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Nemaline myopathy (NM) is a congenital myopathy with generalised muscle weakness, most pronounced in neck flexor, bulbar and respiratory muscles. The aim of this cross-sectional study was to assess the Dutch NM patient cohort. We assessed medical history, physical examination, quality of life (QoL), fatigue severity, motor function (MFM), and respiratory muscle function. We included 18 of the 28 identified patients (13 females (11-67 years old); five males (31-74 years old)) with typical or mild NM and eight different genotypes. Nine patients (50 %) used a wheelchair, eight patients (44 %) used mechanical ventilation, and four patients (22 %) were on tube feeding. Spinal deformities were found in 14 patients (78 %). The median Medical Research Council (MRC) sum score was 38/60 [interquartile range 32-51] in typical and 48/60 [44-50] in mild NM. The experienced QoL was lower and fatigue severity was higher than reference values of the healthy population. The total MFM score was 55 % [49-94] in typical and 88 % [72-93] in mild NM. Most of the patients who performed spirometry had a restrictive lung function pattern (11/15). This identification and characterisation of the Dutch NM patient cohort is important for international collaboration and can guide the design of future clinical trials., Competing Interests: Declaration of competing interest None., (Copyright © 2024. Published by Elsevier B.V.)

We describe three patients with asymmetric congenital myopathy without definite nemaline bodies and one patient with severe nemaline myopathy. In all four patients, the phenotype had been caused by pathogenic missense variants in ACTA1 leading to the same amino acid change, p.(Gly247Arg). The three patients with milder myopathy were mosaic for their variants. In contrast, in the severely affected patient, the missense variant was present in a de novo, constitutional form. The grade of mosaicism in the three mosaic patients ranged between 20 % and 40 %. We speculate that the milder clinical and histological manifestations of the same ACTA1 variant in the patients with mosaicism reflect the lower abundance of mutant actin in their muscle tissue. Similarly, the asymmetry of body growth and muscle weakness may be a consequence of the affected cells being unevenly distributed. The partial improvement in muscle strength with age in patients with mosaicism might be due to an increased proportion over time of nuclei carrying and expressing two normal alleles., Competing Interests: Declaration of Competing Interest The authors declare no competing interests. Author MV is and stockholder of Invitae, and TLW is a former employee of Invitae., (Copyright © 2023 The Author(s). Published by Elsevier B.V. All rights reserved.)

In this cross-sectional study, we comprehensively assessed respiratory muscle function in various clinical forms of nemaline myopathy (NM) including non-volitional tests for diaphragm function. Forty-two patients with NM were included (10 males (25-74 y/o); 32 females (11-76 y/o)). The NM forms were typical (n=11), mild (n=7), or childhood-onset with slowness of movements (n=24). Forced vital capacity (FVC) and maximal inspiratory pressure were decreased in typical NM in comparison with childhood-onset NM with slowness (32.0 [29.0-58.5] vs 81.0 [75.0-87.0]%, p<0.01, and 35.0 [24.0-55.0] vs 81.0 [65.0-102.5] cmH 2 O, p<0.01). Eight patients with childhood-onset NM with slowness had respiratory muscle weakness. There was a low correlation between FVC and Motor Function Measure scores (r=0.48, p<0.01). End-inspiratory diaphragm thickness and twitch mouth pressure were decreased in patients requiring home mechanical ventilation compared to non-ventilated patients with normal lung function (1.8 [1.5-2.4] vs 3.1 [2.0-4.6] mm, p=0.049, and -7.9 [-10.9- -4.0] vs -14.9 [-17.3- -12.6], p=0.04). Our results show that respiratory muscle weakness is present in all NM forms, including childhood-onset NM with slowness, and may be present irrespective of the degree of general motor function impairment. These findings highlight the importance for screening of respiratory function in patients with NM to guide respiratory management., Competing Interests: Declarations of interest None., (Copyright © 2022 The Authors. Published by Elsevier B.V. All rights reserved.)

