The longterm goal of this project is to understand the development of myofibrils in striated muscle. To achieve part of that goal, we have focused on the assembly and alignment of thick myofilaments during sarcomere formation. Attention has been drawn to a group of poorly understood myosin-binding proteins MyBP-C and MyBP-H (formerly termed C- and H-protein), respectively, which are hypothesized to be regulators of A-band assembly. To test this hypothesis a set of structural and functional experiments are proposed to characterize the myosin-binding sites on chicken MyBP-C and MyBP-H, and define the complementary binding site(s) on the myosin rod. Based on studies with the avian proteins, we will later extend the research to comparable human isoforms. The experiments will: a) identify the amino acid residues in the myosin- binding domain (MyBD) which are required for the association of MyBP-C with myosin; b) define the region of light meromyosin (LMM) which contains the binding site for MyBP-C; c) identify the MyBD in MyBP-H; d) correlate mutational assays of in vitro myosin binding with myosin heavy chain cable formation in COS cell transfectants; e) establish the tertiary structure of the MyBD of both MyBP-C and MyBP-H by X-ray crystallography; g) test the physiological properties of recombinant MyBP-C in chemically skinned muscle fiber bundles; h) probe the function of the MyBPs during myofibrillogenesis in vivo using retroviral expression vectors; i) establish other functional domains of MyBP-C which interact with titin and actin. This research will provide the first detailed biochemical information on the interaction of an intracellular immunoglobulin/fibronectin-type adhesion protein with a coiled-coil rod- shaped molecule. The results should have relevance to many other protein- protein interactions both in- and outside of cells, and shed new light on the assembly of myofibrils in developing and regenerating striated muscle. Additionally, a collaboration is proposed with J.G. and C.E. Seidman on the potential role of MyBP-H in a human genetic disease, familial hypertrophic cardiomyopathy (FHC). The gene for human skeletal MyBP-H has been mapped to chromosome 1g32.1. Since mutations in several families with FHC map to this same chromosome band , and no recombination has been observed between this gene and the disease locus, a molecular genetic study is outlined to test whether an altered MyBP-H gene, or one closely linked to it, underlies the pathogenesis of FHC in families that map to the first chromosome.

Agency
National Institute of Health (NIH)
Institute
National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS)
Type
Research Project (R01)
Project #
5R01AR032147-16
Application #
2390490
Study Section
Molecular Cytology Study Section (CTY)
Project Start
1982-07-01
Project End
1999-03-31
Budget Start
1997-04-01
Budget End
1999-03-31
Support Year
16
Fiscal Year
1997
Total Cost
Indirect Cost
Name
Weill Medical College of Cornell University
Department
Anatomy/Cell Biology
Type
Schools of Medicine
DUNS #
201373169
City
New York
State
NY
Country
United States
Zip Code
10065
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