The proper function of striated muscle cells depends on the coordinated integration of cytoskeletal systems to the physiological requirements of individual organs. Inherited mutations in desmin, a major cytoskeletal intermediate filament (IF) protein of myocytes, lead to desminopathy, a disorder that affects cardiac, skeletal, and smooth muscles. Significantly, it was recently estimated that 73% of patients with desminopathy manifest severe forms of human cardiomyopathies. Desmin IF links to sarcomeres, basic contractile units of a myofibril, by binding directly to nebulin, a giant actin-thin filament binding protein. The central hypothesis for this proposal is that sarcomere contractile dysfunction in these desminopathies involves critical perturbations in the interaction of desmin with sarcomeres, particularly through nebulin. The structural changes in sarcomeres caused by disease-associated mutant desmin will be directly ascertained in this proposal by a combination of biochemical, molecular and cell biology approaches coupled to high resolution imaging. We specifically aim to: 1) Determine the in vitro nebulin-binding properties of cardiomyopathy-linked desmin mutants; 2) Assess the impact of disease-associated desmin mutations on sarcomere structure-function in live primary cultures of chick cardiac and skeletal myocytes; and 3) Dissect the contribution of nebulin and actin-capping proteins to actin thin filament disarray in myocytes expressing the cardiomyopathy-linked desmin E245D mutation. We anticipate that our results will reveal significant insights into the molecular mechanisms underlying desminopathies and the effect that desmin has on sarcomere function. This understanding will ultimately lead to new strategies to treat cardiomyopathies linked to mutations in desmin.