The nemaline myopathies constitute a large proportion of the congenital or structural myopathies. Common to all patients is muscle weakness and the presence in the muscle biopsy of nemaline rods. The causative genes are at least twelve, encoding structural or regulatory proteins of the thin filament, and the clinical picture as well as the histological appearance on muscle biopsy vary widely. Here, we suggest a renewed clinical classification to replace the original one, summarise what is known about the pathogenesis from mutations in each causative gene to the forms of nemaline myopathy described to date, and provide perspectives on pathogenetic mechanisms possibly open to therapeutic modalities., Competing Interests: Declarations of Competing Interest None., (Copyright © 2021 The Authors. Published by Elsevier B.V. All rights reserved.)

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., (Copyright © 2019 Elsevier B.V. All rights reserved.)

We describe two Finnish siblings in whom an incidentally detected elevated creatine kinase activity eventually led to a diagnosis of limb-girdle muscular dystrophy-dystroglycanopathy (Type C12; MDDGC12). When diagnosed at age 10 and 13 years, they were mildly affected with a slow or non-progressive disease course. The main symptoms comprised infrequent hip cramps triggered by flexion, neck cramps triggered by yawning, transient growing pains, calf hypertrophy and mild proximal muscle weakness. Their cognitive and motor developments were unremarkable and they were physically active. Whole-exome sequencing revealed compound heterozygous mutations, both of which were novel, in the protein O-mannosyl kinase (POMK) gene in both siblings; a missense mutation, p.Pro322Leu (c.965C > T), and a nonsense mutation, p.Arg46Ter (c.136C > T). The results were confirmed by Sanger sequencing, showing that the parents were heterozygous carriers of one mutation each. This report adds to the literature by providing phenotype and genotype data on this ultra-rare POMK-related dystroglycanopathy., (Copyright © 2018 Elsevier B.V. All rights reserved.)

We present here a Finnish nemaline myopathy family with a dominant mutation in the skeletal muscle α-actin gene, p.(Glu85Lys), segregating in three generations. The index patient, a 5-year-old boy, had the typical form of nemaline myopathy with congenital muscle weakness and motor milestones delayed but reached, while his mother never had sought medical attention for her very mild muscle weakness, and his maternal grandmother had been misdiagnosed as having myotonic dystrophy. This illustrates the clinical variability in nemaline myopathy., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Nemaline myopathy (NM) constitutes a heterogeneous group of congenital myopathies. Mutations in the nebulin gene (NEB) are the main cause of recessively inherited NM. NEB is one of the most largest genes in human. To date, 68 NEB mutations, mainly small deletions or point mutations have been published. The only large mutation characterized is the 2.5 kb deletion of exon 55 in the Ashkenazi Jewish population. To investigate any copy number variations in this enormous gene, we designed a novel custom comparative genomic hybridization microarray, NM-CGH, targeted towards the seven known genes causative for NM. During the validation of the NM-CGH array we identified two novel deletions in two different families. The first is the largest deletion characterized in NEB to date, (∼53 kb) encompassing 24 exons. The second deletion (1 kb) covers two exons. In both families, the copy number change was the second mutation to be characterized and shown to have been inherited from one of the healthy carrier parents. In addition to these novel mutations, copy number variation was identified in four samples in three families in the triplicate region of NEB. We conclude that this method appears promising for the detection of copy number variations in NEB., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Mutations in the nebulin gene are the main cause of autosomal recessive nemaline myopathy, with clinical presentations ranging from mild to severe disease. We have previously reported a nonspecific distal myopathy caused by homozygous missense mutations in the nebulin gene in six Finnish patients from four different families. Here we describe three non-Finnish patients in two unrelated families with distal nemaline myopathy caused by four different compound heterozygous nebulin mutations, only one of which is a missense mutation. One of the mutations has previously been identified in one family with the severe form of nemaline myopathy. We conclude that nemaline myopathy and distal myopathy caused by nebulin mutations form a clinical and histological continuum. Nemaline myopathy should be considered as a differential diagnosis in patients presenting with an early-onset predominantly distal myopathy., (Copyright © 2011 Elsevier B.V. All rights reserved.)

