Calpain 3 is a modulator of the dysferlin protein complex in skeletal muscle

Center for Human and Clinical Genetics, Leiden University Medical Center, Leiden, The Netherlands.
Human Molecular Genetics (Impact Factor: 6.39). 07/2008; 17(12):1855-66. DOI: 10.1093/hmg/ddn081
Source: PubMed


Muscular dystrophies comprise a genetically heterogeneous group of degenerative muscle disorders characterized by progressive
muscle wasting and weakness. Two forms of limb-girdle muscular dystrophy, 2A and 2B, are caused by mutations in calpain 3
(CAPN3) and dysferlin (DYSF), respectively. While CAPN3 may be involved in sarcomere remodeling, DYSF is proposed to play
a role in membrane repair. The coexistence of CAPN3 and AHNAK, a protein involved in subsarcolemmal cytoarchitecture and membrane
repair, in the dysferlin protein complex and the presence of proteolytic cleavage fragments of AHNAK in skeletal muscle led
us to investigate whether AHNAK can act as substrate for CAPN3. We here demonstrate that AHNAK is cleaved by CAPN3 and show
that AHNAK is lost in cells expressing active CAPN3. Conversely, AHNAK accumulates when calpain 3 is defective in skeletal
muscle of calpainopathy patients. Moreover, we demonstrate that AHNAK fragments cleaved by CAPN3 have lost their affinity
for dysferlin. Thus, our findings suggest interconnectivity between both diseases by revealing a novel physiological role
for CAPN3 in regulating the dysferlin protein complex.

