MINI REVIEW ARTICLE published: 04 May 2012 doi: 10.3389/fphys.2012.00122 Defects in cytoskeletal signaling pathways, arrhythmia, and sudden cardiac death

Sakima Smith1,2, Jerry Curran1,Thomas J. Hund 1,2,3 and Peter J. Mohler 1,2,4*

1 Dorothy M. Davis Heart and Lung Research Institute, The Ohio State University Medical Center, Columbus, OH, USA 2 Division of Cardiovascular Medicine, Department of Internal Medicine, The Ohio State University Medical Center, Columbus, OH, USA 3 Department of Biomedical Engineering, The Ohio State University Medical Center, Columbus, OH, USA 4 Department of Physiology and Cell Biology, The Ohio State University Medical Center, Columbus, OH, USA

Edited by: polypeptides are cellular adapter that tether integral membrane proteins to Carol Ann Remme, University of the in a host of human organs. Initially identified as integral components of the Amsterdam, Netherlands cytoskeleton in erythrocytes, a recent explosion in ankyrin research has demonstrated that Reviewed by: Hugues Abriel, University of these proteins play prominent roles in cytoskeletal signaling pathways and membrane pro- Lausanne, Switzerland tein trafficking/regulation in a variety of excitable and non-excitable cells including heart and Heather S. Duffy, Beth Israel brain. Importantly, ankyrin research has translated from bench to bedside with the discov- Deaconess Medical Center, Harvard ery of human variants associated with ventricular arrhythmias that alter ankyrin–based Medical School, USA pathways. Ankyrin polypeptides have also been found to play an instrumental role in various *Correspondence: Peter J. Mohler, Dorothy M. Davis forms of sinus node disease and atrial fibrillation (AF). Mouse models of ankyrin-deficiency Heart and Lung Research Institute, have played fundamental roles in the translation of ankyrin-based research to new clinical The Ohio State University Medical understanding of human sinus node disease, AF,and ventricular tachycardia. Center, 473 W. 12th Avenue, 110G DHLRI, Columbus, OH 43210, USA. Keywords: ankyrin, , arrhythmia, cytoskeleton, mouse model e-mail: [email protected]

INTRODUCTION cell-specific binding partners and function. For example, ANK1 Ankyrin polypeptides are a family of adapter proteins that contains over 40 exons resulting in gene products ranging from link integral membrane proteins with the submembranous −30 to +200kD (Bennett and Chen, 2001). The ANK2 gene spans /spectrin-based cytoskeleton. Ankyrin-R was identified in over 560 kb with >50 exons and diverse splice forms ranging from the late 1970s as a critical link between several anion exchanger −50 to 440 kD in brain,heart,skeletal muscle and thymus (Bennett isoforms and β-spectrin in red blood cells (Bennett, 1979). Over and Chen, 2001). ANK3 encodes numerous ankyrin-G isoforms 30 years later, are now widely regarded as key players in broadly expressed in epithelial tissue, kidney, skeletal and cardiac the formation of complexes including ion channels and muscle, and brain (Bennett and Chen, 2001). While the specific transporters, cell adhesion molecules (CAMs), signaling proteins, role of alternatively spliced ankyrin isoforms is unknown, it is and cytoskeletal elements. In heart, ankyrin-associated protein likely that variability in expression contributes to organization and complexes organize specialized membrane-domains with distinct maintenance of distinct subcellular domains in eukaryotic cells. electrical and structural properties in cardiac sinus node, atrial, Canonical ankyrins are comprised of four functional domains: and ventricular cardiomyocytes. With particular emphasis given to the membrane-binding domain (MBD), the spectrin-binding ankyrin-B and ankyrin-G, this review provides important insight domain (SBD), the death domain (DD), and the C-terminal into the role of ankyrins in cardiac physiology as well as pro- domain (CTD; Figure 1; Hashemi et al., 2009). The MBD medi- viding information on the link between ankyrin dysfunction and ates the majority of ankyrin interactions with integral membrane human arrhythmias based primarily on data garnered from mouse proteins including voltage-gated Na+ and K+ channels, the Na/Ca models. (NCX) exchanger, the Na+/K+ ATPase (NKA), the ammonium transporter, the KATP channel, the inositol 1,4,5 trisphosphate ANKYRIN STRUCTURE AND FUNCTION receptor (IP3R), and the anion exchanger (Table 1; Bennett and ANK1, ANK2, and ANK3 encode three classes of ankyrin Healy, 2009; Kline et al., 2009). Furthermore, the MBD interacts polypeptides (ankyrin-R,ankyrin-B,and ankyrin-G,respectively). with CAMs including CD44, E-cadherin, and L1-CAMs. Ankyrin While ankyrin-R expression is primarily limited to erythrocytes, membrane-domains are multivalent for integral membrane pro- neurons, and skeletal muscle, both ankyrin-B and ankyrin-G are tein binding, and therefore have the ability to coordinate large widely expressed (Bennett and Chen, 2001; Cunha and Mohler, membrane-associated protein complexes. For example, in car- 2006). Although each gene is alternatively spliced to produce pro- diomyocytes, a single ankyrin-B polypeptide can form a ternary tein products of variable size, canonical ankyrin gene products complex including the NCX, NKA, and IP3R include 210 kD ankyrin-R,220 kD ankyrin-B,and 190 kD ankyrin- (Mohler et al., 2005). While there is a homology in the MBD G(Figure 1; Bennett and Chen, 2001; Cunha and Mohler, 2006). across members of the ankyrin family, each ankyrin possesses a Ankyrin genes are large, and complex splicing events determine unique set of binding partners (e.g., ankyrin-G and voltage-gated

