X-linked Dystrophy Research Articles

Clinical and Genetic Characterization of Female Dystrophinopathy

Background and PurposeDuchenne and Becker muscular dystrophies are the most common X-linked recessive muscular dystrophies. Dystrophin gene mutations usually affect men, but reportedly 2.5-7.8% of women are affected and are classified as symptomatic carriers. The aim of this study was to clinically and genetically characterize symptomatic female dystrophinopathy carriers.MethodsThe clinical and genetic data of 11 female dystrophinopathy carriers among 285 patients who underwent multiplex ligation-dependent probe amplification (MLPA) analysis for the dystrophin gene were reviewed. Women with muscle weakness and/or dilated cardiomyopathy were classified as symptomatic carriers, while subjects with high serum creatine kinase (CK) levels and/or minor myopathic signs such as muscle cramps and myalgia were classified as asymptomatic.ResultsTwelve female carriers were identified, but 1 symptomatic carrier who also had Turner syndrome was excluded from the study. Of the 11 included female carriers, 4 were symptomatic and 7 were asymptomatic. The age at symptom onset in the symptomatic female carriers ranged from 15 to 31 years (mean, 30.6 years), and the age at diagnosis for asymptomatic carriers ranged from 4 to 38 years (mean, 24.5 years). Serum CK levels were markedly elevated (mean, 1,301 IU/mL) in three of the four (75%) symptomatic female carriers, and mildly elevated in three of the seven (42%) asymptomatic female carriers. Symptomatic female carriers typically presented with asymmetric bilateral leg weakness as the initial symptom, with aggravated symptoms after labor.ConclusionsFemale dystrophinopathy is not uncommon, and it is an important factor with respect to males with dystrophinopathy who may be born to such patients. Screening with MLPA is useful because it can aid in early diagnosis and appropriate management.

Domain Wise Distribution of Mutations in Dystrophin Protein and Duchenne Muscular Dystrophy

Duchenne muscular dystrophy, the most common inherited X-linked recessive muscular dystrophy, affects 20000 new borns per year globally. Since its discovery in 1860, extensive research works have been carried out to understand the complex architecture of the disease formation. Reason behind the onset of the disease has been mapped back to the set of mutations of different types in dystrophin gene (DMD, 2.4 million bp), the largest gene in the body. Dystrophin (Dp), the cytosolic protein acts as the root of the complex which was primarily thought to link extracellular matrix with cellular actin cytoskeleton but later on has been associated with the stability of the cells, signal transduction as well as in proper development. In this review, we gathered the details of all the mutations occurring in DMD gene and observed that majority of the mutations are present in the N terminal Actin Binding Domain. Some of the mutations were found to be present in the Cysteine Rich Domain of the protein, reflecting the point that these two domains are the most mutation prone regions contributing to Duchenne Muscular Dystrophy (DMD) onset. This review therefore gives an integrative view describing the involvements of Dp in regulating the complexity of DMD disease with a future aspect to study the structural details of DMD genes along with its genetic variations.

Update of Emerinopathies’ clinical-genetic spectrum: the French network experience

Emerinopathies include diseases caused by EMD gene mutations localized on chromosome X and encoding emerin, an integral protein of the nuclear envelope. The most frequent emerinopathy is the X-linked EmeryDreifuss muscular dystrophy (X-EDMD) that was first reported in the 60ths by Emery and Dreifuss [1]. The disease is characterized by muscle weakness and wasting usually with a humeroperoneal distribution in the first stages, early joint contractures involving Achilles tendons, elbows, neck and the whole spine, and cardiac involvement featuring conduction defects, arrhythmias, subsequent cardiomyopathy (usually dilated) and frequently responsible of sudden death. Bione et al. [2] identified the first mutations of EMD gene encoding emerin to be responsible of X-EDMD. These mutations usually lead to absence or reduced level of emerin in different tissues of affected males including skeletal muscle, skin, oral mucosa cells and lymphocytes as it is demonstrated by immunocytochemical and histochemical methods [3,4]. Female carriers exhibit mosaic expression patterns with usually normal emerin amounts [3]. These methods may thus be used in the diagnostic strategy of X-EDMD prior to EMD gene analysis. In a recent study (unpublished work from the French network and LBGM) aiming to assess the diagnostic utility of emerin study by western blot on lymphoblastoid cell lines, we looked at EMD mutation rate observed in a cohort of 269 male and female patients with variable emerin amounts. In male patients, absence or severe reduction of emerin (<5%) lead to EMD mutation identification in all cases, while moderate emerin amount reduction revealed EMD mutation in 75% of the patients. Interestingly, in all cases where emerin amounts were considered as normal, no EMD mutations were found. In female cases, all cases with emerin moderate or severe reduction harbored EMD mutation. When emerin is normal, EMD mutation was found in 58% of female cases. These results suggest that a diagnostic rate of 100% may be reached if emerin study by western blot is performed prior to EMD gene screening in male patients. Moreover, emerin gene mutations have been rarely observed in rare cases of isolated cardiac disease [5,6] and limb girdle muscular dystrophies with cardiac disease and without joint contractures [7,8].

