Dystrophin muscle enhancer 1 is implicated in the activation of non-muscle isoforms in the skeletal muscle of patients with X-linked dilated cardiomyopathy.

X-linked dilated cardiomyopathy (XLDC) is a dystrophinopathy characterized by severe cardiomyopathy with no skeletal muscle involvement. Several XLDC patients have been described with mutations that abolish dystrophin muscle (M) isoform expression. The absence of skeletal muscle degeneration normally associated with loss of dystrophin function was shown to be due to increased expression of brain (B) and cerebellar Purkinje (CP) isoforms of the gene exclusively in the skeletal muscle of these patients. This suggested that the B and CP promoters have an inherent capacity to function in skeletal muscle or that they are up-regulated by a skeletal muscle-specific enhancer unaffected by the mutations in these patients. In this work we have analyzed the deletion breakpoints of two XLDC patients with deletions removing the M promoter and exon 1, but not affecting the B and CP promoters. Despite the presence of several muscle-specific regulatory motifs, the B and CP promoters were found to be essentially inactive in muscle cell lines and primary cultures. As dystrophin muscle enhancer 1 (DME1), the only known muscle-specific enhancer within the dystrophin gene, is preserved in these patients, we tested its ability to up-regulate the B and CP promoters in muscle cells. B and CP promoter activity was significantly increased in the presence of DME1, and more importantly, activation was observed exclusively in cells presenting a skeletal muscle phenotype. These results point to a role for DME1 in the induction of B and CP isoform expression in the skeletal muscle of XLDC patients defective for M isoform expression.     

The mouse dystrophin muscle enhancer-1 imparts skeletal muscle, but not cardiac muscle, expression onto the dystrophin Purkinje promoter in transgenic mice

A subset of patients harboring mutations in the dystrophin gene suffer from X-linked dilated cardiomyopathy (XLCM), a familial heart disease that is not accompanied by any clinical signs of skeletal muscle myopathy. As the muscle (M) isoform of dystrophin is not expressed in these patients, the absence of skeletal muscle symptoms has been attributed to expression of the brain (B) and cerebellar Purkinje (CP) isoforms of dystrophin in skeletal, but not cardiac, muscles of XLCM patients. The compensatory mechanism of dystrophin B and CP promoter upregulation is not known but it has been suggested that the dystrophin muscle enhancer from intron 1, DME-1, may be important in this activity. Previous studies have shown that the presence of the DME-1 is essential for a significant increase in dystrophin B and CP promoter activity in skeletal muscle cells in culture. Here, we demonstrate that the mouse dystrophin CP promoter drives expression of a lacZ reporter gene specifically to the cerebellar Purkinje cell layer but not to skeletal or cardiac muscle of transgenic mice. However, if the mouse counterpart of DME-1 is present in the transgene construct, the dystrophin CP promoter is now activated in skeletal muscle, but not in cardiac muscle. Our findings provide in vivo evidence for the importance of the dystrophin muscle enhancer sequences in activating the dystrophin CP promoter in skeletal muscle. Furthermore, they provide support for the model in which muscle enhancers, like DME-1, activate the dystrophin B and CP promoters in skeletal muscle, but not in cardiac muscle, of XLCM patients.     

Mutations in the dystrophin gene are associated with sporadic dilated cardiomyopathy