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FIGURE 1 | Canonical ankyrin (R, B, G) domain organization.

Table 1 | Select ankyrin (R, B, G) interacting proteins and site of domain associates with the ankyrin-BMBD (Abdi et al., 2006). protein binding. Relevant for this review, the ankyrin-B “regulatory domain” is the site of the majority of human gene variants linked with arrhythmia Interacting protein Ankyrin Ankyrin domain (if known) phenotypes. ION CHANNELS/TRANSPORTERS ANKYRIN-B AND VENTRICULAR TACHYCARDIA Na/Ca exchanger B MBD Defects in ankyrin have been linked to a variety of human diseases, Voltage-gated Nav channel G MBD and play an integral role in cardiovascular disease and arrhythmia. Na/K ATPase B,G MBD In heart, a growing number of inherited arrhythmia syndromes Anion exchanger (1, 2, 3) R MBD have been linked to ankyrin dysfunction with mounting data that Kir6.2 B MBD ankyrin defects may also play a role in common forms of acquired IP3 receptor B MBD heart disease. The first such disorder linked to ankyrin dysfunc- Kv7 G MBD tion was inherited long-QT syndrome (LQTS), characterized by Ammonium transporter R, G abnormalities on the electrocardiogram (ECG), and increased ADHESION MOLECULES susceptibility to polymorphic ventricular arrhythmias and sud- Cadherin (N, E) G den death. Mutations within ion channels that are involved in L1-CAMs R, B, G MBD the generation and termination of action potentials make up the CD44 R MBD majority of inherited LQTS (Splawski et al., 2000). However, work CYTOSKELETON nearly a decade ago identified the first loss-of-function gene vari- Beta-spectrin R, B, G SBD ant in ANK2 in large pedigrees with LQTS (Mohler et al., 2003). R, B, G CTD Since this initial discovery, ANK2 loss-of-function variants are MEMBRANETRAFFICKING now well documented to cause dominantly inherited type 4 LQTS Clathrin R MBD (LQT4) and have been identified throughout the world in cohorts EHD3 B MBD with phenotypes linked with LQT4 (Mohler et al., 2004b, 2007). R, B MBD Beyond loss-of-function mutations, analysis of large human pop- -4 B SBD ulations has linked ANK2 single-nucleotide polymorphisms with MISCELLANEOUS alterations in the cardiac QT interval (Sedlacek et al., 2008) further PP2A B SBD supporting the role of ankyrin-B in cardiac electrical regulation. Hdj1 B CTD The development a mouse model heterozygous for a null muta- Fas G CTD tion in ankyrin-B (ankyrin-B+/−) was critical for understanding the pathophysiology of the human disease. In fact, ankyrin-B+/− MBD, membrane-binding domain; SBD, spectrin-binding domain; CTD, C-terminal mice phenocopy human LQT4. It is also imperative to high- domain. light that mice homozygous for null mutations in ankyrin-B (ankyrin-B−/−) also demonstrate abnormalities on ECG that mir- Na+ and K+ channels, ankyrin-B and NCX, NKA, IP3R). The rors those of LQT4 with abnormal QT rate adaptation resulting ankyrin SBD links ankyrin to the actin-based cytoskeleton via in greater QT prolongation with heart rate deceleration (Chauhan β-spectrin isoforms (Mohler et al., 2004c) thereby conferring et al., 2000 #2). In regards to the heterozygous model, ankyrin- structural stability to larger protein networks. Apart from struc- B+/−mice and human ANK2 E1425G carriers display bradycardia, tural capacities, the SBD has also been demonstrated to bind other prolonged QT interval, heart rate variability, and episodes of proteins including the regulatory subunit (B56alpha) of the PP2A isorhythmic atrio-ventricular dissociation (Mohler et al., 2003; family (Bhasin et al., 2007). Lastly, the DD and CTD together com- Le Scouarnec et al., 2008). Furthermore, similar to their human prise the ankyrin “regulatory domain.” This domain regulates the counterparts, ankyrin-B+/− mice display polymorphic ventricu- binding of ankyrin isoforms to large muscle proteins including lar tachycardia, syncope, and sudden cardiac death in response to obscurin (Kontrogianni-Konstantopoulos et al., 2003; Cunha and exercise and increased circulating catecholamines. In light of the Mohler, 2008). However, beyond intermolecular interactions, this complex phenotype presented by human ANK2 variant carriers