Identification of a novel human LAP1 isoform that is regulated by protein phosphorylation.

Lamina associated polypeptide 1 (LAP1) is an integral protein of the inner nuclear membrane that is ubiquitously expressed. LAP1 binds to lamins and chromatin, probably contributing to the maintenance of the nuclear envelope architecture. Moreover, LAP1 also interacts with torsinA and emerin, proteins involved in DYT1 dystonia and X-linked Emery-Dreifuss muscular dystrophy disorder, respectively. Given its relevance to human pathological conditions, it is important to better understand the functional diversity of LAP1 proteins. In rat, the LAP1 gene (TOR1AIP1) undergoes alternative splicing to originate three LAP1 isoforms (LAP1A, B and C). However, it remains unclear if the same occurs with the human TOR1AIP1 gene, since only the LAP1B isoform had thus far been identified in human cells. In silico analysis suggested that, across different species, potential new LAP1 isoforms could be generated by alternative splicing. Using shRNA to induce LAP1 knockdown and HPLC-mass spectrometry analysis the presence of two isoforms in human cells was described and validated: LAP1B and LAP1C; the latter is putatively N-terminal truncated. LAP1B and LAP1C expression profiles appear to be dependent on the specific tissues analyzed and in cultured cells LAP1C was the major isoform detected. Moreover, LAP1B and LAP1C expression increased during neuronal maturation, suggesting that LAP1 is relevant in this process. Both isoforms were found to be post-translationally modified by phosphorylation and methionine oxidation and two LAP1B/LAP1C residues were shown to be dephosphorylated by PP1. This study permitted the identification of the novel human LAP1C isoform and partially unraveled the molecular basis of LAP1 regulation.

Laparoscopic guided local injection in the X-linked muscular dystrophy mouse (mdx) diaphragm. An advance in experimental therapies for Duchenne Muscular Dystrophy.

To investigate the development of a laparoscopy technique for local injection into the X-linked muscular dystrophy (mdx) diaphragm. It was used 10 mice Balb/C57 and 5 mdx mice and three differents decubitus type were tested: the right lateral, supine, and supine decubitus with 20 degrees elevation of the forelimb. Abdominal caudal face and the 10 intercostal space were tested as spot to introduce the needle into the diaphragm. Supine position with elevation of 20 degrees forelimb and the 10th intercostal space are the beneficial position to apply a local injection. It was proved to be possible to perform the laparoscopy technique in the X-linked muscular dystrophy diaphragm and this requires a specific position and technique during the surgery.

Characterization of dystrophin deficient rats: a new model for Duchenne muscular dystrophy.

A few animal models of Duchenne muscular dystrophy (DMD) are available, large ones such as pigs or dogs being expensive and difficult to handle. Mdx (X-linked muscular dystrophy) mice only partially mimic the human disease, with limited chronic muscular lesions and muscle weakness. Their small size also imposes limitations on analyses. A rat model could represent a useful alternative since rats are small animals but 10 times bigger than mice and could better reflect the lesions and functional abnormalities observed in DMD patients. Two lines of Dmd mutated-rats (Dmdmdx) were generated using TALENs targeting exon 23. Muscles of animals of both lines showed undetectable levels of dystrophin by western blot and less than 5% of dystrophin positive fibers by immunohistochemistry. At 3 months, limb and diaphragm muscles from Dmdmdx rats displayed severe necrosis and regeneration. At 7 months, these muscles also showed severe fibrosis and some adipose tissue infiltration. Dmdmdx rats showed significant reduction in muscle strength and a decrease in spontaneous motor activity. Furthermore, heart morphology was indicative of dilated cardiomyopathy associated histologically with necrotic and fibrotic changes. Echocardiography showed significant concentric remodeling and alteration of diastolic function. In conclusion, Dmdmdx rats represent a new faithful small animal model of DMD.

