Ca2+/calmodulin kinase II-dependent regulation of ßIV-spectrin modulates cardiac fibroblast gene expression, proliferation, and contractility.

Fibrosis is a pronounced feature of heart disease and the result of dysregulated activation of resident cardiac fibroblasts (CFs). Recent work identified stress-induced degradation of the cytoskeletal protein ßIV-spectrin as an important step in CF activation and cardiac fibrosis. Furthermore, loss of ßIV-spectrin was found to depend on Ca2+/calmodulin-dependent kinase II (CaMKII). Therefore, we sought to determine the mechanism for CaMKII-dependent regulation of ßIV-spectrin and CF activity. Computational screening and MS revealed a critical serine residue (S2250 in mouse and S2254 in human) in ßIV-spectrin phosphorylated by CaMKII. Disruption of ßIV-spectrin/CaMKII interaction or alanine substitution of ßIV-spectrin Ser2250 (ßIV-S2254A) prevented CaMKII-induced degradation, whereas a phosphomimetic construct (ßIV-spectrin with glutamic acid substitution at serine 2254 [ßIV-S2254E]) showed accelerated degradation in the absence of CaMKII. To assess the physiological significance of this phosphorylation event, we expressed exogenous ßIV-S2254A and ßIV-S2254E constructs in ßIV-spectrin-deficient CFs, which have increased proliferation and fibrotic gene expression compared with WT CFs. ßIV-S2254A but not ßIV-S2254E normalized CF proliferation, gene expression, and contractility. Pathophysiological targeting of ßIV-spectrin phosphorylation and subsequent degradation was identified in CFs activated with the profibrotic ligand angiotensin II, resulting in increased proliferation and signal transducer and activation of transcription 3 nuclear accumulation. While therapeutic delivery of exogenous WT ßIV-spectrin partially reversed these trends, ßIV-S2254A completely negated increased CF proliferation and signal transducer and activation of transcription 3 translocation. Moreover, we observed ßIV-spectrin phosphorylation and associated loss in total protein within human heart tissue following heart failure. Together, these data illustrate a considerable role for the ßIV-spectrin/CaMKII interaction in activating profibrotic signaling.

Cardiac inflammation in COVID-19: Lessons from heart failure.

Cardiovascular disease (CVD) is the most common co-morbidity associated with COVID-19 and the fatality rate in COVID-19 patients with CVD is higher compared to other comorbidities, such as hypertension and diabetes. Preliminary data suggest that COVID-19 may also cause or worsen cardiac injury in infected patients through multiple mechanisms such as 'cytokine storm', endotheliosis, thrombosis, lymphocytopenia etc. Autopsies of COVID-19 patients reveal an infiltration of inflammatory mononuclear cells in the myocardium, confirming the role of the immune system in mediating cardiovascular damage in response to COVID-19 infection and also suggesting potential causal mechanisms for the development of new cardiac pathologies and/or exacerbation of underlying CVDs in infected patients. In this review, we discuss the potential underlying molecular mechanisms that drive COVID-19-mediated cardiac damage, as well as the short term and expected long-term cardiovascular ramifications of COVID-19 infection in patients.

Fibroblast growth factor-inducible 14 mediates macrophage infiltration in heart to promote pressure overload-induced cardiac dysfunction.

AIMS: Heart failure (HF) is characterized by compromised cardiac structure and function. Previous work has identified a link between upregulation of pro-inflammatory cytokines and HF. Tumor necrosis factor (TNF)-like weak inducer of apoptosis (TWEAK) is a pro-inflammatory cytokine, which binds to fibroblast growth factor inducible 14 (Fn14), a ubiquitously expressed cell-surface receptor. The objective of this study was to investigate the role of TWEAK/Fn14 pathway in promoting cardiac inflammation under non ischemic stress conditions. MAIN METHODS: Wild type (WT) and Fn14 knock out (Fn14-/-) mice were subjected to pressure overload [transaortic constriction (TAC)] for 1 or 6 weeks. A subset of WT TAC animals were treated with the Fn14 antagonist L524-0366. Cardiac function was measured by echocardiography. Cardiac fibrosis and macrophage infiltration were quantified using immunohistochemistry and flow cytometry, respectively. Cardiac fibroblasts were isolated for quantifying TWEAK-induced chemokine release. KEY FINDINGS: Fn14-/- mice displayed improved cardiac function, reduced fibrosis and lower macrophage infiltration in heart compared to WT following TAC. L524-0366 mitigated maladaptive remodeling with TAC. TWEAK induced secretion of the pro-inflammatory chemokine, monocyte chemoattractant protein 1 from WT but not Fn14-/- fibroblasts in vitro, in part through activation of non-canonical NF-κB signaling. Finally, Fn14 expression was increased in mouse following TAC and in human failing hearts. SIGNIFICANCE: Our findings support an important role for the TWEAK/Fn14 promoting macrophage infiltration and fibrosis in heart under non-ischemic stress, with potential for therapeutic intervention to improve cardiac function in the setting of HF.

