Digenic inheritance involving a muscle-specific protein kinase and the giant titin protein causes a skeletal muscle myopathy.

In digenic inheritance, pathogenic variants in two genes must be inherited together to cause disease. Only very few examples of digenic inheritance have been described in the neuromuscular disease field. Here we show that predicted deleterious variants in SRPK3, encoding the X-linked serine/argenine protein kinase 3, lead to a progressive early onset skeletal muscle myopathy only when in combination with heterozygous variants in the TTN gene. The co-occurrence of predicted deleterious SRPK3/TTN variants was not seen among 76,702 healthy male individuals, and statistical modeling strongly supported digenic inheritance as the best-fitting model. Furthermore, double-mutant zebrafish (srpk3-/-; ttn.1+/-) replicated the myopathic phenotype and showed myofibrillar disorganization. Transcriptome data suggest that the interaction of srpk3 and ttn.1 in zebrafish occurs at a post-transcriptional level. We propose that digenic inheritance of deleterious changes impacting both the protein kinase SRPK3 and the giant muscle protein titin causes a skeletal myopathy and might serve as a model for other genetic diseases.

CACNA1S Variant Associated With a Myalgic Myopathy Phenotype.

BACKGROUND AND OBJECTIVES: This study aimed to characterize the phenotype of a novel myalgic myopathy encountered in a Finnish family. METHODS: Four symptomatic and 3 asymptomatic individuals from 2 generations underwent clinical, neurophysiologic, imaging, and muscle biopsy examinations. Targeted sequencing of all known myopathy genes was performed. RESULTS: A very rare CACNA1S gene variant c.2893G>C (p.E965Q) was identified in the family. The symptomatic patients presented with exercise-induced myalgia, cramping, muscle stiffness, and fatigue and eventually developed muscle weakness. Examinations revealed mild ptosis and unusual muscle hypertrophy in the upper limbs. In the most advanced disease stage, muscle weakness and muscle atrophy of the limbs were evident. In some patients, muscle biopsy showed mild myopathic findings and creatine kinase levels were slightly elevated. DISCUSSION: Myalgia is a very common symptom affecting quality of life. Widespread myalgia may be confused with other myalgic syndromes such as fibromyalgia. In this study, we show that variants in CACNA1S gene may be one cause of severe exercise-induced myalgia.

Extension of the DNAJB2a isoform in a dominant neuromyopathy family.

Recessive mutations in the DNAJB2 gene, encoding the J-domain co-chaperones DNAJB2a and DNAJB2b, have previously been reported as the genetic cause of progressive peripheral neuropathies, rarely involving pyramidal signs, parkinsonism and myopathy. We describe here a family with the first dominantly acting DNAJB2 mutation resulting in a late-onset neuromyopathy phenotype. The c.832 T > G p.(*278Glyext*83) mutation abolishes the stop codon of the DNAJB2a isoform resulting in a C-terminal extension of the protein, with no direct effect predicted on the DNAJB2b isoform of the protein. Analysis of the muscle biopsy showed reduction of both protein isoforms. In functional studies, the mutant protein mislocalized to the endoplasmic reticulum due to a transmembrane helix in the C-terminal extension. The mutant protein underwent rapid proteasomal degradation and also increased the turnover of co-expressed wild-type DNAJB2a, potentially explaining the reduced protein amount in the patient muscle tissue. In line with this dominant negative effect, both wild-type and mutant DNAJB2a were shown to form polydisperse oligomers.

HSPB8 frameshift mutant aggregates weaken chaperone-assisted selective autophagy in neuromyopathies.