Attenuation of Wnt/β-catenin activity reverses enhanced generation of cardiomyocytes and cardiac defects caused by the loss of emerin.

Mutations in EMD, encoding emerin cause skeletal muscle and heart defects in patients with X-linked Emery-Dreifuss muscular dystrophy (X-EDMD) but the underlying mechanisms leading to cardiac defects are poorly understood. Here, we investigated the role of emerin in controlling cardiomyocyte proliferation and cardiac remodeling and explored its function in regulation of the Wnt/β-catenin pathway. We observed a remarkable increase of cardiomyocytes in emerin-null adult mice accompanied with decreased numbers of multinucleated cells. Depletion of emerin in mouse ES cell-derived cardiomyocytes by shRNA caused hyperactivation of Wnt/β-catenin signaling, increased proliferation and abrogated timely cardiac differentiation. Likewise, emerin-null mice exhibited increased Wnt/β-catenin signaling, cardiac dysfunction and perturbed hypertrophic remodeling following pressure overload. Pharmacological inhibition of β-catenin normalized proliferation and differentiation of ES cell-derived cardiomyocytes while inactivation of a single allele of β-catenin efficiently rescued cardiac dysfunction in emerin-null mice. We conclude that emerin constrains β-catenin signaling in the heart providing tight control of cardiomyocyte numbers. Enhanced Wnt/β-catenin signaling seems to contribute to cardiac defects observed in X-EDMD. Hence, therapeutic inhibition of Wnt/β-catenin signaling might be beneficial for X-EDMD patients.

G.P.142: A new EMD gene missense mutation in exon 1 leads to absence of emerin and is responsible for X-linked dilated cardiomyopathy with conduction defects and arrhythmias and almost elusive skeletal muscle features

Mutations in the EMD gene, encoding emerin, lead to X-linked Emery-Dreifuss muscular dystrophy (EDMD). Although always associated with EDMD, an almost exclusive cardiac involvement was found in exceptional patients carrying EMD gene mutations. We here report 2 brothers belonging to a family where 9 affected male subjects developed dilated cardiomyopathy (DCM) with conduction defects and arrhythmias. The oldest brother of 53years developed DCM since 49years and requiring implantable cardioverter defibrillator (ICD) implantation. He subsequently developed atrio-ventricular and ventricular conduction defects as well as ventricular arrhythmias at 52years that required ICD upgrading and additional pacemaker implantation. His youngest brother aged 45years was also affected. The family history showed several affected members: a third brother who died after cardiac transplantation at 38years, a fourth brother who died suddenly at 50years, two maternal uncles and cousins died in their 5th or 6th decades among them 2 were diagnosed as arrhythmogenic right ventricular dysplasia. An X-linked mode of inheritance was therefore suspected and DMD gene analysis did not show any exonic deletion or duplication or exon 1 point mutation. Further neurological assessment of the oldest brother showed neither muscle weakness nor joint contractures whereas hands and legs were slightly hypotrophic. CPK were at normal level and needle electromyography examination showed normal pattern. Deltoid muscle biopsy was normal as were protein immunohistochemical studies including dystrophin, desmin, myotilin and caveolin 3 while emerin staining was totally absent. Subsequent EMD gene analysis revealed a new missense mutation in exon 1 (c.65C>T, p.Pro22Leu). In addition to the 2 previously reported families with a close emerin mutation (p.delK37) and an exclusive cardiac disease, our family further indicates that emerin protein is crucial for correct cardiac function.

Generation of muscular dystrophy model rats with a CRISPR/Cas system.

