Regulation of cardiac ion channels by signaling complexes: role of modified leucine zipper motifs

doi:10.1016/j.yjmcc.2004.04.014

Journal of Molecular and Cellular Cardiology

Volume 37, Issue 3, September 2004, Pages 625-631

https://doi.org/10.1016/j.yjmcc.2004.04.014 Get rights and content

Abstract

Modulation of ion channels by protein phosphorylation is a dynamic process precisely controlled by the opposing actions of protein kinases and phosphoprotein phosphatases. It is well accepted that the targeting and localization of such signaling enzymes to discrete subcellular compartments or substrates is an important regulatory mechanism ensuring specificity of signaling events in response to local stimuli. Compartmentalization of these enzymes is achieved through association with anchoring or adaptor proteins that target them to subcellular organelles or tether them directly to target substrates via protein–protein interactions. Recently, a novel role for modified leucine zipper motifs in targeting kinases and phosphatases via anchoring proteins has been described for three families of cardiac ion channels: ryanodine-sensitive calcium (Ca²⁺) release channels, voltage-gated Ca²⁺ channels, and delayed rectifier potassium (K⁺) channels. This review will summarize the recent advances made on the regulation of cardiac ion channels by these macromolecular signaling complexes in the normal and diseased heart.

Introduction

To meet changing hemodynamic demands placed upon the heart, excitation–contraction (EC) coupling is constantly modulated by multiple signaling pathways. The most important regulator of cardiac function on a beat-to-beat basis is the autonomic nervous system. Release of catecholamines from the autonomic nervous system activates adrenoceptors. Cardiac β-adrenergic receptors (β-ARs) are the primary targets for sympathetic neurotransmitters (e.g. norepinephrine) and adrenal hormones (e.g. epinephrine). Stimulation of the β-AR signaling pathway increases the chronotropic (heart rate), inotropic (strength of contraction during systole), and lusitropic (rate and extent of relaxation during diastole) states of the heart. At the cellular level, stimulation of β-ARs activates the cAMP-dependent protein kinase A (PKA) signaling pathway resulting in the phosphorylation of a number of target proteins. These include sarcolemmal and transverse tubule L-type calcium (Ca²⁺) channels, which conduct the trigger Ca²⁺ currents that initiate EC coupling; the sarcoplasmic reticulum (SR) ryanodine receptor (RyR)-sensitive Ca²⁺ release channels; and sarcolemmal potassium (K⁺) channels responsible for the slowly activating outward current (I_Ks) [1], [2], [3], [4], [5].

Modulation of ion channels by protein phosphorylation is a dynamic process precisely controlled by the opposing actions of protein kinases and phosphoprotein phosphatases. It is well accepted that the targeting and localization of such signaling enzymes to discrete subcellular compartments or substrates is an important regulatory mechanism ensuring specificity of signaling events in response to local stimuli [6], [7]. At the molecular level, compartmentalization of these enzymes is achieved through the association with anchoring or adaptor proteins that target them to subcellular organelles or tether them to substrates via protein–protein interactions. To date, three families of cardiac ion channels have been shown to form kinase–phosphatase signaling complexes. This review summarizes the recent advances made on the regulation of cardiac ion channels by macromolecular signaling complexes in the normal and diseased heart.

Section snippets

A-kinase-anchoring proteins

A well-characterized example of protein kinase targeting is the compartmentalization of PKA. In its inactive form, PKA is a heterotetramer composed of a regulatory (R) subunit dimer and two catalytic (C) subunits [8]. Upon binding cAMP, the C subunits are released from the R subunit dimer and phosphorylate appropriate substrates. The subcellular location of PKA is maintained through protein–protein interactions between the R subunit dimer and A-kinase-anchoring proteins (AKAPs) [9], [10] a

Regulation of ion channels by signaling complexes

Regulation of ion channels by the β-AR/cAMP signaling pathway requires the targeting of PKA and its counterbalancing protein phosphatases to the vicinity of channels through association with AKAPs and adaptor proteins [17]. Insight into the molecular mechanisms underlying the targeting of kinases and phosphatases to ion channels has recently emerged. Marx et al. [18] identified a novel role for modified leucine zipper (mLZ) motifs in targeting kinases and phosphatases to the cardiac RyR2.

