Molecular mechanisms of plectin-related muscular dystrophy

Plectin, a multifunctional cytoskeletal linker protein, interlinks intermediate filaments (IFs), essential structural elements of the cytoskeleton, with each other and anchors them to sites of strategic importance for the organization and performance of cells. Mutations in the human plectin gene (PLEC) cause epidermolysis bullosa simplex with muscular dystrophy (EBS-MD), a skin blistering disorder associated with progressive muscle weakness. In addition, PLEC mutations have been shown to lead to EBS-MD with myasthenic syndrome, EBS with pyloric atresia, or limb-girdle muscular dystrophy type 2Q. The only dominant mutation identified in PLEC so far causes EBS-Ogna, a disease characterized by fragile skin without any muscular symptoms. With muscles harboring pathologic protein aggregates, degenerative changes of the myofibrils, and mitochondrial abnormalities, most plectinopathies can be annotated among the expanding group of myofibrillar myopathies (MFM). All MFM show a progressive clinical course, lead to severe physical disability and premature death. The knowledge of the precise molecular mechanisms that translate MFM-causing gene mutations into the myopathic phenotype is still limited, but critical for the understanding of patients needs and the development of treatments. In skeletal muscle, the interplay between plectin and desmin IFs is essential for fiber integrity and cytoarchitecture. Accordingly, the loss of IF network function and the concomitant increased mechanical vulnerability of myofibers are supposed to be underlying mechanisms of MFMs. Altered signaling and protein degradation pathways are likely contributing to these effects. This research project aims at investigating molecular mechanisms that lead to muscle dysfunction in plectin-related MFM, with a focus on IF network alterations and protein degradation. Rescue experiments reconstituting affected signaling cascades will be done in an effort to lay the groundwork for therapies. In addition, this will be the first study investigating a dominant plectinopathy which causes muscular dystrophy without skin involvement, adding a to-date unknown disease entity to the emerging group of plectinopathies. The combined genetic, biochemical, cellular, molecular, and physiological approaches proposed here are bound to yield a more integrated picture of how the skeletal muscle copes with stress under normal conditions and in disease. In order to perform structural and functional analyses of skeletal muscle, tissue samples of several different mutant mouse lines, either lacking plectin or the skeletal muscle IF desmin, will be used in combination with muscle cell cultures and muscle samples derived from patients. The results obtained during this project will help to comprehend the sequential steps that lead to cellular dysfunctions in plectinopathies as well as in MFM, and clarify the molecular pathways that lead from aberrant IF arrangement and resulting cellular stress to weakness and damage.

Plectin, a multifunctional cytoskeletal linker protein, interlinks intermediate filaments (IFs), essential structural elements of the cytoskeleton, with each other and anchors them to sites of strategic importance for the organization and performance of cells. Mutations in the human plectin gene (PLEC) cause epidermolysis bullosa simplex with muscular dystrophy (EBS-MD), a skin blistering disorder associated with progressive muscle weakness. In addition, PLEC mutations have been shown to lead to EBS-MD with myasthenic syndrome (EB-MD-MyS), EBS with pyloric atresia (EB-PA), or limb-girdle muscular dystrophy type 2Q (LGMD2Q), skin-only EBS, as well as the autosomal-dominant variant EBS-Ogna. With muscles harboring pathologic protein aggregates, degenerative changes of the myofibrils, and mitochondrial abnormalities, most plectinopathies can be annotated among the expanding group of myofibrillar myopathies (MFM). All MFM show a progressive clinical course, lead to severe physical disability and premature death. The knowledge of the precise molecular mechanisms that translate MFM-causing gene mutations into the myopathic phenotype is still limited, but critical for the understanding of patients' needs and the development of treatments. In this project, we addressed the molecular pathophysiology of plectin-deficient muscles and performed a preclinical evaluation of a treatment for plectinopathies. Our studies on the skeletal muscle pathology demonstrated that the ablation of plectin caused a multi-level pathology, which primarily affects the extrasarcomeric desmin IF cytoskeleton and subsequently leads to disturbances of the alignment and orientation of myofibrils, and the biomechanical stress resistance of muscle fibers. Notably, one specific plectin isoform, namely plectin d (P1d), is specifically tethering desmin IFs to the Z-disk in myofibrils and functions as scaffolding platform for the chaperone-assisted selective autophagy (CASA) machinery by directly interacting with HSC70 and synpo2. Here, our research identified a specific role of P1d in tension-induced proteolysis activated upon high loads of physical exercise and muscle contraction. Moreover, we identified that autophagic flux was significantly impaired in skeletal muscles from EBS-MD patients, plectin-deficient mice and myoblasts, indicating that the characteristic accumulation of desmin in EBS-MD skeletal muscle results from impaired protein turnover. Treatment of plectin-deficient mice with an autophagy-boosting compound revealed moderate improvement of their muscle function. In extension to our initial workplan, we evaluated downstream pathogenic effects in EBS-MD patient-derived dermal fibroblasts, highlighting the role of plectin in IF and organelle morphology, cell migration and adhesion. Taken together, our project provided substantial new insights into the molecular pathophysiology of plectin-related MFM and, moreover, paved the way towards novel therapeutic approaches for the treatment of plectinopathies.