Chaperone-assisted selective autophagy (CASA) is a highly selective pathway for the disposal of misfolding and aggregating proteins. In muscle, CASA assures muscle integrity by favoring the turnover of structural components damaged by mechanical strain. In neurons, CASA promotes the removal of aggregating substrates. A crucial player of CASA is HSPB8 (heat shock protein family B (small) member 8), which acts in a complex with HSPA, their cochaperone BAG3, and the E3 ubiquitin ligase STUB1. Recently, four novel HSPB8 frameshift (fs) gene mutations have been linked to neuromyopathies, and encode carboxy-terminally mutated HSPB8, sharing a common C-terminal extension. Here, we analyzed the biochemical and functional alterations associated with the HSPB8_fs mutant proteins. We demonstrated that HSPB8_fs mutants are highly insoluble and tend to form proteinaceous aggregates in the cytoplasm. Notably, all HSPB8 frameshift mutants retain their ability to interact with CASA members but sequester them into the HSPB8-positive aggregates together with two autophagy receptors SQSTM1/p62 and TAX1BP1. This copartitioning process negatively affects the CASA capability to remove its clients and causes a general failure in proteostasis response. Further analyses revealed that the aggregation of the HSPB8_fs mutants occurs independently of the other CASA members or from the autophagy receptors interaction, but it is an intrinsic feature of the mutated amino acid sequence. HSPB8_fs mutants aggregation alters the differentiation capacity of muscle cells and impairs sarcomere organization. Collectively, these results shed light on a potential pathogenic mechanism shared by the HSPB8_fs mutants described in neuromuscular diseases.Abbreviations : ACD: α-crystallin domain; ACTN: actinin alpha; BAG3: BAG cochaperone 3; C: carboxy; CASA: chaperone-assisted selective autophagy; CE: carboxy-terminal extension; CLEM: correlative light and electron microscopy; CMT2L: Charcot-Marie-Tooth type 2L; CTR: carboxy-terminal region; dHMNII: distal hereditary motor neuropathy type II; EV: empty vector; FRA: filter retardation assay; fs: frameshift; HSPA/HSP70: heat shock protein family A (Hsp70); HSPB1/Hsp27: heat shock protein family B (small) member 1; HSPB8/Hsp22: heat shock protein family B (small) member 8; HTT: huntingtin; KO: knockout; MAP1LC3B/LC3: microtubule associated protein 1 light chain 3 beta; MD: molecular dynamics; MTOC: microtubule organizing center; MYH: myosin heavy chain; MYOG: myogenin; NBR1: NBR1 autophagy cargo receptor; CALCOCO2/NDP52: calcium binding and coiled-coil domain 2; NSC34: Neuroblastoma X Spinal Cord 34; OPTN: optineurin; polyQ: polyglutamine; SQSTM1/p62: sequestosome 1; STUB1/CHIP: STIP1 homology and U-box containing protein 1; TARDBP/TDP-43: TAR DNA binding protein; TAX1BP1: Tax1 binding protein 1; TUBA: tubulin alpha; WT: wild-type.

Comprehensive transcriptomic analysis shows disturbed calcium homeostasis and deregulation of T lymphocyte apoptosis in inclusion body myositis.

OBJECTIVE: Inclusion body myositis (IBM) has an unclear molecular etiology exhibiting both characteristic inflammatory T-cell activity and rimmed-vacuolar degeneration of muscle fibers. Using in-depth gene expression and splicing studies, we aimed at understanding the different components of the molecular pathomechanisms in IBM. METHODS: We performed RNA-seq on RNA extracted from skeletal muscle biopsies of clinically and histopathologically defined IBM (n = 24), tibial muscular dystrophy (n = 6), and histopathologically normal group (n = 9). In a comprehensive transcriptomics analysis, we analyzed the differential gene expression, differential splicing and exon usage, downstream pathway analysis, and the interplay between coding and non-coding RNAs (micro RNAs and long non-coding RNAs). RESULTS: We observe dysregulation of genes involved in calcium homeostasis, particularly affecting the T-cell activity and regulation, causing disturbed Ca2+-induced apoptotic pathways of T cells in IBM muscles. Additionally, LCK/p56, which is an essential gene in regulating the fate of T-cell apoptosis, shows increased expression and altered splicing usage in IBM muscles. INTERPRETATION: Our analysis provides a novel understanding of the molecular mechanisms in IBM by showing a detailed dysregulation of genes involved in calcium homeostasis and its effect on T-cell functioning in IBM muscles. Loss of T-cell regulation is hypothesized to be involved in the consistent observation of no response to immune therapies in IBM patients. Our results show that loss of apoptotic control of cytotoxic T cells could indeed be one component of their abnormal cytolytic activity in IBM muscles.