LZ motifs

LZs were originally identified as highly conserved motifs mediating the interaction of protein components of transcription factors [22]. A role for LZ motifs in promoting homo- and heterodimerization of transcription factors [23], protein oligomerization [24], and specific protein–protein interactions [25], [26] has been described. The LZ is an α-helical structure comprised of heptad repeats (abcdefg)_n with hydrophobic residues at positions ‘a’ and ‘d’ that form coiled-coils through

SR Ca²⁺ release channels

RyR2 are the major SR Ca²⁺ release channels required for EC coupling in the heart [33]. These channels are tetrameric structures composed of four RyR2 polypeptides that bind four FK506-binding proteins (FKBP12.6). FKBP12s are regulatory subunits that stabilize RyR channel function [34] and coordinate activation and inactivation of neighboring RyRs during EC coupling—a process known as coupled gating [35], [36]. FKBP12.6 binding to RyR2 can be regulated by PKA phosphorylation of RyR2 at serine

Conclusion

As reviewed in this article, recent research has demonstrated that direct interactions between ion channels and kinase–phosphatase-anchoring proteins are required for normal ion channel regulation by PKA signaling pathways. Remarkably, these protein–protein interactions are mediated by mLZ motifs and serve to target the kinase and phosphatase close to its sites of phosphorylation in the channel protein. As these examples illustrate, local signaling by ion channels is locally regulated by

References (87)

R.W. Tsien et al.
Mechanisms of calcium channel modulation by beta-adrenergic agents and dihydropyridine calcium agonists
J Mol Cell Cardiol
(1986)
M. Colledge et al.
AKAPs: from structure to function
Trend Cell Biol
(1999)
M.G. Newlon et al.
The A-kinase anchoring domain of type II alpha cAMP-dependent protein kinase is highly helical
J Biol Chem
(1997)
K.A. Tasken et al.
Phosphodiesterase 4D and protein kinase a type II constitute a signaling unit in the centrosomal area
J Biol Chem
(2001)
P.C. Gray et al.
Regulation of ion channels by cAMP-dependent protein kinase and A-kinase anchoring proteins
Curr Opin Neurobiol
(1998)
J.T. Hulme et al.
A novel leucine zipper targets AKAP15 and cyclic AMP-dependent protein kinase to the C terminus of the skeletal muscle Ca²⁺ channel and modulates its function
J Biol Chem
(2002)
H.K. Simmerman et al.
A leucine zipper stabilizes the pentameric membrane domain of phospholamban and forms a coiled-coil pore structure
J Biol Chem
(1996)
A. Lupas
Coiled coils: new structures and new functions
Trend Biochem Sci
(1996)
I.W. Leung et al.
Dimerization via tandem leucine zippers is essential for the activation of the mitogen-activated protein kinase kinase kinase, MLK-3
J Biol Chem
(1998)
C.A. Syme et al.
Trafficking of the Ca²⁺-activated K⁺ channel, hIK1, is dependent upon a C-terminal leucine zipper
J Biol Chem
(2003)

D.L. Galvan et al.

Subunit oligomerization, and topology of the inositol 1,4,5-trisphosphate receptor

J Biol Chem

(1999)

A.B. Brillantes et al.

Stabilization of calcium release channel (ryanodine receptor) function by FK506-binding protein

Cell

(1994)

S.O. Marx et al.

PKA phosphorylation dissociates FKBP12.6 from the calcium release channel (ryanodine receptor): defective regulation in failing hearts

Cell

(2000)

J. Hain et al.

Phosphorylation modulates the function of the calcium release channel of sarcoplasmic reticulum from cardiac muscle

J Biol Chem

(1995)

M.L. Ruehr et al.

Targeting of protein kinase A by muscle A kinase-anchoring protein (mAKAP) regulates phosphorylation and function of the skeletal muscle ryanodine receptor

J Biol Chem

(2003)

E.I. Rotman et al.

Specific phosphorylation of a COOH-terminal site on the full-length form of the α1 subunit of the skeletal muscle calcium channel by cAMP-dependent protein kinase

J Biol Chem

(1992)

E.I. Rotman et al.

Sites of selective cAMP-dependent phosphorylation of the L-type calcium channel α1 subunit from intact rabbit skeletal muscle myotubes

J Biol Chem

(1995)

P.C. Gray et al.

