Plectin, a high-molecular-weight cytoskeletal linker protein, binds with high affinity to intermediate filaments of all types and connects them to junctional complexes, organelles, and inner membrane systems. In addition, it interacts with actomyosin structures and microtubules. As a multifunctional protein, plectin has been implicated in several multisystemic diseases, the most common of which is epidermolysis bullosa simplex with muscular dystrophy (EBS-MD). A great part of our knowledge about plectin\'s functional diversity has been gained through the analysis of a unique collection of transgenic mice that includes a full (null) knockout (KO), several tissue-restricted and isoform-specific KOs, three double KOs, and two knock-in lines. The key molecular features and pathological phenotypes of these mice will be discussed in this review. In summary, the analysis of the different genetic models indicated that a functional plectin is required for the proper function of striated and simple epithelia, cardiac and skeletal muscle, the neuromuscular junction, and the vascular endothelium, recapitulating the symptoms of humans carrying plectin mutations. The plectin-null line showed severe skin and muscle phenotypes reflecting the importance of plectin for hemidesmosome and sarcomere integrity; whereas the ablation of individual isoforms caused a specific phenotype in myofibers, basal keratinocytes, or neurons. Tissue-restricted ablation of plectin rendered the targeted cells less resilient to mechanical stress. Studies based on animal models other than the mouse, such as zebrafish and C. elegans, will be discussed as well.

BACKGROUND: Myofibrillar myopathies (MFM) are a subgroup of protein aggregate myopathies (PAM) characterized by a common histological picture of myofibrillar dissolution, Z-disk disintegration, and accumulation of degradation products into inclusions. Mutations in genes encoding components of the Z-disk or Z-disk-associated proteins occur in some patients whereas in most of the cases, the causative gene defect is still unknown. We aimed to search for pathogenic mutations in genes not previously associated with MFM phenotype.
METHODS: We performed whole-exome sequencing in four patients from three unrelated families who were diagnosed with PAM without aberrations in causative genes for MFM.
RESULTS: In the first patient and her affected daughter, we identified a heterozygous p.(Arg89Cys) missense mutation in LMNA gene which has not been linked with PAM pathology before. In the second patient, a heterozygous p.(Asn4807Phe) mutation in RYR1 not previously described in PAM represents a novel, candidate gene with a possible causative role in the disease. Finally, in the third patient and his symptomatic daughter, we found a previously reported heterozygous p.(Cys30071Arg) mutation in TTN gene that was clinically associated with cardiac involvement.
CONCLUSIONS: Our study identifies a new genetic background in PAM pathology and expands the clinical phenotype of known pathogenic mutations.

Myofibrillar myopathy (MFM) is a group of inherited muscular disorders characterized by myofibril dissolution and abnormal accumulation of degradation products. The diagnosis of muscular disorders based on clinical presentation is difficult due to phenotypic heterogeneity and overlapping symptoms. In addition, precise diagnosis does not always explain the disease etiopathology or the highly variable clinical course even among patients diagnosed with the same type of myopathy. The advent of high-throughput next-generation sequencing (NGS) has provided a successful and cost-effective strategy for identification of novel causative genes in myopathies, including MFM. So far, pathogenic mutations associated with MFM phenotype, including atypical MFM-like cases, have been identified in 17 genes: DES, CRYAB, MYOT, ZASP, FLNC, BAG3, FHL1, TTN, DNAJB6, PLEC, LMNA, ACTA1, HSPB8, KY, PYROXD1, and SQSTM + TIA1 (digenic). Most of these genes are also associated with other forms of muscle diseases. In addition, in many MFM patients, numerous genomic variants in muscle-related genes have been identified. The various myopathies and muscular dystrophies seem to form a single disease continuum; therefore, gene identification in one disease impacts the genetic etiology of the others. In this review, we describe the heterogeneity of the MFM genetic background focusing on the role of rare variants, the importance of whole genome sequencing in the identification of novel disease-associated mutations, and the emerging concept of variant load as the basis of the phenotypic heterogeneity.

Myofibrillar myopathies (MFMs) are muscular disorders involving proteins that play a role in the structure, maintenance processes and protein quality control mechanisms closely related to the Z-disc in the muscular fibers. MFMs share common histological characteristics including progressive disorganization of the interfibrillar network and protein aggregation. Currently no treatment is available. In this review, we describe first clinical symptoms associated with mutations of the six genes (DES, CRYAB, MYOT, ZASP, FLNC and BAG3) primary involved in MFM and defining the origin of this pathology. As mechanisms determining the aetiology of the disease remain unclear yet, several research teams have developed animal models from invertebrates to mammalians species. Thus we describe here these different models that often recapitulate human clinical symptoms. Therefore they are very useful for deeper studies to understand early molecular and progressive mechanisms determining the pathology. Finally in the last part, we emphasize on the potential therapeutic approaches for MFM that could be conducted in the future. In conclusion, this review offers a link from patients to future therapy through the use of MFMs animal models.

Myofibrillar myopathies (MFMs) are genetically heterogeneous dystrophies characterized by the disintegration of Z-disks and myofibrils and are associated with mutations in genes encoding Z-disk or Z-disk-related proteins. The c.626 C > T (p.P209L) mutation in the BAG3 gene has been described as causative of a subtype of MFM. We report a sporadic case of a 26-year-old Italian woman, affected by MFM with axonal neuropathy, cardiomyopathy, rigid spine, who carries the c.626 C > T mutation in the BAG3 gene. The patient and her non-consanguineous healthy parents and brother were studied with whole exome sequencing (WES) to further investigate the genetic basis of this complex phenotype. In the patient, we found that the BAG3 mutation is associated with variants in the NRAP and FHL1 genes that encode muscle-specific, LIM domain containing proteins. Quantitative real time PCR, immunohistochemistry and Western blot analysis of the patient\'s muscular biopsy showed the absence of NRAP expression and FHL1 accumulation in aggregates in the affected skeletal muscle tissue. Molecular dynamic analysis of the mutated FHL1 domain showed a modification in its surface charge, which could affect its capability to bind its target proteins. To our knowledge this is the first study reporting, in a BAG3 MFM, the simultaneous presence of genetic variants in the BAG3 and FHL1 genes (previously described as independently associated with MFMs) and linking the NRAP gene to MFM for the first time.

Myofibrillar myopathies (MFMs) are a group of inherited or sporadic neuromuscular disorders morphologically characterized by foci of myofibril dissolution, disintegration of the Z-disk, and insoluble protein aggregates within the muscle fibers. The diagnosis is based on muscle biopsy. Light and electron microscopy has a central role in the diagnostic work up, and immunohistochemistry shows abnormal deposition of several proteins including αB-crystallin, desmin, and myotilin. In contrast, immunoblotting does not have any diagnostic value because it does not highlight differences in the amount of involved proteins. We investigated the pattern and level expression of desmin, αB-crystallin, myotilin, and ZASP (Z-band alternatively spliced PDZ motif-containing protein) in muscle of seven patients with MFMs by immunoblotting after SDS-PAGE and 2D-PAGE using two different solubilizing solutions, one radioimmunoprecipitation assay (RIPA) buffer, and the other urea-containing buffer. Our data demonstrated that urea-containing buffer improves the solubilization and recovery of desmin, αB-crystallin, myotilin, and ZASP as compared with RIPA buffer and that the total content of these proteins is increased in muscles of patients. The present results provide evidence that immunoblotting is an additional tool for confirming diagnosis of MFMs.

OBJECTIVE: Myofibrillar myopathies (MFMs) are a group of inherited or sporadic neuromuscular disorders characterized morphologically by foci of myofibril dissolution, disintegration of the Z-disk and insoluble protein aggregates within the muscle fibres. The sequential events leading to muscle fibre damage remains largely unknown.
RESULTS: We investigated the expression and the cellular localization of RNA polymerase II (RNAPII)-associated proteins (RPAPs) in muscle biopsies from patients with genetically proven and sporadic MFMs. Our data demonstrated that RPAP2, and to a lesser extent GPN1/RPAP4, are accumulated focally in the cytoplasm of MFM muscle fibres in which they co-localize with POLR2A/RPB1, the largest subunit of RNAPII, and correspond to αB-cystallin deposits in distribution and staining intensity. No abnormal staining for RPAP2 has been observed in muscle of patients with central cores, minicores and neurogenic target fibres.
CONCLUSIONS: Together, these findings could provide new insights into the molecular pathogenesis of MFMs and suggest that RPAP2 immunostaining can be a useful diagnostic tool to depict protein aggregates in MFMs.