Sabine Sasse-Klaassen

Mutations in the desmosomal protein plakophilin-2 are common in arrhythmogenic right ventricular cardiomyopathy

Arrhythmogenic right ventricular cardiomyopathy (ARVC) is associated with fibrofatty replacement of cardiac myocytes, ventricular tachyarrhythmias and sudden cardiac death. In 32 of 120 unrelated individuals with ARVC, we identified heterozygous mutations in PKP2, which encodes plakophilin-2, an essential armadillo-repeat protein of the cardiac desmosome. In two kindreds with ARVC, disease was incompletely penetrant in most carriers of PKP2 mutations.

Mutations of TTN , encoding the giant muscle filament titin, cause familial dilated cardiomyopathy

Congestive heart failure (CHF) can result from various disease states with inadequate cardiac output. CHF due to dilated cardiomyopathy (DCM) is a familial disease in 20–30% of cases and is associated with mutations in genes encoding cytoskeletal, contractile or inner–nuclear membrane proteins. We show that mutations in the gene encoding giant-muscle filament titin (TTN) cause autosomal dominant DCM linked to chromosome 2q31 (CMD1G; MIM 604145). Titin molecules extend from sarcomeric Z-discs to M-lines, provide an extensible scaffold for the contractile machinery and are crucial for myofibrillar elasticity and integrity. In a large DCM kindred, a segregating 2-bp insertion mutation in TTN exon 326 causes a frameshift, truncating A-band titin. The truncated protein of approximately 2 mD is expressed in skeletal muscle, but western blot studies with epitope-specific anti-titin antibodies suggest that the mutant protein is truncated to a 1.14-mD subfragment by site-specific cleavage. In another large family with DCM linked to CMD1G, a TTN missense mutation (Trp930Arg) is predicted to disrupt a highly conserved hydrophobic core sequence of an immunoglobulin fold located in the Z-disc–I-band transition zone. The identification of TTN mutations in individuals with CMD1G should provide further insights into the pathogenesis of familial forms of CHF and myofibrillar titin turnover.

Mutant Desmocollin-2 Causes Arrhythmogenic Right Ventricular Cardiomyopathy

Arrhythmogenic right ventricular cardiomyopathy (ARVC) is a genetically heterogeneous heart-muscle disorder characterized by progressive fibrofatty replacement of right ventricular myocardium and an increased risk of sudden cardiac death. Mutations in desmosomal proteins that cause ARVC have been previously described; therefore, we investigated 88 unrelated patients with the disorder for mutations in human desmosomal cadherin desmocollin-2 (DSC2). We identified a heterozygous splice-acceptor-site mutation in intron 5 (c.631-2A-->G) of the DSC2 gene, which led to the use of a cryptic splice-acceptor site and the creation of a downstream premature termination codon. Quantitative analysis of cardiac DSC2 expression in patient specimens revealed a marked reduction in the abundance of the mutant transcript. Morpholino knockdown in zebrafish embryos revealed a requirement for dsc2 in the establishment of the normal myocardial structure and function, with reduced desmosomal plaque area, loss of the desmosome extracellular electron-dense midlines, and associated myocardial contractility defects. These data identify DSC2 mutations as a cause of ARVC in humans and demonstrate that physiologic levels of DSC2 are crucial for normal cardiac desmosome formation, early cardiac morphogenesis, and cardiac function.

Isolated noncompaction of the left ventricular myocardium in the adult is an autosomal dominant disorder in the majority of patients

Isolated noncompaction of the ventricular myocardium (INVM, MIM 300183 and 604169) is a congenital unclassified cardiomyopathy with numerous prominent trabeculations and deep intertrabecular recesses in a hypertrophied and hypokinetic myocardium. Mutations in the G4.5 gene result in a wide spectrum of severe infantile X‐linked cardiomyopathic phenotypes including Barth syndrome with dilated cardiomyopathy and INVM. Molecular genetic analysis of INVM has only been performed in pediatric patients. Although adult INVM patients show similar cardiac abnormalities, the influence of genetic factors, especially of mutations in G4.5, is unknown. We analyzed 25 adult INVM patients for the presence of mutations in the G4.5 gene and performed a pedigree analysis of probands. Mutations were not found in the coding sequence or splice sites of G4.5. Systematic analysis of relatives from seven of nine probands showed multiple affected members consistent with an autosomal dominant pattern of inheritance in the majority of cases. We conclude that INVM in the adult is an autosomal dominant disorder rarely caused by mutations in G4.5 and therefore genetically distinct from infantile X‐linked cases.

Novel Gene Locus for Autosomal Dominant Left Ventricular Noncompaction Maps to Chromosome 11p15

Background—Left ventricular noncompaction (LVNC) is a congenital unclassified cardiomyopathy with numerous prominent trabeculations and deep intertrabecular recesses in a hypertrophied and hypokinetic myocardium. It has been reported to occur in isolation or in association with congenital heart disease. Mutations in the X-linked G4.5 gene are responsible for cases of isolated LVNC in male infants, but G4.5 mutations were not found in patients with clinical onset of disease in adulthood. In addition, several families with LVNC and an autosomal dominant pattern of inheritance suggest genetic heterogeneity. Methods and Results—We performed a genome-wide linkage analysis in a family with autosomal dominant LVNC and show that a locus containing the LVNC disease gene maps to chromosome 11p15. A peak 2-point logarithm of odds score of 5.06 was obtained with marker D11S902 at &thetas;=0. Haplotype analysis defined a critical interval of 6.4 centimorgan between D11S1794 and D11S928 corresponding to a physical distance of 6.8 megabases. No disease-causing mutation was identified in 2 prime positional candidate genes, muscle LIM protein (MLP) and SOX6. Conclusions—We have mapped a locus for autosomal dominant LVNC to a 6.8-megabase region on human chromosome 11p15. Identification of the disease gene will allow genetic screening and provide fundamental insight into the understanding of myocardial morphogenesis.