Computational tools for automated histological image analysis and quantification in cardiac tissue.

Image processing and quantification is a routine and important task across disciplines in biomedical research. Understanding the effects of disease on the tissue and organ level often requires the use of images, however the process of interpreting those images into data which can be tested for significance is often time intensive, tedious and prone to inaccuracy or bias. When working within resource constraints, these different issues often present a trade-off between time invested in analysis and accuracy. To address these issues, we present two novel open source and publically available tools for automated analysis of histological cardiac tissue samples: •Automated Fibrosis Analysis Tool (AFAT) for quantifying fibrosis; and•Macrophage Analysis Tool (MAT) for quantifying infiltrating macrophages.

ßIV-Spectrin/STAT3 complex regulates fibroblast phenotype, fibrosis, and cardiac function.

Increased fibrosis is a characteristic remodeling response to biomechanical and neurohumoral stress and a determinant of cardiac mechanical and electrical dysfunction in disease. Stress-induced activation of cardiac fibroblasts (CFs) is a critical step in the fibrotic response, although the precise sequence of events underlying activation of these critical cells in vivo remain unclear. Here, we tested the hypothesis that a ßIV-spectrin/STAT3 complex is essential for maintenance of a quiescent phenotype (basal nonactivated state) in CFs. We reported increased fibrosis, decreased cardiac function, and electrical impulse conduction defects in genetic and acquired mouse models of ßIV-spectrin deficiency. Loss of ßIV-spectrin function promoted STAT3 nuclear accumulation and transcriptional activity, and it altered gene expression and CF activation. Furthermore, we demonstrate that a quiescent phenotype may be restored in ßIV-spectrin-deficient fibroblasts by expressing a ßIV-spectrin fragment including the STAT3-binding domain or through pharmacological STAT3 inhibition. We found that in vivo STAT3 inhibition abrogates fibrosis and cardiac dysfunction in the setting of global ßIV-spectrin deficiency. Finally, we demonstrate that fibroblast-specific deletion of ßIV-spectrin is sufficient to induce fibrosis and decreased cardiac function. We propose that the ßIV-spectrin/STAT3 complex is a determinant of fibroblast phenotype and fibrosis, with implications for remodeling response in cardiovascular disease (CVD).

Novel Mechanistic Roles for Ankyrin-G in Cardiac Remodeling and Heart Failure.

Ankyrin polypeptides are intracellular proteins responsible for targeting cardiac membrane proteins. Here, the authors demonstrate that ankyrin-G plays an unexpected role in normal compensatory physiological remodeling in response to myocardial stress and aging; the authors implicate disruption of ankyrin-G in human heart failure. Mechanistically, the authors illustrate that ankyrin-G serves as a key nodal protein required for cardiac myofilament integration with the intercalated disc. Their data define novel in vivo mechanistic roles for ankyrin-G, implicate ankyrin-G as necessary for compensatory cardiac physiological remodeling under stress, and implicate disruption of ankyrin-G in the development and progression of human heart failure.

ßIV-Spectrin regulates STAT3 targeting to tune cardiac response to pressure overload.

Heart failure (HF) remains a major source of morbidity and mortality in the US. The multifunctional Ca2+/calmodulin-dependent kinase II (CaMKII) has emerged as a critical regulator of cardiac hypertrophy and failure, although the mechanisms remain unclear. Previous studies have established that the cytoskeletal protein ßIV-spectrin coordinates local CaMKII signaling. Here, we sought to determine the role of a spectrin-CaMKII complex in maladaptive remodeling in HF. Chronic pressure overload (6 weeks of transaortic constriction [TAC]) induced a decrease in cardiac function in WT mice but not in animals expressing truncated ßIV-spectrin lacking spectrin-CaMKII interaction (qv3J mice). Underlying the observed differences in function was an unexpected differential regulation of STAT3-related genes in qv3J TAC hearts. In vitro experiments demonstrated that ßIV-spectrin serves as a target for CaMKII phosphorylation, which regulates its stability. Cardiac-specific ßIV-spectrin-KO (ßIV-cKO) mice showed STAT3 dysregulation, fibrosis, and decreased cardiac function at baseline, similar to what was observed with TAC in WT mice. STAT3 inhibition restored normal cardiac structure and function in ßIV-cKO and WT TAC hearts. Our studies identify a spectrin-based complex essential for regulation of the cardiac response to chronic pressure overload. We anticipate that strategies targeting the new spectrin-based "statosome" will be effective at suppressing maladaptive remodeling in response to chronic stress.

Spectrin-based pathways underlying electrical and mechanical dysfunction in cardiac disease.

INTRODUCTION: In the heart, pathways that transduce extracellular environmental cues (e.g. mechanical force, inflammatory stress) into electrical and/or chemical signals at the cellular level are critical for the organ-level response to chronic biomechanical/neurohumoral stress. Specifically, a diverse array of membrane-bound receptors and stretch-activated proteins converge on a network of intracellular signaling cascades that control gene expression, protein translation, degradation and/or regulation. These cellular reprogramming events ultimately lead to changes in cell excitability, growth, proliferation, and/or survival. Areas covered: The actin/spectrin cytoskeleton has emerged as having important roles in not only providing structural support for organelle function but also in serving as a signaling 'superhighway,' linking signaling events at/near the membrane to distal cellular domains (e.g. nucleus, mitochondria). Furthermore, recent work suggests that the integrity of the actin/spectrin cytoskeleton is critical for canonical signaling of pathways involved in cellular response to stress. This review discusses these emerging roles for spectrin and consider implications for heart function and disease. Expert commentary: Despite growth in our understanding of the broader roles for spectrins in cardiac myocytes and other metazoan cells, there remain important unanswered questions, the answers to which may point the way to new therapies for human cardiac disease patients.

A recessive mutation in beta-IV-spectrin (SPTBN4) associates with congenital myopathy, neuropathy, and central deafness.

Congenital myopathies are a heterogeneous group of muscle disorders that are often genetically determined. Here, we investigated a boy with congenital myopathy, deafness, and neuropathy from a consanguineous Kurdish family by autozygosity mapping and whole exome sequencing. We found a homozygous nonsense mutation in SPTBN4 [c.1597C>T, NM_020971.2; p.(Q533*), NP_066022.2; ClinVar SUB2292235] encoding ßIV-spectrin, a non-erythrocytic member of the ß-spectrin family. Western blot confirmed the absence of the full-length 288 kDa isoform in muscle and of a specific 72 kDa isoform in fibroblasts. Clinical symptoms of the patient largely corresponded to those described for the quivering mouse, a loss-of-function animal model. Since the human phenotype of ßIV-spectrin deficiency included a myopathy with incomplete congenital fiber-type disproportion, we investigated muscle of the quivering (qv4J) mouse and found complete absence of type 1 fibers (fiber-type 2 uniformity). Immunohistology confirmed expression of ßIV-spectrin in normal human and mouse muscle at the sarcolemma and its absence in patient and quivering (qv4J) mouse. SPTBN4 mRNA-expression levels in healthy skeletal muscle were found in the range of other regulatory proteins. More patients have to be described to confirm the triad of congenital myopathy, neuropathy and deafness as the defining symptom complex for ßIV-spectrin deficiency.

Neuronal Na+ Channels Are Integral Components of Pro-arrhythmic Na+/Ca2+ Signaling Nanodomain That Promotes Cardiac Arrhythmias During ß-adrenergic Stimulation.