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that includes ventricular and supraventricular defects, the disease using optical mapping (Glukhov et al., 2010). As expected in wild- name was revised to the moniker “Ankyrin-B Syndrome.” type mice, isoproterenol accelerated the sinoatrial rate and shifted Great strides have been made over the past 10 years to establish the leading pacemaker site whereas acetylcholine had the oppo- the link between ankyrin-deficiency and increased susceptibility site effect. In contrast, following isoproterenol, ankyrin-B+/− mice to cardiac arrhythmia. At the level of the single myocyte, loss of exhibited significant beat-to-beat variability (bradycardia followed ankyrin-B results in aberrant membrane targeting and regulation by abrupt bursts of tachycardia), a disorganized shift of the leading of the NKA,NCX,KATP ,and IP3R. Loss of these proteins ultimately pacemaker, and multiple competing pacemaker foci- phenotypes produces defects in intracellular Na+ and Ca2+ regulation, that in consistent with the human sinus node disease. Ankyrin-B+/− the cardiomyocyte leads to increased sarcoplasmic reticulum cal- mouse hearts also displayed reduced sensitivity to acetylcholine. cium load, inappropriate calcium release, and early and delayed Collectively these data highlight the importance of the functional after depolarizations in the presence of high levels of circulating anatomy of the entire sinoatrial node pacemaker complex, and catecholamines. Subsequent computational modeling has further clearly demonstrate the role of ankyrin-B in cardiac automaticity supported the link between ankyrin-deficiency, loss of membrane in humans and in mice. function of NCX and NKA, intracellular Na+ and Ca2+ accumula- tion, and increased susceptibility of myocytes to develop SR Ca2+ ANKYRIN-B AND ATRIAL FIBRILLATION overload (Wolf et al.,2010). In particular,the ankyrin-deficient cel- This past year, the role of ankyrin-B in cardiovascular electro- lular phenotype was illustrated to produce frequent spontaneous physiology and disease has continued to expand by the linkage of Ca2+ release events and increased the incidence of pro-arrhythmic ankyrin-B dysfunction in human atrial fibrillation (AF; Cunha after depolarizations during rapid pacing and/or elevated cate- et al., 2011). Specifically, individuals harboring ANK2 loss-of- cholamines (Wolf et al., 2010). This biophysical phenotype was function alleles presented with a high incidence of early-onset independently confirmed in a recent report investigating Ca2+ AF, commonly progressing to permanent AF. In line with this handling in intact cardiomyocytes isolated from ankyrin-B+/− finding, continuous telemetry ECG recordings from ankyrin-B- mice (Camors et al., 2012). deficient mice revealed atrial arrhythmias similar to those observed in human carriers of ANK2 variants, including bradycardia with ANKYRIN-B AND SINUS NODE DISEASE erratic atrial activity, lack of discrete P waves, and variable ven- Beyond ventricular arrhythmias, loss-of-function variants in tricular response (Cunha et al., 2011). Ankyrin-B+/− mice also ankyrin-B have more recently been linked with sinus node dys- showed increased susceptibility to induction of AFin response to function (Le Scouarnec et al., 2008). Similar to the discovery atrial burst pacing. Primary atrial myocytes from ankyrin-B+/− of the link between ANK2 and human ventricular phenotypes, mice displayed shortened action potentials, a hallmark of AF, and large human pedigrees were initially utilized to provide genetic a sharp decrease of l-type Ca2+ current. Subsequent molecular linkage between sinus node disease and the ANK2 . Related studies determined that loss of l-type calcium current was due directly to sinus node disease, two large European cohorts mutant to loss of Cav1.3, but not Cav1.2 in atrial cardiomyocytes also ANK2 alleles were shown to display highly penetrant bradycardia explaining previously discovered reductions in I Ca,L in sinoatrial and exaggerated heart rate variability. Immunoblotting of mus- node myocytes (Le Scouarnec et al., 2008). Moreover, Cav1.3 was cle biopsies in affected individuals confirmed loss of ankyrin-B identified as a novel ankyrin-B-interacting protein, and this direct expression associated with the phenotype (Le Scouarnec et al., interaction was shown to be mediated by a short motif in the 2008). The degree of penetrance and the age of onset of sinus C-terminus. As a final link between ankyrin-B and AF, analysis node dysfunction were truly remarkable in affected individuals. of human paroxysmal AF showed decreased ankyrin-B, as well as Sinus node dysfunction is largely a disease of the elderly popu- Cav1.3 expression. Together, these data provided a new molecu- lation (Bernstein and Parsonnet, 1992). However, carriers of the lar mechanism underlying congenital AF, a new binding partner ANK2 mutation received pacemakers (devices implanted under- for ankyrin-B in heart, and implicate ankyrin-B in the pathogen- neath the skin of the clavicle with leads attached to the heart esis of common acquired forms of AF. These studies also provide designed to pace the heart at a safe heart rate if the native con- insight into how defects in a single protein can result in a broad duction system falters) at a mean age of 30-extraordinarily rare in spectrum clinical disease with unique regional manifestations. In clinical practice. Akin to these patients with ankyrin-B-associated other words, the complexity of “ankyrin-B syndrome” arises from sinus node disease, mice lacking ankyrin-B display severe sinus the fact that ankyrin-B is both broadly expressed in the heart and node dysfunction, with reduced expression of NKA, NCX, and forms distinct, region-specific macromolecular complexes. IP3R (Le Scouarnec et al., 2008). However, unlike ventricular cardiomyocytes, I Ca,L was also significantly reduced in isolated ANKYRIN-B, ISCHEMIA, AND ARRHYTHMIAS ankyrin-B+/− sinoatrial node cells, further altering calcium han- In addition to roles in congenital arrhythmias, ankyrin-B has dling and sinoatrial node automaticity (Le Scouarnec et al., 2008). more recently been linked with more common forms of heart Together these data link ankyrin-B dysfunction to sinus node dis- disease. Electrical and structural remodeling in the peri-infarct ease and provide key mechanistic insights into molecular basis for zone creates a substrate favorable for the initiation and mainte- the disease at the cellular level. Subsequent studies have provided nance of reentrant arrhythmias following myocardial infarction, new insight into the role of ankyrin-B in cardiac pacing at the in particular, ventricular fibrillation and tachycardia. An essential level of the whole heart. This past year, regulation of automatic- component of this remodeling process involves the redistribution ity in wild-type and ankyrin-B+/− mouse hearts was examined of ion channels and transporters in surviving myocytes near the

