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Contains fulltext : 231705.pdf (Publisher’s version ) (Open Access), The extracellular matrix comprises a network of macromolecules such as collagens, proteoglycans and glycoproteins. VWA1 (von Willebrand factor A domain containing 1) encodes a component of the extracellular matrix that interacts with perlecan/collagen VI, appears to be involved in stabilizing extracellular matrix structures, and demonstrates high expression levels in tibial nerve. Vwa1-deficient mice manifest with abnormal peripheral nerve structure/function; however, VWA1 variants have not previously been associated with human disease. By interrogating the genome sequences of 74 180 individuals from the 100K Genomes Project in combination with international gene-matching efforts and targeted sequencing, we identified 17 individuals from 15 families with an autosomal-recessive, non-length dependent, hereditary motor neuropathy and rare biallelic variants in VWA1. A single disease-associated allele p.(G25Rfs*74), a 10-bp repeat expansion, was observed in 14/15 families and was homozygous in 10/15. Given an allele frequency in European populations approaching 1/1000, the seven unrelated homozygote individuals ascertained from the 100K Genomes Project represents a substantial enrichment above expected. Haplotype analysis identified a shared 220 kb region suggesting that this founder mutation arose >7000 years ago. A wide age-range of patients (6-83 years) helped delineate the clinical phenotype over time. The commonest disease presentation in the cohort was an early-onset (mean 2.0 ± 1.4 years) non-length-dependent axonal hereditary motor neuropathy, confirmed on electrophysiology, which will have to be differentiated from other predominantly or pure motor neuropathies and neuronopathies. Because of slow disease progression, ambulation was largely preserved. Neurophysiology, muscle histopathology, and muscle MRI findings typically revealed clear neurogenic changes with single isolated cases displaying additional myopathic process. We speculate that a few findings of my

BACKGROUND AND PURPOSE: Nemaline myopathy (NEM) has been associated with mutations in 12 genes to date. However, for some patients diagnosed with NEM, definitive mutations are not identified in the known genes, suggesting that there are other genes involved. This study describes compound heterozygosity for rare variants in ryanodine receptor type 3 (RYR3) gene in one such patient. METHODS AND RESULTS: Clinical examination of the patient at 22 years of age revealed a long narrow face, high arched palate and bilateral facial weakness. She had proximal weakness in all four limbs, mild scapular winging but no scoliosis. Muscle biopsy revealed wide variation in fibre size with type 1 fibre predominance and atrophy. Abundant nemaline bodies were located in perinuclear and subsarcolemmal areas, and within the cytoplasm. No likely pathogenic mutations in known NEM genes were identified. Copy number variation in known NEM genes was excluded by NEM-targeted comparative genomic hybridization array. Next-generation sequencing revealed compound heterozygous missense variants in the RYR3 gene. RYR3 transcripts are expressed in human fetal and adult skeletal muscle as well as in human brain and cauda equina samples. Immunofluorescence of human skeletal muscle revealed a 'single-row' appearance of RYR3, interspersed between the 'double rows' of ryanodine receptor type 1 (RYR1) at each A-I junction. CONCLUSION: The results suggest that variants in RYR3 may cause a recessive muscle disease with pathological features including nemaline bodies. We characterize the expression pattern of RYR3 in human skeletal muscle and brain, and the subcellular localization of RYR1 and RYR3 in human skeletal muscle.

Hereditary myopathy with early respiratory failure (HMERF) and extensive myofibrillar lesions have been described in sporadic and familial cases and linked to various chromosomal regions. We describe the clinical manifestations, muscle histopathology and genetics in eight individuals from three apparently unrelated families with clinical and pathological features of HMERF. All patients had muscle weakness in the pelvic girdle, neck flexors, respiratory and trunk muscles, and the majority had prominent calf hypertrophy. Examination of pulmonary function showed decreased vital capacity. No signs of cardiac muscle involvement were found. Muscle histopathological features included marked muscle fibre size variation, fibre splitting, numerous internal nuclei and fatty infiltration. Frequent groups of fibres showed eosinophilic inclusions and deposits. At the ultrastructural level there were extensive myofibrillar lesions with marked Z-disc alterations. Whole exome sequencing in four individuals from one family revealed a missense mutation, g.274375, T>C; p.Cys30071Arg, in the titin gene, TTN. The mutation, which changes a highly conserved residue in the myosin binding A-band titin, was demonstrated to segregate with the disease in all three families. High density single nucleotide polymorphism arrays covering the entire genome demonstrated sharing of a 699 Mb haplotype, located in chromosome region 2q31 including TTN, indicating common ancestry of this novel and first disease-causing mutation in A-band titin associated with HMERF.

