Journal article
Identification of Residues within Tropomodulin-1 Responsible for Its Localization at the Pointed Ends of the Actin Filaments in Cardiac Myocytes
The Journal of biological chemistry, Vol.286(3), pp.2194-2204
01/21/2011
Handle:
https://hdl.handle.net/2376/113996
PMCID: PMC3023515
PMID: 21078668
Abstract
Tropomodulin is a tropomyosin-dependent actin filament capping protein involved in the structural formation of thin filaments and in the regulation of their lengths through its localization at the pointed ends of actin filaments. The disordered N-terminal domain of tropomodulin contains three functional sites: two tropomyosin-binding and one tropomyosin-dependent actin-capping sites. The C-terminal half of tropomodulin consists of one compact domain containing a tropomyosin-independent actin-capping site. Here we determined the structural properties of tropomodulin-1 that affect its roles in cardiomyocytes. To explore the significance of individual tropomyosin-binding sites, GFP-tropomodulin-1 with single mutations that destroy each tropomyosin-binding site was expressed in cardiomyocytes. We demonstrated that both sites are necessary for the optimal localization of tropomodulin-1 at thin filament pointed ends, with site 2 acting as the major determinant. To investigate the functional properties of the tropomodulin C-terminal domain, truncated versions of GFP-tropomodulin-1 were expressed in cardiomyocytes. We discovered that the leucine-rich repeat (LRR) fold and the C-terminal helix are required for its proper targeting to the pointed ends. To investigate the structural significance of the LRR fold, we generated three mutations within the C-terminal domain (V232D, F263D, and L313D). Our results show that these mutations affect both tropomyosin-independent actin-capping activity and pointed end localization, most likely by changing local conformations of either loops or side chains of the surfaces involved in the interactions of the LRR domain. Studying the influence of these mutations individually, we concluded that, in addition to the tropomyosin-independent actin-capping site, there appears to be another regulatory site within the tropomodulin C-terminal domain.
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Details
- Title
- Identification of Residues within Tropomodulin-1 Responsible for Its Localization at the Pointed Ends of the Actin Filaments in Cardiac Myocytes
- Creators
- Takehiro Tsukada - From theLucy Kotlyanskaya - Department of Neuroscience and Cell Biology, Robert Wood Johnson Medical School, Piscataway, New Jersey 08854, andRobert Huynh - Department of Neuroscience and Cell Biology, Robert Wood Johnson Medical School, Piscataway, New Jersey 08854, andBrinda Desai - Department of Neuroscience and Cell Biology, Robert Wood Johnson Medical School, Piscataway, New Jersey 08854, andStefanie M Novak - From theAndrey V Kajava - Centre de Recherches de Biochimie Macromoléculaire, CNRS, Université of Montpellier 1 and 2, 1919 Route de Mende, 34293 Montpellier, FranceCarol C Gregorio - From theAlla S Kostyukova - Department of Neuroscience and Cell Biology, Robert Wood Johnson Medical School, Piscataway, New Jersey 08854, and
- Publication Details
- The Journal of biological chemistry, Vol.286(3), pp.2194-2204
- Academic Unit
- Chemical Engineering and Bioengineering, School of
- Publisher
- American Society for Biochemistry and Molecular Biology; 9650 Rockville Pike, Bethesda, MD 20814, U.S.A
- Grant note
- HL083146; 5R25GM62584; GM081688; 0825870G / National Institutes of Health
- Identifiers
- 99900548081201842
- Language
- English
- Resource Type
- Journal article