Dilated cardiomyopathy (DCM) is the major indication for heart transplantation. Approximately 30% of all DCM is thought to be inherited, while 70% is sporadic. Mutations in the dystrophin gene have been associated with the uncommon X-linked form of DCM. We hypothesized that missense mutations and other less severe mutations of the dystrophin gene might predispose to the common form of sporadic DCM. To test this hypothesis, 22kb of genomic dystrophin DNA was scanned with DOVAM-S in each of the 22 patients with sporadic DCM, including all 79 coding sequences and splice junctions, as well as six alternative exon 1 dystrophin isoforms (484kb, total). Three putative new mutations (IVS5+1 G>T, K18N, and F3228L) and seven polymorphisms were identified. The splice site mutation IVS5+1 is predicted to cause skipping of exon 5, which is within a region containing an actin binding site. The missense mutations occur at amino acids that display substantial evolutionary conservation. Screening of 236 control individuals failed to identify these three mutations. The three patients with putative mutations had CK-MM (creatine kinase, skeletal muscle) levels greater than 250 units while the 14 patients without mutations for which CK-MM were available had values ranging from 20 to 200. The first comprehensive mutation scanning of the exons and splice junctions of the dystrophin gene in patients with sporadic DCM presents the evidence that point mutations are associated with sporadic DCM without clinical evidence of skeletal myopathy. It may be prudent to measure CK-MM in all patients with dilated cardiomyopathy to identify candidates at high risk for dystrophin mutations.     

Dilated cardiomyopathy and new 16 bp deletion in exon 44 of the Dystrophin gene: the possible role of repeated motifs in mutation generation

Here we report a boy with dilated cardiomyopathy and severe Duchenne muscular dystrophy (DMD). The disease-causing mutation was a new 16 bp deletion in exon 44 of the dystrophin gene, which led to frameshifting and premature translation termination. This deletion in exon 44 was associated with dilated cardiomyopathy. The dystrophin region in exon 44 might be considered as one of the high-risk regions in which mutations could lead to myocardial damage, dilated cardiomyopathy, and early death. The abundance of repeated motifs was detected within the deleted segment and in the region. These sequence motifs might be involved in secondary structure formation and thus they could participate in the mutation generation.     

Dystrophin analysis in idiopathic dilated cardiomyopathy.

Idiopathic dilated cardiomyopathy (DCM) is characterised by ventricular dilatation and impaired systolic function resulting in congestive heart failure and frequently death. A dilated cardiomyopathy is common in patients with symptomatic Duchenne/Becker muscular dystrophy, a disease caused by dystrophin gene defects. However, cardiomyopathy is rarely the predominant clinical feature of this form of muscular dystrophy. To determine whether dystrophin gene defects might account for a significant number of patients with apparently isolated idiopathic DCM, we performed dystrophin gene analysis in 27 DCM patients, who were ascertained as part of a prospective study on idiopathic DCM. No dystrophin gene defects were found in our patients, whose average age was 50 years. These data suggest that dystrophin defects are not a common cause of idiopathic DCM in this age group in the absence of skeletal muscle cramps or weakness.     

Danon disease presenting with dilated cardiomyopathy and a complex phenotype

X-linked dilated cardiomyopathy (XLCM) was first described in 1987 and associated with dystrophin gene (DMD) mutations a decade later in one of the original two families. Here we report long-term follow-up of the second family (XLCM-2), for which a DMD mutation was never found. Analysis of the lysosome-associated membrane protein-2 (LAMP-2) gene detected a novel mutation, confirming a diagnosis of Danon disease. The broad phenotype in this family included dilated and hypertrophic cardiomyopathy, cardiac pre-excitation, skeletal myopathy with high serum creatine kinase, cognitive impairment (in males), and a pigmentary retinopathy in affected females. Cardiac biopsy specimens showed extensive vacuolar changes in an affected adult male. Remarkably, the skeletal muscle biopsy in a 13-month-old mutation-carrying male showed no vacuolization by standard histology. We conclude that XLCM may be the presenting sign of Danon disease and, in the presence of a familial history of HCM, pre-excitation, skeletal muscle involvement and retinal pigmentary dystrophy should prompt LAMP-2 clinical testing. Furthermore, the absence of vacuolar myopathy in biopsies from young patients may not exclude Danon disease. Supported by American Heart Association (0150453N), Muscular Dystrophy Association (PN0007-056) and NIH (1RO1HL69071-01 and 1K23Hl67915-01A1).     

Mouse dystrophin enhancer preferentially targets lacZ expression in skeletal and cardiac muscle.