Tropomyosin (TM), a sarcomeric thin-filament protein, plays an essential part in muscle contraction by regulating actin-myosin interaction. We describe two patients, a woman and her daughter, with muscle weakness and distal arthrogryposis (DA) type 2B, caused by a heterozygous missense mutation, R133W, in TPM2, the gene encoding beta-TM. Our results demonstrate the involvement of muscle dysfunction in the pathogenesis of DA and the fact that DA2B may be caused by mutations in TPM2.

Objective: To describe a three-generation family with distal arthrogryposis associated with myopathy and caused by a mutation in the gene encoding for sarcomeric thin filament protein troponin I, TNNI2. Methods: The authors performed clinical investigations and reviewed medical records. Muscle biopsy specimens were obtained for morphologic analysis. Genomic DNA was extracted from blood and analyzed for mutations in TNNI2. Results: The five affected individuals had predominantly distal congenital joint contractures, mild facial involvement (mild micrognathia, narrow palpebral fissures), and no detectable muscle weakness. The four affected adults had slightly increased levels of creatine kinase in blood, and muscle biopsy specimens showed findings of myopathy with changes restricted to type 2 fibers. These included variability of muscle fiber size, internalized nuclei, and increased interstitial connective tissue. Analysis of TNNI2 encoding the troponin I isoform expressed in type 2 muscle fibers disclosed a heterozygous three-base in-frame deletion, 2,918-2,920del, skipping the highly conserved lysine at position 176. The mutation was present in all 5 affected individuals but was not identified in any of the 11 unaffected family members. Conclusion: Distal arthrogryposis type 1 is genetically heterogeneous, and myopathy due to sarcomeric protein dysfunction may be one underlying cause of the disease.

The pathogenic events leading to the progressive muscle weakness in patients with a E706K mutation in the head of the myosin heavy chain (MyHC) IIa were analyzed at the muscle cell and motor protein levels. Contractile properties were measured in single muscle fiber segments using the skinned fiber preparation and a single muscle fiber in vitro motility assay. A dramatic impairment in the function of the IIa MyHC isoform was observed at the motor protein level. At the single muscle fiber level, on the other hand, a general decrease was observed in the number of preparations where the specific criteria for acceptance were fulfilled irrespective of MyHC isoform expression. Our results provide evidence that the pathogenesis of the MyHC IIa E706K myopathy involves defective function of the mutated myosin as well as alterations in the structural integrity of all muscle cells irrespective of MyHC isoform expression.

We report the observation of an 18-year-old girl, whose clinical presentation was very suggestive of a congenital myopathy with neonatal onset. A congenital myopathy had been already diagnosed in her brother and in addition her half-cousin died diagnosed with a severe nemaline myopathy at age 4 years. A muscle biopsy performed on both siblings revealed histological and ultrastructural features of 'cap myopathy'. This case report suggests that 'cap myopathy' and some cases of nemaline myopathy with neonatal onset might be two phenotypic expressions of the same genetic disorder. These two entities could therefore, perhaps, be regarded as 'Z-line disorders' possibly caused by defective myofibrillogenesis.

BACKGROUND: The authors recently described a new autosomal dominant myopathy (OMIM 605637 inclusion body myopathy 3) associated with a missense mutation in the myosin heavy chain (MyHC) IIa gene (MyHC IIa, Human Gene Map [HGM] locus MYH2). Young patients showed minor changes in their muscle biopsies, although dystrophic alterations and rimmed vacuoles with 15- to 20-nm tubulofilaments identical to those in sporadic inclusion body myositis (s-IBM) were observed in some of the adult (especially older) patients. The current study was undertaken to investigate the relation between expression of the mutant MyHC IIa and pathologic changes in muscle. METHODS: The expression of MyHC IIa in nine muscle specimens from six individuals carrying the mutation was analyzed by immunohistochemistry, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and a new reverse transcriptase--PCR method to measure the relative abundance of the various MyHC transcripts. RESULTS: Young patients with muscle weakness and minor pathologic changes in muscle expressed MyHC IIa at undetectable levels. MyHC IIa was expressed at high levels in adults with a progressive clinical course and dystrophic muscle changes. In these cases, a large number of muscle fibers were hybrids with expression of more than one MyHC isoform. Both MyHC IIa alleles were equally expressed. The relative level of MyHC IIa transcripts exceeded that of the corresponding protein, indicating an increased turnover of mutated protein. MyHC IIa expression was a consistent finding in muscle fibers with rimmed vacuoles. CONCLUSIONS: The clear correlation between pathologic changes and expression of MyHC IIa indicates that defects in MyHC may lead not only to muscle weakness but also to muscle degeneration. The consistent expression of MyHC IIa in muscle fibers with rimmed vacuoles indicates that the breakdown of sarcomeric proteins is a key element in the pathogenesis of rimmed vacuoles of s-IBM type.