The arrangement of myosin and accessory protein such as C- protein in the thick filaments of fish, avian, and mammalian skeletal muscles is still not clearly elucidated. Recent work in our laboratory on the structure of frog thick filaments however, has demonstrated that the structure of vertebrate muscles may be amenable to analysis by a combination of electron microscopy, optical diffraction analysis, and computer image analysis. Additionally, it has been shown that thick filaments from fich, avian, and mammalian muscle can be isolated with a similarly well ordered array of crossbridges; thus making these filaments suitable for analysis by the same type of techniques which have proven useful for the frog thick filaments.
The specific aim of the proposed research is to extend the techniques used for the frog thick filaments to the determination of the structure of the thick filaments of fish, avian, and mammalian skeletal muscles. The major objective of theses studies will be the determination of the arrangement of the myosin and accessory proteins in the filaments of these vertebrate classes. The studies to accomplish these aims will include: 1) the use of a combination of negative staining, platinum shadowing, rapid-freeze deep-etch cryo studies, and optical diffraction analysis to elucidate the major ultrastructural features of the filaments and to assess the preservation of the filaments as compared to the structural parameters expected from X-ray diffraction patterns of living muscle; 2) the use of computer image analysis of the filament images in the best electron micrographs to determine the rotational symmetry and arrangement of the myosin heads, and to produce a three-dimensional reconstruction of the structure of the filament; 3) the use of immunolabelling studies to determine the probable location of accessory proteins such as C-protein and X-protein on the filaments, and to correlate this information with the structure of the filament as seen in the three dimensional reconstructions. These studies will provide basic scientific information about the structure of muscle, and will thus provide a better foundation for understanding how pathological conditions may affect the functioning of this tissue.

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Type
Research Project (R01)
Project #
5R01AR030442-10
Application #
3155808
Study Section
Biophysical Chemistry Study Section (BBCB)
Project Start
1988-10-01
Project End
1992-12-31
Budget Start
1990-01-01
Budget End
1990-12-31
Support Year
10
Fiscal Year
1990
Total Cost
Indirect Cost
Name
University of Missouri Kansas City
Department
Type
Schools of Medicine
DUNS #
800772162
City
Kansas City
State
MO
Country
United States
Zip Code
64110
Kensler, R W; Woodhead, J L (1995) The chicken muscle thick filament: temperature and the relaxed cross-bridge arrangement. J Muscle Res Cell Motil 16:79-90
Kensler, R W; Peterson, S; Norberg, M (1994) The effects of changes in temperature or ionic strength on isolated rabbit and fish skeletal muscle thick filaments. J Muscle Res Cell Motil 15:69-79
Kensler, R W; Stewart, M (1993) The relaxed crossbridge pattern in isolated rabbit psoas muscle thick filaments. J Cell Sci 105 ( Pt 3):841-8
Kensler, R W; Stewart, M (1989) An ultrastructural study of crossbridge arrangement in the fish skeletal muscle thick filament. J Cell Sci 94 ( Pt 3):391-401
Levine, R J; Chantler, P D; Kensler, R W (1988) Arrangement of myosin heads on Limulus thick filaments. J Cell Biol 107:1739-47