Bi-allelic loss-of-function OBSCN variants predispose individuals to severe recurrent rhabdomyolysis.

Rhabdomyolysis is the acute breakdown of skeletal myofibres in response to an initiating factor, most commonly toxins and over exertion. A variety of genetic disorders predispose to rhabdomyolysis through different pathogenic mechanisms, particularly in patients with recurrent episodes. However, most cases remain without a genetic diagnosis. Here we present six patients who presented with severe and recurrent rhabdomyolysis, usually with onset in the teenage years; other features included a history of myalgia and muscle cramps. We identified 10 bi-allelic loss-of-function variants in the gene encoding obscurin (OBSCN) predisposing individuals to recurrent rhabdomyolysis. We show reduced expression of OBSCN and loss of obscurin protein in patient muscle. Obscurin is proposed to be involved in sarcoplasmic reticulum function and Ca2+ handling. Patient cultured myoblasts appear more susceptible to starvation as evidenced by a greater decreased in sarcoplasmic reticulum Ca2+ content compared to control myoblasts. This likely reflects a lower efficiency when pumping Ca2+ back into the sarcoplasmic reticulum and/or a decrease in Ca2+ sarcoplasmic reticulum storage ability when metabolism is diminished. OSBCN variants have previously been associated with cardiomyopathies. None of the patients presented with a cardiomyopathy and cardiac examinations were normal in all cases in which cardiac function was assessed. There was also no history of cardiomyopathy in first degree relatives, in particular in any of the carrier parents. This cohort is relatively young, thus follow-up studies and the identification of additional cases with bi-allelic null OBSCN variants will further delineate OBSCN-related disease and the clinical course of disease.

Dominant Distal Myopathy 3 (MPD3) Caused by a Deletion in the HNRNPA1 Gene.

BACKGROUND AND OBJECTIVES: To determine the genetic cause of the disease in the previously reported family with adult-onset autosomal dominant distal myopathy (myopathy, distal, 3; MPD3). METHODS: Continued clinical evaluation including muscle MRI and muscle pathology. A linkage analysis with single nucleotide polymorphism arrays and genome sequencing were used to identify the genetic defect, which was verified by Sanger sequencing. RNA sequencing was used to investigate the transcriptional effects of the identified genetic defect. RESULTS: Small hand muscles (intrinsic, thenar, and hypothenar) were first involved with spread to the lower legs and later proximal muscles. Dystrophic changes with rimmed vacuoles and cytoplasmic inclusions were observed in muscle biopsies at advanced stage. A single nucleotide polymorphism array confirmed the previous microsatellite-based linkage to 8p22-q11 and 12q13-q22. Genome sequencing of three affected family members combined with structural variant calling revealed a small heterozygous deletion of 160 base pairs spanning the second last exon 10 of the heterogeneous nuclear ribonucleoprotein A1 (HNRNPA1) gene, which is in the linked region on chromosome 12. Segregation of the mutation with the disease was confirmed by Sanger sequencing. RNA sequencing showed that the mutant allele produces a shorter mutant mRNA transcript compared with the wild-type allele. Immunofluorescence studies on muscle biopsies revealed small p62 and larger TDP-43 inclusions. DISCUSSION: A small exon 10 deletion in the gene HNRNPA1 was identified as the cause of MPD3 in this family. The new HNRNPA1-related phenotype, upper limb presenting distal myopathy, was thus confirmed, and the family displays the complexities of gene identification.

Out-of-Frame Mutations in ACTN2 Last Exon Cause a Dominant Distal Myopathy With Facial Weakness.