We report a family with autosomal dominant centronuclear (myotubular) myopathy caused by a novel mutation, p.A618D, in dynamin 2 (DNM2). The 64-year-old mother and 26-year-old daughter had neonatal onset with hypotonia and weak suckling, followed by improvement, then slowly progressive muscle weakness and respiratory restriction. Muscle biopsy showed radial sarcoplasmic strands around the frequent central nuclei. Electrophysiology revealed predominantly myopathic patterns without peripheral nerve involvement. Centronuclear myopathy with neonatal onset caused by a DNM2 mutation in the C-terminal part of the pleckstrin homology domain may have a favorable prognosis and follow a course similar to adult-onset centronuclear myopathy. We advise respiratory follow-up in these patients.

In 2004, Anderson et al. reported a homozygous 2502 bp deletion including exon 55 of the nebulin gene in five Ashkenazi Jewish probands with nemaline myopathy. We determined the occurrence of this deletion in a world-wide series of 355 nemaline myopathy probands with no previously known mutation in other genes and found the mutation in 14 probands, two of whom represented families previously ascertained by Anderson et al. Two of the families were not of known Ashkenazi Jewish descent but they had the haplotype known to segregate with this mutation. In all but two of eight homozygous patients, the clinical picture was more severe than in typical nemaline myopathy.

"Cap myopathy" or "cap disease" is a congenital myopathy characterised by cap-like structures at the periphery of muscle fibres, consisting of disarranged thin filaments with enlarged Z discs. Here we report a deletion in the beta-tropomyosin (TPM2) gene causing cap disease in a 36-year-old male patient with congenital muscle weakness, myopathic facies and respiratory insufficiency. The mutation identified in this patient is an in-frame deletion (c.415&#95;417delGAG) of one codon in exon 4 of TPM2 removing a single glutamate residue (p.Glu139del) from the beta-tropomyosin protein. This is expected to disrupt the seven-amino acid repeat essential for making a coiled coil, and thus to impair tropomyosin-actin interaction. Missense mutations in TPM2 have previously been found to cause rare cases of nemaline myopathy and distal arthrogryposis. This mutation is one not previously described and the first genetic cause identified for cap disease.

We report muscle MRI findings of 10 patients from 8 families with nemaline myopathy. Patients with involvement of the nebulin (NEB) gene showed a consistent pattern of selective muscle involvement corresponding to clinical severity. In mild cases, there was complete sparing of thigh muscles and selective involvement of tibialis anterior and soleus. In moderate cases, there was predominant involvement of rectus femoris, vastus lateralis and hamstring muscles and diffuse involvement of anterior compartment and soleus. Patients with nemaline myopathy secondary to mutations in the skeletal muscle alpha-actin (ACTA1) gene showed diffuse involvement of thigh and leg muscles with relative sparing of the gastrocnemii. Selective muscle involvement in both genetic categories was distinct from what has been reported in other congenital myopathies. We conclude that muscle MRI may be applied to distinguish nemaline myopathy from other conditions with similar clinical and histopathological features, to supplement clinical assessment in individual patients and to help direct genetic testing.

We present comparisons of the clinical pictures in a series of 60 patients with nemaline myopathy in whom mutations had been identified in the genes for nebulin or skeletal muscle alpha-actin. In the patients with nebulin mutations, the typical form of nemaline myopathy predominated, while severe, mild or intermediate forms were less frequent. Autosomal recessive inheritance had been verified or appeared likely in all nebulin cases. In the patients with actin mutations, the severe form of nemaline myopathy was the most common, but some had the mild or typical form, and a few showed other associated features such as intranuclear rods or actin accumulation. Most cases were sporadic, but in addition there were instances of both autosomal dominant and autosomal recessive inheritance, while two families showed mosaicism for dominant mutations. Although no specific phenotype was found to be associated with mutations in either gene, clinical and histological features together with pedigree data may be used in guiding mutation detection. Finding the causative mutation(s) determines the mode of inheritance and permits prenatal diagnosis if requested, but will not as such permit prognostication.

