Alternative Versions of the Myosin Relay Domain Differentially Respond to Load to Influence Drosophila Muscle Kinetics

We measured the influence of alternative versions of the Drosophila melanogaster myosin heavy chain relay domain on muscle mechanical properties. We exchanged relay domain regions (encoded by alternative versions of exon 9) between an embryonic (EMB) isoform and the indirect flight muscle isoform (IFI) of myosin. Previously, we observed no effect of exchanging the EMB relay domain region into the flight muscle isoform (IFI-9b) on in vitro actin motility velocity or solution ATPase measurements compared to IFI. However, in indirect flight muscle fibers, IFI-9b exhibited decreased maximum power generation (Pmax) and optimal frequency of power generation (fmax) to 70% and 83% of IFI fiber values. The decrease in muscle performance reduced the flight ability and wing-beat frequency of IFI-9b Drosophila compared to IFI Drosophila. Previously, we found that exchanging the flight muscle specific relay domain into the EMB isoform (EMB-9a) prevented actin movement in the in vitro motility assay compared to EMB, which does support actin movement. However, in indirect flight muscle fibers EMB-9a was a highly effective motor, increasing Pmax and fmax 2.5-fold and 1.4-fold, respectively, compared to fibers expressing EMB. We propose that the oscillatory load EMB-9a experiences in the muscle fiber reduces a high activation energy barrier between two strongly bound states of the cross-bridge cycle, thereby promoting cross-bridge cycling. The IFI relay domain's enhanced sensitivity to load increases cross-bridge kinetics, whereas the EMB version is less load-sensitive.