BACKGROUND AND OBJECTIVES: To clinically, genetically, and histopathologically characterize patients presenting with an unusual combination of distal myopathy and facial weakness, without involvement of upper limb or shoulder girdle muscles. METHODS: Two families with a novel form of actininopathy were identified. Patients had been followed up over 10 years. Their molecular genetic diagnosis was not clear after extensive investigations, including analysis of candidate genes and FSHD1-related D4Z4 repeats. RESULTS: Patients shared a similar clinical phenotype and a common pattern of muscle involvement. They presented with a very slowly progressive myopathy involving anterior lower leg and facial muscles. Muscle MRI finding showed complete fat replacement of anterolateral compartment muscles of the lower legs with variable involvement of soleus and gastrocnemius but sparing thigh muscles. Muscle biopsy showed internalized nuclei, myofibrillar disorganization, and rimmed vacuoles. High-throughput sequencing identified in each proband a heterozygous single nucleotide deletion (c.2558del and c.2567del) in the last exon of the ACTN2 gene. The deletions are predicted to lead to a novel but unstructured slightly extended C-terminal amino acid sequence. DISCUSSION: Our findings indicate an unusual form of actininopathy with specific molecular and clinical features. Actininopathy should be considered in the differential diagnosis of distal myopathy combined with facial weakness.

Missense mutations in small muscle protein X-linked (SMPX) cause distal myopathy with protein inclusions.

Using deep phenotyping and high-throughput sequencing, we have identified a novel type of distal myopathy caused by mutations in the Small muscle protein X-linked (SMPX) gene. Four different missense mutations were identified in ten patients from nine families in five different countries, suggesting that this disease could be prevalent in other populations as well. Haplotype analysis of patients with similar ancestry revealed two different founder mutations in Southern Europe and France, indicating that the prevalence in these populations may be higher. In our study all patients presented with highly similar clinical features: adult-onset, usually distal more than proximal limb muscle weakness, slowly progressing over decades with preserved walking. Lower limb muscle imaging showed a characteristic pattern of muscle involvement and fatty degeneration. Histopathological and electron microscopic analysis of patient muscle biopsies revealed myopathic findings with rimmed vacuoles and the presence of sarcoplasmic inclusions, some with amyloid-like characteristics. In silico predictions and subsequent cell culture studies showed that the missense mutations increase aggregation propensity of the SMPX protein. In cell culture studies, overexpressed SMPX localized to stress granules and slowed down their clearance.

Congenital asymmetric distal myopathy with hemifacial weakness caused by a heterozygous large de novo mosaic deletion in nebulin.

We report the first mosaic mutation, a deletion of exons 11-107, identified in the nebulin gene in a Finnish patient presenting with a predominantly distal congenital myopathy and asymmetric muscle weakness. The female patient is ambulant and currently 26 years old. Muscle biopsies showed myopathic features with type 1 fibre predominance, strikingly hypotrophic type 2 fibres and central nuclei, but no nemaline bodies. The deletion was detected in a copy number variation analysis based on next-generation sequencing data. The parents of the patient did not carry the deletion. Mosaicism was detected using a custom, targeted comparative genomic hybridisation array. Expression of the truncated allele, less than half the size of full-length nebulin, was confirmed by Western blotting. The clinical and histological picture resembled that of a family with a slightly smaller deletion, and that in patients with recessively inherited distal forms of nebulin-caused myopathy. Asymmetry, however, was a novel feature.

Genotype-phenotype correlations in recessive titinopathies.

