The high wing-beat frequencies of many insect species allows them to fly with remarkable precision, land virtually anywhere (sometimes upside down) and even hover! The key to their flying skills lies in the asynchronous flight muscle, a peculiar type of muscle exclusive to some insects. Unlike ordinary skeletal muscle, which is directly regulated by cyclic calcium concentration changes in the muscle fibres, asynchronous flight muscle is relatively insensitive to calcium and is fully activated by stretching, resulting in asynchrony between electrical and mechanical events. Following an initial calcium stimulus, the asynchronous flight muscle contracts in an oscillatory fashion that produces resonant changes in the shape of the thorax concomitant with the wing movements, at frequencies that can reach 1000 Hz in midges!

Although the molecular mechanisms that control stretch activation of asynchronous flight muscle are not yet known, the role of the troponin complex in skeletal muscle contraction is well understood. The troponin complex is composed of several components: the inhibitory subunit troponin I, troponin H and the regulatory subunit troponin C (TnC). The binding of TnC to two calcium ions induces a conformational change of the troponin, which exposes the myosin-binding site of actin and allows the actin and myosin filaments to form cross-bridges, causing the muscle to contract. Since TnC has a fundamental role in vertebrate skeletal muscle contraction, Qiu and collaborators decided to investigate whether TnC might also be involved in the regulation of the asynchronous flight muscle.

Qiu and his team started by isolating TnC from the asynchronous flight muscle of the giant water bug Lethocerus and found that the water bug's asynchronous flight muscle is unusual, as it expresses two distinct isoforms of TnC: TnC1 and TnC4. Moreover, when they looked at the relative amounts of each isoform in flight muscle, they realised that the levels of TnC4 were fivefold higher than those of TnC1. Aligning the sequence of the Lethocerus TnC isoforms with TnC sequences from the fruitfly and malarial mosquito revealed that both TnC1 and TnC4 isoforms are present in all three insects.

But the team were in for a surprise when they looked for calcium-binding sites in the TnC isoforms. Although TnC1 has two calcium-binding sites like other arthropod TnCs, the water bug's TnC4 is only capable of chelating one calcium ion! Hence, TnC4 is not expected to be regulated by calcium because it lacks the essential N-terminal calcium-binding site.

Knowing that the N-terminal calcium-binding site of vertebrate TnC is required to revert the inhibitory effect of troponin I and permit muscle contraction, Qiu et al. suggest that the initial stretch activation that triggers asynchronous contractions is regulated by Ca2+ binding to the two-sited TnC1. The absence of an N-terminal calcium-binding site renders TnC4 relatively insensitive to calcium. Therefore, the team proposes that the troponin complexes which contain the single-site TnC4 may be involved in the full activation of asynchronous flight muscle by the action of stretch involving other proteins.

Even though TnC is just one piece of the troponin-tropomyosin jigsaw, the presence of a major, asynchronous flight muscle-specific TnC with a single Ca2+ binding site in these three species of insects indicates that this TnC isoform is an essential regulatory element of this high-frequency muscle. Next time a midge bites you and flies away in the blink of an know that TnC4 is to blame!

Qiu, F., Lakey, A., Agianian, B., Hutchings, A., Butcher, G. W.,Labeit, S., Leonard, K. and Bullard, B. (
). Troponin C in different insect muscle types: identification of two isoforms in Lethocerus, Drosophila and Anopheles that are specific to asynchronous flight muscle in the adult insect.
Biochem. J.