Identification of a novel frameshift mutation in the giant muscle filament titin in a large Australian family with dilated cardiomyopathy

Dilated cardiomyopathy (DCM) is an etiologically heterogeneous cardiac disease characterized by left ventricular dilation and systolic dysfunction. Approximately 25–30% of DCM patients show a family history of mainly autosomal dominant inheritance. We and others have previously demonstrated that mutations in the giant muscle filament titin (TTN) can cause DCM. However, the prevalence of titin mutations in familial DCM is unknown. In this paper, we report a novel heterozygous 1-bp deletion mutation (c.62890delG) in TTN that cosegregates with DCM in a large Australian pedigree (A3). The TTN deletion mutation c.62890delG causes a frameshift, thereby generating a truncated A-band titin due to a premature stop codon (p.E20963KfsX10) and the addition of ten novel amino acid residues. The clinical phenotype of DCM in kindred A3 demonstrates incomplete penetrance and variable expressivity. Finally, protein analysis of a skeletal muscle biopsy sample from an affected member did not reveal the predicted truncated titin isoform although the aberrant mRNA was present, suggesting posttranslational modification and degradation of the truncated protein. The identification of a novel disease-causing mutation in the giant titin gene in a third large family with DCM indicates that mutations in titin may account for a significant portion of the genetic etiology in familial DCM.

Response to correspondence by Dr. Finsterer and Dr. Stöllberger: Heterogenous myopathic background of left ventricular hypertrabeculation/noncompaction

Left ventricular hypertrabeculation (LVHT) and isolated noncompaction of the left ventricular myocardium (INVM) are different and distinct entities. The presence of more than three prominent trabeculations in the left ventricle has been defined as ‘‘left ventricular hypertrabeculation’’ by the authors Stöllberger and Finsterer [2004], a definition that has not been used by others than the authors themselves. INVM as described in our article by Sasse-Klaassen et al. [2003], is characterized by hypertrophied left ventricular (LV) myocardium with hypokinetic segments consisting of two layers. There is a noncompacted endocardial layer which is thicker than the compacted epicardial layer (ratio >/1⁄42) and consists of prominent and excessive trabeculations and deep recesses filled with blood from the ventricular cavity as visualized by color Doppler imaging. In our article, we did not analyze patients with LVHT. We strongly suggest that the term INVM, which has been introduced by Chin et al. [1990] and has been accepted as ‘‘noncompacted myocardium’’ by the WHO classification on cardiomyopathies [Richardson et al., 1996], is used in the right context and not transformed to or mixed with LVHT, which is not mentioned by the WHO classification. LVHT was reported to be frequently associated with underlying extracardiac, particularly neuromuscular or metabolic disorders [Stöllberger et al., 2002; Stöllberger and Finsterer, 2004]. Heart muscle diseases, however, that are associated with neuromuscular or metabolic disorders are classified as specific cardiomyopathies [Richardson et al., 1996]. Most of the questions raised by the authors were not objectives of our study but relate to the article published previously by Oechslin et al. [2000]. We briefly respond to some concerns that are raised: our patient population of 25 probands was derived from and, thus, is not completely identical with the one presented in Oechslin et al. [2000]. Therefore, their clinical and echocardiographic characteristics are described in Table II of our article. An end-diastolic still frame picture (parasternal short axis view) was chosen for Figure 1 aswell as for the colorDoppler apical long-axis view of the same patient for the best visualization of the intertrabecular recesses. Criteria for the best visual differentiation of the two layers during end systole have been described earlier [Jenni et al., 2001]. In our study, all noncompacted segments were hypokinetic. The normally compacted segments were occasionally also hypokinetic—despite normal wall thickness— [Oechslin et al., 2000], which may be partly due to coronary microcirculatory dysfunction in association with INVM [Jenni et al., 2002]. The high rate of thromboembolism in our series of patients can be explained by the poor systolic LV function in our patients (Table II). In their patients with LVHT and neurologic abnormalities, metabolicmyopathies were foundmost frequently [Stöllberger et al., 2002]. In our patients with INVM, there were no clinical signs of a skeletal myopathy asmentioned in our article. As we had no patients with skeletal myopathies, disease-causing genes, e.g., dystrophin (DMD),were not relevant to us. There is frequent progressive cardiac involvement in different forms of muscular and myotonic dystrophies but characterized mainly by the development of dilated cardiomyopathy [Bushby et al., 2003]. The gene encoding for taffazin is calledG4.5. In our families with several affected members only the proband was screened for amutation in theG4.5gene, since it canbeassumed that the genetic cause for this rare autosomal dominant disorder in one kindred is the same.