Duchenne muscular dystrophy (DMD) is an X-linked lethal muscle disorder caused by mutations in the Dmd gene encoding Dystrophin12. DMD model animals, such as mdx mice and canine X-linked muscular dystrophy dogs, have been widely utilized in the development of a treatment for DMD3. Here, we demonstrate the generation of Dmd-mutated rats using a clustered interspaced short palindromic repeats (CRISPR)/Cas system, an RNA-based genome engineering technique that is also adaptive to rats. We simultaneously targeted two exons in the rat Dmd gene, which resulted in the absence of Dystrophin expression in the F0 generation. Dmd-mutated rats exhibited a decline in muscle strength, and the emergence of degenerative/regenerative phenotypes in the skeletal muscle, heart, and diaphragm. These mutations were heritable by the next generation, and F1 male rats exhibited similar phenotypes in their skeletal muscles. These model rats should prove to be useful for developing therapeutic methods to treat DMD.

Whole exome sequencing identifies a novel EMD mutation in a Chinese family with dilated cardiomyopathy.

BackgroundVariants in the emerin gene (EMD) were implicated in X-linked recessive Emery-Dreifuss muscular dystrophy (EDMD), characterized by early-onset contractures of tendons, progressive muscular weakness and cardiomyopathy. To date, 223 mutations have been reported in EMD gene and the majority of them caused a predominant skeletal muscular phenotype. In this study, we identified a novel deletion mutation in EMD exon 1, which results in almost a complete loss of emerin protein in a large Chinese family. However, the patients suffered severe dilated cardiomyopathy (DCM) but very mild skeletal muscle disorder.Case presentationWhole exome sequencing (WES) and linkage analysis were performed to identify the underlying mutation in a Chinese DCM family spanning five generations. A missense variation in the GPR50 gene was found co-segregated with the disease phenotype, whereas no functional alteration was detected in the variant GPR50 protein. When analyzing the failure sequences in the exome sequencing data, a novel deletion mutation (c.26_39delATACCGAGCTGACC) in EMD exon 1, was identified in this family. Different from the typical clinical features caused by most reported EMD mutations, patients in our study presented very mild skeletal muscle degeneration that had not been diagnosed until the mutation was found.ConclusionWe described a family with rare clinical presentations caused by a novel EMD deletion mutation. Our findings broaden the heterogeneous spectrum of phenotypes attributed to EMD mutations and provide new insight to explain the genotype-phenotype correlations between EMD mutations and EDMD symptoms.

New pathobiochemical insights into dystrophinopathy from the proteomics of senescent mdx mouse muscle.

Primary abnormalities in the dystrophin gene cause X-linked muscular dystrophy, a highly progressive muscle wasting disorder of childhood. A spontaneous animal model of Duchenne muscular dystrophy is the mdx mouse, which presents a highly interesting phenotype that exhibits considerable variations in the degree of fiber degeneration in different subtypes of muscles. The idea that aging exacerbates the dystrophic mdx phenotype, as previously indicated by a large number of biochemical and cell biological studies, was clearly confirmed by comparative muscle proteomics. Here we outline recent findings of age-dependent changes in the dystrophin-deficient muscle proteome and contrast these results with the previously established proteomic profile of sarcopenic muscle. Besides comparable perturbations of various biochemical functions, especially striking are similarities in the cellular stress response associated with a drastic up-regulation of small αB-crystallin-like heat shock proteins. Hence, the comparison of large-scale proteomic data sets of natural muscle aging with dystrophic sarcopenia promises to shed light on the differential effect of sarcopenia of old age vs. senescent abnormalities on a mutant dystrophic background.

Blunted cardiac beta-adrenergic response as an early indication of cardiac dysfunction in Duchenne muscular dystrophy