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infarct (Baba et al., 2005). Recently, ankyrin-B mRNA and pro- ANKYRIN-G AND BRUGADA SYNDROME tein levels, along with downstream ankyrin-B-associated channels While much focus has been placed on the role of ankyrin-B in were identified as differentially regulated following coronary artery heart due in part to the identified cohorts of human patients with occlusion in the canine (Hund et al., 2009). Specifically, a marked ankyrin-B mutations, the influence of ankyrin polypeptides on increase in ankyrin-B mRNA levels 5 days post-occlusion were heart function extends beyond ankyrin-B. Specifically, ankyrin- seen, presumably in response to the observed decrease in protein G has been shown to play roles in heart health and disease. levels of ankyrin-B at the same time. Protein levels and/or mem- Importantly, similar to its role in neurons, ankyrin-G has been brane expression of NCX, NKA, and IP3R were also decreased at shown to target Nav1.5, for maintenance of normal cardiac cell 5 days post-occlusion, in line with their known role as ankyrin- excitability. Ankyrin-G and Na channels directly associate and B-binding partners (Hund et al., 2009). These changes were are co-localized in heart at the intercalated disk, where neigh- observed in absence of any changes to β2-spectrin. By 14 days post- boring cells are electrically and mechanically coupled. Silencing occlusion, ankyrin-B protein levels had returned to baseline, and of ankyrin-G in myocytes results in loss of Nav1.5 expression, the recovery of NCX, NKA, and IP3R expression were, also, under- membrane targeting, and membrane current (Lowe et al., 2008). way, consistent with the normalization of electrical remodeling Defects in ankyrin-G-based targeting pathway have been linked observed in this model.While these studies focus on a chronic stage to excitable cell disease. Interestingly, ankyrin-G has emerged as a of myocardial infarction, ankyrin-B likely is involved in heart’s candidate in multiple GWAS studies as associated with bipolar dis- response to acute ischemia. Specifically, ankyrin-B plays a pivotal order (Ferreira et al., 2008 #2309; Scott et al., 2009 #5597). In heart, + role in ATP-sensitive K channel (KATP ) regulation. Ankyrin-B disruption of ankyrin-G/Nav1.5 interaction has been linked to the associates with the KATP channel via the major subunit, Kir6.2, Brugada Syndrome, an autosomal-dominant, cardiac arrhythmia a key component of the cellular apparatus required for intrinsic syndrome characterized by ST segment elevation in the precordial cardioprotection from ischemia (Li et al., 2010). Hearts and iso- leads associated with right bundle branch block and T-wave inver- lated cardiomyocytes lacking ankyrin-B display a loss of Kir6.2 sion (Antzelevitch et al., 2005). SCN5A encodes Nav1.5, and gene membrane expression resulting in decreased membrane I K,ATP variants account for nearly a third of Brugada Syndrome cases. (Li et al., 2010). KATP channels serve a cardioprotective role in SCN5A variants largely lead to biophysical defects that disrupt the ischemia via a KATP channel-mediated shortening of the action inward sodium current, which is a key current in cellular depo- potential (Suzuki et al., 2001), hence decreased expression of KATP larization during the action potential. A SCN5A variant (E1053K) could increase the susceptibility to arrhythmias. More recently, that is localized to the ankyrin-G-binding motif in Nav1.5 and dis- we have observed similar changes in ankyrin and related proteins rupts ankyrin-G/Nav1.5 association results in Brugada Syndrome (e.g., EHD family of trafficking proteins) in a variety of animal (Mohler et al., 2004a). Mechanistically, this variant disrupts nor- models of ischemic and non-ischemic disease and in human heart mal membrane targeting of Nav1.5 resulting in loss of membrane failure (Gudmundsson et al., 2010, 2012). In summary, these data sodium channel current density and impaired excitability (Mohler suggest potential new roles for ankyrin-B in the processes of elec- et al., 2004a). trical and structural remodeling in common forms of heart disease Beyond Nav1.5 membrane targeting, ankyrin-G plays addi- (myocardial infarction/heart failure). tional roles in Nav1.5 membrane regulation via recruitment of Clearly, loss of membrane ion channels and transporters is an Nav1.5 regulatory proteins. Specifically, Hund et al. (2010) recently important link between ankyrin dysfunction and disease. How- demonstrated that βIV-spectrin is co-localized and interacts with ever, recent findings suggest that ankyrin-B regulation extends ankyrin-G and Nav1.5 at the cardiac intercalated disk. More- beyond these integral membrane proteins to cardiac signaling in over, βIV-spectrin through direct binding, targets CaMKII to the health and disease. For example, in ventricular myocytes, ankyrin- intercalated disk to directly phosphorylate Nav1.5 at a specific B has also been identified as a binding partner of B56α, the residue (571) in the DI–DII linker. This study revealed that the regulatory subunit of PP2A (Bhasin et al., 2007). PP2A is a βIV-spectrin/CaMKII complex has a direct effect on membrane multifunctional serine/threonine phosphatase known to regulate excitability, and mutant βIV-spectrin mice lacking CaMKII bind- cardiac β-adrenergic signaling (Klein et al., 2003; De Arcangelis ing activity (qv3J) displayed defects in Nav1.5 phosphorylation, et al., 2008). Cardiomyocytes lacking ankyrin-B expression dis- activity, excitability, and heart function (Hund et al., 2010). Based play a marked loss of B56α (Bhasin et al., 2007). As inhibition on these initial findings, it is interesting to consider the poten- of B56α in myocytes results in RyR2 hyper-phosphorylation [by tial role of βIV-spectrin/ankyrin-G-based signaling platform in calcium/calmodulin-dependent kinase II (CaMKII)] and myocyte regulating excitable cell function in human disease (Hund et al., electrical instability, it is reasonable to speculate that defects 2010). in CaMKII-based signaling pathways may contribute to human Apart from regulating membrane excitability, ankyrin-G was ankyrin-B-linked arrhythmias (Terentyev et al., 2009). In sum- recently proposed to play secondary roles in regulation of interca- mary, work from the past decade has illustrated complex roles lated disk structural integrity. Delmar and colleagues identified for ankyrin-B in myocyte function and in disease. The multi- ankyrin-G as member of a larger complex of Nav1.5 and gap factorial nature of ankyrin-B and its broad distribution present junctions at the cardiac desmosome (Sato et al., 2011). Loss of unique challenges and opportunities as we consider the devel- ankyrin-G caused significant changes in the subcellular distribu- opment of therapies to treat patients with inherited or acquired tion and abundance of desmosomal proteins. This complex could ankyrin-deficiency and associated disease. potentially play key roles in arrhythmia susceptibility as human

