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Novel mutations in NEB cause abnormal nebulin expression and markedly impaired muscle force generation in severe nemaline myopathy

Division of Genetics and Program in Genomics, The Manton Center for Orphan Disease Research, Children's Hospital Boston, Harvard Medical School, 300 Longwood Avenue, CLSB 15026, Boston, MA 02115, USA.
Skeletal muscle 06/2011; 1(1):23. DOI: 10.1186/2044-5040-1-23
Source: PubMed

ABSTRACT Nemaline myopathy (NM) is a congenital muscle disease associated with weakness and the presence of nemaline bodies (rods) in muscle fibers. Mutations in seven genes have been associated with NM, but the most commonly mutated gene is nebulin (NEB), which is thought to account for roughly 50% of cases.
We describe two siblings with severe NM, arthrogryposis and neonatal death caused by two novel NEB mutations: a point mutation in intron 13 and a frameshift mutation in exon 81. Levels of detectable nebulin protein were significantly lower than those in normal control muscle biopsies or those from patients with less severe NM due to deletion of NEB exon 55. Mechanical studies of skinned myofibers revealed marked impairment of force development, with an increase in tension cost.
Our findings demonstrate that the mechanical phenotype of severe NM is the consequence of mutations that severely reduce nebulin protein levels and suggest that the level of nebulin expression may correlate with the severity of disease.

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Available from: Henk L Granzier, Jul 23, 2015
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    • "Nebulin-deficient myofibrils also have a reduced Ca 2+ sensitivity of force production (Witt et al., 2006), a slower rate of tension redevelopment and an elevated tension cost, meaning more ATP has to be expended to generate a given tension. This suggests that there is a reduced fraction of force-generating cross-bridges, which, in turn, contributes to muscle weakness (Ottenheijm et al., 2010; Lawlor et al., 2011a). Recently studies have shown that in vitro treatment of nebulin-deficient myofibres with a fast skeletal muscle troponin activator (CK-2066260) enhanced force production at submaximal activating Ca 2+ levels (de Winter et al., 2013; Lee et al., 2013; Ottenheijm et al., 2013). "
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    ABSTRACT: The congenital myopathies are a diverse group of genetic skeletal muscle diseases, which typically present at birth or in early infancy. There are multiple modes of inheritance and degrees of severity (ranging from foetal akinesia, through lethality in the newborn period to milder early and later onset cases). Classically, the congenital myopathies are defined by skeletal muscle dysfunction and a non-dystrophic muscle biopsy with the presence of one or more characteristic histological features. However, mutations in multiple different genes can cause the same pathology and mutations in the same gene can cause multiple different pathologies. This is becoming ever more apparent now that, with the increasing use of next generation sequencing, a genetic diagnosis is achieved for a greater number of patients. Thus, considerable genetic and pathological overlap is emerging, blurring the classically established boundaries. At the same time, some of the pathophysiological concepts underlying the congenital myopathies are moving into sharper focus. Here we explore whether our emerging understanding of disease pathogenesis and underlying pathophysiological mechanisms, rather than a strictly gene-centric approach, will provide grounds for a different and perhaps complementary grouping of the congenital myopathies, that at the same time could help instil the development of shared potential therapeutic approaches. Stemming from recent advances in the congenital myopathy field, five key pathophysiology themes have emerged: defects in (i) sarcolemmal and intracellular membrane remodelling and excitation-contraction coupling; (ii) mitochondrial distribution and function; (iii) myofibrillar force generation; (iv) atrophy; and (v) autophagy. Based on numerous emerging lines of evidence from recent studies in cell lines and patient tissues, mouse models and zebrafish highlighting these unifying pathophysiological themes, here we review the congenital myopathies in relation to these emerging pathophysiological concepts, highlighting both areas of overlap between established entities, as well as areas of distinction within single gene disorders. Published by Oxford University Press on behalf of the Guarantors of Brain 2014. This work is written by US Government employees and is in the public domain in the US.
    Brain 12/2014; 138(2). DOI:10.1093/brain/awu368 · 10.23 Impact Factor
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    • "Indeed, myofibers force production was severely decreased in a NM patient with $10% of the normal nebulin level due to compound heterozygous NEB mutations. This impaired contractile performance largely exceeded what has been reported in patients for whom the nebulin protein level was roughly 28% of control values as a result of a deletion of exon 55 [25] [26]. On the contrary, a mild muscle weakness has been reported in a patient with 70% of the control protein level while single muscle fibers analysis showed no changes in both the force-sarcomere length relationship and in the calcium-sensitivity of force generation [27]. "
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    ABSTRACT: Nemaline myopathy is the most common congenital skeletal muscle disease, and mutations in the nebulin gene account for 50% of all cases. Recent studies suggest that the disease severity might be related to the nebulin expression levels. Considering that mutations in the nebulin gene are typically recessive, one would expect that a single functional nebulin allele would maintain nebulin protein expression which would result in preserved skeletal muscle function. We investigated skeletal muscle function of heterozygous nebulin knock-out (i.e., nebulin +/À) mice using a multidisciplinary approach including protein and gene expression analysis and combined in vivo and in vitro force measurements. Skeletal muscle anatomy and energy metabolism were studied strictly non-invasively using magnetic resonance imaging and 31P-magnetic resonance spectroscopy. Maximal force production was reduced by around 16% in isolated muscle of nebulin +/À mice while in vivo force generating capacity was preserved. Muscle weakness was associated with a shift toward a slower proteomic phenotype, but was not related to nebulin protein deficiency or to an impaired energy metabolism. Further studies would be warranted in order to determine the mechanisms leading to a mild skeletal muscle phenotype resulting from the expression of a single nebulin allele.
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    • "Indeed, myofibers force production was severely decreased in a NM patient with $10% of the normal nebulin level due to compound heterozygous NEB mutations. This impaired contractile performance largely exceeded what has been reported in patients for whom the nebulin protein level was roughly 28% of control values as a result of a deletion of exon 55 [25] [26]. On the contrary, a mild muscle weakness has been reported in a patient with 70% of the control protein level while single muscle fibers analysis showed no changes in both the force-sarcomere length relationship and in the calcium-sensitivity of force generation [27]. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Nemaline myopathy is the most common congenital skeletal muscle disease, and mutations in the nebulin gene account for 50% of all cases. Recent studies suggest that the disease severity might be related to the nebulin expression levels. Considering that mutations in the nebulin gene are typically recessive, one would expect that a single functional nebulin allele would maintain nebulin protein expression which would result in preserved skeletal muscle function. We investigated skeletal muscle function of heterozygous nebulin knock-out (i.e., nebulin(+/-)) mice using a multidisciplinary approach including protein and gene expression analysis and combined in vivo and in vitro force measurements. Skeletal muscle anatomy and energy metabolism were studied strictly non-invasively using magnetic resonance imaging and 31P-magnetic resonance spectroscopy. Maximal force production was reduced by around 16% in isolated muscle of nebulin(+/-) mice while in vivo force generating capacity was preserved. Muscle weakness was associated with a shift toward a slower proteomic phenotype, but was not related to nebulin protein deficiency or to an impaired energy metabolism. Further studies would be warranted in order to determine the mechanisms leading to a mild skeletal muscle phenotype resulting from the expression of a single nebulin allele.
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