribution to journalArticlepeer-review

13 Scopus citations

Abstract

Background The reduced form of nicotinamide adenine dinucleotide (NADH) increases in cardiomyopathy, activates protein kinase C (PKC), up-regulates mitochondrial reactive oxygen species (mitoROS), and down-regulates the cardiac Na+ channel (NaV1.5). Objective The purpose of this study was to determine how NADH signals down-regulation of NaV1.5. Methods Isolated mouse cardiomyocytes were used for patch-clamp recording and for monitoring mitoROS with MitoSOX Red. HEK293 cells were used for transient transfections. HEK293 cells stably expressing human NaV1.5 were used for single channel recording, whole-cell patch-clamp recording, activity measurements of phospholipase C and phospholipase D (PLD), channel protein purification, and co-immunoprecipitation with PKC isoforms. HL-1 cells were used for mitochondria isolation. Results NADH enhanced PLD activity (1.6- ± 0.1-fold, P <.01) and activated PKCδ. Activated PKCδ translocated to mitochondria and up-regulated mitoROS (2.8- ± 0.3-fold, P <.01) by enhancing the activities of mitochondrial complexes I, II, and IV (1.1- to 1.5-fold, P <.01). PKCδ also interacted with NaV1.5 to down-regulate Na+ current (INa). Reduction in INa by activated PKCδ was prevented by antioxidants and by mutating the known PKC phosphorylation site S1503. At the single channel level, the mechanism of current reduction by PKC and recovery by protein kinase A was a change in single channel conductance. Conclusion NADH activated PKCδ by enhancing PLD activity. PKCδ modulated both mitoROS and NaV1.5. PKCδ elevated mitoROS by enhancing mitochondrial oxidative phosphorylation complex activities. PKCδ-mediated channel phosphorylation and mitoROS were both required to down-regulate NaV1.5 and alter single channel conductance.

Original languageEnglish (US)
Pages (from-to)440-447
Number of pages8
JournalHeart Rhythm
Volume14
Issue number3
DOIs
StatePublished - Mar 1 2017

Bibliographical note

Publisher Copyright:
© 2016 Heart Rhythm Society

Keywords

  • Arrhythmia
  • Cardiomyopathy
  • Channel phosphorylation
  • Metabolism
  • Mitochondria
  • Nicotinamide adenine dinucleotide
  • Protein kinase Cδ

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