Identification of a 15-kDa cAMP-dependent protein kinase-anchoring protein associated with skeletal muscle L-type calcium channels

J Biol Chem

(1997)

P.C. Gray et al.

Primary structure and function of an A kinase anchoring protein associated with calcium channels

Neuron

(1998)

J.W. Hell et al.

Differential phosphorylation of two size forms of the neuronal class C L-type calcium channel α1 subunit

J Biol Chem

(1993)

T. Gao et al.

cAMP-dependent regulation of cardiac L-type Ca²⁺ channels requires membrane targeting of PKA and phosphorylation of channel subunits

Neuron

(1997)

M.A. Davare et al.

The A-kinase anchor protein MAP2B and cAMP-dependent protein kinase are associated with class C L-type calcium channels in neurons

J Biol Chem

(1999)

C. Altier et al.

Trafficking of L-type calcium channels mediated by the postsynaptic scaffolding protein AKAP79

J Biol Chem

(2002)

Y. Chen-Izu et al.

G_i-dependent localization of beta-2-adrenergic receptor signaling to L-type Ca²⁺ channels

Biophys J

(2000)

L. Tian et al.

Leucine zipper domain targets cAMP-dependent protein kinase to mammalian BK channels

J Biol Chem

(2003)

S. Nakade et al.

Cyclic AMP-dependent phosphorylation of an immunoaffinity-purified homotetrameric inositol 1,4,5-trisphosphate receptor (type I) increases Ca²⁺ flux in reconstituted lipid vesicles

J Biol Chem

(1994)

R.J. Wojcikiewicz et al.

Phosphorylation of inositol 1,4,5-trisphosphate receptors by cAMP-dependent protein kinase. Type I, II, and III receptors are differentially susceptible to phosphorylation and are phosphorylated in intact cells

J Biol Chem

(1998)

N. Desouza et al.

Protein kinase A and two phosphatases are components of the inositol 1,4,5-trisphosphate receptor macromolecular signaling complex

J Biol Chem

(2002)

D.H. Schulze et al.

Sodium/calcium exchanger (NCX1) macromolecular complex

J Biol Chem

(2003)

M.T. Keating et al.

Molecular and cellular mechanisms of cardiac arrhythmias

Cell

(2001)

K. Piippo et al.

A founder mutation of the potassium channel KCNQ1 in long QT syndrome: implications for estimation of disease prevalence and molecular diagnostics

J Am Coll Cardiol

(2001)

H. Reuter

Calcium channel modulation by neurotransmitters, enzymes and drugs

Nature

(1983)

W.A. Catterall

Structure and regulation of voltage-gated calcium channels

Annu Rev Cell Dev Biol

(2000)

A.R. Marks

Ryanodine receptors, FKBP12, and heart failure

Front Biosci

(2002)

C.E. Clancy et al.

Defective cardiac ion channels: from mutations to clinical syndromes

J Clin Invest

(2002)

D. Schechtman et al.

Adaptor proteins in protein kinase C-mediated signal transduction

Oncogene

(2001)

T. Pawson et al.

Signaling through scaffold, anchoring, and adaptor proteins

Science

(1997)

E.G. Krebs et al.

Phosphorylation–dephosphorylation of enzymes

Annu Rev Biochem

(1979)

C.S. Rubin

A kinase anchor proteins and the intracellular targeting of signals carried by cyclic AMP

Biochim Biophys Acta

(1994)

M.G. Newlon et al.

A novel mechanism of PKA anchoring revealed by solution structures of anchoring complexes

EMBO J

(2001)

C. Rosenmund et al.

Anchoring of protein kinase A is required for modulation of AMPA/kainate receptors on hippocampal neurons

Nature

(1994)

M.B. Kennedy

Signal-processing machines at the postsynaptic density

Science

(2000)

K.L. Dodge et al.

mAKAP assembles a protein kinase A/PDE4 phosphodiesterase cAMP signaling module

EMBO J

(2001)

Cited by (19)

