A nemaline myopathy mutation in α-tropomyosin causes defective regulation of striated muscle force production

Michele, Daniel E.; Albayya, Faris; Metzger, Joseph M.

doi:10.1172/jci7842

Cited by 50 publications

(45 citation statements)

References 33 publications

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“…Sprague-Dawley rats were anesthetized and hearts were removed. Cardiac myocytes were isolated enzymatically as previously described with minor modifications (40). A total of 20,000 cells in 200 l of MEM with 5% FBS were plated on glass cover slips or Bioflex plates (Flex Cell International BF-3001U) that were coated with 200 l of 50 g/ml EHS laminin 50 (Sigma Aldrich L2020).…”

Section: Methodsmentioning

confidence: 99%

“…Alternatively, 1,000,000 total cells in 1 ml MEM with 5% FBS were plated on 100-mm culture dishes coated with 1 ml of 50 g/ml EHS laminin. Cardiac myocytes were transduced by adenovirus in culture media as described previously (40), and cells were cultured for 24 -72 h to express ␦SG.…”

Section: Methodsmentioning

confidence: 99%

“…Immunofluorescence staining on permeabilized myocytes was performed with fixation, and cells were permeabilized with 1% Triton-X 100 in PBS as described previously (40,41). For live-cell staining, media were aspirated, and cardiac myocytes were incubated with primary antibody suspended in MEM for 2 h at 37°C.…”

Section: Methodsmentioning

confidence: 99%

“…Cardiac myocytes were washed with 5 ml MEM for 5 min three times. After final aspiration of media cardiac myocytes were fixed with 2 ml of 3% paraformaldehyde in DPBS for 10 min and then permeabilized with 1% TX-100 in PBS followed by blocking with 5% BSA in PBS and secondary antibody staining, as described previously (40,41). Primary antibodies included: A-14 rabbit polyclonal anti-myc (Santa Cruz Biotechnology; 1:100), goat anti-human ␦-sarcoglycan (R&D Systems; 1:1,000), IIH6 anti-␣ dystroglycan (Millipore; 1:1,000), and mouse anti-␤ dystroglycan (Vector Laboratories; 1:1,000).…”

Section: Methodsmentioning

confidence: 99%

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Dilated cardiomyopathy mutations in δ-sarcoglycan exert a dominant-negative effect on cardiac myocyte mechanical stability

Campbell

Witcher

Gopal

et al. 2016

American Journal of Physiology-Heart and Circulatory Physiology

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Campbell MD, Witcher M, Gopal A, Michele DE. Dilated cardiomyopathy mutations in ␦-sarcoglycan exert a dominant-negative effect on cardiac myocyte mechanical stability. Am J Physiol Heart Circ Physiol 310: H1140 -H1150, 2016. First published March 11, 2016 doi:10.1152/ajpheart.00521.2015.-Delta-sarcoglycan is a component of the sarcoglycan subcomplex within the dystrophin-glycoprotein complex located at the plasma membrane of muscle cells. While recessive mutations in ␦-sarcoglycan cause limb girdle muscular dystrophy 2F, dominant mutations in ␦-sarcoglycan have been linked to inherited dilated cardiomyopathy (DCM). The purpose of this study was to investigate functional cellular defects present in adult cardiac myocytes expressing mutant ␦-sarcoglycans harboring the dominant inherited DCM mutations R71T or R97Q. This study demonstrates that DCM mutant ␦-sarcoglycans can be stably expressed in adult rat cardiac myocytes and traffic similarly to wild-type ␦-sarcoglycan to the plasma membrane, without perturbing assembly of the dystrophin-glycoprotein complex. However, expression of DCM mutant ␦-sarcoglycan in adult rat cardiac myocytes is sufficient to alter cardiac myocyte plasma membrane stability in the presence of mechanical strain. Upon cyclical cell stretching, cardiac myocytes expressing mutant ␦-sarcoglycan R97Q or R71T have increased cell-impermeant dye uptake and undergo contractures at greater frequencies than myocytes expressing normal ␦-sarcoglycan. Additionally, the R71T mutation creates an ectopic N-linked glycosylation site that results in aberrant glycosylation of the extracellular domain of ␦-sarcoglycan. Therefore, appropriate glycosylation of ␦-sarcoglycan may also be necessary for proper ␦-sarcoglycan function and overall dystrophin-glycoprotein complex function. These studies demonstrate that DCM mutations in ␦-sarcoglycan can exert a dominant negative effect on dystrophin-glycoprotein complex function leading to myocardial mechanical instability that may underlie the pathogenesis of ␦-sarcoglycan-associated DCM.Listen to this article's corresponding podcast at http://ajpheart. podbean.com/e/dilated-cardiomyopathy-delta-sarcoglycan-mutationscause-cardiomyocyte-membrane-instability/. dilated cardiomyopathy; ␦-sarcoglycan; dystrophin-glycoprotein complex; mechanical strain; dye uptake THE DYSTROPHIN-GLYCOPROTEIN complex (DGC) is a large complex of transmembrane and membrane-associated proteins that resides in the lateral sarcolemma of cardiac and skeletal muscle cells. The DGC contributes to an important functional linkage between the extracellular matrix (ECM) and cytoskeletal proteins in muscle cells (20,21). The DGC attaches and interacts with the cytoskeleton through the binding of dystrophin to F-actin (62). The central transmembrane protein dystroglycan completes the link of dystrophin and the DGC to the extracellular matrix and requires the functional glycosylation of ␣-dystroglycan to serve as a high-affinity matrix receptor (38,42,62). The critical functional roles of the DGC in musc...

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Dilated cardiomyopathy mutations in δ-sarcoglycan exert a dominant-negative effect on cardiac myocyte mechanical stability

Campbell

Witcher

Gopal

et al. 2016

American Journal of Physiology-Heart and Circulatory Physiology

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“…Nemaline myopathy is a rare, genetically heterogeneous, autosomal dominant or recessive myopathy characterized by severe, generalized weakness and hypotonia from birth or infancy [84, 85]. Nemaline myopathy may be due to mutations in the α-actin, α-tropomyosin or nebulin gene (table 1).…”

Section: Myopathies With Cardiac Involvementmentioning

confidence: 99%

Cardiac Involvement in Primary Myopathies

Finsterer

Stöllberger

2000

Cardiology

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Simultaneous or temporarily staggered affection of both the skeletal as well as the cardiac muscle (cardiac involvement, CI) is a frequent finding in primary myopathies (MPs). CI leads to impulse generation defects, impulse conduction defects, thickened myocardium, left ventriculalr hypertrabeculation, dilatation of the cardiac cavities, secondary valve insufficiency, reduction of coronary vasodilative reserve, intracardial thrombus formation, and heart failure with systolic and diastolic dysfunction. CI has been found in Duchenne muscular dystrophy (MD), Becker MD, Emery-Dreifuss MD, facioscapulohumeral MD, sarcoglycanopathies, myotubular congenital MD, myotonic dystrophies type 1 and 2, proximal myotonic myopathy, myoadenylate deaminase deficiency, glycogenosis type II, III, IV, VII and IX, carnitine deficiency, mitochondriopathy, desmin MP, nemaline MP, central core disease, multicore MP, congenital fiber-type disproportion MP, Barth syndrome, McLeod syndrome and Bethlem MP. Patients with any of the above-mentioned myopathies should be cardiologically investigated as soon as their diagnosis is established, since sufficient cardiac therapy improves CI in MPs and since management of these patients is influenced by the degree of CI.

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Thin Filament Proteins: Nemaline and Related Congenital Myopathies

Lawlor

Beggs

2013

Muscle Disease

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A nemaline myopathy mutation in α-tropomyosin causes defective regulation of striated muscle force production

Cited by 50 publications

References 33 publications

Dilated cardiomyopathy mutations in δ-sarcoglycan exert a dominant-negative effect on cardiac myocyte mechanical stability

Dilated cardiomyopathy mutations in δ-sarcoglycan exert a dominant-negative effect on cardiac myocyte mechanical stability

Cardiac Involvement in Primary Myopathies

Thin Filament Proteins: Nemaline and Related Congenital Myopathies

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