The contractile system of vertebrate skeletal and cardiac muscle is activated by the binding of calcium to troponin C. Considerable information is available on the relationship between free Ca2+ concentration and mechanical response in skinned fiber preparations, as well as on the biochemical properties of the regulatory proteins (troponin C, troponin I, troponin T, tropomyosin) which mediate Ca2+ activation. Less is known about the relationship between bound Ca2+ and mechanical response in the intact myofilament system. Recent investigations have shown that not only is contraction activated by binding of Ca2+ to troponin C but the properties of troponin C are influenced by the mechanical state of the muscle. This is best illustrated by recent studies in this laboratory and elsewhere showing that the steep relationship between contractile force and sarcomere length in cardiac muscle (Frank-Starling relation) is largely determined by a length dependence of Ca2+-troponin C affinity. The goal of this research will be to define the cooperative interactions between contractile and regulatory proteins which determine the Ca2+ binding properties of troponin C in the intact myofilament system and determine where these interactions differ in skeletal and cardiac muscle. A combination of mechanical measurements, isotopic Ca2+ binding, and fluorescence spectroscopy will be used to address the following questions: 1) What are the contributions of cross-bridge interactions and length-dependent Ca2+ binding to the shapes of the ascending limb of the force-length curves in skeletal and cardiac muscle? 2) Are apparent length dependent changes in Ca2+ sensitivity or Ca2+ binding based on changes in length per se, cross-bridge attachments, or interfilament spacing? 3) What are the quantitative differences between skeletal and cardiac troponin C with respect to cooperative effects of cross-bridge interaction and shortening? 4) In interactions between adjacent troponin units play a role in cross-bridge mediated changes in the Ca2+-troponin C interaction, and 5) What structural changes take place in cardiac troponin C in association with Ca2+ binding and cross-bridge interactions?

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Type
Research Project (R01)
Project #
5R01AR010551-29
Application #
2078262
Study Section
Physiology Study Section (PHY)
Project Start
1978-05-01
Project End
1998-08-31
Budget Start
1994-09-01
Budget End
1998-08-31
Support Year
29
Fiscal Year
1994
Total Cost
Indirect Cost
Name
University of Pittsburgh
Department
Physiology
Type
Schools of Medicine
DUNS #
053785812
City
Pittsburgh
State
PA
Country
United States
Zip Code
15213
Wang, Y P; Fuchs, F (2000) Length-dependent effects of osmotic compression on skinned rabbit psoas muscle fibers. J Muscle Res Cell Motil 21:313-9
Fuchs, F; Wang, Y P (1997) Length-dependence of actin-myosin interaction in skinned cardiac muscle fibers in rigor. J Mol Cell Cardiol 29:3267-74
Bukatina, A E; Fuchs, F; Watkins, S C (1996) A study on the mechanism of phalloidin-induced tension changes in skinned rabbit psoas muscle fibres. J Muscle Res Cell Motil 17:365-71
Fuchs, F; Wang, Y P (1996) Sarcomere length versus interfilament spacing as determinants of cardiac myofilament Ca2+ sensitivity and Ca2+ binding. J Mol Cell Cardiol 28:1375-83
Wang, Y P; Fuchs, F (1995) Osmotic compression of skinned cardiac and skeletal muscle bundles: effects on force generation, Ca2+ sensitivity and Ca2+ binding. J Mol Cell Cardiol 27:1235-44
Bukatina, A E; Fuchs, F; Brandt, P W (1995) Thin filament activation by phalloidin in skinned cardiac muscle. J Mol Cell Cardiol 27:1311-5
Wang, Y P; Fuchs, F (1994) Length, force, and Ca(2+)-troponin C affinity in cardiac and slow skeletal muscle. Am J Physiol 266:C1077-82
Bukatina, A E; Fuchs, F (1994) Effect of phalloidin on the ATPase activity of striated muscle myofibrils. J Muscle Res Cell Motil 15:29-36
Liou, Y M; Fuchs, F (1993) Energy-transfer measurements of the Cys35-Cys84 distance in bovine cardiac troponin C. Biochim Biophys Acta 1202:92-8
Liou, Y M; Fuchs, F (1992) Pyrene-labeled cardiac troponin C. Effect of Ca2+ on monomer and excimer fluorescence in solution and in myofibrils. Biophys J 61:892-901

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