Mutations in the skeletal muscle alpha-actin gene (ACTA1) associated with congenital myopathy with excess of thin myofilaments, nemaline myopathy and intranuclear rod myopathy were first described in 1999. At that time, only 15 different missense mutations were known in ACTA1. More than 60 mutations have now been identified. This review analyses this larger spectrum of mutations in ACTA1. It investigates the molecular consequences of the mutations found to date, provides a framework for genotype-phenotype correlation and suggests future studies in light of results of investigation of normal and mutant actin in other systems, notably the actin specific to the indirect flight muscles of Drosophila. The larger series confirms that the majority of ACTA1 mutations are dominant, a small number are recessive and most isolated cases with no previous family history have de novo dominant mutations. The severity of the disease caused ranges from lack of spontaneous movements at birth requiring immediate mechanical ventilation, to mild disease compatible with life to adulthood. Overall, the mutations within ACTA1 are randomly distributed throughout the protein. However, the larger series of mutations now available indicates that there may be clustering of mutations associated with some phenotypes, e.g. actin myopathy. This would suggest that interference with certain actin functions may be more associated with certain phenotypes, though the exact pathophysiology of the actin mutations remains unknown.

X-linked myotubular myopathy is a rare severe muscle disorder in affected male neonates. Most female carriers are free from symptoms. Skewed X inactivation has been proposed to be responsible for the affected phenotype seen in some carriers. We have compared the X inactivation patterns in blood DNA with the clinical phenotype in carriers of X-linked myotubular myopathy. The X-inactivation analysis was performed using HpaII predigestion of DNA followed by polymerase chain reaction of the highly polymorphic CAG repeat of the androgen receptor (AR) gene. The frequency of skewed X inactivation was similar in the X-linked myotubular myopathy carriers (22%) and in 235 controls (18%). Three overtly affected carriers had skewed X inactivation with the mutated X as the predominantly active X in at least two of them. Four females with mild symptoms had random X inactivation. The unaffected X-linked myotubular myopathy carriers had either skewed X inactivation in favour of expression from the normal X or random X-inactivation. Thus, there was a tendency for females with a more severe phenotype to have a skewed pattern of X inactivation, while females with an intermediate phenotype had a random pattern of X-inactivation.

X-linked myotubular myopathy is a severe congenital myopathy in males, caused by mutations in the myotubularin (MTM1) gene on chromosome Xq28. In heterozygous carriers of MTM1 mutations, clinical symptoms are usually absent or only mild. We report a 6-year-old girl presenting at birth with marked hypotonia and associated feeding and respiratory difficulties. A muscle biopsy performed at 5 months suggested a diagnosis of myotubular myopathy. On examination at 6 years she had marked facial weakness with bilateral ptosis and external ophthalmoplegia, severe axial and proximal weakness and a mild scoliosis. Muscle magnetic resonance imaging showed a distinctive pattern of muscle involvement. Molecular genetic investigation of the MTM1 gene identified a heterozygous mutation in exon 12. X-inactivation studies in lymphocytes showed an extremely skewed pattern (97:3). This case emphasizes that investigation of the MTM1 gene and X-inactivation studies are indicated in isolated females with histopathological and clinical findings suggestive of myotubular myopathy.

We describe an atypical case of nemaline myopathy with an unusual distribution of muscle weakness who presented at 14 years of age with kyphoscoliosis. In this patient, we demonstrate heterozygosity for a de novo CGT-CAT (Arg167His) mutation in a constitutively expressed exon (exon 5) of slow alpha-tropomyosin (TPM3). This is the first mutation identified in a constitutively expressed exon of TPM3 in a nemaline myopathy patient, but is similar to recently described mutations in beta-tropomyosin (TPM2) associated with nemaline myopathy and mutations in fast alpha-tropomyosin (TPM1) which cause hypertrophic cardiomyopathy.

X-linked myotubular myopathy is a severe congenital myopathy that presents in the neonatal period with profound hypotonia and an inability to establish spontaneous respiration. Usually death occurs in infancy from respiratory failure. However, there is phenotypic variability; a number of affected boys have achieved respiratory independence and become ambulatory. Disease-causing mutations have been identified throughout the MTM1 gene on Xq28. MTM1 encodes the protein myotubularin, which is expressed ubiquitously. The main objectives of this study were to establish whether the nature or site of the mutation in the MTM1 gene could predict severity of the disease and to investigate whether early intensive clinical intervention facilitated survival until spontaneous improvement occurred. An association was demonstrated between the presence of a non-truncating mutation of the MTM1 gene and the mild phenotype. However, many non-truncating mutations were also seen in association with the severe phenotype and these were not confined to recognized functional domains of the protein. This suggests that the use of mutation analysis to predict prognosis in the early period following diagnosis is limited. Unexpectedly, over 50 patients surviving for more than 1 year were identified in this study. Further information obtained on 40 of these cases revealed that 50% were receiving 24-h ventilatory support, while 27% were ventilated at night only. The high survival rate for this disorder therefore reflects intensive medical intervention without which the majority of these boys would not survive.