PURPOSE: High throughput sequencing analysis has facilitated the rapid analysis of the entire titin (TTN) coding sequence. This has resulted in the identification of a growing number of recessive titinopathy patients. The aim of this study was to (1) characterize the causative genetic variants and clinical features of the largest cohort of recessive titinopathy patients reported to date and (2) to evaluate genotype-phenotype correlations in this cohort. METHODS: We analyzed clinical and genetic data in a cohort of patients with biallelic pathogenic or likely pathogenic TTN variants. The cohort included both previously reported cases (100 patients from 81 unrelated families) and unreported cases (23 patients from 20 unrelated families). RESULTS: Overall, 132 causative variants were identified in cohort members. More than half of the cases had hypotonia at birth or muscle weakness and a delayed motor development within the first 12 months of life (congenital myopathy) with causative variants located along the entire gene. The remaining patients had a distal or proximal phenotype and a childhood or later (noncongenital) onset. All noncongenital cases had at least one pathogenic variant in one of the final three TTN exons (362-364). CONCLUSION: Our findings suggest a novel association between the location of nonsense variants and the clinical severity of the disease.

Mutations in the J domain of DNAJB6 cause dominant distal myopathy.

Eight patients from five families with undiagnosed dominant distal myopathy underwent clinical, neurophysiological and muscle biopsy examinations. Molecular genetic studies were performed using targeted sequencing of all known myopathy genes followed by segregation of the identified mutations in the affected families using Sanger sequencing. Two novel mutations in DNAJB6 J domain, c.149C>T (p.A50V) and c.161A>C (p.E54A), were identified as the cause of disease. The muscle involvement with p.A50V was distal calf-predominant, and the p.E54A was more proximo-distal. Histological findings were similar to those previously reported in DNAJB6 myopathy. In line with reported pathogenic mutations in the glycine/phenylalanine (G/F) domain of DNAJB6, both the novel mutations showed reduced anti-aggregation capacity by filter trap assay and TDP-43 disaggregation assays. Modeling of the protein showed close proximity of the mutated residues with the G/F domain. Myopathy-causing mutations in DNAJB6 are not only located in the G/F domain, but also in the J domain. The identified mutations in the J domain cause dominant distal and proximo-distal myopathy, confirming that mutations in DNAJB6 should be considered in distal myopathy cases.

Panorama of the distal myopathies.

Distal myopathies are genetic primary muscle disorders with a prominent weakness at onset in hands and/or feet. The age of onset (from early childhood to adulthood), the distribution of muscle weakness (upper versus lower limbs) and the histological findings (ranging from nonspecific myopathic changes to myofibrillar disarrays and rimmed vacuoles) are extremely variable. However, despite being characterized by a wide clinical and genetic heterogeneity, the distal myopathies are a category of muscular dystrophies: genetic diseases with progressive loss of muscle fibers. Myopathic congenital arthrogryposis is also a form of distal myopathy usually caused by focal amyoplasia. Massive parallel sequencing has further expanded the long list of genes associated with a distal myopathy, and contributed identifying as distal myopathy-causative rare variants in genes more often related with other skeletal or cardiac muscle diseases. Currently, almost 20 genes (ACTN2, CAV3, CRYAB, DNAJB6, DNM2, FLNC, HNRNPA1, HSPB8, KHLH9, LDB3, MATR3, MB, MYOT, PLIN4, TIA1, VCP, NOTCH2NLC, LRP12, GIPS1) have been associated with an autosomal dominant form of distal myopathy. Pathogenic changes in four genes (ADSSL, ANO5, DYSF, GNE) cause an autosomal recessive form; and disease-causing variants in five genes (DES, MYH7, NEB, RYR1 and TTN) result either in a dominant or in a recessive distal myopathy. Finally, a digenic mechanism, underlying a Welander-like form of distal myopathy, has been recently elucidated. Rare pathogenic mutations in SQSTM1, previously identified with a bone disease (Paget disease), unexpectedly cause a distal myopathy when combined with a common polymorphism in TIA1. The present review aims at describing the genetic basis of distal myopathy and at summarizing the clinical features of the different forms described so far.

Recurrent TTN metatranscript-only c.39974-11T>G splice variant associated with autosomal recessive arthrogryposis multiplex congenita and myopathy.

