Skip to main content
PMC home page
Biochemical Journal logoLink to Biochemical Journal
. 1997 Oct 15;327(Pt 2):335–340. doi: 10.1042/bj3270335

Troponin I and troponin T interact with troponin C to produce different Ca2+-dependent effects on actin-tropomyosin filament motility.

W Bing 1, I D Fraser 1, S B Marston 1
PMCID: PMC1218798  PMID: 9359398

Abstract

We have developed an in vitro motility assay to make a detailed quantitative analysis of Ca2+ control of skeletal-muscle troponin-tropomyosin control of actin-filament movement over immobilized myosin. Ca2+ regulates both filament velocity and the fraction of filaments that are motile. We have demonstrated that the two effects are due to separate interactions of troponin C with troponin I and troponin T. When 64nM of the complex actin-tropomyosin-troponin I-troponin C was added at pCa 5, more than 80% of filaments were moving and their velocity did not change. At pCa 9, more than 20% of the filaments were moving. When 20nM of the complex actin-tropomyosin-troponin T+troponin I+troponin C was added at pCa 5, filament motility remained high, whereas velocity increased. The 30% increase in velocity observed when troponin T was present was also observed when heavy meromyosin fragment 1 labelled with N-ethylmaleimide (NEM S-1) was added after actin-tropomyosin filaments. The NEM S-1 effect was not additive with the troponin T-dependent velocity increase. The pattern of motile behaviour is characteristic of myosin on silicone-treated glass and different from the behaviour on nitrocellulose-coated glass.

