| 1. |
- Abdul-Hussein, Saba, et al.
(författare)
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Expression profiles of muscle disease-associated genes and their isoforms during differentiation of cultured human skeletal muscle cells
- 2012
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Ingår i: BMC Musculoskeletal Disorders. - : BioMed Central. - 1471-2474. ; 13
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: The formation of contractile myofibrils requires the stepwise onset of expression of muscle specific proteins. It is likely that elucidation of the expression patterns of muscle-specific sarcomeric proteins is important to understand muscle disorders originating from defects in contractile sarcomeric proteins.METHODS: We investigated the expression profile of a panel of sarcomeric components with a focus on proteins associated with a group of congenital disorders. The analyses were performed in cultured human skeletal muscle cells during myoblast proliferation and myotube development.RESULTS: Our culture technique resulted in the development of striated myotubes and the expression of adult isoforms of the sarcomeric proteins, such as fast TnI, fast TnT, adult fast and slow MyHC isoforms and predominantly skeletal muscle rather than cardiac actin. Many proteins involved in muscle diseases, such as beta tropomyosin, slow TnI, slow MyBPC and cardiac TnI were readily detected in the initial stages of muscle cell differentiation, suggesting the possibility of an early role for these proteins as constituent of the developing contractile apparatus during myofibrillogenesis. This suggests that in disease conditions the mechanisms of pathogenesis for each of the mutated sarcomeric proteins might be reflected by altered expression patterns, and disturbed assembly of cytoskeletal, myofibrillar structures and muscle development.CONCLUSIONS: In conclusion, we here confirm that cell cultures of human skeletal muscle are an appropriate tool to study developmental stages of myofibrillogenesis. The expression of several disease-associated proteins indicates that they might be a useful model system for studying the pathogenesis of muscle diseases caused by defects in specific sarcomeric constituents.
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| 2. |
- Abdul-Hussein, Saba, et al.
(författare)
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Phenotypes of Myopathy-Related Beta-Tropomyosin Mutants in Human and Mouse Tissue Cultures
- 2013
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Ingår i: Plos One. - : Public Library of Science (PLoS). - 1932-6203. ; 8:9
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Tidskriftsartikel (refereegranskat)abstract
- Mutations in TPM2 result in a variety of myopathies characterised by variable clinical and morphological features. We used human and mouse cultured cells to study the effects of beta-TM mutants. The mutants induced a range of phenotypes in human myoblasts, which generally changed upon differentiation to myotubes. Human myotubes transfected with the E41K-beta-TMEGFP mutant showed perinuclear aggregates. The G53ins-beta-TMEGFP mutant tended to accumulate in myoblasts but was incorporated into filamentous structures of myotubes. The K49del-beta-TMEGFP and E122K-beta-TMEGFP mutants induced the formation of rod-like structures in human cells. The N202K-beta-TMEGFP mutant failed to integrate into thin filaments and formed accumulations in myotubes. The accumulation of mutant beta-TMEGFP in the perinuclear and peripheral areas of the cells was the striking feature in C2C12. We demonstrated that human tissue culture is a suitable system for studying the early stages of altered myofibrilogenesis and morphological changes linked to myopathy-related beta-TM mutants. In addition, the histopathological phenotype associated with expression of the various mutant proteins depends on the cell type and varies with the maturation of the muscle cell. Further, the phenotype is a combinatorial effect of the specific amino acid change and the temporal expression of the mutant protein.
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| 3. |
- Olive, M., et al.
(författare)
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New cardiac and skeletal protein aggregate myopathy associated with combined MuRF1 and MuRF3 mutations
- 2015
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Ingår i: Human Molecular Genetics. - : Oxford University Press (OUP). - 0964-6906 .- 1460-2083. ; 24:13, s. 3638-3650
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Tidskriftsartikel (refereegranskat)abstract
- Protein aggregate myopathies (PAMs) define muscle disorders characterized by protein accumulation in muscle fibres. We describe a new PAM in a patient with proximal muscle weakness and hypertrophic cardiomyopathy, whose muscle fibres contained inclusions containing myosin and myosin-associated proteins, and aberrant distribution of microtubules. These lesions appear as intact A- and M-bands lacking thin filaments and Z-discs. These features differ from inclusions in myosin storage myopathy (MSM), but are highly similar to those in mice deficient for the muscle-specific RING finger proteins MuRF1 and MuRF3. Sanger sequencing excluded mutations in the MSM-associated gene MYH7 but identified mutations in TRIM63 and TRIM54, encoding MuRF1 and MuRF3, respectively. No mutations in other potentially disease-causing genes were identified by Sanger and whole exome sequencing. Analysis of seven family members revealed that both mutations segregated in the family but only the homozygous TRIM63 null mutation in combination with the heterozygous TRIM54 mutation found in the proband caused the disease phenotype. Both MuRFs are microtubule-associated proteins localizing to sarcomeric M-bands and Z-discs. They are E3 ubiquitin ligases that play a role in degradation of sarcomeric proteins, stabilization of microtubules and myogenesis. Lack of ubiquitin and the 20S proteasome subunit in the inclusions found in the patient suggested impaired turnover of thick filament proteins. Disruption of microtubules in cultured myotubes was rescued by transient expression of wild-type MuRF1. The unique features of this novel myopathy point to defects in homeostasis of A-band proteins in combination with instability of microtubules as cause of the disease.
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| 4. |
- Pokrzywa, Malgorzata, 1977, et al.
(författare)
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Developmental MYH3 Myopathy Associated with Expression of Mutant Protein and Reduced Expression Levels of Embryonic MyHC
- 2015
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Ingår i: PLoS One. - : Public Library of Science (PLoS). - 1932-6203. ; 10:11
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Tidskriftsartikel (refereegranskat)abstract
- Objective An essential role for embryonic MyHC in foetal development has been found from its association with distal arthrogryposis syndromes, a heterogeneous group of disorders characterised by congenital contractions. The latter probably result from severe myopathy during foetal development. Lack of embryonic muscle biopsy material and suitable animal models has hindered study of the pathomechanisms linking mutations in MYH3 to prenatal myopathy. We determined the pathomechanisms of developmental myopathy caused by recurrent p. Thr178Ile MYH3 heterozygosity, using patient-derived skeletal muscle cells in culture as an experimental disease model to emulate early embryonic development. These cultured cells were processed for discrimination and quantitative analysis of mutant and wild-type MYH3 alleles and MyHC transcripts, real-time RT-qPCR, sequence analysis, immunofluorescence microscopy, immunoblot, and proteomic assessments. Involvement of the ubiquitin proteasome system was investigated in patients with p. Thr178Ile mutations in MYH3 and MYH2. We found equal overall expression of mutant and wild-type MyHC mRNAs and proteins. Compared to the controls, however, expression of embryonic MyHC transcripts and proteins was reduced whereas expression of myosin-specific E3 ubiquitin ligase (MuRF1) was increased. We also found delayed myofibrillogenesis and atrophic myotubes but structured sarcomeres. In conclusion, this study suggests that developmental p.Thr178Ile MYH3 myopathy is associated with a combined pathomechanism of insufficient dosage of functional embryonic MyHC and production of mutant protein.
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| 5. |
- Tajsharghi, Homa, et al.
(författare)
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Unexpected myopathy associated with a mutation in MYBPC3 and misplacement of the cardiac myosin binding protein C
- 2010
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Ingår i: Journal of Medical Genetics. - : BMJ Publishing Group. - 0022-2593 .- 1468-6244. ; 47:8, s. 575-577
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Tidskriftsartikel (refereegranskat)abstract
- BACKGROUND: Myosin binding protein C (MyBPC) is essential for the structure of the sarcomeres in striated muscle. There is one cardiac specific isoform and two skeletal muscle specific isoforms. Mutations in MYBPC3 encoding the cardiac isoform cause cardiomyopathy.METHODS AND RESULTS: We have identified an infant with fatal cardiomyopathy due to a homozygous mutation, p.R943X, in MYBPC3. The patient also had an unexpected skeletal myopathy. The patient expressed the cardiac specific MyBPC isoform in skeletal muscle at transcript and protein levels. Numerous muscle fibres expressing the mutant cardiac isoform showed structural abnormalities with disorganisation of sarcomeres and depletion of myosin thick filaments.CONCLUSIONS: The surprising identification of a skeletal myopathy in this patient was due to aberrant expression of mutant cardiac MyBPC in skeletal muscle.
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