We present eight families with arthrogryposis multiplex congenita and myopathy bearing a TTN intron 213 extended splice-site variant (NM_001267550.1:c.39974-11T>G), inherited in trans with a second pathogenic TTN variant. Muscle-derived RNA studies of three individuals confirmed mis-splicing induced by the c.39974-11T>G variant; in-frame exon 214 skipping or use of a cryptic 3' splice-site effecting a frameshift. Confounding interpretation of pathogenicity is the absence of exons 213-217 within the described skeletal muscle TTN N2A isoform. However, RNA-sequencing from 365 adult human gastrocnemius samples revealed that 56% specimens predominantly include exons 213-217 in TTN transcripts (inclusion rate ≥66%). Further, RNA-sequencing of five fetal muscle samples confirmed that 4/5 specimens predominantly include exons 213-217 (fifth sample inclusion rate 57%). Contractures improved significantly with age for four individuals, which may be linked to decreased expression of pathogenic fetal transcripts. Our study extends emerging evidence supporting a vital developmental role for TTN isoforms containing metatranscript-only exons.

New family with HSPB8-associated autosomal dominant rimmed vacuolar myopathy.

OBJECTIVE: We clinically and molecularly characterize a new family with autosomal dominant rimmed vacuolar myopathy (RVM) caused by mutations in the HSPB8 gene. METHODS: We performed whole-exome and whole-genome sequencing in the family. Western blot and immunocytochemistry were used to analyze 3 patient fibroblasts, and findings were compared with their age- and sex-matched controls. RESULTS: Affected patients have distal and proximal myopathy, with muscle biopsy showing rimmed vacuoles, muscle fiber atrophy, and endomysial fibrosis typical of RVM. Muscle MRI showed severe relatively symmetric multifocal fatty degenerative changes of the lower extremities. We identified a duplication of C at position 515 of the HSPB8 gene (c.515dupC) by whole-genome sequencing, which caused a frameshift with a predicted alternate stop codon p.P173SFS*43 in all affected individuals, resulting in an elongated protein product. Western blot and immunocytochemistry studies revealed reduced expression of heat shock protein beta 8 in patient fibroblasts compared with control fibroblasts, in addition to disrupted autophagy pathology. CONCLUSIONS: We report a novel family with autosomal dominant RVM caused by the c.515dupC mutation of the HSPB8 gene, causing a translational frameshift that results in an elongated protein. Understanding the mechanism for the RVM pathology caused by mutated chaperone will permit novel targeted strategies to alter the natural history progression. As next-generation sequencing becomes more available, additional myopathic families will be identified with HSPB8 mutations.

Novel mutation in TNPO3 causes congenital limb-girdle myopathy with slow progression.

OBJECTIVE: We report a second family with autosomal dominant transportinopathy presenting with congenital or early-onset myopathy and slow progression, causing proximal and less pronounced distal muscle weakness. METHODS: Patients had clinical examinations, muscle MRI, EMG, and muscle biopsy studies. The MYOcap gene panel was used to identify the gene defect in the family. Muscle biopsies were used for histopathologic and protein expression studies, and TNPO3 constructs were used to study the effect of the mutations in transfected cells. RESULTS: We identified a novel heterozygous mutation, c.2757delC, in the last part of the transportin-3 (TNPO3) gene in the affected family members. The mutation causes an almost identical frameshift affecting the stop codon and elongating the C-term protein product of the TNPO3 transcript, as was previously reported in the first large Spanish-Italian LGMD1F kindred. TNPO3 protein was increased in the patient muscle and accumulated in the subsarcolemmal and perinuclear areas. At least one of the cargo proteins, the splicing factor SRRM2 was normally located in the nucleus. Transiently transfected mutant TNPO3 constructs failed to localize to cytoplasmic annulate lamellae pore complexes in cells. CONCLUSIONS: We report the clinical, molecular genetic, and histopathologic features of the second transportinopathy family. The variability of the clinical phenotype together with histopathologic findings suggests that several molecular pathways may be involved in the disease pathomechanism, such as nucleocytoplasmic shuttling, protein aggregation, and defective protein turnover.

A novel COL6A2 mutation causing late-onset limb-girdle muscular dystrophy.

