The Effect of Truncated Troponin Components on Activation of Lethocerus Flight Muscle

Indirect flight muscle (IFM) of Lethocerus is activated by periodic stretches at a constant priming concentration of calcium. The muscle is unusually stiff and stress is transmitted to the thick and thin filaments by kettin, which reinforces links between both filaments and the Z-disc. The activating effect of stress on thin filaments is likely to affect troponin. The isoforms of troponin in IFM differ from those in other muscles. TnT has a C-terminal extension not present in vertebrate TnT; TnH is an isoform of TnI with a C-terminal extension rich in Pro and Ala; TnC is present in two isoforms: F1 binds a single calcium in the C-lobe and is needed for stretch-activation; F2 binds one calcium in both N- and C-lobes and is needed for isometric force. Under conditions of low ionic strength, native fibres have a force-pCa curve that shows high calcium-sensitivity and low cooperativity (pCa50 = 6.2, nH = 1.3). Fibres with F2 alone have a pCa curve similar to that of cardiac muscle, (pCa50 = 5.8, nH = 3.2). A fragment of F1 without the N-lobe (F1-Ct) inhibits stretch-activation; therefore the N-lobe of F1 is necessary, although it does not bind calcium or TnH. F1-Ct is displaced by F2 and isometric force is restored, but not stretch-activation. We hope to show the effect of replacing endogenous troponin in fibres with a complex containing TnT truncated at the C-terminus, TnH with TnI sequence but without the Pro-Ala extension, and either F1 or F2. This will show how important the IFM isoforms of troponin are to the stretch-activation response.