Molecular pathogenesis of myfibrillar myopathies

The most common disease caused by plecfin deficiency, epidermolysis bullosa (EB)-MD, is characterized by muscular dystrophy and severe skin blistering. Our previous studies on skeletal muscle suggest that the 4 major plectin isoforms expressed in muscle are cmcial for the integrity of myofibers by specifically targeting and anchoring desmin IF networks to Zdisks, costameres, mitochondria, and the nuclear/ER membrane system. EB-MD patients and plectin-deficient mice display massive desmin aggregation and severe mitochondrial dysfunction, the hallmark of MFM. The major new objectives of this proposal are to i) analyze the mechanistic link between desmin network collapse, protein aggregation, and mitochondrial dysfuncfion due to plecfin (isoform) deficiencies, ii) define desmin`s molecular interface wilh plectin and its differential targeting lo dislincl myofiber substmctures via disiinct isoforms or plectin on the molecular level, and iii) dissect the molecular mechanism of IF network collapse and mitochondrial dysfimction effected by the plectin Ins 16 mutation in vitro, and generate a corresponding knock-in mouse model, to be myopathologically characterized and compared with other MFMs and related myopathies.

The ubiquitously expressed multifunctional cytolinker protein plectin which we discovered some 30 years ago, is essential for muscle cell integrity and myofiber cytoarchitecture. Patients suffering from plectinopathy-associated epidermolysis bullosa simplex with muscular dystrophy (EBS-MD) and mice lacking plectin in skeletal muscle display pathological protein aggregation and misalignment of the contractile apparatus, which are hallmarks of myofibrillar myopathies (MFMs). One of the major goals of the project was to develop immortalized murine myoblast cell lines to examine the pathogenesis of plectinopathies at the molecular and single-cell level. We succeeded to generate such cell lines and found that plectin-deficient differentiated myotubes were fully functional and mirrored the pathological features of EBS-MD myofibers, including the presence of protein aggregates and a concurrent disarrangement of the myofibrillar apparatus. Using this cell model, we demonstrated that plectin deficiency leads to increased intermediate filament (IF) network and sarcomere dynamics, marked upregulation of heat shock proteins, and reduced myotube resilience following mechanical stretch. Since currently no specific therapy or treatment is available to improve plectin-related or other forms of MFMs, we assessed the therapeutic potential of chemical chaperones to relieve plectinopathies. Treatment of cells with one of them, 4-phenylbutyrate, resulted in remarkable amelioration of the pathological phenotypes in plectin-deficient myotubes as well as in plectin-deficient mice. Our study demonstrated the biological relevance of MFM cell models and suggested that such models have potential use for the development of therapeutic approaches for EBS-MD. (A report of this study was published in the prestigious Journal of Clinical Investigation). Several other groups of the research consortium, of which this project was part of, as well as a number of external groups, are using our newly developed cell systems, or are applying our methodology. Other highlights of the project were our findings that IFs and plectin are involved in cytoskeleton-regulated mechanosensing, a feature that is fundamental for controlled cell mobility and tumor progression (published in FASEB Journal), and a collaborative study in which we could show that plectin has an important function for the pathogenesis of pancreatic ductal adenocarcinoma and potentially for other types of cancer (published in Proc. Natl. Acad. Sci. USA). During the course of the project (2010-2014) we published in total 12 original articles (~2/3rd were directly related to the project), and 8 review articles/book chapters.