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Available from: Antoine de Morree, Dec 16, 2014
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    • "Accordingly, DYSF is considered to act with MG53, annexins and other proteins in the accumulation of vesicles at the site of damage following membrane disruption (Fig. 3). In addition to MG53, annexins and caveolin-3, human neuroblast differentiation-associated protein (AHNAK)33, affixin34, S100A1035, calpain-336, tubulin37, and dihydropyridine receptor (DHPR)38 have been reported to interact with DYSF. Figure 4 shows a schema of the proteins that interact with DYSF at the sarcolemma in skeletal muscle. "
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    ABSTRACT: Dysferlin (DYSF) is involved in the membrane-repair process, in the intracellular vesicle system and in T-tubule development in skeletal muscle. It interacts with mitsugumin 53, annexins, caveolin-3, AHNAK, affixin, S100A10, calpain-3, tubulin and dihydropyridine receptor. Limb-girdle muscular dystrophy 2B (LGMD2B) and Miyoshi myopathy (MM) are muscular dystrophies associated with recessively inherited mutations in the DYSF gene. The diseases are characterized by weakness and muscle atrophy that progress slowly and symmetrically in the proximal muscles of the limb girdles. LGMD2B and MM, which are collectively termed "dysferlinopathy", both lead to abnormalities in vesicle traffic and membrane repair at the plasma membrane in muscle fibers. SJL/J (SJL) and A/J mice are naturally occurring animal models for dysferlinopathy. Since there has been no an approach to therapy for dysferlinopathy, the immediate development of a therapeutic method for this genetic disorder is desirable. The murine models are useful in verification experiments for new therapies and they are valuable tools for identifying factors that accelerate dystrophic changes in skeletal muscle. It could be possible that the genetic or immunological background in SJL or A/J mice could modify muscle damage in experiments involving these models, because SJL and A/J mice show differences in the progress and prevalent sites of skeletal muscle lesions as well as in the gene-expression profiles of their skeletal muscle. In this review, we provide up-to-date information on the function of dysferlin, the development of possible therapies for muscle dystrophies (including dysferlinopathy) and the detection of new therapeutic targets for dysferlinopathy by means of experiments using animal models for dysferlinopathy.
    Full-text · Article · Jun 2012 · Journal of Toxicologic Pathology
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    • "myosin light chain 1, suggesting a role for calpain 3 in sarcomere remodeling [5,8,9]. Another important in vivo calpain 3 substrates that has been described is AHNAK, a very large protein involved in subsarcolemmal cytostructure and part of the dysferlin membrane repair complex, requires calpain 3 for it to be cleaved and the membrane repair to proceed [10]. Hence, it is a key component in the repair of the wear and tear of skeletal muscle tissue. "
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    ABSTRACT: Limb girdle muscular dystrophy (LGMD) type 2A is caused by mutations in the CAPN3 gene and complete lack of functional calpain 3 leads to the most severe muscle wasting. Calpain 3 is suggested to be involved in maturation of contractile elements after muscle degeneration. The aim of this study was to investigate how mutations in the four functional domains of calpain 3 affect muscle regeneration. We studied muscle regeneration in 22 patients with LGMD2A with calpain 3 deficiency, in five patients with LGMD2I, with a secondary reduction in calpain 3, and in five patients with Becker muscular dystrophy (BMD) with normal calpain 3 levels. Regeneration was assessed by using the developmental markers neonatal myosin heavy chain (nMHC), vimentin, MyoD and myogenin and counting internally nucleated fibers. We found that the recent regeneration as determined by the number of nMHC/vimentin-positive fibers was greatly diminished in severely affected LGMD2A patients compared to similarly affected patients with LGMD2I and BMD. Whorled fibers, a sign of aberrant regeneration, was highly elevated in patients with a complete lack of calpain 3 compared to patients with residual calpain 3. Regeneration is not affected by location of the mutation in the CAPN3 gene. Our findings suggest that calpain 3 is needed for the regenerative process probably during sarcomere remodeling as the complete lack of functional calpain 3 leads to the most severe phenotypes.
    Full-text · Article · Mar 2012 · BMC Musculoskeletal Disorders
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    • "Furthermore, ahnak has been identified as a constituent of the dysferlin protein complex that is important for sarcolemmal integrity (Huang et al. 2007, 2008; de Morree et al. 2010). Interestingly, the interaction between ahnak1 and dysferlin is lost after proteolytic processing of ahnak1 by calpain3 (Huang et al. 2008). Because of the complex interactions between ahnak1, dysferlin, and calpain3 and their role in muscle repair and regeneration we compared ahnak1 expression in patients with LGMD2B and LGMD2A. "
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    ABSTRACT: Ahnak1 is a giant, ubiquitously expressed, plasma membrane support protein whose function in skeletal muscle is largely unknown. Therefore, we investigated whether ahnak would be influenced by alterations of the sarcolemma exemplified by dysferlin mutations known to render the sarcolemma vulnerable or by mutations in calpain3, a protease known to cleave ahnak. Human muscle biopsy specimens obtained from patients with limb girdle muscular dystrophy (LGMD) caused by mutations in dysferlin (LGMD2B) and calpain3 (LGMD2A) were investigated for ahnak expression and localization. We found that ahnak1 has lost its sarcolemmal localization in LGMD2B but not in LGMD2A. Instead ahnak1 appeared in muscle connective tissue surrounding the extracellular site of the muscle fiber in both muscular dystrophies. The entire giant ahnak1 molecule was present outside the muscle fiber and did only partially colocalize with CD45-positive immune cell infiltration and the extracelluar matrix proteins fibronectin and collagenVI. Further, vesicles shedded in response to Ca2+ by primary human myotubes were purified and their protein content was analysed. Ahnak1 was prominently present in these vesicles. Electron microscopy revealed a homogenous population of vesicles with a diameter of about 150 nm. This is the first study demonstrating vesicle release from human myotubes that may be one mechanism underlying abnormally localized ahnak1. Taken together, our results define ahnak1 in muscle connective tissue as a novel feature of two genetically distinct muscular dystrophies that might contribute to disease pathology. Electronic supplementary material The online version of this article (doi:10.1007/s10974-011-9271-8) contains supplementary material, which is available to authorized users.
    Full-text · Article · Nov 2011 · Journal of Muscle Research and Cell Motility
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