Phosphoproteomics and molecular cardiology: Techniques, applications and challenges
2012, Journal of Molecular and Cellular Cardiology
Citation Excerpt :
For example, cardiac contractile machinery contains phosphorylated proteins such as troponin I [210,211], myosin light chain 2 [212], and myosin-binding protein C [213,214], with an altered phosphorylation pattern found in many pathophysiological conditions [215]. Functions of major ion channels in cardiomyocytes such as ryanodine-sensitive Ca2+ channels [216,217], voltage-gated L-type Ca2+ channels [218], and delayed rectifier potassium (K+) channels [219] are also regulated via protein phosphorylation [220]. Given the breadth of protein phosphorylation in many critical cardiac functions, as well as the implication of protein phosphorylation in cardiac disease development, kinase modulators have emerged as an important class of cardiovascular drugs [4,221,222].
Protein phosphorylation has been widely documented as a key regulatory and signaling mechanism associated with many cardiac diseases. Recent advances in phosphoproteomic technologies such as phosphopeptide enrichment, novel mass spectrometry applications, and bioinformatic tools have resulted in high-throughput identification and quantitation of protein phosphorylation in a global manner. This review summarizes mainstream phosphoproteomic workflows and highlights the most recent applications of phosphoproteomics used in a range of molecular cardiology research.
Deletion of the distal C terminus of Ca<inf>v</inf>1.2 channels leads to loss of β-adrenergic regulation and heart failure in Vivo
2011, Journal of Biological Chemistry
Citation Excerpt :
This result is consistent with our recent studies showing that the DCT is required for the normal enhancement of voltage-dependent inactivation of CaV1.2 channels by physiological levels of intracellular Mg2+ (37). Upon β-adrenergic activation of cardiac myocytes, PKA phosphorylates α1 and β subunits of the CaV1.2 channel (15, 23, 38, 39), which increases peak Ca2+ currents, increases the maximal probability of channel opening and negatively shifts the voltage dependence of channel activation (6, 8, 40). The β-adrenergic receptor agonist isoproterenol increases Ca2+ current of embryonic cardiomyocytes isolated at E16-E18 by ∼2-fold and shifts the voltage dependence of activation to more negative membrane potential (data not shown), indicating that the fetal cardiomyocyte is a valid system to examine the regulation of β-adrenergic signaling pathway.
L-type calcium currents conducted by Ca_V1.2 channels initiate excitation-contraction coupling in cardiac and vascular smooth muscle. In the heart, the distal portion of the C terminus (DCT) is proteolytically processed in vivo and serves as a noncovalently associated autoinhibitor of Ca_V1.2 channel activity. This autoinhibitory complex, with A-kinase anchoring protein-15 (AKAP15) bound to the DCT, is hypothesized to serve as the substrate for β-adrenergic regulation in the fight-or-flight response. Mice expressing Ca_V1.2 channels with the distal C terminus deleted (DCT^−/−) develop cardiac hypertrophy and die prematurely after E15. Cardiac hypertrophy and survival rate were improved by drug treatments that reduce peripheral vascular resistance and hypertension, consistent with the hypothesis that Ca_V1.2 hyperactivity in vascular smooth muscle causes hypertension, hypertrophy, and premature death. However, in contrast to expectation, L-type Ca²⁺ currents in cardiac myocytes from DCT^−/− mice were dramatically reduced due to decreased cell-surface expression of Ca_V1.2 protein, and the voltage dependence of activation and the kinetics of inactivation were altered. Ca_V1.2 channels in DCT^−/− myocytes fail to respond to activation of adenylyl cyclase by forskolin, and the localized expression of AKAP15 is reduced. Therefore, we conclude that the DCT of Ca_V1.2 channels is required in vivo for normal vascular regulation, cell-surface expression of Ca_V1.2 channels in cardiac myocytes, and β-adrenergic stimulation of L-type Ca²⁺ currents in the heart.
Ryanodine receptor-mediated arrhythmias and sudden cardiac death
2009, Pharmacology and Therapeutics
Citation Excerpt :
However, there may be other determinants for CICR termination, such as intrinsic changes occurring within the RyR2 such as inactivation, adaptation (Cheng et al., 1995; Sitsapesan et al., 1995; Laver & Lamb, 1998; Fill et al., 2000) and stochastic attrition (Stern et al., 1999) or allostery between RyRs (Yin et al., 2008). Kinase/phosphatase signalling complexes have been described for three cardiac ion channels; LCC, K+ channels and RyR2, created by the association of phosphatases and kinases with anchoring proteins held together by leucine zipper protein/protein interaction domains (Marx et al., 2001b; Hulme et al., 2004). These complexes enable the compartmentalisation of protein kinase A (PKA), protein phosphatase 1 (PPI), protein phosphatase 2A (PP2A) (Marx et al., 2001b), CaMKII (Ai et al., 2005), and phosphodiesterase 4D3 (PDE4D3) (Dodge et al., 2001), so that phosphorylation/dephosphorylation can be spatially and temporally regulated (Hulme et al., 2004).
The cardiac ryanodine receptor-Ca²⁺ release channel (RyR2) is an essential sarcoplasmic reticulum (SR) transmembrane protein that plays a central role in excitation–contraction coupling (ECC) in cardiomyocytes. Aberrant spontaneous, diastolic Ca²⁺ leak from the SR due to dysfunctional RyR2 contributes to the formation of delayed after-depolarisations, which are thought to underlie the fatal arrhythmia that occurs in both heart failure (HF) and in catecholaminergic polymorphic ventricular tachycardia (CPVT). CPVT is an inherited disorder associated with mutations in either the RyR2 or a SR luminal protein, calsequestrin. RyR2 shows normal function at rest in CPVT but the RyR2 dysfunction is unmasked by physical exercise or emotional stress, suggesting abnormal RyR2 activation as an underlying mechanism. Several potential mechanisms have been advanced to explain the dysfunctional RyR2 observed in HF and CPVT, including enhanced RyR2 phosphorylation status, altered RyR2 regulation at luminal/cytoplasmic sites and perturbed RyR2 intra/inter-molecular interactions. This review considers RyR2 dysfunction in the context of the structural and functional modulation of the channel, and potential therapeutic strategies to stabilise RyR2 function in cardiac pathology.
Scaffolding and docking proteins of the heart, an introduction
2004, Journal of Molecular and Cellular Cardiology
Something old, something new: Changing views on the cellular mechanisms of heart failure
2005, Cardiovascular Research
Ion channels as part of macromolecular multiprotein complexes: Clinical significance
2018, Herzschrittmachertherapie und Elektrophysiologie

