Many degenerative neuromuscular diseases are associated with cytoplasmic accumulation of protein aggregates. These aggregates are widely considered to contribute disease and thus are thought to be toxic to the cell. Recently, we discovered similar aggregates, termed myo-granules, in healthy regenerating and differentiating skeletal muscle. Myo-granules contain RNA and RNA-binding proteins. One prominent RNA- binding protein found in myo-granules, TDP-43, is of particular interest due to its presence in aggregates observed in degenerative disease. Further, myo-granules localize to developing sarcomeres and contain an abundance of sarcomeric mRNAs that bear TDP-43 binding-sites. As the muscle regenerates and sarcomeres are replaced, myo-granules are cleared and TDP-43 re-localizes to the nuclei. Together, these findings suggest that TDP-43 may play a role in facilitating sarcomere formation and that accumulation of TDP-43 containing- aggregates found in disease may not be the cause but instead the result of recurring attempts at skeletal regeneration. In pre-liminary work, I found that TDP-43 deletion in muscle stem cells prevents differentiation, identifying an essential role for TDP-43 in skeletal muscle.
Specific Aim 1 will evaluate the role of TDP-43 in myofiber formation and maturation during regeneration. To accomplish this, I will delete TDP-43 at regenerative checkpoints for fusion of differentiated cells and for maturation of the regenerating muscle. The results from aim 1 will be used to inform experiments in Specific Aim 2 where I will analyze the role of TDP- 43 in sarcomere formation. In pre-liminary work, TDP-43 deletion in differentiating cells during regeneration results in muscle that is improperly repaired and myofiber size is drastically reduced. Additionally, myofiber maturation is reduced when TDP-43 is deleted as indicated by the continued expression of developmental sarcomeric proteins. Thus, is TDP-43 required for sarcomere regeneration and repair? I will ask this question by taking an in vivo approach where TDP-43 will be deleted in both healthy and regenerating muscle to evaluate the effect on sarcomere formation, maturation, and degradation. Delineating the function of TDP-43 in sarcomere formation should lead to a better understanding of the relationship between myo-granules and disease-associated TDP-43 aggregates.
Cytoplasmic aggregates that contain the RNA-binding protein TDP-43 are found in the skeletal muscle of patients with degenerative neuromuscular diseases and are assumed to be pathological. The experiments outlined in this proposal will delineate the function of TDP-43 in skeletal muscle formation. Identifying the function of TDP-43 in normal skeletal muscle function may shape our understanding of TDP-43 pathology in and how we treat patients with degenerative disease.