We report mutational analysis of the last 42 exons of the nebulin gene (NEB) in 77 patients with various forms of nemaline myopathy. In addition to the previously described six mutations in five families, we identified 12 novel recessive mutations in 13 families. Affected individuals were homozygous for the mutations in five families and compound heterozygous in two, while in the remaining cases only one heterozygous mutation was identified. The majority of the mutations were frameshifts due to small deletions or insertions; also common were point mutations causing premature stop codons or abnormal splicing, while missense mutations appeared rare. There were no obvious mutational hotspots, although four unrelated patients showed mutations in the differentially expressed exon 177d, and another three showed mutations in exon 184. Most of the mutations are predicted to result in truncated or internally deleted proteins. Mutations in the differentially expressed exons are expected to reduce the nebulin isoform diversity necessary for normal muscle development.

Previously, we reported results indicating that nebulin was the gene causing the typical form of autosomal recessive nemaline (rod) myopathy. Here we describe the identification of mutations in the nebulin gene in seven offspring of five families affected by the severe congenital form of nemaline myopathy. One pregnancy was terminated on the grounds of foetal abnormality, while six affected infants died at ages ranging from the first day of life to 19 months. Only three of the six neonates were able to establish spontaneous respiration. Three had arthrogryposis. In three of the five families, the mutations were located in exon 184. These mutations are predicted to cause absence of the C-terminal part of nebulin.

Nemaline myopathy is a clinically and genetically heterogeneous muscle disorder. In the nebulin gene we have detected a number of autosomal recessive mutations. Both autosomal dominant and recessive mutations have been detected in the genes for alpha-actin and alpha-tropomyosin 3. A recessive mutation causing nemaline myopathy among the Old Order Amish has recently been identified in the gene for slow skeletal muscle troponin T. As linkage studies had shown that at least one further gene exists for nemaline myopathy, we investigated another tropomyosin gene expressed in skeletal muscle, the beta-tropomyosin 2 gene. Screening 66 unrelated patients, using single strand conformation polymorphism analysis and sequencing, we found four polymorphisms and two heterozygous missense mutations. Both mutations affect conserved amino acids, and in both cases, the mutant allele is expressed. We speculate that the observed mutations affect the formation of the tropomyosin dimer and its actin-binding properties.

Nemaline myopathy is clinically and genetically heterogeneous. The most common autosomal recessive form affecting infants (NEM2) links to chromosome 2q, and is caused by mutations in the gene for nebulin. We have examined the immunocytochemical expression of nebulin in skeletal muscle in 11 cases of nemaline myopathy, from ten families, with linkage compatible to chromosome 2q.22, the locus for nebulin. Mutations in the gene for nebulin have been found in eight of these cases. Immunolabelling with polyclonal antibodies to C-terminal regions of nebulin was compared with antibodies to fibre-type-specific myofibrillar proteins, including myosin heavy chain isoforms and alpha-actinin isoforms. No cases showed a complete absence of C-terminal nebulin, and no enhancement of labelling of the rods was seen with conventional fluorescence microscopy. In control muscle an antibody to the M176-181 repeat region of nebulin showed higher expression in fibres with slow myosin, while ones to the serine-rich domain and to the SH3 domain showed uniform expression. In some cases of nemaline myopathy differences in these patterns were observed. Two siblings with a homozygous mutation in exon 185, that produces a stop codon, showed an absence of labelling only with the SH3 antibody, and other cases showed uneven labelling with this antibody or some fibres devoid of label. Fibre type correlations also showed differences from controls, as some fibres had a fast isoform of one protein but a slow isoform of another. These results indicate that analysis of nebulin expression may detect abnormalities in some cases linked to the corresponding locus and may help to direct molecular analysis. In addition, they may also be relevant to studies of fibre type plasticity and diversity in nemaline myopathy.