Full Text

The Full Text of this article is available as a PDF (552.3 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Brenner B. Effect of Ca2+ on cross-bridge turnover kinetics in skinned single rabbit psoas fibers: implications for regulation of muscle contraction. Proc Natl Acad Sci U S A. 1988 May;85(9):3265–3269. doi: 10.1073/pnas.85.9.3265. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Brenner B., Jacob R. Calcium activation and maximum unloaded shortening velocity. Investigation on glycerinated skeletal and heart muscle preparations. Basic Res Cardiol. 1980 Jan-Feb;75(1):40–46. doi: 10.1007/BF02001392. [DOI] [PubMed] [Google Scholar]
  3. Chalovich J. M., Eisenberg E. Inhibition of actomyosin ATPase activity by troponin-tropomyosin without blocking the binding of myosin to actin. J Biol Chem. 1982 Mar 10;257(5):2432–2437. [PMC free article] [PubMed] [Google Scholar]
  4. Chalovich J. M., Yu L. C., Brenner B. Involvement of weak binding crossbridges in force production in muscle. J Muscle Res Cell Motil. 1991 Dec;12(6):503–506. doi: 10.1007/BF01738438. [DOI] [PubMed] [Google Scholar]
  5. Eisenberg E., Kielley W. W. Troponin-tropomyosin complex. Column chromatographic separation and activity of the three, active troponin components with and without tropomyosin present. J Biol Chem. 1974 Aug 10;249(15):4742–4748. [PubMed] [Google Scholar]
  6. Fraser I. D., Marston S. B. In vitro motility analysis of actin-tropomyosin regulation by troponin and calcium. The thin filament is switched as a single cooperative unit. J Biol Chem. 1995 Apr 7;270(14):7836–7841. doi: 10.1074/jbc.270.14.7836. [DOI] [PubMed] [Google Scholar]
  7. Fraser I. D., Marston S. B. In vitro motility analysis of smooth muscle caldesmon control of actin-tropomyosin filament movement. J Biol Chem. 1995 Aug 25;270(34):19688–19693. doi: 10.1074/jbc.270.34.19688. [DOI] [PubMed] [Google Scholar]
  8. Gordon A. M., Ridgway E. B. Cross-bridges affect both TnC structure and calcium affinity in muscle fibers. Adv Exp Med Biol. 1993;332:183–194. doi: 10.1007/978-1-4615-2872-2_17. [DOI] [PubMed] [Google Scholar]
  9. Hill T. L., Eisenberg E., Greene L. Theoretical model for the cooperative equilibrium binding of myosin subfragment 1 to the actin-troponin-tropomyosin complex. Proc Natl Acad Sci U S A. 1980 Jun;77(6):3186–3190. doi: 10.1073/pnas.77.6.3186. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Homsher E., Kim B., Bobkova A., Tobacman L. S. Calcium regulation of thin filament movement in an in vitro motility assay. Biophys J. 1996 Apr;70(4):1881–1892. doi: 10.1016/S0006-3495(96)79753-9. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Kołakowski J., Makuch R., Stepkowski D., Dabrowska R. Interaction of calponin with actin and its functional implications. Biochem J. 1995 Feb 15;306(Pt 1):199–204. doi: 10.1042/bj3060199. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Kron S. J., Spudich J. A. Fluorescent actin filaments move on myosin fixed to a glass surface. Proc Natl Acad Sci U S A. 1986 Sep;83(17):6272–6276. doi: 10.1073/pnas.83.17.6272. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kron S. J., Toyoshima Y. Y., Uyeda T. Q., Spudich J. A. Assays for actin sliding movement over myosin-coated surfaces. Methods Enzymol. 1991;196:399–416. doi: 10.1016/0076-6879(91)96035-p. [DOI] [PubMed] [Google Scholar]
  14. Lehman W., Craig R., Vibert P. Ca(2+)-induced tropomyosin movement in Limulus thin filaments revealed by three-dimensional reconstruction. Nature. 1994 Mar 3;368(6466):65–67. doi: 10.1038/368065a0. [DOI] [PubMed] [Google Scholar]
  15. Lehman W., Vibert P., Uman P., Craig R. Steric-blocking by tropomyosin visualized in relaxed vertebrate muscle thin filaments. J Mol Biol. 1995 Aug 11;251(2):191–196. doi: 10.1006/jmbi.1995.0425. [DOI] [PubMed] [Google Scholar]
  16. Lehrer S. S. The regulatory switch of the muscle thin filament: Ca2+ or myosin heads? J Muscle Res Cell Motil. 1994 Jun;15(3):232–236. doi: 10.1007/BF00123476. [DOI] [PubMed] [Google Scholar]
  17. Lin D., Bobkova A., Homsher E., Tobacman L. S. Altered cardiac troponin T in vitro function in the presence of a mutation implicated in familial hypertrophic cardiomyopathy. J Clin Invest. 1996 Jun 15;97(12):2842–2848. doi: 10.1172/JCI118740. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Margossian S. S., Lowey S. Preparation of myosin and its subfragments from rabbit skeletal muscle. Methods Enzymol. 1982;85(Pt B):55–71. doi: 10.1016/0076-6879(82)85009-x. [DOI] [PubMed] [Google Scholar]
  19. Marston S. B., Fraser I. D., Bing W., Roper G. A simple method for automatic tracking of actin filaments in the motility assay. J Muscle Res Cell Motil. 1996 Aug;17(4):497–506. doi: 10.1007/BF00123365. [DOI] [PubMed] [Google Scholar]
  20. Marston S. B., Fraser I. D., Huber P. A. Smooth muscle caldesmon controls the strong binding interaction between actin-tropomyosin and myosin. J Biol Chem. 1994 Dec 23;269(51):32104–32109. [PubMed] [Google Scholar]
  21. McKillop D. F., Geeves M. A. Regulation of the acto.myosin subfragment 1 interaction by troponin/tropomyosin. Evidence for control of a specific isomerization between two acto.myosin subfragment 1 states. Biochem J. 1991 Nov 1;279(Pt 3):711–718. doi: 10.1042/bj2790711. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. McKillop D. F., Geeves M. A. Regulation of the interaction between actin and myosin subfragment 1: evidence for three states of the thin filament. Biophys J. 1993 Aug;65(2):693–701. doi: 10.1016/S0006-3495(93)81110-X. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Potter J. D. Preparation of troponin and its subunits. Methods Enzymol. 1982;85(Pt B):241–263. doi: 10.1016/0076-6879(82)85024-6. [DOI] [PubMed] [Google Scholar]
  25. Potter J. D., Sheng Z., Pan B. S., Zhao J. A direct regulatory role for troponin T and a dual role for troponin C in the Ca2+ regulation of muscle contraction. J Biol Chem. 1995 Feb 10;270(6):2557–2562. doi: 10.1074/jbc.270.6.2557. [DOI] [PubMed] [Google Scholar]
  26. Shirinsky V. P., Biryukov K. G., Hettasch J. M., Sellers J. R. Inhibition of the relative movement of actin and myosin by caldesmon and calponin. J Biol Chem. 1992 Aug 5;267(22):15886–15892. [PubMed] [Google Scholar]
  27. Swartz D. R., Moss R. L. Influence of a strong-binding myosin analogue on calcium-sensitive mechanical properties of skinned skeletal muscle fibers. J Biol Chem. 1992 Oct 5;267(28):20497–20506. [PubMed] [Google Scholar]
  28. Tobacman L. S. Thin filament-mediated regulation of cardiac contraction. Annu Rev Physiol. 1996;58:447–481. doi: 10.1146/annurev.ph.58.030196.002311. [DOI] [PubMed] [Google Scholar]
  29. Wolff M. R., McDonald K. S., Moss R. L. Rate of tension development in cardiac muscle varies with level of activator calcium. Circ Res. 1995 Jan;76(1):154–160. doi: 10.1161/01.res.76.1.154. [DOI] [PubMed] [Google Scholar]
  30. el-Saleh S. C., Warber K. D., Potter J. D. The role of tropomyosin-troponin in the regulation of skeletal muscle contraction. J Muscle Res Cell Motil. 1986 Oct;7(5):387–404. doi: 10.1007/BF01753582. [DOI] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES