Duchenne Muscular dystrophy (DMD) is a group of lethal degenerative muscle diseases that affect roughly 1 in 3,500 males. The disease is caused by mutations in the gene encoding dystrophin, a highly conserved protein that links the muscle cell membrane and the contractile machinery within it. In humans loss of functional dystrophin protein is linked to muscle degeneration, leading to death. While traditional analyses of worms and mice modeling the disease, through loss-of-function mutations in the dystrophin gene, have resulted in great advances, they have also only produced mild muscular and behavioral phenotypes. The ability to model the acute muscle degeneration observed in DMD humans in a model system amenable to genetics would allow the screening for, and characterization of, molecular targets able to mitigate the muscle degeneration characteristic of this disease. Although previous studies suggested that the strength of muscle contraction plays an important role in the progression of the disease, most assays currently used do not control the physical difficulty of the assessed behaviors. We therefore developed a burrowing assay where substrate density can be modulated to increase the force required by animals to locomote. Our assay elicits strong behavioral and cellular phenotypes in nematodes with loss-of-function mutations on the worm dystrophin gene (dys-1) to a degree not previously attained in other systems. After conducting the first forward screen on a worm modeling DMD behaviorally and cellularly, we isolated several suppressor mutants capable of preventing the behavioral decline associated with the dys-1 mutation. The identification of the loci of these suppressor mutations, and the characterization of their mechanisms of action, would present an important advance in the search for novel molecular targets capable of mitigating the progression of this disease. The first specific aim of this project is to fully characterize the kinematics and muscular integrity of wild-type and dys-1 mutants burrowing as a function of substrate density. This will provide us with a library of behavioral and cellular phenotypes across a series of tasks of increasing difficulty.
The second aim i s to quantify the degree of rescue displayed by the different suppressor mutants by comparing their behavior and cellular integrity to those of wild-type and dys-1 animals. Mutants capable of preventing cellular decline will be sequenced to identify the loci of their mutation. In the third aim of this proposal we will conduct RNAi silencing of genes that are likely to be functionally linked to the mitigation of the behavioral and muscular decay in animals modeling DMD. Completion of these aims should identify new molecular targets and pathways that can be used to mitigate the muscular and behavioral degeneration that are hallmarks of Duchenne muscular dystrophy in humans.
Relevance to Public Health: Duchenne Muscular Dystrophy is a degenerative muscle disease affecting up to 1 in 3,500 males and resulting in death. Although no cure currently exists, muscular dystrophy is known to develop as a result of mutations on the gene encoding the protein dystrophin, required for normal muscle function and survival. This project will support the development of new treatments for individuals suffering from Duchenne Muscular Dystrophy by identifying new molecular targets capable of mitigating the degenerative progression of this disease.
