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provided by Elsevier - Publisher Connector Journal of the American College of Cardiology Vol. 54, No. 4, 2009 © 2009 by the American College of Cardiology Foundation ISSN 0735-1097/09/$36.00 Published by Elsevier Inc. doi:10.1016/j.jacc.2009.04.029

Because was previously found to be associated with EDITORIAL COMMENT both hypertrophic (HCM) and (DCM) (6–8), CARP, as part of the titin complex, was also hypothesized to play a role in cardiomy- Phenotypic Heterogeneity of opathies. In this issue of the Journal, 2 reports (1,2) confirm this hypothesis and show that in fact ANKRD1 mutations Sarcomeric Mutations can cause both DCM and HCM. Arimura et al. (2) report the results of the ANKRD1 A Matter of Gain and Loss?* mutation screening in a large HCM population collected in Japan and in the U.S. In 384 index patients, they found 3 Luisa Mestroni, MD missense mutations (ANKRD1 Pro52Ala, Thr123Met, and Ϸ Aurora, Colorado; and Trieste, Italy Ile280Val), accounting for 1% of HCM cases. Interest- ingly, they also investigated the N2A CARP-binding do- main of titin, and found 2 additional mutations (TTN Arg8500 and Arg8604Gln) in their HCM cohort. Moulik After several decades of intense research and various at- et al. (1) investigated a series of 208 DCM index patients of tempts at definition and classification, Japanese and U.S. origin, and found 3 missense mutations still remain disorders of remarkable and intriguing complex- (ANKRD1 Pro105Ser, which was recurrent in 2 families, ity. Once more, this aspect is elicited by the recent discovery Val107Leu, and Met1841Ile) accounting for 2% of DCM that mutations in the cardiac cases, further supporting a role of the titin mechanotrans- (CARP), a protein functionally part of the , can duction complex in the pathogenesis of cardiomyopathies. cause different types of cardiomyopathies, as reported by But how to explain 2 different cardiomyopathies with Moulik et al. (1) and Arimura et al. (2) in this issue of the opposite pathophysiology caused by the same gene? Journal, as well as congenital heart disease (3). Phenotypic heterogeneity in cardiomyopathies. Pheno- typic heterogeneity (also called “allelic variants” in OMIM) See pages 325 and 334 (9) is a well-known and common phenomenon in genetics, referring to the occurrence of more than 1 phenotype caused ANKRD1 in normal heart and disease. CARP is a 36 kD by allelic mutations at a single locus (10); examples familiar protein encoded by the cardiac ankyrin repeat domain 1 to cardiologists are Duchenne and Becher muscular dystro- gene ANKRD1, which maps on 10. ANKRD1 phies caused by the same dystrophin gene, laminopathies is a member of a conserved gene family, coding for muscle ranging from progeria to lipodystrophy due to lamin A/C ankyrin repeat (MARPs), involved in muscle stress gene, and LQT syndrome and congenital conduction defect response such as stretch, injury, and hypertrophy (4). CARP caused by the cardiac sodium channel gene SCN5A. is a nuclear cofactor, a signaling molecule The reason for the clinical variability in allelic disorders predominantly expressed in the heart. CARP is found in the lies in the different function of the mutant proteins. In the sarcomere, where it colocalizes with the N2A domain of case of sarcomeric , it appears that a “gain” of function titin and myopalladin in the I-band of the Z disk (Fig. 1), usually results in increased energy demand, inefficient aden- and in the nucleus (4). The expression of CARP is con- osine triphosphate utilization, and hypertrophy, whereas a “loss” of function results in decreased contractility (Table 1). trolled, at least in part, by the titin-based mechanotrans- The 2 studies published in this issue seem to follow the rule. duction signaling pathway, and it is increased in heart Arimura et al. (2) show that ANKRD1 mutations in HCM development and conditions of injury and stress. In heart increase binding of CARP to titin and myopalladin, and development, CARP acts as a transcriptional repressor of that titin mutations at the CARP-binding site have the myocyte contractile elements. In , CARP is same effect. Conversely, Moulik et al. (1) show that overexpressed, suggesting a role in the “fetal gene program” ANKRD1 mutations in DCM cause a loss of CARP characteristic of the molecular remodeling of the failing binding to talin 1, potentially leading to loss of stretch- heart (5). sensing, disruption of the link between titin complex and cytoskeletal network, and transcriptional deregulation of genes involved in cell cycle and other pathways. *Editorials published in the Journal of the American College of Cardiology reflect the However, gain and loss are not the only mechanisms views of the authors and do not necessarily represent the views of JACC or the American College of Cardiology. involved in the phenotypic heterogeneity of ANKRD1. From the University of Colorado Cardiovascular Institute and Adult Medical Indeed, a recent publication by Cinquetti et al. (3) reports Genetics Program, University of Colorado at Denver and Health Sciences Center, the identification of increased CARP expression or pro- Aurora, Colorado; and Cluster in BioMedicine, Trieste, Italy. Supported by the National Institutes of Health (HL69071-01, MO1 RR00051-1575), American Heart tein stability in 3 cases with total anomalous pulmonary Association 0150453N, and Association PN0007-056. venous return, a rare congenital heart defect characterized 344 Mestroni JACC Vol. 54, No. 4, 2009 ANKRD1 and Heart Disease July 21, 2009:343–5