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mutations in desmosomal genes have been linked with the devel- regulation and cardiac signaling. While ankyrins are linked with opment of arrhythmogenic right ventricular dysplasia (Pilichou congenital ventricular and atrial arrhythmias, current data sug- et al., 2006), characterized by inflammation, fibrosis or adiposis, gest that these proteins serve central nodal points of regulation and susceptibility to cardiac arrhythmia and sudden cardiac death for common forms of acute and chronic cardiovascular dis- (Sen-Chowdhry et al., 2010). In summary, based on the ability to ease. Moreover, based on expression, it is not unreasonable that regulate both critical electrical and structural signaling, ankyrin- ankyrins likely play key roles in other types of human cardiovas- G-based pathways have been shown to play key roles in myocyte cular disease including inflammation and metabolic and vascular physiology and likely are a central key node for human structural disease. and electrical heart disease. ACKNOWLEDGMENTS SUMMARY This work was supported in part by a grant from the Saving Tiny The past decade has illustrated critical roles for ankyrins in Hearts Society (Peter J. Mohler). This work was also supported by cardiac physiology and in disease. Unlike their classic roles in the National Institutes of Health (HL084583, HL083422 to Peter the erythrocyte, ankyrins play dynamic roles in protein target- J Mohler; HL096805 to Thomas J. Hund); and American Heart ing and stability, and secondary unexpected roles in protein Association Established Investigator Award (Peter J. Mohler).