Duchenne muscular dystrophy is a muscle wasting disease that results from a dystrophin deficiency in skeletal and cardiac muscle. Studies concerning the regulatory elements that govern dystrophin gene expression in skeletal and/or cardiac muscle in both mouse and human have identified a promoter and an enhancer located in intron 1. In transgenic mice, the muscle promoter alone targets the expression of a lacZ reporter gene only to the right ventricle of the heart, suggesting the need for other regulatory elements to target skeletal muscle and the rest of the heart. Here we report that the mouse dystrophin enhancer from intron 1 can target the expression of a lacZ reporter gene in skeletal muscle as well as in other heart compartments of transgenic mice. Our results also suggest that sequences surrounding the mouse dystrophin enhancer may affect its function throughout mouse development.     

心肌病患者线粒体DNA异常的探讨

心肌病的病因至今未明,它被认为是一种多原因、多因素的疾病。其中,线粒体DNA缺失也是原因之一。本文采用PCR法,对12例心肌病者的血细胞以及其中4例患者骨骼肌细胞的线粒体DNA,进行了在第11012—11980位片段内的检测,均未见有缺失,对此结果进行了初步讨论,为从分子水平进一步探讨心肌病的病因打下基础。     

Two related Dutch families with a clinically variable presentation of cardioskeletal myopathy caused by a novel S13F mutation in the desmin gene

Desmin-related myopathy is characterised by skeletal muscle weakness often combined with cardiac involvement. Mutations in the desmin gene have been described as a cause of desmin-related myopathy (OMIM 601419). We report here on two distantly related Dutch families with autosomal dominant inheritance of desmin-related myopathy affecting 15 family members. A highly heterogeneous clinical picture is apparent, varying from isolated dilated cardiomyopathy to a more generalised skeletal myopathy and mild respiratory problems. Morphological analysis of muscle biopsies revealed intracytoplasmic desmin aggregates (desmin and p62 staining). In both families we identified an identical novel pathogenic heterozygous missense mutation, S13F, in the 'head' domain of the desmin gene which cosegregates with the disease phenotype. This is the 5th reported missense mutation located at the 'head' domain of the desmin gene and the first reported Dutch family with desmin-related myopathy. This article illustrates the importance of analysing the desmin gene in patients with (familial) cardiac conduction disease, dilated cardiomyopathy and/or a progressive skeletal myopathy resembling limb-girdle muscular dystrophy.     

HnRNP G and Tra2beta: opposite effects on splicing matched by antagonism in RNA binding

The hnRNP G family comprises three closely related proteins, hnRNP G, RBMY and hnRNP G-T. We showed previously that they interact with splicing activator proteins, particularly hTra2beta, and suggested that they were involved in regulating Tra2-dependent splicing. We show here that hnRNP G and hTra2beta have opposite effects upon the incorporation of several exons, both being able to act as either an activator or a repressor. HnRNP G acts via a specific sequence to repress the skeletal muscle-specific exon (SK) of human slow skeletal alpha-tropomyosin, TPM3, and stimulates inclusion of the alternative non-muscle exon. The binding of hnRNP G to the exon is antagonized by hTra2beta. The two proteins also have opposite effects upon a dystrophin pseudo-exon. This exon is incorporated in a patient to a higher level in heart muscle than skeletal muscle, causing X-linked dilated cardiomyopathy. It is included to a higher level after transfection of a mini-gene into rodent cardiac myoblasts than into skeletal muscle myoblasts. Co-transfection with hnRNP G represses incorporation in cardiac myoblasts, whereas hTra2beta increases it in skeletal myoblasts. Both the cell specificity and the protein responses depend upon exon sequences. Since the ratio of hnRNP G to Tra2beta mRNA in humans is higher in skeletal muscle than in heart muscle, we propose that the hnRNP G/Tra2beta ratio contributes to the cellular splicing preferences and that the higher proportion of hnRNP G in skeletal muscle plays a role in preventing the incorporation of the pseudo-exon and thus in preventing skeletal muscle dystrophy.     