Figure 1 Model for the Titin-N2A Signaling Complex

The sequence of N2A titin interacts with muscle ankyrin repeat proteins (MARPs) such as cardiac ankyrin repeat protein (CARP), ANKRD2, or diabetes-related ankyrin repeat protein. Myopalladin associates with the MARP/N2A complex by interacting with the N-terminal domains of MARPs. CAPN3 ϭ calpain 3; Ig ϭ immu- noglobulin; IS ϭ muscle specific sequence; NH2 ϭ N-terminal domain; PEVK ϭ sequence element rich in (P), glutamic acid (E), lysine (K), and (V) residues. Reprinted with permission from Miller et al. (4). by failure of the pulmonary veins to connect to the left Reprint requests and correspondence: Dr. Luisa Mestroni, atrium during development. In this case, CARP overex- Molecular Genetics, CUCVI, 12700 East 19th Avenue, Mail Stop pression or its increased activity are believed to repress F442, Aurora, Colorado 80045-6511. E-mail: Luisa.Mestroni@ normal cardiac , leading to abnormal ucdenver.edu. heart development. Impact of ANKRD1 mutations discovery in clinical REFERENCES care. The discovery of ANKRD1 mutations in cardiomy- opathies has several implications. First, it contributes to fill 1. Moulik M, Vatta M, Witt SH, et al. ANKRD1, the gene encoding the gap of the large number of patients in whom the cause cardiac ankyrin repeat protein, is a novel dilated cardiomyopathy gene. Ϸ J Am Coll Cardiol 2009;54:325–33. of cardiomyopathy is still unknown, 40% of cases in 2. Arimura T, Bos JM, Sato A, et al. Cardiac ankyrin repeat protein gene HCM and probably Ϸ70% in DCM (11). Second, it (ANKRD1) mutations in hypertrophic cardiomyopathy. J Am Coll expands our knowledge of the mechanisms leading to Cardiol 2009;54:334–42. 3. Cinquetti R, Badi I, Campione M, et al. Transcriptional deregulation hypertrophy and heart failure to include abnormal stretch- and a missense mutation define ANKRD1 as a candidate gene for total based signaling in response to force: this appears to be anomalous pulmonary venous return. Hum Mutat 2008;29:468–74. another “common pathway” for HCM and DCM that could 4. Miller MK, Bang ML, Witt CC, et al. The muscle ankyrin repeat proteins: CARP, ankrd2/Arpp and DARP as a family of titin filament- be targeted by novel therapeutic strategies. Finally, it raises based stress response molecules. J Mol Biol 2003;333:951–64. the question of clinical genetic testing of ANKRD1 in 5. Zolk O, Frohme M, Maurer A, et al. Cardiac ankyrin repeat protein, HCM and DCM patients. Although the low prevalence of a negative regulator of cardiac gene expression, is augmented in human heart failure. Biochem Biophys Res Commun 2002;293:1377–82. mutations may currently limit routine screening for the 6. Itoh-Satoh M, Hayashi T, Nishi H, et al. Titin mutations as the ANKRD1 gene, we may expect that the implementation of molecular basis of dilated cardiomyopathy. Biochem Biophys Res resequencing technology will allow a systematic screening Commun 2002;291:385–91. 7. Gerull B, Gramlich M, Atherton J, et al. Mutations of TTN, encoding for rare cardiomyopathy genes in these patients in the near the giant muscle filament titin, cause familial dilated cardiomyopathy. future. Nature Genet 2002;30:201–4. JACC Vol. 54, No. 4, 2009 Mestroni 345 July 21, 2009:343–5 ANKRD1 and Heart Disease