Nemaline myopathy is a structural congenital myopathy which may show both autosomal dominant and autosomal recessive inheritance patterns. Mutations in three different genes have been identified as the cause of nemaline myopathy: the gene for slow alpha-tropomyosin 3 (TPM3) at 1q22-23, the nebulin gene (NEB) at 2q21.1-q22, and the actin gene (ACTA1) at 1q42. The typical autosomal recessive form appears to be the most common one and is caused by mutations in the nebulin gene. We have studied the pattern of nebulin labeling, in patients with the typical congenital form (ten patients), the severe congenital form (two patients) or the mild, childhood-onset form (one patient), using antibodies against three different domains of nebulin. A qualitative and quantitative nebulin analysis in muscle tissue showed the presence of nebulin in myofibers from all patients. Some differences relating to the rod structure were observed. The majority of the largest subsarcolemmal rods were not labeled with the N2 nebulin antibody (I-band epitope) and showed an indistinct pattern with the two antibodies directed to the Z-band portion of nebulin (epitopes M176-181 and serine-rich domain). Diffuse rods were not revealed using the three antibodies. A discordant pattern of nebulin N2 epitope labeling was found in two affected sisters with a mutation in the nebulin gene, suggesting that modifications in nebulin distribution inside the rods might occur with the progression of the disease. Western blot analysis showed no direct correlation with immunofluorescence data. In nine patients, the band had a molecular weight comparable to the normal control, while in one patient, it was detected with a higher molecular weight. Our results suggest that presence/absence of specific nebulin Z-band epitopes in rod structures is variable and could depend on the degree of rod organization.

Nemaline myopathy is a clinically and genetically heterogeneous condition. The clinical spectrum ranges from severe cases with antenatal or neonatal onset and early death to late onset cases with only slow progression. Three genes are known to cause nemaline myopathy: the genes for nebulin (NEB) on chromosome 2q22, slow alpha-tropomyosin (TPM3) on chromosome 1q21 and skeletal muscle alpha-actin (ACTA1) on chromosome 1q42. We present a 39-year-old lady with a mild form of nemaline myopathy, whom we have followed over a period of 25 years. She presented at the age of 7 years with symptoms of mild axial and proximal muscle weakness. The overall course was essentially static, but at 36 years, she went into life-threatening respiratory failure, for which she is currently treated with night-time ventilation. Muscle biopsies at 12, 17 and 39 years of age showed typical nemaline rods, particularly in type 1 fibres. Areas with unevenness of oxidative stain were present in the second and third biopsies. The presence of rods and core-like areas was confirmed on electron microscopy. There was no detectable alteration in actin expression immunocytochemically. A dominant missense mutation in the skeletal muscle alpha-actin gene (ACTA1) was found. This case illustrates the clinical and genetic heterogeneity of nemaline myopathy, and one phenotype of the wide spectrum of severity caused by mutations in the skeletal muscle alpha-actin (ACTA1) gene. In addition, it shows the diversity of pathological features that can occur in congenital myopathies due to mutations in the same gene.

The nemaline myopathies are muscle disorders of variable severity and age of onset, with characteristic nemaline bodies in the sarcoplasm. Genes for dominant (NEM1) and recessive (NEM2A) nemaline myopathy have been localised to chromosomes one and two, respectively. A missense mutation in the alpha-tropomyosin gene (TPM3) has been associated with NEM1 in one family. Probands from 76 other nemaline myopathy families have now been screened for TPM3 mutations. One proband, who was not noted to have any weakness neonatally, but who died at 21 months of age, was shown to be homozygous for a single strand conformation polymorphism (SSCP) in skeletal-muscle-specific exon 1 of TPM3. Sequencing revealed homozygosity for a nonsense mutation at codon 31 (CAG to TAG). The patient should have no functioning alpha-tropomyosin slow protein. The nemaline bodies in this patient were exclusively in type one fibres, consistent with the expression of TPM3 only in type one fibres.

