High Resolution Live Cell Fluorescence Imaging of Dysferlin
Skeletal muscle is amongst tissues subject to a high level of mechanical stress. Dysferlin is highly expressed in skeletal muscle, where it is important for maintenance of the plasma membrane integrity. Despite numerous efforts, the mechanism(s) responsible to resolve the acute membrane damage generated in muscle fibers is still unknown.
Membrane repair is a favorable system for analysis, because events at the cell surface are especially accessible to advanced live-cell imaging methods that make it possible to observe at defined locations individual molecular events directly monitored by observing the recruitment dynamics of fluorescently labeled proteins like dysferlin.
With support from the Foundation, we successfully used gene editing to generate, for the first time, muscle cells expressing physiological amounts of fluorescently tagged dysferlin. With this unique cellular reagent in hand, we will use our unique advanced live-cell imaging methods with high temporal and spatial resolution to study the molecular mechanisms underlying dysferlinopathy, particularly in the context of the intracellular traffic of dysferlin and its role(s) for membrane repair.
Complementary to this goal, we will use our imaging methods to adress the general question of whether the muscles in zebrafish embryos can be considered a model system for muscle membrane repair mediated by dysferlin.
Final Project Summary: