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Functional characterization of the human ?-cardiac actin mutations Y166C and M305L involved in hypertrophic cardiomyopathy

Müller, Mirco and Mazur, Antonina Joanna and Behrmann, Elmar and Diensthuber, Ralph P. and Radke, Michael B. and Qu, Zheng and Littwitz, Christoph and Raunser, Stefan and Schoenenberger, Cora-Ann and Manstein, Dietmar J. and Mannherz, Hans Georg. (2012) Functional characterization of the human ?-cardiac actin mutations Y166C and M305L involved in hypertrophic cardiomyopathy. Cellular and molecular life sciences, Vol. 69, no. 20. pp. 3457-3479.

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Official URL: http://edoc.unibas.ch/dok/A6043627

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Abstract

Inherited cardiomyopathies are caused by point mutations in sarcomeric gene products, including ?-cardiac muscle actin (ACTC1). We examined the biochemical and cell biological properties of the ?-cardiac actin mutations Y166C and M305L identified in hypertrophic cardiomyopathy (HCM). Untagged wild-type (WT) cardiac actin, and the Y166C and M305L mutants were expressed by the baculovirus/Sf9-cell system and affinity purified by immobilized gelsolin G4-6. Their correct folding was verified by a number of assays. The mutant actins also displayed a disturbed intrinsic ATPase activity and an altered polymerization behavior in the presence of tropomyosin, gelsolin, and Arp2/3 complex. Both mutants stimulated the cardiac ?-myosin ATPase to only 50 % of WT cardiac F-actin. Copolymers of WT and increasing amounts of the mutant actins led to a reduced stimulation of the myosin ATPase. Transfection of established cell lines revealed incorporation of EGFP- and hemagglutinin (HA)-tagged WT and both mutant actins into cytoplasmic stress fibers. Adenoviral vectors of HA-tagged WT and Y166C actin were successfully used to infect adult and neonatal rat cardiomyocytes (NRCs). The expressed HA-tagged actins were incorporated into the minus-ends of NRC thin filaments, demonstrating the ability to form hybrid thin filaments with endogenous actin. In NRCs, the Y166C mutant led after 72 h to a shortening of the sarcomere length when compared to NRCs infected with WT actin. Thus our data demonstrate that a mutant actin can be integrated into cardiomyocyte thin filaments and by its reduced mode of myosin interaction might be the basis for the initiation of HCM.
Faculties and Departments:05 Faculty of Science > Departement Biozentrum > Former Organization Units Biozentrum > Structural Biology (Schoenenberger)
UniBasel Contributors:Schoenenberger, Cora-Ann
Item Type:Article, refereed
Article Subtype:Research Article
Bibsysno:Link to catalogue
Publisher:Birkhäuser
ISSN:1420-9071
Note:Publication type according to Uni Basel Research Database: Journal article
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Last Modified:01 Feb 2013 08:46
Deposited On:01 Feb 2013 08:39

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