Tropomyosin as a Stretch Sensor in the Troponin Bridges of Insect Flight Muscle

Abstract

Listen

Translate article

The flight muscles of many insects contract at high frequency and nearly constant concentrations of calcium. The fibres are activated by rapid stretches, and bridges between thick and thin filaments have been identified as possible stretch sensors. The bridges are spaced at 38.7 nm at the position of troponin on the thin filament, alternating with force-producing crossbridges at target sites in actin. The tropomyosin-troponin complex (Tm-Tn) in Lethocerus flight muscle has two isoforms of tropomyosin (Tm1 and Tm2). Tm-Tn and Tm1 bind to thick filaments or myosin, whereas Tm2 does not. The Tm isoforms exist as homodimers in situ and could act independently: Tm1 forming part of a Tn bridge and Tm2 acting as conventional Tm. Electron micrographs of Tm-Tn incubated with isolated thick filaments showed the complex following the helical arrangement of myosin molecules, with projections every ∼40 nm. Thick filaments were sometimes crosslinked by Tm-Tn or by Tm1 alone. Atomic force microscopy (AFM) of thick filaments in relaxing buffer showed crossbridge crowns every ∼43 nm that were ∼30 nm in height above the mica surface, and intermediate crowns ∼19 nm above the surface. The higher crowns would be more likely to interact with Tm-Tn on thin filaments. Thick filaments crosslinked by Tm1 were also observed by tapping mode AFM. Direct binding of Tm1 to myosin confirms the suggestion that non-force producing crossbridges could activate the thin filament directly when fibres are stretched, pulling Tm from an inhibitory position on actin every cycle of oscillatory contraction, without changes in calcium concentration. We will describe the differences in biophysical properties of Tm1 and Tm2.