Autosomal recessive nemaline (rod) myopathy is clinically and genetically heterogeneous. A clinically distinct, typical form, with onset in infancy and a non-progressive or slowly progressive course, has been assigned to a region on chromosome 2q22 harbouring the nebulin gene Mutations have now been found in this gene, confirming its causative role. The gene for slow tropomyosin TPM3 on chromosome 1q21, previously found to cause a dominantly inherited form, has recently been found to be homozygously mutated in one severe consanguineous case. Here we wished to determine the degree of genetic homogeneity or heterogeneity of autosomal recessive nemaline myopathy by linkage analysis of 45 families from 10 countries. Forty-one of the families showed linkage results compatible with linkage to markers in the nebulin region, the highest combined lod scores at zero recombination being 14.13 for the marker D2S2236. We found no indication of genetic heterogeneity for the typical form of nemaline myopathy. In four families with more severe forms of nemaline myopathy, however, linkage to both the nebulin and the TPM3 locus was excluded. Our results indicate that at least three genetic loci exist for autosomal recessive nemaline myopathy. Studies of additional families are needed to localise the as yet unknown causative genes, and to fully elucidate genotype-phenotype correlations.

We describe a family with an autosomal dominant, multisystem disorder, consisting of late-onset proximal muscular dystrophy, electrophysiological myotonia, cataracts, late-onset deafness and male hypogonadism. Four patients were available for clinical examinations. Examination of asymptomatic family members revealed another patient with bilateral cataracts but without definite muscle disorder. Five deceased members of the family had proximal muscle weakness, reportedly or confirmed in medical records. Molecular examination of genomic DNA showed no expansion of the unstable (CTG)n trinucleotide repeat on chromosome 19q13.3 associated with myotonic dystrophy (DM). Linkage to two loci implicated in other myotonic disorders, the muscle chloride channel (CLCN1) gene, and the muscle sodium channel (SCN4A) gene, was assessed and excluded. The clinical findings differ from those described in proximal myotonic myopathy (PROMM), in terms of the more severe muscle involvement with atrophy of affected muscles and the hearing loss. These findings suggest phenotypic and probably genetic heterogeneity among the proximal myotonic syndromes.

Clinical genetic evidence suggests the existence of an autosomal recessive form of congenital nemaline myopathy in addition to the autosomal dominant one(s). One mutation in an Australian kindred has been identified as causing an autosomal dominant form of the disease. This mutation in the alpha-tropomyosin gene TPM3 has previously been excluded as causing autosomal recessive nemaline myopathy. We searched systematically for genetic linkage to autosomal recessive nemaline myopathy (NEM2) by studying microsatellite marker alleles in seven multiplex families from Finland, Denmark, Wales, England and The Netherlands. Significant evidence of linkage was found to markers of chromosome 2q, the highest multipoint lod score value being 5.34 for the marker D2S151. Recombinant genotypes in affected individuals demarcate the the region in which the NEM2 gene is likely to reside as a 13 cM region between the markers D2S150 and D2S142. These results confirm the existence of at least one distinctive form of autosomal recessive nemaline myopathy and provide a basis for the identification of its gene.

To elucidate the protein composition of the nemaline bodies present in the muscle fibres of patients with congenital nemaline myopathy (CNM), we studied muscle biopsies with monoclonal antibodies against alpha-actinin and desmin in combination with a modified Gomori trichrome method. Electron microscopy of immunolabelled resin embedded sections was used for cytochemical localisation of alpha-actinin and desmin. Light microscopy of sections immunolabelled for alpha-actinin showed a cross-striation of the muscle fibres corresponding to the Z band pattern, focal thickening of the Z bands and additional reactivity with a granular pattern corresponding to the presence of nemaline bodies. Labelling of desmin did not show a similar pattern. Electron microscopy confirmed the presence of alpha-actinin in the nemaline bodies and Z bands, whereas desmin was only found in intermediate filaments around the Z bands. Western blots showed single, sharp alpha-actinin bands indistinguishable from normal. Our results provide direct evidence for the presence of alpha-actinin in nemaline bodies and a lack of quantitative or qualitative differences between the alpha-actinin of normal and CNM muscle.