Modulation of contractile activation in skeletal muscle by a calcium-insensitive troponin C mutant

Calcium controls the level of muscle activation via interactions with the t roponin complex. Replacement of the native, skeletal calcium-binding subuni t of troponin, troponin C, with mixtures of functional cardiac and mutant c ardiac troponin C insensitive to calcium and permanently inactive provides a novel method to alter the number of myosin cross-bridges capable of bindi ng to the actin filament. Extraction of skeletal troponin C and replacement with functional and mutant cardiac troponin C were used to evaluate the re lationship between the extent of thin filament activation (fractional calci um binding), isometric force, and the rate of force generation in muscle fi bers independent of the calcium concentration. The experiments showed a dir ect, linear relationship between force and the number of cross-bridges atta ching to the thin filament. Further, above 35% maximal isometric activation , following partial replacement with mixtures of cardiac and mutant troponi n C, the rate of force generation was independent of the number of actin si tes available for cross-bridge interaction at saturating calcium concentrat ions. This contrasts with the marked decrease in the rate of force generati on when force was reduced by decreasing the calcium concentration. The resu lts are consistent with hypotheses proposing that calcium controls the tran sition between weakly and strongly bound cross-bridge states.