The N-terminal tropomyosin- and actin-binding sites are important for leiomodin 2's function.
Leiomodin is a potent actin nucleator related to tropomodulin, a capping protein localized at the pointed end of the thin filaments. Mutations in leiomodin-3 are associated with lethal nemaline myopathy in humans, and leiomodin-2-knockout mice present with dilated cardiomyopathy. The arrangement of the N-terminal actin- and tropomyosin-binding sites in leiomodin is contradictory and functionally not well understood. Using one-dimensional nuclear magnetic resonance and the pointed-end actin polymerization assay, we find that leiomodin-2, a major cardiac isoform, has an N-terminal actin-binding site located within residues 43-90. Moreover, for the first time, we obtain evidence that there are additional interactions with actin within residues 124-201. Here we establish that leiomodin interacts with only one tropomyosin molecule, and this is the only site of interaction between leiomodin and tropomyosin. Introduction of mutations in both actin- and tropomyosin-binding sites of leiomodin affected its localization at the pointed ends of the thin filaments in cardiomyocytes. On the basis of our new findings, we propose a model in which leiomodin regulates actin poly-merization dynamics in myocytes by acting as a leaky cap at thin filament pointed ends.
Voiland School of Chemical Engineering and Bioengineering, Washington State University, Pullman, WA 99164-6515.
Thu
T
Ly
Voiland School of Chemical Engineering and Bioengineering, Washington State University, Pullman, WA 99164-6515.
Natalia
N
Moroz
Department of Cellular and Molecular Medicine, Sarver Molecular Cardiovascular Research Program, University of Arizona, Tucson, AZ 85721.
Christopher T
CT
Pappas
Department of Cellular and Molecular Medicine, Sarver Molecular Cardiovascular Research Program, University of Arizona, Tucson, AZ 85721.
Stefanie M
SM
Novak
Voiland School of Chemical Engineering and Bioengineering, Washington State University, Pullman, WA 99164-6515.
Dmitri
D
Tolkatchev
Voiland School of Chemical Engineering and Bioengineering, Washington State University, Pullman, WA 99164-6515.
Dayton
D
Wooldridge
Department of Cellular and Molecular Medicine, Sarver Molecular Cardiovascular Research Program, University of Arizona, Tucson, AZ 85721.
Rachel M
RM
Mayfield
Center for NMR Spectroscopy, Washington State University, Pullman, WA 99164-4630.
Gregory
G
Helms
Department of Cellular and Molecular Medicine, Sarver Molecular Cardiovascular Research Program, University of Arizona, Tucson, AZ 85721.
Carol C
CC
Gregorio
Voiland School of Chemical Engineering and Bioengineering, Washington State University, Pullman, WA 99164-6515 alla.kostyukova@wsu.edu.
Alla S
AS
Kostyukova
D008841:Actin Cytoskeleton / Q000378:metabolism; D000199:Actins / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002352:Carrier Proteins / Q000378:metabolism; D002645:Chickens; D003598:Cytoskeletal Proteins / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008840:Microfilament Proteins / Q000378:metabolism; D009124:Muscle Proteins / Q000235:genetics / Q000378:metabolism*; D032383:Myocytes, Cardiac / Q000378:metabolism; D011485:Protein Binding; D000072417:Protein Domains; D012518:Sarcomeres / Q000378:metabolism; D051348:Tropomodulin / Q000235:genetics / Q000378:metabolism; D014335:Tropomyosin / Q000378:metabolism
D000199:Actins; D002352:Carrier Proteins; D003598:Cytoskeletal Proteins; D008840:Microfilament Proteins; D009124:Muscle Proteins; D051348:Tropomodulin; D014335:Tropomyosin; C523689:leiomodin protein, mouse