To determine whether altered beta-adrenergic responses contribute to early cardiac dysfunction in mdx (X-linked muscular dystrophy) mice, an animal model for human Duchenne muscular dystrophy. Replacement fibrosis in mdx hearts gradually increased with age, suggesting a gradual loss of cardiomyocytes. Echocardiography and intra-left ventricular haemodynamic measurements detected baseline cardiac dysfunction in mdx mice at ≥8 months. However, a reduction of cardiac beta-adrenergic response to isoproterenol (ISO) was already present in mdx mice at 4 months. Ventricular myocytes (VMs) isolated from 4- and 8-month-old mdx mice had greater baseline contractile function {fractional shortening, [Ca(2+)]i, and sarcoplasmic reticulum (SR) Ca(2+) content} and ICa-L than age-matched control VMs and than myocytes isolated from 2-month-old mdx mice. ISO increased myocyte function in the VMs of 4- and 8-month-old mdx mice to the same level as in age-matched control VMs. In the VMs of 12-month-old mdx mice, ISO failed to increase myocyte function to the level in VMs of 12-month-old control mice and could not further increaseICa-L. No differences were observed in the expression of Cav1.2α1c, Cav1.2β1, Cav1.2β2, sarco/endoplasmic reticulum Ca(2+) ATPase (SERCA), and the Na(+)/Ca(2+) exchanger. In contrast, total ryanodine receptor 2 (RyR2) and basal phosphorylation of RyR2, phospholamban, and Cav1.2α1c were found to be increased in hearts of 4-month-old mdx mice; baseline protein kinase A activity was also increased. After ISO treatment, phosphorylation levels were the same in mdx and control hearts. VMs of 4-month-old mdx mice had reduced beta1-adrenergic receptor (β1-AR) density and beta-adrenergic sensitivity. In young mdx mice, the myocyte increases its contractile function to compensate for myocyte loss. However, these myocytes with enhanced baseline function have reduced potential for stimulation, decreased β1-AR density/sensitivity, leading to blunted cardiac beta-adrenergic response.

Proteomic profiling of the dystrophin-deficient mdx phenocopy of dystrophinopathy-associated cardiomyopathy.

Cardiorespiratory complications are frequent symptoms of Duchenne muscular dystrophy, a neuromuscular disorder caused by primary abnormalities in the dystrophin gene. Loss of cardiac dystrophin initially leads to changes in dystrophin-associated glycoproteins and subsequently triggers secondarily sarcolemmal disintegration, fibre necrosis, fibrosis, fatty tissue replacement, and interstitial inflammation. This results in progressive cardiac disease, which is the cause of death in a considerable number of patients afflicted with X-linked muscular dystrophy. In order to better define the molecular pathogenesis of this type of cardiomyopathy, several studies have applied mass spectrometry-based proteomics to determine proteome-wide alterations in dystrophinopathy-associated cardiomyopathy. Proteomic studies included both gel-based and label-free mass spectrometric surveys of dystrophin-deficient heart muscle from the established mdx animal model of dystrophinopathy. Comparative cardiac proteomics revealed novel changes in proteins associated with mitochondrial energy metabolism, glycolysis, signaling, iron binding, antibody response, fibre contraction, basal lamina stabilisation, and cytoskeletal organisation. This review summarizes the importance of studying cardiomyopathy within the field of muscular dystrophy research, outlines key features of the mdx heart and its suitability as a model system for studying cardiac pathogenesis, and discusses the impact of recent proteomic findings for exploring molecular and cellular aspects of cardiac abnormalities in inherited muscular dystrophies.

Mass Spectrometry-Based Identification of Muscle-Associated and Muscle-Derived Proteomic Biomarkers of Dystrophinopathies

The optimization of large-scale screening procedures of pathological specimens by genomic, proteomic and metabolic methods has drastically increased the bioanalytical capability for swiftly identifying novel biomarkers of inherited disorders, such as neuromuscular diseases. X-linked muscular dystrophy represents the most frequently inherited muscle disease and is characterized by primary abnormalities in the membrane cytoskeletal protein dystrophin. Mass spectrometry-based proteomics has been widely employed for the systematic analysis of dystrophin-deficient muscle tissues, using patient samples and animal models of dystrophinopathy. Both, gel-based methods and label-free mass spectrometric techniques have been applied in comparative analyses and established a large number of altered proteins that are associated with muscle contraction, energy metabolism, ion homeostasis, cellular signaling, the cytoskeleton, the extracellular matrix and the cellular stress response. Although these new indicators of muscular dystrophy have increased our general understanding of the molecular pathogenesis of dystrophinopathy, their application as new diagnostic or prognostic biomarkers would require the undesirable usage of invasive methodology. Hence, to reduce the need for diagnostic muscle biopsy procedures, more recent efforts have focused on the proteomic screening of suitable body fluids, such as plasma, serum or urine, for the identification of changed concentration levels of muscle-derived peptides, protein fragments or intact proteins. The occurrence of muscular dystrophy-related protein species in biofluids will be extremely helpful for the future development of cost-effective and non-invasive diagnostic procedures. Novel biomarker signatures of dystrophinopathies will be indispensible for the swift evaluation of innovative therapeutic approaches, such as exon skipping, codon-read-through or stem cell therapy.