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A., Miki, T., Uemura, 3:122. doi: 10.3389/fphys.2012.00122 a spectrum of activities in cardiomy- nexin43 at the cardiac intercalated H., Sakamoto, N., Ohmoto-Sekine, This article was submitted to Frontiers in ocytes. Circulation 115, 432–441. disc. Circ. Res. 109, 193–201. Y., Tamagawa, M., Ogura, T., Seino, Cardiac Electrophysiology, a specialty of Mohler, P. J., Rivolta, I., Napolitano, Scott, L. J., Muglia, P., Kong, X. Q., S., Marban, E., and Nakaya, H. Frontiers in Physiology. C., Lemaillet, G., Lambert, S., Pri- Guan, W., Flickinger, M., Upmanyu, (2001). Functional roles of car- Copyright © 2012 Smith, Curran, Hund ori, S. G., and Bennett, V. (2004a). R., Tozzi, F., Li, J. Z., Burmeister, diac and vascular ATP-sensitive and Mohler. This is an open-access Nav1.5E. 1053K mutation causing M., Absher, D., Thompson, R. C., potassium channels clarified by article distributed under the terms of Brugada syndrome blocks binding to Francks,C.,Meng,F.,Antoniades,A., Kir6.2-knockout mice. Circ. Res. 88, the Creative Commons Attribution Non ankyrin-G and expression of Nav1.5 Southwick, A. M., Schatzberg, A. F., 570–577. Commercial License, which permits non- on the surface of cardiomyocytes. Bunney, W. E., Barchas, J. D., Jones, Terentyev, D., Belevych, A. E., Teren- commercial use, distribution, and repro- Proc. Natl. Acad. Sci. U.S.A. 101, E. G., Day, R., Matthews, K., McGuf- tyeva, R., Martin, M. M., Malana, duction in other forums, provided the 17533–17538. fin, P., Strauss, J. S., Kennedy, J. L., G. E., Kuhn, D. E., Abdellatif, M., original authors and source are credited.

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