Limb-girdle muscular dystrophies (LGMD) are genetic disorders characterized by weakness of predominantly proximal limb and trunk muscles due to progressive loss of muscle tissue. Collagen VI-related muscular dystrophies usually display more generalized muscle involvement combined with contractures and/or hyperlaxity of distal finger joints. LGMD-like phenotype of collagenopathy has only rarely been described and as reported is usually of childhood onset. We identified a Finnish family with COL6A2-related LGMD with autosomal dominant inheritance and very late onset at 40-60 years of age. Since the mutation was previously unreported, the pathognomonic findings on muscle MRI were the decisive clue for the correct diagnosis.

Actininopathy: A new muscular dystrophy caused by ACTN2 dominant mutations.

OBJECTIVE: To clinically and pathologically characterize a cohort of patients presenting with a novel form of distal myopathy and to identify the genetic cause of this new muscular dystrophy. METHODS: We studied 4 families (3 from Spain and 1 from Sweden) suffering from an autosomal dominant distal myopathy. Affected members showed adult onset asymmetric distal muscle weakness with initial involvement of ankle dorsiflexion later progressing also to proximal limb muscles. RESULTS: In all 3 Spanish families, we identified a unique missense variant in the ACTN2 gene cosegregating with the disease. The affected members of the Swedish family carry a different ACTN2 missense variant. INTERPRETATION: ACTN2 encodes for alpha actinin2, which is highly expressed in the sarcomeric Z-disk with a major structural and functional role. Actininopathy is thus a new genetically determined distal myopathy. ANN NEUROL 2019;85:899-906.

An unusual ryanodine receptor 1 (RYR1) phenotype: Mild calf-predominant myopathy.

OBJECTIVE: To identify the genetic defect causing a distal calf myopathy with cores. METHODS: Families with a genetically undetermined calf-predominant myopathy underwent detailed clinical evaluation, including EMG/nerve conduction studies, muscle biopsy, laboratory investigations, and muscle MRI. Next-generation sequencing and targeted Sanger sequencing were used to identify the causative genetic defect in each family. RESULTS: A novel deletion-insertion mutation in ryanodine receptor 1 (RYR1) was found in the proband of the index family and segregated with the disease in 6 affected relatives. Subsequently, we found 2 more families with a similar calf-predominant myopathy segregating with unique RYR1-mutated alleles. All patients showed a very slowly progressive myopathy without episodes of malignant hyperthermia or rhabdomyolysis. Muscle biopsy showed cores or core-like changes in all families. CONCLUSIONS: Our findings expand the spectrum of RYR1-related disorders to include a calf-predominant myopathy with core pathology and autosomal dominant inheritance. Two families had unique and previously unreported RYR1 mutations, while affected persons in the third family carried 2 previously known mutations in the same dominant allele.

Dominantly inherited distal nemaline/cap myopathy caused by a large deletion in the nebulin gene.

We report the first family with a dominantly inherited mutation of the nebulin gene (NEB). This ∼100 kb in-frame deletion encompasses NEB exons 14-89, causing distal nemaline/cap myopathy in a three-generation family. It is the largest deletion characterized in NEB hitherto. The mutated allele was shown to be expressed at the mRNA level and furthermore, for the first time, a deletion was shown to cause the production of a smaller mutant nebulin protein. Thus, we suggest that this novel mutant nebulin protein has a dominant-negative effect, explaining the first documented dominant inheritance of nebulin-caused myopathy. The index patient, a young man, was more severely affected than his mother and grandmother. His first symptom was foot drop at the age of three, followed by distal muscle atrophy, slight hypomimia, high-arched palate, and weakness of the neck and elbow flexors, hands, tibialis anterior and toe extensors. Muscle biopsies showed myopathic features with type 1 fibre predominance in the index patient and nemaline bodies and cap-like structures in biopsies from his mother and grandmother. The muscle biopsy findings constitute a further example of nemaline bodies and cap-like structures being part of the same spectrum of pathological changes.