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Phenotypes of myopathy-related beta-tropomyosin mutants in human and mouse tissue cultures.

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 ?-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-?-TM(EGFP) mutant showed perinuclear aggregates. The G53ins-?-TM(EGFP) mutant tended to accumulate in myoblasts but was incorporated into filamentous structures of myotubes. The K49del-?-TM(EGFP) and E122K-?-TM(EGFP) mutants induced the formation of rod-like structures in human cells. The N202K-?-TM(EGFP) mutant failed to integrate into thin filaments and formed accumulations in myotubes. The accumulation of mutant ?-TM(EGFP) 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 ?-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.

SUBMITTER: Abdul-Hussein S

PROVIDER: S-EPMC3769345 | biostudies-literature | 2013

REPOSITORIES: biostudies-literature

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Phenotypes of myopathy-related beta-tropomyosin mutants in human and mouse tissue cultures.

Abdul-Hussein Saba S Rahl Karin K Moslemi Ali-Reza AR Tajsharghi Homa H

PloS one 20130910 9

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 β-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-β-TM(EGFP) mutant showed perinuclear aggregates. The G53ins-β-TM(EGFP) mutant tended to accumulate in myoblasts but was incorporated into filamento ...[more]

PMID: 24039757

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Project description:The clinical features of myofibrillar myopathies display a wide phenotypic heterogeneity. To this date, no studies have evaluated this parameter due to the absence of pertinent animal models. By studying two mutants of desmin, which induce subtle phenotypic differences in patients, we address this issue using an animal model based on the use of adeno-associated virus (AAV) vectors carrying mutated desmin cDNA.After preparation of the vectors, they were injected directly into the tibialis anterior muscles of C57BL/6 mice to allow expression of wild-type (WT) or mutated (R406W or E413K) desmin. Measurements of maximal force were carried out on the muscle in situ and then the injected muscles were analyzed to determine the structural consequences of the desmin mutations on muscle structure (microscopic observations, histology and immunohistochemistry).Injection of AAV carrying WT desmin results in the expression of exogenous desmin in 98% of the muscle fibers without any pathological or functional perturbations. Exogenous WT and endogenous desmin are co-localized and no differences were observed compared to non-injected muscle. Expression of desmin mutants in mouse muscles induce morphological changes of muscle fibers (irregular shape and size) and the appearance of desmin accumulations around the nuclei (for R406W) or in subsarcolemmal regions of fibers (for E413K). These accumulations seem to occur and disrupt the Z-line, and a strong regeneration was observed in muscle expressing the R406W desmin, which is not the case for E413K. Moreover, both mutants of desmin studied here induce a decrease in muscle force generation capacity.In this study we show that AAV-mediated expression of desmin mutants in mouse muscles recapitulate the aggregation features, the decrease in contractile function and the morphological changes observed in patients with myofibrillar myopathy. More importantly, our results suggest that the R406W desmin mutant induces a robust muscle regeneration, which is not the case for the E413K mutant. This difference could help to explain the phenotypic differences observed in patients. Our results highlight the heterogeneous pathogenic mechanisms between different desmin mutants and open the way for new advances in the study of myofibrillar myopathies.

Project description:A GWAS study recently demonstrated single nucleotide polymorphisms (SNPs) in the human GLRB gene of individuals with a prevalence for agoraphobia. GLRB encodes the glycine receptor (GlyRs) ? subunit. The identified SNPs are localized within the gene flanking regions (3' and 5' UTRs) and intronic regions. It was suggested that these nucleotide polymorphisms modify GlyRs expression and phenotypic behavior in humans contributing to an anxiety phenotype as a mild form of hyperekplexia. Hyperekplexia is a human neuromotor disorder with massive startle phenotypes due to mutations in genes encoding GlyRs subunits. GLRA1 mutations have been more commonly observed than GLRB mutations. If an anxiety phenotype contributes to the hyperekplexia disease pattern has not been investigated yet. Here, we compared two mouse models harboring either a mutation in the murine Glra1 or Glrb gene with regard to anxiety and startle phenotypes. Homozygous spasmodic animals carrying a Glra1 point mutation (alanine 52 to serine) displayed abnormally enhanced startle responses. Moreover, spasmodic mice exhibited significant changes in fear-related behaviors (freezing, rearing and time spent on back) analyzed during the startle paradigm, even in a neutral context. Spastic mice exhibit reduced expression levels of the full-length GlyRs ? subunit due to aberrant splicing of the Glrb gene. Heterozygous animals appear normal without an obvious behavioral phenotype and thus might reflect the human situation analyzed in the GWAS study on agoraphobia and startle. In contrast to spasmodic mice, heterozygous spastic animals revealed no startle phenotype in a neutral as well as a conditioning context. Other mechanisms such as a modulatory function of the GlyRs ? subunit within glycinergic circuits in neuronal networks important for fear and fear-related behavior may exist. Possibly, in human additional changes in fear and fear-related circuits either due to gene-gene interactions e.g., with GLRA1 genes or epigenetic factors are necessary to create the agoraphobia and in particular the startle phenotype.

Dataset Information

Phenotypes of myopathy-related beta-tropomyosin mutants in human and mouse tissue cultures.

Publications

Phenotypes of myopathy-related beta-tropomyosin mutants in human and mouse tissue cultures.

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