BACKGROUND: Cardiac arrhythmias are a leading cause of death in the US. Vast majority of these arrhythmias including catecholaminergic polymorphic ventricular tachycardia (CPVT) are associated with increased levels of circulating catecholamines and involve abnormal impulse formation secondary to aberrant Ca2+ and Na+ handling. However, the mechanistic link between ß-AR stimulation and the subcellular/molecular arrhythmogenic trigger(s) remains elusive. METHODS AND RESULTS: We performed functional and structural studies to assess Ca2+ and Na+ signaling in ventricular myocyte as well as surface electrocardiograms in mouse models of cardiac calsequestrin (CASQ2)-associated CPVT. We demonstrate that a subpopulation of Na+ channels (neuronal Na+ channels; nNav) that colocalize with RyR2 and Na+/Ca2+ exchanger (NCX) are a part of the ß-AR-mediated arrhythmogenic process. Specifically, augmented Na+ entry via nNav in the settings of genetic defects within the RyR2 complex and enhanced sarcoplasmic reticulum (SR) Ca2+-ATPase (SERCA)-mediated SR Ca2+ refill is both an essential and a necessary factor for the arrhythmogenesis. Furthermore, we show that augmentation of Na+ entry involves ß-AR-mediated activation of CAMKII subsequently leading to nNav augmentation. Importantly, selective pharmacological inhibition as well as silencing of Nav1.6 inhibit myocyte arrhythmic potential and prevent arrhythmias in vivo. CONCLUSION: These data suggest that the arrhythmogenic alteration in Na+/Ca2+ handling evidenced ruing ß-AR stimulation results, at least in part, from enhanced Na+ influx through nNav. Therefore, selective inhibition of these channels and Nav1.6 in particular can serve as a potential antiarrhythmic therapy.

Two-Pore K+ Channel TREK-1 Regulates Sinoatrial Node Membrane Excitability.

BACKGROUND: Two-pore K(+) channels have emerged as potential targets to selectively regulate cardiac cell membrane excitability; however, lack of specific inhibitors and relevant animal models has impeded the effort to understand the role of 2-pore K(+) channels in the heart and their potential as a therapeutic target. The objective of this study was to determine the role of mechanosensitive 2-pore K(+) channel family member TREK-1 in control of cardiac excitability. METHODS AND RESULTS: Cardiac-specific TREK-1-deficient mice (αMHC-Kcnk(f/f)) were generated and found to have a prevalent sinoatrial phenotype characterized by bradycardia with frequent episodes of sinus pause following stress. Action potential measurements from isolated αMHC-Kcnk2(f/f) sinoatrial node cells demonstrated decreased background K(+) current and abnormal sinoatrial cell membrane excitability. To identify novel pathways for regulating TREK-1 activity and sinoatrial node excitability, mice expressing a truncated allele of the TREK-1-associated cytoskeletal protein ßIV-spectrin (qv(4J) mice) were analyzed and found to display defects in cell electrophysiology as well as loss of normal TREK-1 membrane localization. Finally, the ßIV-spectrin/TREK-1 complex was found to be downregulated in the right atrium from a canine model of sinoatrial node dysfunction and in human cardiac disease. CONCLUSIONS: These findings identify a TREK-1-dependent pathway essential for normal sinoatrial node cell excitability that serves as a potential target for selectively regulating sinoatrial node cell function.

Ablation of HRC alleviates cardiac arrhythmia and improves abnormal Ca handling in CASQ2 knockout mice prone to CPVT.

AIMS: Cardiac calsequestrin (CASQ2) and histidine-rich Ca-binding protein (HRC) are sarcoplasmic reticulum (SR) Ca-binding proteins that regulate SR Ca release in mammalian heart. Deletion of either CASQ2 or HRC results in relatively mild phenotypes characterized by preserved cardiac structure and function, although CASQ2 knockout (KO), or Cnull, shows increased arrhythmia burden under conditions of catecholaminergic stress. We hypothesized that given the apparent overlap of functions of CASQ2 and HRC, simultaneous ablation of both would deteriorate the cardiac phenotype compared with the single knockouts. METHODS AND RESULTS: In contrast to this expectation, double knockout (DKO) mice lacking both CASQ2 and HRC exhibited normal cardiac ejection fraction and ultrastructure. Moreover, the predisposition to catecholamine-dependent arrhythmia that characterizes the Cnull phenotype was alleviated in the DKO mice. At the myocyte level, DKO mice displayed Ca transients of normal amplitude; additionally, the frequency of spontaneous Ca waves and sparks in the presence of isoproterenol were decreased markedly compared with Cnull. Furthermore, restitution of SR Ca release was slowed in DKO myocytes compared with Cnull cells. CONCLUSION: Our results suggest that rather than being functionally redundant, CASQ2 and HRC modulate cardiac ryanodine receptor-mediated (RyR2) Ca release in an opposing manner. In particular, while CASQ2 stabilizes RyR2 rendering it refractory in the diastolic phase, HRC enhances RyR2 activity facilitating RyR2 recovery from refractoriness.

