Drosophila jump muscle myofibrils: A new tool for investigating activation and relaxation

Drosophila models have proven invaluable for studying skeletal and cardiac muscle diseases. While permeabilized indirect flight muscle (IFM) and jump muscle fibers from Drosophila yield insightful mechanical data, these larger fiber preparations cannot resolve the kinetics of activation and relaxation because calcium diffusion into the fiber core is rate-limited at this scale. A single myofibril preparation, which is ideal for measuring physiologically relevant activation and relaxation rates, would therefore be a valuable addition to the Drosophila toolkit. However, previous attempts to prepare IFM myofibrils have failed to produce active force. Here, we developed a method to isolate myofibrils from the Drosophila jump muscle. By applying brief, low-amplitude sonication to permeabilized jump muscles, we isolated myofibrils that produced 19.8 ± 10.5 mN/mm2 net active tension with an activation rate of 8.2 ± 4.0 s-1. Jump muscle myofibrils exhibited the typical biphasic relaxation seen in vertebrates: an initial slow, linear phase lasting 82 ± 9 ms, followed by a fast exponential decay with a rate constant of 19.7 ± 9.6 s-1. We further characterized myofibrils from flies expressing an alternative myosin isoform (EMB) known to have slower actin-binding and detachment kinetics. EMB myofibrils produced ~1.6-fold higher active tension and a 38% slower activation rate compared to controls, while relaxation parameters remained unchanged. These results suggest that activation rate is influenced by myosin attachment kinetics, whereas myosin detachment from actin is not likely the rate-limiting step for relaxation. To our knowledge, this is the first report of active myofibril mechanics from an insect muscle, representing a significant addition to the Drosophila experimental toolkit.