Epac and phospholipase Cepsilon regulate Ca2+ release in the heart by activation of protein kinase Cepsilon and calcium-calmodulin kinase II

J Biol Chem. 2009 Jan 16;284(3):1514-22. doi: 10.1074/jbc.M806994200. Epub 2008 Oct 27.


Recently, we identified a novel signaling pathway involving Epac, Rap, and phospholipase C (PLC)epsilon that plays a critical role in maximal beta-adrenergic receptor (betaAR) stimulation of Ca2+-induced Ca2+ release (CICR) in cardiac myocytes. Here we demonstrate that PLCepsilon phosphatidylinositol 4,5-bisphosphate hydrolytic activity and PLCepsilon-stimulated Rap1 GEF activity are both required for PLCepsilon-mediated enhancement of sarcoplasmic reticulum Ca2+ release and that PLCepsilon significantly enhances Rap activation in response to betaAR stimulation in the heart. Downstream of PLCepsilon hydrolytic activity, pharmacological inhibition of PKC significantly inhibited both betaAR- and Epac-stimulated increases in CICR in PLCepsilon+/+ myocytes but had no effect in PLCepsilon-/- myocytes. betaAR and Epac activation caused membrane translocation of PKCepsilon in PLCepsilon+/+ but not PLCepsilon-/- myocytes and small interfering RNA-mediated PKCepsilon knockdown significantly inhibited both betaAR and Epac-mediated CICR enhancement. Further downstream, the Ca2+/calmodulin-dependent protein kinase II (CamKII) inhibitor, KN93, inhibited betaAR- and Epac-mediated CICR in PLCepsilon+/+ but not PLCepsilon-/- myocytes. Epac activation increased CamKII Thr286 phosphorylation and enhanced phosphorylation at CamKII phosphorylation sites on the ryanodine receptor (RyR2) (Ser2815) and phospholamban (Thr17) in a PKC-dependent manner. Perforated patch clamp experiments revealed that basal and betaAR-stimulated peak L-type current density are similar in PLCepsilon+/+ and PLCepsilon-/- myocytes suggesting that control of sarcoplasmic reticulum Ca2+ release, rather than Ca2+ influx through L-type Ca2+ channels, is the target of regulation of a novel signal transduction pathway involving sequential activation of Epac, PLCepsilon, PKCepsilon, and CamKII downstream of betaAR activation.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Benzylamines / pharmacology
  • Calcium / metabolism*
  • Calcium Channels, L-Type / genetics
  • Calcium Channels, L-Type / metabolism
  • Calcium Signaling / physiology*
  • Calcium-Binding Proteins / genetics
  • Calcium-Binding Proteins / metabolism
  • Calcium-Calmodulin-Dependent Protein Kinase Type 2 / antagonists & inhibitors
  • Calcium-Calmodulin-Dependent Protein Kinase Type 2 / genetics
  • Calcium-Calmodulin-Dependent Protein Kinase Type 2 / metabolism*
  • Cell Membrane / enzymology
  • Cell Membrane / genetics
  • Enzyme Activation / physiology
  • Guanine Nucleotide Exchange Factors / genetics
  • Guanine Nucleotide Exchange Factors / metabolism*
  • Mice
  • Mice, Knockout
  • Myocardium / cytology
  • Myocardium / enzymology*
  • Myocytes, Cardiac / cytology
  • Myocytes, Cardiac / enzymology
  • Phosphatidylinositol 4,5-Diphosphate
  • Phosphoinositide Phospholipase C / genetics
  • Phosphoinositide Phospholipase C / metabolism*
  • Phosphorylation / physiology
  • Protein Kinase C-epsilon / genetics
  • Protein Kinase C-epsilon / metabolism
  • Protein Kinase Inhibitors / pharmacology
  • Protein Transport / physiology
  • Receptors, Adrenergic, beta / genetics
  • Receptors, Adrenergic, beta / metabolism
  • Ryanodine Receptor Calcium Release Channel / genetics
  • Ryanodine Receptor Calcium Release Channel / metabolism
  • Sarcoplasmic Reticulum / enzymology
  • Sarcoplasmic Reticulum / genetics
  • Sulfonamides / pharmacology


  • Benzylamines
  • Calcium Channels, L-Type
  • Calcium-Binding Proteins
  • Epac protein, mouse
  • Guanine Nucleotide Exchange Factors
  • Phosphatidylinositol 4,5-Diphosphate
  • Protein Kinase Inhibitors
  • Receptors, Adrenergic, beta
  • Ryanodine Receptor Calcium Release Channel
  • Sulfonamides
  • phospholamban
  • KN 93
  • Prkce protein, mouse
  • Protein Kinase C-epsilon
  • Calcium-Calmodulin-Dependent Protein Kinase Type 2
  • Phosphoinositide Phospholipase C
  • phospholipase C epsilon
  • Calcium