Voltage-Gated Sodium Channel Phosphorylation at Ser571 Regulates Late Current, Arrhythmia, and Cardiac Function In Vivo.

BACKGROUND: Voltage-gated Na(+) channels (Nav) are essential for myocyte membrane excitability and cardiac function. Nav current (INa) is a large-amplitude, short-duration spike generated by rapid channel activation followed immediately by inactivation. However, even under normal conditions, a small late component of INa (INa,L) persists because of incomplete/failed inactivation of a subpopulation of channels. Notably, INa,L is directly linked with both congenital and acquired disease states. The multifunctional Ca(2+)/calmodulin-dependent kinase II (CaMKII) has been identified as an important activator of INa,L in disease. Several potential CaMKII phosphorylation sites have been discovered, including Ser571 in the Nav1.5 DI-DII linker, but the molecular mechanism underlying CaMKII-dependent regulation of INa,L in vivo remains unknown. METHODS AND RESULTS: To determine the in vivo role of Ser571, 2 Scn5a knock-in mouse models were generated expressing either: (1) Nav1.5 with a phosphomimetic mutation at Ser571 (S571E), or (2) Nav1.5 with the phosphorylation site ablated (S571A). Electrophysiology studies revealed that Ser571 regulates INa,L but not other channel properties previously linked to CaMKII. Ser571-mediated increases in INa,L promote abnormal repolarization and intracellular Ca(2+) handling and increase susceptibility to arrhythmia at the cellular and animal level. Importantly, Ser571 is required for maladaptive remodeling and arrhythmias in response to pressure overload. CONCLUSIONS: Our data provide the first in vivo evidence for the molecular mechanism underlying CaMKII activation of the pathogenic INa,L. Relevant for improved rational design of potential therapies, our findings demonstrate that Ser571-dependent regulation of Nav1.5 specifically tunes INa,L without altering critical physiological components of the current.

Role of sinoatrial node architecture in maintaining a balanced source-sink relationship and synchronous cardiac pacemaking.

Normal heart rhythm (sinus rhythm) depends on regular activity of the sinoatrial node (SAN), a heterogeneous collection of specialized myocytes in the right atrium. SAN cells, in general, possess a unique electrophysiological profile that promotes spontaneous electrical activity (automaticity). However, while automaticity is required for normal pacemaking, it is not necessarily sufficient. Less appreciated is the importance of the elaborate structure of the SAN complex for proper pacemaker function. Here, we review the important structural features of the SAN with a focus on how these elements help manage a precarious balance between electrical charge generated by the SAN ("source") and the charge needed to excite the surrounding atrial tissue ("sink"). We also discuss how compromised "source-sink" balance due, for example to fibrosis, may promote SAN dysfunction, characterized by slow and/or asynchronous pacemaker activity and even failure, in the setting of cardiovascular disease (e.g., heart failure, atrial fibrillation). Finally, we discuss implications of the "source-sink" balance in the SAN complex for cell and gene therapies aimed at creating a biological pacemaker as replacement or bridge to conventional electronic pacemakers.

Ankyrin-G coordinates intercalated disc signaling platform to regulate cardiac excitability in vivo.

RATIONALE: Nav1.5 (SCN5A) is the primary cardiac voltage-gated Nav channel. Nav1.5 is critical for cardiac excitability and conduction, and human SCN5A mutations cause sinus node dysfunction, atrial fibrillation, conductional abnormalities, and ventricular arrhythmias. Further, defects in Nav1.5 regulation are linked with malignant arrhythmias associated with human heart failure. Consequently, therapies to target select Nav1.5 properties have remained at the forefront of cardiovascular medicine. However, despite years of investigation, the fundamental pathways governing Nav1.5 membrane targeting, assembly, and regulation are still largely undefined. OBJECTIVE: Define the in vivo mechanisms underlying Nav1.5 membrane regulation. METHODS AND RESULTS: Here, we define the molecular basis of an Nav channel regulatory platform in heart. Using new cardiac-selective ankyrin-G(-/-) mice (conditional knock-out mouse), we report that ankyrin-G targets Nav1.5 and its regulatory protein calcium/calmodulin-dependent kinase II to the intercalated disc. Mechanistically, ßIV-spectrin is requisite for ankyrin-dependent targeting of calcium/calmodulin-dependent kinase II-δ; however, ßIV-spectrin is not essential for ankyrin-G expression. Ankyrin-G conditional knock-out mouse myocytes display decreased Nav1.5 expression/membrane localization and reduced INa associated with pronounced bradycardia, conduction abnormalities, and ventricular arrhythmia in response to Nav channel antagonists. Moreover, we report that ankyrin-G links Nav channels with broader intercalated disc signaling/structural nodes, as ankyrin-G loss results in reorganization of plakophilin-2 and lethal arrhythmias in response to ß-adrenergic stimulation. CONCLUSIONS: Our findings provide the first in vivo data for the molecular pathway required for intercalated disc Nav1.5 targeting/regulation in heart. Further, these new data identify the basis of an in vivo cellular platform critical for membrane recruitment and regulation of Nav1.5.