PhenotypicTable 1 HeterogeneityPhenotypic Heterogeneity of Sarcomeric of Genes Sarcomeric and Differential Genes and Changes Differential of Mutant Changes Protein of Mutant Function Protein Function

Gene Protein DCM Function HCM Function Other Allelic Disorders Thick filament MYH7 Cardiac ␤ myosin heavy Yes 2Maximal force generation Yes 1Maximal force Laing distal , myosin chain 2Contractility generation storage myopathy, 2Velocity of actin sliding 1Caϩϩ sensitivity scapuloperoneal myopathy, left ventricular noncompaction, endocardial fibroelastosis MYH6 Cardiac ␣ myosin heavy Yes * Yes * Atrial septal defect chain MYL2 Regulatory myosin light Not described Yes 1Caϩϩ sensitivity chain MYL3 Essential myosin light Not described Yes 1Caϩϩ sensitivity chain MYBPC3 Cardiac myosin-binding Yes Yes Hypertrophy diastolic protein C dysfunction Thin filament TNNT2 Cardiac troponin T Yes 2Myofibrillar function Yes 1Myofibrillar function Restrictive cardiomyopathy 2Caϩϩ sensitivity 1Caϩϩ sensitivity TNNTI3 Cardiac troponin I Yes 2Myofibrillar function Yes 1Myofibrillar function Restrictive cardiomyopathy 2Binding to cTnT 1Caϩϩ sensitivity _( Caϩϩ sensitivity) TNNC1 Cardiac troponin C Yes 2Myofibrillar function Yes 1Myofibrillar function 2Caϩϩ sensitivity 2PKC effect TPM1 Tropomyosin 1 ␣ chain Yes 2Caϩϩ sensitivity Yes 1Myofibrillar function 2Maximum force 1Caϩϩ sensitivity ACTC1 ␣-cardiac actin Yes 2␣-Cardiac actinin (Z-disk) Yes Impaired actomyosin Restrictive cardiomyopathy affinity binding 2Force transmission Titin filament and Z-disk TTN Titin Yes 2Binding to actinin and Yes 1Binding to actinin and Tibialis muscular dystrophy or Udd Tcap Tcap distal myopathy; hereditary myopathy with early respiratory failure; recessive limb-girdle muscular dystrophy type 2J TCAP Titin-cap or telethonin Yes 2Binding to titin, MLP, and Yes 1Binding to titin and Limb-girdle muscular dystrophy myozenin-2 myozenin-2 type 2G ANKRD1 Cardiac ankyrin repeat Yes 2Binding to talin1 Yes 1Binding to titin and Total anomalous pulmonary venous protein myopalladin return1 ( expression) LDB3 Cypher/ZASP Yes Cytoskeletal disarray, Not described Left ventricular noncompaction, 2PKC affinity myofibrillar myopathy CSRP3 MLP Yes Yes 2Binding to actinin MYOZ2 Myozenin-2 Not described Yes OBSCN Obscurin Not described Yes 2Binding to titin

Data from Matsumoto et al. (8), McKusick (9), Mizra et al. (12), Kaski et al. (13), Debold et al. (14), Rajan et al. (15), and Morimoto (16). *The ␤ myosin heavy chain mutations found in hypertrophic cardiomyopathy (HCM) and dilated cardiomyopathy (DCM) were studied in ␣ myosin heavy chain of knock-in mouse models. MLP ϭ muscle LIM-domain protein; PKC ϭ protein kinase C; ZASP ϭ Z-band alternatively spliced PDZ motif-containing protein.

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