Proteomics of the Dystrophin-glycoprotein Complex and Dystrophinopathy

The largest human gene is represented by the X-chromosomal dystrophin gene of 2.4 million bases, which encodes for the membrane cytoskeletal protein dystrophin. The dystrophin isoform Dp427 has a subsarcolemmal location and forms a supramolecular membrane assembly with a variety of glycoproteins. In healthy muscle fibres, dystrophin acts as an actin-binding protein that links the cytoskeleton via the α/β-dystroglycan complex to the extracellular matrix protein laminin. This trans-sarcolemmal complex is believed to stabilize the muscle surface and thus prevents membrane rupturing during excitation-contraction-relaxation cycles. In the highly progressive muscle wasting disease Duchenne muscular dystrophy, the primary deficiency in dystrophin causes a drastic reduction in dystrophin-associated glycoproteins, which renders muscle fibres more susceptible to necrosis. Following the biochemical and cell biological characterization of the dystrophin-glycoprotein complex, several mass spectrometry-based proteomic studies have investigated global changes in dystrophin-deficient muscle tissues. This review briefly outlines the basic domain structure of Dp427 and the composition of the dystrophin-associated glycoprotein complex from skeletal muscle. A detailed discussion of recent proteomic analyses of the purified dystrophin-glycoprotein complex is included, as well as a summary of mass spectrometric surveys of dystrophic specimens. The study of these new areas of muscle proteomics tends to improve our understanding of the normal function of dystrophin in contractile fibres and better define the molecular mechanism of X-linked muscular dystrophy.

Prenatal Diagnosis of BMD in Morocco: Evolution and Limits

The muscular dystrophy is a group of inherited disorders characterized in the most of cases by progressive muscle weakness. The best known are X-linked disorder Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD). BMD is a milder form of the disease with a later age of onset and a slower clinical progression. The DMD gene, located on Xp21, is the largest human gene in the human genome (2.3 Mb). DMD gene consists of 79 exons and codes for dystrophin protein. A 9-year-old boy, who experienced symptoms of the disease, was admitted to the Casablanca University Children's Hospital. The patient, with no known family history of significant muscle disease, was first examined at 4 years of age because of walking difficulties and a limited hands force. Blood tests revealed elevated serum levels of creatine kinase (7.60 U/L). The electromyogram showed myopathic changes, consisting of polyphasic potentials, and the muscular biopsy revealed dystrophic aspect. Analysis of the dystrophin-encoding gene by PCR deletion analysis of the dystrophin gene was performed by multiplex PCR primer sets of Chamberlain and Beggs. The analysis showed a deletion of exons 45 to 49. Mother genetic testing showed the heterozygosis deletion.

Lamina-Associated Polypeptide-1 Interacts with the Muscular Dystrophy Protein Emerin and Is Essential for Skeletal Muscle Maintenance

X-linked Emery-Dreifuss muscular dystrophy is caused by loss of function of emerin, an integral protein of the inner nuclear membrane. Yet emerin null mice are essentially normal, suggesting the existence of a critical compensating factor. We show that the lamina-associated polypeptide1 (LAP1) interacts with emerin. Conditional deletion of LAP1 from striated muscle causes muscular dystrophy; this pathology is worsened in the absence of emerin. LAP1 levels are significantly higher in mouse than human skeletal muscle, and reducing LAP1 by approximately half in mice also induces muscle abnormalities in emerin null mice. Conditional deletion of LAP1 from hepatocytes yields mice that exhibit normal liver function and are indistinguishable from littermate controls. These results establish that LAP1 interacts physically and functionally with emerin and plays an essential and selective role in skeletal muscle maintenance. They also highlight how dissecting differences between mouse and human phenotypes can provide fundamental insights into disease mechanisms.