Mathematical modeling of physiological systems: an essential tool for discovery.

Mathematical models are invaluable tools for understanding the relationships between components of a complex system. In the biological context, mathematical models help us understand the complex web of interrelations between various components (DNA, proteins, enzymes, signaling molecules etc.) in a biological system, gain better understanding of the system as a whole, and in turn predict its behavior in an altered state (e.g. disease). Mathematical modeling has enhanced our understanding of multiple complex biological processes like enzyme kinetics, metabolic networks, signal transduction pathways, gene regulatory networks, and electrophysiology. With recent advances in high throughput data generation methods, computational techniques and mathematical modeling have become even more central to the study of biological systems. In this review, we provide a brief history and highlight some of the important applications of modeling in biological systems with an emphasis on the study of excitable cells. We conclude with a discussion about opportunities and challenges for mathematical modeling going forward. In a larger sense, the review is designed to help answer a simple but important question that theoreticians frequently face from interested but skeptical colleagues on the experimental side: "What is the value of a model?"

Modeling CaMKII in cardiac physiology: from molecule to tissue.

Post-translational modification of membrane proteins (e.g., ion channels, receptors) by protein kinases is an essential mechanism for control of excitable cell function. Importantly, loss of temporal and/or spatial control of ion channel post-translational modification is common in congenital and acquired forms of cardiac disease and arrhythmia. The multifunctional Ca(2+)/calmodulin-dependent protein kinase II (CaMKII) regulates a number of diverse cellular functions in heart, including excitation-contraction coupling, gene transcription, and apoptosis. Dysregulation of CaMKII signaling has been implicated in human and animal models of disease. Understanding of CaMKII function has been advanced by mathematical modeling approaches well-suited to the study of complex biological systems. Early kinetic models of CaMKII function in the brain characterized this holoenzyme as a bistable molecular switch capable of storing information over a long period of time. Models of CaMKII activity have been incorporated into models of the cell and tissue (particularly in the heart) to predict the role of CaMKII in regulating organ function. Disease models that incorporate CaMKII overexpression clearly demonstrate a link between its excessive activity and arrhythmias associated with congenital and acquired heart disease. This review aims at discussing systems biology approaches that have been applied to analyze CaMKII signaling from the single molecule to intact cardiac tissue. In particular, efforts to use computational biology to provide new insight into cardiac disease mechanisms are emphasized.

ß(IV)-Spectrin regulates TREK-1 membrane targeting in the heart.

AIMS: Cardiac function depends on the highly regulated and co-ordinate activity of a large ensemble of potassium channels that control myocyte repolarization. While voltage-gated K(+) channels have been well characterized in the heart, much less is known about regulation and/or targeting of two-pore K(+) channel (K(2P)) family members, despite their potential importance in modulation of heart function. METHODS AND RESULTS: Here, we report a novel molecular pathway for membrane targeting of TREK-1, a mechano-sensitive K(2P) channel regulated by environmental and physical factors including membrane stretch, pH, and polyunsaturated fatty acids (e.g. arachidonic acid). We demonstrate that ß(IV)-spectrin, an actin-associated protein, is co-localized with TREK-1 at the myocyte intercalated disc, associates with TREK-1 in the heart, and is required for TREK-1 membrane targeting. Mice expressing ß(IV)-spectrin lacking TREK-1 binding (qv(4J)) display aberrant TREK-1 membrane localization, decreased TREK-1 activity, delayed action potential repolarization, and arrhythmia without apparent defects in localization/function of other cardiac potassium channel subunits. Finally, we report abnormal ß(IV)-spectrin levels in human heart failure. CONCLUSIONS: These data provide new insight into membrane targeting of TREK-1 in the heart and establish a broader role for ß(IV)-spectrin in organizing functional membrane domains critical for normal heart function.