Regulation of oscillatory contraction in insect flight muscle by troponin.
Insect indirect flight muscle is activated by sinusoidal length change, which enables the muscle to work at high frequencies, and contracts isometrically in response to Ca(2+). Indirect flight muscle has two TnC isoforms: F1 binding a single Ca(2+) in the C-domain, and F2 binding Ca(2+) in the N- and C-domains. Fibres substituted with F1 produce delayed force in response to a single rapid stretch, and those with F2 produce isometric force in response to Ca(2+). We have studied the effect of TnC isoforms on oscillatory work. In native Lethocerus indicus fibres, oscillatory work was superimposed on a level of isometric force that depended on Ca(2+) concentration. Maximum work was produced at pCa 6.1; at higher concentrations, work decreased as isometric force increased. In fibres substituted with F1 alone, work continued to rise as Ca(2+) was increased up to pCa 4.7. Fibres substituted with various F1:F2 ratios produced maximal work at a ratio of 100:1 or 50:1; a higher proportion of F2 increased isometric force at the expense of oscillatory work. The F1:F2 ratio was 9.8:1 in native fibres, as measured by immunofluorescence, using isoform-specific antibodies. The small amount of F2 needed to restore work to levels obtained for the native fibre is likely to be due to the relative affinity of F1 and F2 for TnH, the Lethocerus homologue of TnI. Affinity of TnC isoforms for a TnI fragment of TnH was measured by isothermal titration calorimetry. The K(d) was 1.01 muM for F1 binding and 22.7 nM for F2. The higher affinity of F2 can be attributed to two TnH binding sites on F2 and a single site on F1. Stretch may be sensed by an extended C-terminal domain of TnH, resulting in reversible dissociation of the inhibitory sequence from actin during the oscillatory cycle.
Krzic
Uros
U
EMBL, Meyerhofstrasse 1, D-69117 Heidelberg, Germany.
Rybin
Vladimir
V
Leonard
Kevin R
KR
Linke
Wolfgang A
WA
Bullard
Belinda
B
D000818:Animals; D002118:Calcium / Q000378:metabolism; D005426:Flight, Animal / Q000502:physiology*; D020062:Heteroptera / Q000502:physiology*; D007537:Isometric Contraction / Q000502:physiology; D009119:Muscle Contraction / Q000502:physiology*; D018485:Muscle Fibers, Skeletal / Q000502:physiology; D009132:Muscles / Q000502:physiology*; D020033:Protein Isoforms / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D019209:Troponin C / Q000378:metabolism*; D019210:Troponin I / Q000378:metabolism
D020033:Protein Isoforms; D019209:Troponin C; D019210:Troponin I; D002118:Calcium