Unclassified polysaccharidosis of the heart and skeletal muscle in siblings

We describe a 15-year-old boy and his 19-year-old sister with progressive dilated cardiomyopathy and mild non-progressive proximal lower limb myopathy, secondary to the accumulation of amylopectin-like fibrillar glycogen, (polyglucosan) bodies, in heart and skeletal muscle. Evidence of idiopathic amylopectinosis or polysaccharidosis was demonstrated in heart and skeletal muscle tissue by histology, electron microscopy, biochemical, and genetic analysis. In both siblings the heart muscle stored PAS-positive, proteinase-k resistant and partly diastase resistant granulo-filamentous material, simulating polyglucosan bodies. Glycogen branching enzyme activity, and phosphofructokinase enzyme activity, measured in skeletal muscle tissue and explanted heart tissue were all within the normal limits, however glycogen content was elevated. Furthermore, GBE1, PRKAG2, desmin, alphabeta-crystallin, ZASP, myotilin, and LAMP-2 gene sequencing revealed no mutation, excluding e.g. glycogen storage disease type 4 and desmin-related myofibrillar cardiomyopathies. In both patients the diagnosis of an idiopathic polysaccharidosis with progressive dilated cardiomyopathy was made, requiring heart transplantation at age 13 and 14, respectively. Both patients belong to an autosomal recessive group of biochemically and genetically unclassified severe vacuolar glycogen storage disease of the heart and skeletal muscle. Up to now unidentified glycogen synthesis or glycogen degradation pathways are supposed to contribute to this idiopathic glycogen storage disease.     

Functional consequences of an LMNA mutation associated with a new cardiac and non-cardiac phenotype

Heritable dilated cardiomyopathy is a genetically highly heterogeneous disease. To date 17 different chromosomal loci have been described for autosomal dominant forms of dilated cardiomyopathy with or without additional clinical manifestations. Among the 10 mutated genes associated with dilated cardiomyopathy, the lamin A/C (LMNA) gene has been reported in forms associated with conduction-system disease with or without skeletal muscle myopathy. For the first time, we report here a French family affected with a new phenotype composed of an autosomal dominant severe dilated cardiomyopathy with conduction defects or atrial/ventricular arrhythmias, and a specific quadriceps muscle myopathy. In all previously reported cases with both cardiac and neuromuscular involvement, neuromuscular disorders preceded cardiac abnormalities. The screening of the coding sequence of the LMNA gene on all family members was performed and we identified a missense mutation (R377H) in the lamin A/C gene that cosegregated with the disease in the family. Cell transfection experiments showed that the R377H mutation leads to mislocalization of both lamin and emerin. These results were obtained in both muscular (C2C12) and non-muscular cells (COS-7). This new phenotype points out the wide spectrum of neuromuscular and cardiac manifestations associated with lamin A/C mutations, with the functional consequence of this mutation seemingly associated with a disorganization of the lamina.     

Clinical features of patients with Becker muscular dystrophy and deletions of the rod domain of dystrophin geneCSCD

Objective: To explore the clinical features of patients carrying deletions of the rod domain of the dystrophin gene. Methods: Clinical data of 12 Chinese patients with Becker muscular dystrophy (BMD) and such deletions was reviewed. Results: Most patients complained of muscle weakness of lower limbs. Two patients had muscle cramps, one had increased creatine kinase (CK) level, and one had dilated cardiomyopathy. Conclusion: Compared with DMD, the clinical features of BMD are much more variable, particularly for those carrying deletions of the rod domain of the dystrophin gene. Muscular weakness may not be the sole complaint of BMD. The diagnosis of BMD cannot be excluded by moderately elevated CK. For male patients with dilated cardiomyopathy, the possibility of BMD should be considered. © 2018 West China University of Medical Sciences. All rights reserved.