The limb girdle muscular dystrophies (LGMDs) are genetically inherited diseases defined by progressive, proximal muscle wasting. Fifteen autosomal recessive LGMD disease genes have been identified, the majority of which link to the dystrophin glycoprotein complex and involve membrane fragility as a primary pathogenic feature. The most prevalent form of LGMD, LGMD2A, results from mutations in the gene encoding the muscle specific, calcium-dependent protease calpain 3 (capn3, C3). Unlike most other LGMDs, calpainopathy does not involve membrane damage as a primary feature; therefore, the mechanisms of this disease are quite different from other LGMDs and have remained mysterious since the gene defect was discovered in 1997. Over the last decade we have generated numerous genetically modified mice, which have been used to elucidate C3's biological role and to understand pathomechanisms of calpainopathy. This work has revealed a role for C3 in muscle growth and adaptation. Furthermore, muscles lacking C3 have reductions in the abundance and activation of the calcium calmodulin kinase (CaMK) signaling pathway at rest and upon muscle loading. The reductions in CaMK correspond to reduced slow fiber associated gene expression and a decreased number of slow fibers. In support of the relevance of these findings to calpainopathy, we also observed a primary involvement of slow fibers in LGMD2A biopsies. Thus, these studies are the first to identify defective CaMK signaling as the basis of the growth and adaptation defect in calpainopathy, and represent the first potential therapeutic target in LGMD2A. To define the relationship between C3 and CaMK, to identify biomarkers and to validate CaMK as a therapeutic target, we will carry out the following aims.
Aim 1 : We will test possible mechanisms by which calpain 3 modulates CaMK signaling.
Aim 2 : We will determine a transcriptional signature induced during muscle adaptation and determine the role of calpain 3 on that signature.
Aim 3 : We will create a bank of human myotubes derived from patients with different LGMD2A mutations, and then use this in vitro model to examine the effect of specific calpain 3 mutations on CaMK signaling and the gene signature identified in aim 2.
Aim 4 : We will determine if defects in CaMK signaling underlie the pathology of calpainopathy, and whether restoring downstream components of this signaling cascade improves the C3KO phenotype. Data generated from these efforts will provide fundamental insights into mechanisms and biomarkers and will reveal a novel therapeutic strategy for LGMD2A and may have far reaching implications for other diseases and conditions that impact muscle health, such as multi-minicore disease and sarcopenia that occurs with aging.

Public Health Relevance

In this proposal, we seek to identify mechanisms of muscle adaptation and growth that occur after a bout of atrophy. We also seek to elucidate the mechanism underlying limb girdle muscular dystrophy type 2A, a genetically inherited muscle disease for which there is no cure. In this application, we propose to exploit mouse models followed by the use of human models in which we will consistently validate our mouse data. These basic and pre-clinical studies will enhance our understanding of how muscle senses loading and responds with growth, which will enable us to identify therapeutic targets for muscular dystrophy and sarcopenia that occurs with aging.

National Institute of Health (NIH)
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Research Project (R01)
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Skeletal Muscle Biology and Exercise Physiology Study Section (SMEP)
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Cheever, Thomas
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University of California Los Angeles
Schools of Medicine
Los Angeles
United States
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Kramerova, Irina; Torres, Jorge A; Eskin, Ascia et al. (2018) Calpain 3 and CaMKII? signaling are required to induce HSP70 necessary for adaptive muscle growth after atrophy. Hum Mol Genet 27:1642-1653
DiFranco, Marino; Kramerova, Irina; Vergara, Julio L et al. (2016) Attenuated Ca(2+) release in a mouse model of limb girdle muscular dystrophy 2A. Skelet Muscle 6:11
Kramerova, Irina; Ermolova, Natalia; Eskin, Ascia et al. (2016) Failure to up-regulate transcription of genes necessary for muscle adaptation underlies limb girdle muscular dystrophy 2A (calpainopathy). Hum Mol Genet 25:2194-2207
Mokhonova, Ekaterina I; Avliyakulov, Nuraly K; Kramerova, Irina et al. (2015) The E3 ubiquitin ligase TRIM32 regulates myoblast proliferation by controlling turnover of NDRG2. Hum Mol Genet 24:2873-83
Ermolova, Natalia; Kramerova, Irina; Spencer, Melissa J (2015) Autolytic activation of calpain 3 proteinase is facilitated by calmodulin protein. J Biol Chem 290:996-1004
Kudryashova, Elena; Kramerova, Irina; Spencer, Melissa J (2012) Satellite cell senescence underlies myopathy in a mouse model of limb-girdle muscular dystrophy 2H. J Clin Invest 122:1764-76
Kramerova, I; Kudryashova, E; Ermolova, N et al. (2012) Impaired calcium calmodulin kinase signaling and muscle adaptation response in the absence of calpain 3. Hum Mol Genet 21:3193-204
Jaka, Oihane; Kramerova, Irina; Azpitarte, Margarita et al. (2012) C3KO mouse expression analysis: downregulation of the muscular dystrophy Ky protein and alterations in muscle aging. Neurogenetics 13:347-57
Ermolova, Natalia; Kudryashova, Elena; DiFranco, Marino et al. (2011) Pathogenity of some limb girdle muscular dystrophy mutations can result from reduced anchorage to myofibrils and altered stability of calpain 3. Hum Mol Genet 20:3331-45
Kudryashova, Elena; Struyk, Arie; Mokhonova, Ekaterina et al. (2011) The common missense mutation D489N in TRIM32 causing limb girdle muscular dystrophy 2H leads to loss of the mutated protein in knock-in mice resulting in a Trim32-null phenotype. Hum Mol Genet 20:3925-32

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