Generating SM(a)RTer Compounds for Translation Termination Suppression in A-T and Other Genetic Disorders

Reading of the genetic code through nonsense mutations to restore protein function is a concept that dates back 50 years, with the discovery that streptomycin was capable of correcting defective genotypes.1 In the intervening years, suppression of premature termination codons (PTCs) by aminoglycoside antibiotics and other compounds has been employed to express full-length normal protein in a range of disease states. This approach has met with success in restoring at least partial function, and several of these compounds are at an advanced stage in clinical trials. Important targets include genes encoding the cystic fibrosis (CF) fibrosis transmembrane conductance regulator (CFTR) and dystrophin, which is defective in the X-linked neuromuscular disorder Duchenne muscular dystrophy. However, this approach can be applied to many less common genetic disorders caused by nonsense mutations that lead to premature truncation of a protein.2 In this issue, Du et al. build on previous efforts using high-throughput screening of small-molecule libraries to identify novel readthrough compounds (RTCs) that suppress all three stop codons (TGA, TAG, and TAA) in ataxia telangiectasia mutated (ATM), which is defective in the human genetic disorder ataxia telangiectasia (A-T).3 The authors refer to these as small-molecular readthrough (SMRT) compounds and reveal that they have structures that are quite different from those of two previously described compounds (RTC 13 and RTC 14)4 but are comparable or superior to them in efficiency in their readthrough capacity for ATM. Although successful treatment of this rare genetic disorder is unlikely to have a major impact on community health, the authors point to a “one drug fits all” model that would have broad translational potential for several other disorders.

The emerin-binding transcription factor Lmo7 is regulated by association with p130Cas at focal adhesions

Loss of function mutations in the nuclear inner membrane protein, emerin, cause X-linked Emery-Dreifuss muscular dystrophy (X-EDMD). X-EDMD is characterized by contractures of major tendons, skeletal muscle weakening and wasting, and cardiac conduction system defects. The transcription factor Lmo7 regulates muscle- and heart-relevant genes and is inhibited by binding to emerin, suggesting Lmo7 misregulation contributes to EDMD disease. Lmo7 associates with cell adhesions and shuttles between the plasma membrane and nucleus, but the regulation and biological consequences of this dual localization were unknown. We report endogenous Lmo7 also associates with focal adhesions in cells, and both co-localizes and co-immunoprecipitates with p130Cas, a key signaling component of focal adhesions. Lmo7 nuclear localization and transcriptional activity increased significantly in p130Cas-null MEFs, suggesting Lmo7 is negatively regulated by p130Cas-dependent association with focal adhesions. These results support EDMD models in which Lmo7 is a downstream mediator of integrin-dependent signaling that allows tendon cells and muscles to adapt to and withstand mechanical stress.

Comparative proteomic profiling of soleus, extensor digitorum longus, flexor digitorum brevis and interosseus muscles from the mdx mouse model of Duchenne muscular dystrophy

Duchenne muscular dystrophy is due to genetic abnormalities in the dystrophin gene and represents one of the most frequent genetic childhood diseases. In the X-linked muscular dystrophy (mdx) mouse model of dystrophinopathy, different subtypes of skeletal muscles are affected to a varying degree albeit the same single base substitution within exon 23 of the dystrophin gene. Thus, to determine potential muscle subtype-specific differences in secondary alterations due to a deficiency in dystrophin, in this study, we carried out a comparative histological and proteomic survey of mdx muscles. We intentionally included the skeletal muscles that are often used for studying the pathomechanism of muscular dystrophy. Histological examinations revealed a significantly higher degree of central nucleation in the soleus and extensor digitorum longus muscles compared with the flexor digitorum brevis and interosseus muscles. Muscular hypertrophy of 20–25% was likewise only observed in the soleus and extensor digitorum longus muscles from mdx mice, but not in the flexor digitorum brevis and interosseus muscles. For proteomic analysis, muscle protein extracts were separated by fluorescence two-dimensional (2D) gel electrophoresis. Proteins with a significant change in their expression were identified by mass spectrometry. Proteomic profiling established an altered abundance of 24, 17, 19 and 5 protein species in the dystrophin-deficient soleus, extensor digitorum longus, flexor digitorum brevis and interosseus muscle, respectively. The key proteomic findings were verified by immunoblot analysis. The identified proteins are involved in the contraction-relaxation cycle, metabolite transport, muscle metabolism and the cellular stress response. Thus, histological and proteomic profiling of muscle subtypes from mdx mice indicated that distinct skeletal muscles are differentially affected by the loss of the membrane cytoskeletal protein, dystrophin. Varying degrees of perturbed protein expression patterns in the muscle subtypes from mdx mice may be due to dissimilar downstream events, including differences in muscle structure or compensatory mechanisms that counteract pathophysiological processes. The interosseus muscle from mdx mice possibly represents a naturally protected phenotype.