View all citing articles on Scopus

^☆: Guest Editor, Meredith Bond, Ph. D., handled the review of this paper.

View full text

Scaffolding and Docking Proteins of the HeartRegulation of cardiac ion channels by signaling complexes: role of modified leucine zipper motifs☆

Abstract

Introduction

Section snippets

A-kinase-anchoring proteins

Regulation of ion channels by signaling complexes

LZ motifs

SR Ca2+ release channels

Conclusion

J Mol Cell Cardiol

Trend Cell Biol

J Biol Chem

J Biol Chem

Curr Opin Neurobiol

J Biol Chem

J Biol Chem

Trend Biochem Sci

J Biol Chem

J Biol Chem

J Biol Chem

Cell

Cell

J Biol Chem

J Biol Chem

J Biol Chem

J Biol Chem

J Biol Chem

Neuron

J Biol Chem

Neuron

J Biol Chem

J Biol Chem

Biophys J

J Biol Chem

J Biol Chem

J Biol Chem

J Biol Chem

J Biol Chem

Cell

J Am Coll Cardiol

Calcium channel modulation by neurotransmitters, enzymes and drugs

Nature

Structure and regulation of voltage-gated calcium channels

Annu Rev Cell Dev Biol

Ryanodine receptors, FKBP12, and heart failure

Front Biosci

Defective cardiac ion channels: from mutations to clinical syndromes

J Clin Invest

Adaptor proteins in protein kinase C-mediated signal transduction

Oncogene

Signaling through scaffold, anchoring, and adaptor proteins

Science

Phosphorylation–dephosphorylation of enzymes

Annu Rev Biochem

A kinase anchor proteins and the intracellular targeting of signals carried by cyclic AMP

Biochim Biophys Acta

A novel mechanism of PKA anchoring revealed by solution structures of anchoring complexes

EMBO J

Anchoring of protein kinase A is required for modulation of AMPA/kainate receptors on hippocampal neurons

Nature

Signal-processing machines at the postsynaptic density

Science

mAKAP assembles a protein kinase A/PDE4 phosphodiesterase cAMP signaling module

EMBO J

Scaffolding and Docking Proteins of the Heart
Regulation of cardiac ion channels by signaling complexes: role of modified leucine zipper motifs☆

SR Ca²⁺ release channels